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The effects of used motor oil on Myzus persicae and Arabidopsis thaliana YICHANG GE, DHVANIL JOSHI, CHRISTOPHER CHAN, SARA MAR, BEATRICE CO ...... 13
Investigating the link between Elite-level gymnastics and short stature VARSHA JAYASANKAR ...... 19
How a medical miracle evolved into a human hazard: The history of antibiotics AILEEN LIU, MEGHAN CRAUGHWELL ...... 28
Ethics of clinical research: A historical review LIZ BALINT, ASHLYN LEUNG ...... 34
Adverse effects associated with monosodium glutamate consumption: A brief review JAMES LAI ...... 41 f Contents
Redefining invasive species using a stage-based process PASCALE BIDER, TANYA DANIEL, ADAM TWEEDLE ...... 45 able o T The effects of water salinity on Myzus persicae and Arabidopsis thaliana MONICA TAKAHASHI, PASCALE BIDER, ARIANA FRASCHETTI, CAMILLA GILLIS- ADELMAN, AUDREY TAM ...... 54
Testing the effects of nitrogen on the interaction of M. persicae and A. thaliana LIZ BALINT, LEENA ALSHENAIBER, NOEL KIM, ISOBEL SHARPE, MEGAN SWING ...... 61
Effects of anthropogenic acetylsalicylic acid contamination on ecological interactions ANGELA DITTRICH, JOSEPHINE AGUECI, TAYLOR LUU, MEGH RATHOD, NADIA AL HASHEMI...... 66 ISCIENTIST | 2018
2 ORIGINAL RESEARCH Star formation and morphology of galaxies in local compact groups
JESSICA SPEEDIE1, LAURA PARKER2 1. Integrated Science Program, Class of 2020, McMaster University 2. Department of Physics and Astronomy, McMaster University
SUMMARY We present a study on the observed properties of galaxies in a large sample of compact groups identified in the Sloan Digital Sky Survey. Compact groups are extremely dense collections of galaxies, with 3 to 8 member galaxies within a very small volume. These systems are known to have frequent galaxy-galaxy interactions. In this work, we propose a new definition for the radius of a compact group and match our member galaxies to previous catalogues containing information about the masses, star formation rates and morphologies of these galaxies. We find that that both star formation and morphology depend strongly on the galaxy’s stellar mass but more weakly on their host environment: the member galaxy properties show very little radial dependence within their group, but they do depend on their group mass, as traced by
the velocity dispersion. Galaxies in compact groups show similar properties to those Astronom in non-compact groups, despite being in fundamentally different environments.
Received: 11/04/2017 Accepted: 11/29/2017 Published: 01/11/2018 URL: https://theiscientist.com/2018/01/11/SpeedieStar.html
Keywords: compact groups, galaxies: star formation, galaxies: morphology y
INTRODUCTION and galaxy-galaxy interactions are slow. Compact groups are therefore environments in which many Large scale structure in the universe is comprised of galaxy evolution processes occur, including mergers aggregations of galaxies. Gravitational interactions and other dynamical interactions (Brasseur et al. 2008; assemble galaxies into groups (containing 10-100 McConnachie et al. 2009). Previous work has shown members) and clusters (100-10,000 members). When in that the observed properties of galaxies, such as star close proximity, galaxies can fall onto one another and formation and morphology, are influenced by the group affect the motion and formation rate of the stars within environment (Mcconnachie et al. 2008). Additionally, them. The environment in which a galaxy lives –group,
the extremity of the compact group environment puts ISCIENTIST | 2018 cluster or field– has observable effects on its properties constraints on galaxy evolution models, which aim (Conselice, 2014). to predict average global trends in galaxy properties, A special class of galaxy group is the so-called compact but that must reproduce the observations in all group (Figure 1). Compact groups are comprised of 3-8 environments if they are to be successful. member galaxies situated very close together; typical The effect of mergers or gravitational interactions on galaxy-galaxy separation is on the order of kiloparsecs star formation rate (SFR) has been explored for close (kpc), similar to the size of the galaxies themselves pairs of galaxies and for regular groups (Carlberg et al. (Sohn et al. 2016). Compared to “regular” groups, 1994; Ellison et al. 2008) as well as for clusters (Moore compact groups have low velocity dispersions ( 200 et al. 1996; Gomez et al. 2003; Lewis et al. 2002). As tidal km/s, Hickson & P. 1982; McConnachie et al. 2009), interactions disturb and redistribute gas from which meaning the deviation in the radial velocities of the stars may form, enhanced SFR would be expected. The constituent member galaxies is small. observations for close pairs are consistent with this High spatial densities and low velocity dispersions picture, but decreased SFRs are found for galaxies in imply that compact groups have short crossing times,
3 Figure 1: Images of four example compact groups as identified by Sohn et al. (2016). Left to right: MLCG1428 ( = 37 8km/s), MLCG1492 ( = 49 12km/s), MLCG1569 ( = 302 57km/s), MLCG1577 ( = 273 69km/s). Images are from the DR12 Science Archive Server (SAS). clusters when local density is high (Brasseur et al. 2008). been released to the public every few years since data Where compact groups will fall on this spectrum is not collection began in 1998. immediately obvious. From the galaxy properties provided by the SDSS (such There are special challenges involved in the study as luminosity, colour, size, stellar mass and velocity of compact groups, stemming from these groups dispersion), further information can be derived. containing very few member galaxies. Group velocity Analysis of emission lines yields galaxy star formation dispersion is difficult to quantify; the fewer galaxy rates, and galaxy morphology can be estimated directly velocities known, the less accurately their dispersion from telescope images. Independent collaborations can be estimated. Additionally, the geometrical centre perform these analyses and publish them in separate of the group is not a well-defined quantity for an catalogues. We use star formation data from the Max aggregation of only a few galaxies. In larger groups and Planck for Astrophysics and Johns Hopkins University clusters, it is clear by their radial position in the group (MPA JHU) catalogue, which estimates star formation which galaxies live in the densest areas. rates based on the technique discussed in Brinchmann The goal of this work is to study the properties of et al. (2004) and Salim et al. (2007). We use the galaxies in this dense environment - specifically, galaxy morphological data provided in the Galaxy Image 2D star formation rate and morphology. We explore (GIM2D) catalogue by Simard et al. (2011). whether galaxy properties exhibit dependence on their radial position in the group or on the group velocity THE MAGNITUDE-LIMITED COMPACT dispersion (environmental metrics), as well on as stellar
Astronomy GROUP CATALOGUE (MLCG) mass (a property internal to the galaxy). Studying the properties of galaxies in compact groups The paper is structured as follows: In Section 2we requires reliable identification of these systems. Several present the properties of the galaxies and compact algorithms exist for identifying groups in galaxy groups within our data sample. In Section 3 we show surveys, but they all involve linking together galaxies our results for the dependence of galaxy properties that are close together on the sky. The original criteria (star formation and morphology) on environment for identifying compact groups were specified by (radial position and velocity dispersion) and on stellar Hickson & P. (1982), who compiled the first collection mass. We discuss our results in Section 4 and present (consisting of around 100 compact groupss) and started our conclusions in Section 5. Throughout, we adopt the quantifying their properties. Hickson’s approach flat CDM cosmological parameters ofM = 0.3, _ = 0.7 and involved three criteria based on the relative brightness, -1 -1 H0 = 70 km s Mpc . compactness and isolation of the galaxies. For our work, we employ the Magnitude-Limited Compact Group DATA (MLCG) catalogue created by Sohn et al. (2016) from In order to address questions related to galaxy evolution the enhanced SDSS Data Release 12 (DR12; Alam et al. in compact groups we need a large population of these 2015). Their sample was derived using the friends-of- groups and the member galaxies within them. We friends (FoF) algorithm of Barton et al. (1996) and two take advantage of the largest ever survey of galaxies, of the three Hickson criteria. They omit the isolation
ISCIENTIST | 2018 the Sloan Digital Sky Survey (SDSS) (Alam et al., criterion, arguing that it artificially selects against 2015). The SDSS is a wide-field multi-spectral imaging nearby groups. survey covering just over 35% of the sky. It contains The MLCG (column 1 of Table 1) consists of 1588 photometric measurements for around 500 million compact groups composed of a total of 5178 member objects, and spectral information (and thus redshifts) galaxies, covering a range in redshift 0.01 z 0.20 for more than 3 million. New SDSS data sets have (distances of 0.1 to 2.5 billion light years). The majority
4 of groups, 1276, have three member galaxies and the emission lines from hot gas around young stars. To remaining 312 have four or more. Galaxies have absolute control for the dependence of SFR on stellar mass -1 r-band magnitudes down to r < 17.77. (Mstar), we use the specific star formation rate, yr . Table 1: The number of galaxies contained in our To obtain star formation and morphological data on the sample and subsamples. We match the MLCG galaxies in our sample of compact groups, we matched of Sohn et al. (2016) to the GIM2D morphology catalogue and the MPA JHU star formation the MLCG to MPA JHU and GIM2D by the galaxies’ catalogue. coordinates on the sky (Right Ascension, Declination). The MLCG Match to GIM2D Matched to MPA Not all galaxies in our compact group sample had JHU matches in the other catalogues. This was partly due 5178 4272 3656 to the fact that MPA JHU and GIM2D originate from the SDSS Data Release (DR) 7 while the MLCG was GALAXY PROPERTIES constructed from DR12, although 1450 of the 1755 lost The morphology of a galaxy ranges from disk- galaxies do appear in DR7. Figure 2 shows the relevant dominated spiral galaxies (such as the Milky Way) to properties of the galaxies successfully matched and of bulge-dominated elliptical galaxies, whose structure is those not. We find that the galaxies retained in our supported by stars undergoing random motion rather sample have lower stellar mass, lower luminosity in the than rotational motion. Morphology can be described r-band and are at slightly lower redshifts. It is likely qualitatively by eye (using the Hubble “tuning that the unmatched galaxies suffer from fiber collisions fork”; Nair & Abraham 2010) or it can be quantified (the inability to resolve objects that are close together numerically using the generalized de Vaucouleurs on the sky due to the finite size of the fiber plugs in profile (also known as the Sèrsic profile). Weare the telescope; Guo et al. 2011) or that they have low interested in the latter. The GIM2D catalogue (Simard signal-to-noise ratios, making it difficult to determine et al. 2011) provides this metric of galaxy morphology star formation and morphology. Whenever possible, we by fitting a surface brightness radial profile using a conduct our analyses on the largest subsample with the Astronom single parameter, the Sèrsic index n, as follows: required data; this means 3656 galaxies for plots with star formation data (column 3 in Table 1), 4272 with (1) morphology (column 2), and 5178 galaxies otherwise (column 1). where re is the effective radius (the projected radius within which half of the galaxy’s light is emitted) and is In Figure 3, we show specific star formation rate plotted e against galaxy stellar mass. Galaxies fall into one of the surface brightness at re. A value of n = 1 corresponds y to a pure disc, and morphology becomes increasingly two populations: star-forming or non star-forming. bulge-dominated with increasing n. In the optical, these populations appear blue and red, A second important observed property of a galaxy is its respectively. The break in the bimodal distribution -11 -1 star formation rate. The star formation rate of a galaxy occurs at log(sSFR) 10 yr . A galaxy’s location on depends on how large a galaxy is and how much gas it this plot is not static. Various processes can “quench” a has available to form stars. The SFR can be estimated galaxy (drive it off the star-forming sequence) or induce in a number of ways; in the SDSS it is done using the star formation activity. On a cosmological scale, the migration occurs rapidly (Wetzel et al. 2011). Processes ISCIENTIST | 2018
Figure 2: A histogram of three galaxy properties (Left panel: stellar mass; centre panel: magnitude in the r-band - note that brightness decreases with higher r; right panel: redshift). Of the 5188 galaxies in the MLCG by Sohn et al. (2016), only 4372 of them are present in GIM2D morphology catalogue and 3656 in MPA JHU star formation catalogue. Shown in red is the sample remaining after matching to both, and in blue are the galaxies lost in the match.
5 of individual galaxy velocities about their mean. It is derived for a group by measuring the line-of-sight velocities of the galaxies via spectroscopic redshifts. Observational incompleteness (due to the difficulty in resolving spectroscopic measurements for close- by galaxies) combines with the statistics to make velocity dispersion a very uncertain quantity for compact groups. Figure 5 shows the uncertainty in as a proportion of the reported value. The mean uncertainty for the whole MLCG catalogue is 22%. Two groups even have an uncertainty fraction greater than 1 (not shown).
GALAXY AND GROUP RADII We are interested in how the properties of galaxies are Figure 3: Specific star formation rate (SFR normalized by stellar mass) as a function of galaxy stellar mass for all 3656 galaxies in our sample with data in MPA JHU. Two galaxy populations are visible: star-forming and non star-forming (quenched). Like the total SDSS population, ours exhibits a bimodal distribution, with a break at sSFR ≈ 10-11 yr-1. are a function of the environment in which the galaxy lives in addition to the galaxy’s intrinsic properties.
GROUP PROPERTIES y As mentioned in the introduction, some properties of compact groups make them challenging to study: particularly, group richness (the number of member galaxies). As aforementioned, 1276 of 1588 compact groups have only 3 members, and in Figure 4 we show a Figure 5: The distribution of observational uncertainty in compact group velocity dispersion (σ) as a fraction histogram of group richness for our smallest subsample of the reported value. Two groups (not shown) have Astronom of the MLCG (3656 galaxies matched to MPA JHU). an uncertainty fraction greater than 1 (185%, 511%). Reliable statistics cannot be run on a sample of so few The mean for our sample of 5188 galaxies is 22%. member galaxies. Low group richness has implications for related to their distance from the centre of the group. another crucial group property: velocity dispersion. However, investigations of radial trends cannot be Similar to standard deviation, this is the statistical done with simply the group radius as the independent dispersion variable. Compact galaxy groups come in different sizes (differing by a factor of 15), and so it is necessary to normalize the galaxy’s radial distance R to a proportion
of the group size, RGr. The calculation of galaxy radial distance R involves three steps (refer to Figure 6) and was done as outlined in Hogg (2000). The distance to the group (D) was calculated using the cosmological parameters assumed (described in the Introduction) and the group redshift (z). By the small angle approximation, the distance between a galaxy and its group centre is simply D times the angular separation . ISCIENTIST | 2018 One commonly practiced way to define group radius is to fit a surface brightness or number density radial Figure 4: Log-histogram of compact group richness profile and identify where the curve falls to some (the number of member galaxies) for the subset of our sample with data in MPAJHU and GIM2D (3656 chosen value; however, fitting radial profiles requires galaxies). The vast majority of groups contain three many galaxies, which compact groups do not have. The
galaxies, the minimum number to qualify as a group. MLCG by Sohn et al. (2016) defines group radius GrR
6 Figure 6: The radial distance of each galaxy (R) from its group centre (black dot) is approximated as the angular separation (θ) times the angular distance (D). The calculation of D involves cosmological parame- Figure 7: A visual comparison between two defini- tions of compact group radius. Points are coordinates ters (ΩM, Ω_ Λ, ΩK) and was done as outlined in Hogg (2000). on the sky (Right Ascension, Declination) represent- ing the projected location of two groups and their as the radius of smallest circle encompassing all member galaxies, as appear in the MLCG by Sohn group members. In other words, the group radius is et al. (2016). The group MLCG0001 (yellow) contains the same as the largest galaxy radial distance. 3 members, the minimum number to be deemed Although the motivation for this definition is a group; the group MLCG0093 (purple) contains 6, understandable, it is not physical. one of the richest groups in the catalogue. In both An alternative way, often used in the panels, the radius of the large circle corresponds to literature, to calculate the radius of a system of RGr. As a result, a galaxy always lies on the edge. The galaxies is to assume that the system is spherical radius of the smaller inset circle corresponds to Rrms, and in dynamical equilibrium. With these the root mean square of all galaxies’ radial distance assumptions, we can relate group mass (M ), radius from the geometric centre; a galaxy does not neces- Gr sarily lie on the edge. (RvirGr) and velocity dispersion using Equation 2. Unfortunately, due to the large uncertainties in In the left panel of Figure 8 we plot our definition
(Figure 5), we are not able to reliably estimate group Astronom of compact group radius, Rrms against that of Sohn et radius in this way. al. (2016), RGr. As is to be expected based on how it is
calculated, Rrms is always smaller than RGr, and all the points lie below the one-to-one line. For small, tight- (2) knit groups, the two definitions do not differ greatly. Instead, we follow the example of Mok et al. (2013) and However, for groups with a galaxy at a large radial
define compact group radius as the root mean square of y distance (corresponding to a large RGr), the root mean the constituent member galaxy radial distances: square Rrms is significantly smaller than GrR . We expect
to see radial trends with Rrms that are similar but not (3) identical to those of RGr. In the right panel of Figure 8 we see what the two different definitions of group radius mean for the where n is the number of member galaxies and Rn is the galaxies’ normalized radial distance (“specific radius”). distance between the nth galaxy and group centre (we In this plot, the problem with the RGr definition is do not redefine the centre). This definition is similar clearly visible. At R/RGr 1 is an artificial pile-up of to a mean, but it takes into account the variation in galaxies that live on the edge of the group. The cut-off the values of R . Because it uses the radial distances of n is not strictly at R/RGr = 1 because we normalized to all member galaxies, it is more physical than the R of ISCIENTIST | 2018 Gr the RGr of the MLCG instead of to our own max(RN); Sohn et al. (2016). For comparison, we show these two the assumed cosmological parameters and details of definitions of group radius, GrR and Rrms, for two compact numerical integration involved in our calculation of groups in Figure 7. The inner circle, representing rmsR , angular diameter distance likely differs slightly to that is necessarily smaller than that representing RGr, and undertaken by Sohn et al. There is nothing physically does not necessarily have a galaxy located on its edge. significant about a specific radius of 1,rms andR Considering that galaxy sizes themselves are roughly rightfully extends past 1 on its axis. on the scale of tens to hundreds of kpc, this figure also demonstrates the compactness of these groups. MASS SEGREGATION Previous work has shown that galaxy properties depend RESULTS most strongly on their own stellar mass, therefore any trend in galaxy properties needs to be independent of CORRELATION OF GROUP RADIUS radial trends in stellar mass in order to be significant. DEFINITIONS We explore trends of stellar mass with specific radius 7 (sSFR) is strongly dependent on stellar mass, and so we have divided our sample into four stellar mass bins. We also show the trend for the total sample (in red). We find a strong dependence of sSFR on stellar mass, with the lowest mass galaxies exhibiting the highest specific star formation rates. However, we see little to no trend with specific radius; the lines are consistent with being flat for all bins. In Figure 11 we differentiate compact groups based Figure 8: A comparison between two definitions of on their velocity dispersion. Since is such an compact group radius. Left panel: R is the radius Gr uncertain quantity (Figure 5), we make cuts for of the smallest circle enclosing all group member only the highest (300km/s) and lowest (100km/s), galaxies, as used in the MLCG by Sohn et al. (2016). In other words, the projected distance to the mem- instead of dividing the sample in half. Furthermore, ber farthest from the centre defines the radius of the we control for mixing of galaxies with different stellar masses by taking only galaxies with 9.5 group. Rrms is the root mean square of the projected distance of all members from their geometrical cen- log(Mstar/Mdot) 10.5 tre. Right panel: The specific radius of each member galaxy (distance from the group centre as a propor- tion of its group’s radius) is plotted for the two group
radius definitions, GrR and Rrms. in Figure 9. Galaxy stellar mass is plotted as a function of normalized radial distance from the group centre for the two group definitions. In both cases the trend is clear: galaxies with higher stellar mass tend to live in the centre of the group, and those with lower stellar mass on the outskirts. The
range on the y-axis, however, is small (log(Mstar/Mdot) 0.25). We have binned the x-axis by frequency, making
apparent the pile-up at R/RGr 1, which does not occur
in the right panel where specific radius is R/Rrms. For the remainder of the paper we fully adopt the R rms Figure 10: Mean galaxy specific star formation rate as Astronomy definition of group radius for use in exploring radial a function of specific radius (normalized radial dis- trends. tance from group centre) for all 3656 galaxies in our sample with data in MPA JHU. In greens and blues RADIAL TRENDS IN STAR FORMATION are four stellar mass bins, and in red is the total un- Figure 10 shows how the rate at which a galaxy forms binned mean. The x-axis is binned by frequency and error bars correspond to 1 SE. stars is related to its radial position in the group. As described in Figure 3, specific star formation rate (Figure 9 estimates the issue of mass segregation). he stellar mass distributions of the two velocity dispersion
samples are different (low, log(Mstar/Mdot) of 9.8
compared to high, log(Mstar/Mdot) of 10.1) and we found that neglecting to account for this yields a misleading radial trend in the low sample. he mean log stellar mass of the resulting galaxies plotted in Figure 11 are different by a factor of only 0.04 dex. We find that there is a segregation of galaxies by group velocity dispersion. he galaxies in groups with low have consistently higher specific star formation Figure 9: Mean galaxy stellar mass as a function of specific radius (normalized radial distance from rates than those with high. We find no significant ISCIENTIST | 2018 group centre) for all 5178 galaxies in our sample. The trend with radial distance. x-axis is binned by frequency and error bars corre- spond to 1 SE. Left panel: Galaxy radial distance nor- RADIAL TRENDS IN MORPHOLOGY malized by R , the group radius definition used in Gr Sèrsic index, our quantitative metric of the MLCG by Sohn et. al (2016). Right panel: Galaxy galaxy morphology, is shown as a function of specific radial distance normalized by Rrms, the root mean square of all galaxy radii of the group. radius in
8 Figure 12. We find again a strong dependence on stellar mass, indicative of the known correlation between galaxy stellar mass and morphology: disks are lighter in stellar mass than elliptical galaxies. The highest two stellar mass bins are consistent with being flat; high mass galaxies are more likely tobe ellipticals and are found with equal probability at all radial positions. However, we do see a dependence of morphology on radial distance for galaxies low in stellar
Figure 12: Mean galaxy Sèrsic index (n) as a function of specific radius (normalized radial distance from group centre) for all 4372 galaxies in our sample with data in GIM2D. In greens and blues are four stellar mass bins, and in red is the total unbinned mean. The x-axis is binned by frequency and error bars corres- pond to 1 SE. Solid lines correspond to a least squares best fit to the means.) star formation. Secondly, with respect to stellar mass, Mean galaxy sSFR as a function of specif- our sample is not complete. Data cannot be collected Figure 11: Astronom ic radius (normalized radial distance from group for the entire population of low mass galaxies; only the centre) for the 2137 galaxies with 9.5 ≤ log(Mstar/Mdot) closest ones are bright enough to be measured. This ≤ 10.5. Galaxies are separated by group velocity dis- introduces a potential bias with redshift, known as the persion: low, σ ≤ 100km/s (479 galaxies); and high, σ ≤ 300km/s (638 galaxies). Solid lines correspond to a Malmquist bias. In the future, we intend to estimate least squares best fit to the means and both slopes this incompleteness and weight our low stellar mass are consistent with being flat. The x-axis is binned by bins appropriately. It is important to note however that frequency and error bars correspond to 1 SE. 10 y the sample galaxies with Mstar > 10 is complete. mass. Statistically significant slopes exist for the lowest and second lowest bins, and for the unbinned samever, the radial dependence of the lowest two stellar mass DEPENDENCE ON ENVIRONMENT bins is robust. The bulge component of low mass disk There are many ways to describe the nature of the galaxies on the group outskirts is less than that of those environment in which a galaxy exists, although we closer to the group centre. focus on two common metrics. Equation 2 shows that group velocity dispersion is proportional to the mass DISCUSSION of the group. A galaxy’s mass takes two forms: ordinary
matter (stars, gas) and dark matter. The mass of the ISCIENTIST | 2018 Our work investigates two properties of galaxies (star combined group dark matter halo in which the visible formation and morphology) as functions of both stellar mass sits, M , can be proxied by the group velocity mass (another galaxy property) and environmental halo dispersion, . Because the value of is so uncertain (see properties (radial position and velocity dispersion) Figure 5), we do not have a well-determined measure in compact groups. In this section, we discuss and of this first environmental property, halo mass. Our interpret our results in the context of previous second environmental measure is normalized galaxy astronomical work. radial position, R/R . Galaxies living near the centre There are two quantities essential to our work whose rms are in much denser environments. The problem with this limitations are important to keep in mind. Due to metric is introduced by the necessary normalization of the nature of the process for calculating sSFR from galaxy radial position by group radius, defined as R , the observed data, values of log(sSFR) -11.5 have high rms which again is affected by low N . uncertainty (Brinchmann et al. 2008); it is easy to Members Controlling for stellar mass, a given galaxy has the same calculate the SFR of a galaxy forming lots of stars but specific star formation rate regardless of where it lives difficult to constrain that of a galaxy with very little
9 in its compact group (Figure 10). This is consistent mass and we look at velocity dispersion, because Mhalo with previous work by Ziparo et al. (2012), which and are related (Equation 2), the agreement suggests finds a marginal dependence of sSFR on normalized that this is another aspect in which compact groups are radial distance in non-compact galaxy groups. The similar to non-compact groups. intergalactic separation of non-compact groups is on Considering the extreme differences between these the order of hundreds of kpc and group richness 10 two types of galaxy systems (namely, galaxy-galaxy
NMembers 100, in contrast to the compact groups’ mean separation), it would not be surprising if they exhibited
separation of tens of kpc and 3 NMembers 8. These two very different galaxy properties. We find instead that types of galaxy systems are very different, yet they they are in fact similar, in the ways we have outlined in exhibit similar radial trends with star formation. this work. The radial trend with morphology of Figure 12 exists only for galaxies of low stellar mass; their bulge component DEPENDENCE ON INTERNAL increases closer to the group centre. In addition to low PROPERTIES mass, these galaxies have rotation-supported structure In addition to environmental dependencies, we also and tend to be gas-rich; they are easily disrupted by explore how star formation and morphology depend gravitational interactions. Our results suggest that on stellar mass; we do this by binning our data by low mass galaxies “feel” their environment more than frequency into quartiles. In both Figure 10 and 12, high mass ellipticals. This is consistent with the work there is clear segregation between the bins. That Sèrsic of Roberts et al. (2015), which finds that the effect of index correlates strongly with stellar mass is a well- an X-ray-rich environment (which is to increase disk known global trend (Conselice 2014). The result that fraction) is strongest for low stellar mass galaxies. sSFR depends strongly on stellar mass is consistent It is common practice in astronomy to understand with previous work. In addition to finding a strong galaxy environment by considering the group’s dependence on Mhalo, Wetzel et al. (2011) also find a gravitational potential well. For an aggregation of a sSFR distribution predominantly more star-forming sufficient number of galaxies, it is possible to trace out for galaxies of low stellar mass. It is not clear in the the potential well with the velocities of the galaxies. literature whether the mechanisms for quenching star It is unknown whether compact groups have similar formation seen in large galaxy clusters (such as ram- dynamics and velocity distributions to regular groups pressure stripping or strangulation) could apply to the and clusters. As shown in Figure 4 and 5, group velocity smaller regime of groups or compact groups (Wetzel et dispersion is very uncertain as compact groups have al. 2011).
Astronomy very few members. Understanding the precise shape of their potential well is challenging. GALAXY PAIR INTERACTIONS In this context, two main ideas can explain the lack of The clear segregation in sSFR between galaxies in radial dependence for star formation rates. The first Figure 11 shows increased star formation in groups hinges on a relative scale concept: with the separation with lower group velocity dispersion. Although not between galaxies in compact groups being comparable exactly equivalent, the group velocity dispersion is to the size of the galaxies themselves, the environmental an effective indicator for galaxy-galaxy flyby speeds. gradient between the group centre and edge may too Within the constituent member galaxies, stars have small to be noticed by whole galaxies. A second idea, internal velocities on the order of 100-200km/s. If the perhaps more plausible (also suggested by Hickson, trajectories of two galaxies in a group bring them past 1997), is simply that the centre of a compact group is each other at a speed comparable to their internal stellar not a well-defined quantity. We can speculate on what velocities, gas dynamics are more affected. Interactions the observational work suggests, but a more complete at these speeds are also generally longer in duration. understanding will come from theoretical work using Shock fronts (high density areas) are created, making simulated galaxy groups. for effective radiative cooling of intragalactic dust and However, we can look at groups with the largest gas. It is this material that collapses to form stars. and smallest potential wells in our sample, which Features indicative of galaxy pair interactions are visible we effectively do with the 300km/s and 100km/s in the images of four compact groups in Figure 1. Tidal ISCIENTIST | 2018 group samples in Figure 11. Our result that galaxies tails, shock fronts and regions of high star formation in groups with low have consistently higher specific are created as the galaxies slosh about in the potential star formation rates is consistent with previous work. well. Compact groups are ideal systems in which to Wetzel et al. (2011) find a strong dependence of galaxy study intergalaxy interactions. sSFR on the mass of the dark matter halo in which the galaxy lives. Although their study looks directly at halo CONCLUSIONS
10 Our main findings are as follows. their SDSS SFRs. This research would not have been • The star formation rates and morphologies of possible without access to these public catalogues. galaxies in compact groups depends strongly Funding for SDSS-III has been provided by the Alfred on their internal stellar mass. Low stellar mass P. Sloan Foundation, the Participating Institutions, the galaxies are more star forming and diskier. National Science Foundation, and the U.S. Department • Galaxies in compact groups with low velocity of Energy Office of Science. The SDSS-III web site is dispersion, and therefore lower halo masses, http://www.sdss3.org/. are more star forming. SDSS-III is managed by the Astrophysical Research • No radial trend exists for star formation rates Consortium for the Participating Institutions of the of galaxies in compact groups. A weak radial SDSS-III Collaboration including the University of trend exists for morphology at low stellar Arizona, the Brazilian Participation Group, Brookhaven mass; the bulge component of low mass disk National Laboratory, Carnegie Mellon University, galaxies living on the outskirts of the group is University of Florida, the French Participation Group, less than that of those in the group centre. the German Participation Group, Harvard University, In future work, we would like to explore galaxy the Instituto de Astrofisica de Canarias, the Michigan X-ray profiles to define compact group centres based State/Notre Dame/JINA Participation Group, Johns on a peak in that wavelength. Additionally, with Hopkins University, Lawrence Berkeley National simulations, we could trace out the potential well Laboratory, Max Planck Institute for Astrophysics, and quantify the effects of uncertain centres and test Max Planck Institute for Extraterrestrial Physics, different definitions of group radius. New Mexico State University, New York University, Ohio State University, Pennsylvania State University, ACKNOWLEDGEMENTS University of Portsmouth, Princeton University, the Spanish Participation Group, University of Tokyo, We thank J. Sohn et al. for making their SDSS DR12
University of Utah, Vanderbilt University, University Astronom compact group catalogue publicly available, L. Simard of Virginia, University of Washington, and Yale et al. for the publication of their SDSS DR7 morphology University. catalogue, and J. Brinchmann et al. for publication of y ISCIENTIST | 2018
11 12 ISCIENTIST | 2018 Astronomy Release oftheSloanDigitalSkySurvey. Data Early the in Environment of Function a as tion Forma Star Galaxy 2003. York,D.G., and M. baRao, Schneider, Lamb,D.Q., Sub D.P.,A.J., Brinkmann,J., Castander, F.J., Connolly, Hopkins,A.M., Sheth, R., M., Bernardi, A.K., Romer, A.I., Zabludoff, T., Goto, M.L., Balogh, C.J., Miller, R.C., Nichol, P.L., Gomez, AstronomicalThe Journal fraction, andtheluminosity/mass-metallicity relation. nucleus galactic active formation, Star I. Survey. Sky Digital Sloan the McCon in pairs and Galaxy A.W.,2008. nachie, L. Simard, D.R., Patton, S.L., Ellison, and Astrophysics Time. Cosmic Over ture Struc Galaxy of Evolution The 2014. C.J., Conselice, 351(4), pp.1151-1179. universe. redshift low the in galaxies forming star of properties physical The 2003. J., Brinkmann, and T. Heckman, G., Kauffmann, C., Tremonti, S.D.M., White, S., Charlot, J., Brinchmann, tron. Soc. nature ofgalaxies incompact groups. predicted and observed the and Comparing -II. theory practice in groups Compact 2008. D.R., Patton, and S.L. A.W., Ellison, Mcconnachie, C.M., Brasseur, p.871. Surveys. Redshift From Selection Group Compact 1996. L.N., Costa, da Barton, Geller, E., Ramella, Marzke, M.J., M., R.O. and dx.doi.org/10.1088/0067-0049/219/1/12. Available at: Mon. Not. R. As R. Not. Mon. The Astrophysi 112, , ------ical Journal Series Supplement Galaxies in the Sloan Digital Sky Survey. 14034 for Classifications Morphological Visual tailed De of Catalog A 2010. R.G., Abraham, P.B.Nair,and clusters ofgalaxies. of evolution the harassmentand Galaxy 1996. ler,A.J., Oem and A. Dressler, G., Lake, N., Katz, B., Moore, Not. R.Astron. Soc the GEEC2 spectroscopic survey of galaxy 1 from groups. ∼ z at quenching satellite Efficient 2013. L.C., Mulchaey, J.S.andParker, Hou,A., Connelly, J.L., Bower,R.G., S., Giodini, Tanaka, M., Finoguenov,A., Wilman, D.J., Mcgee, S.L., Balogh,M.L., Mok, A., tron. Soc spatial properties ofcompact groups. The -I. practice and theory in D.R., groups Compact 2008. Patton, and S.L. Ellison, A.W., Mcconnachie, 395(1),pp.255-268. Soc., of theSloanDigitalSkySurvey. Compact groups ofgalaxies intheSixthDataRelease III. - practice and theory in groups Compact 2009. L., Simard, and S. Ellison, D., Patton, A., McConnachie, Y Myzus on Myzus oil motor used of ffects 11/04/2017 focusing on efficient and effective methods of clean-up. This study serves to runoff contaminants on the environment have been studied extensively, with ords: ated Science Program, Class of 2020, McMaster University 2020, McMaster Class of Program, Science ated cumulation of c yw egr he e SUMMAR There was also a statistically sig-nificant effect (F organism due to its significant effect high (F proliferation rate. The results interaction showed of oil a and day statistically contaminants can on affect multiple tro-phic the levels, which herbivore should of be considered the when plant. the contamination of natural environments. looking at possible issues regarding The results indicate that research off from roadways various and other urbanized impermeable surfaces. The examine effects the effects of of impacts on varying herbi-vores. volumes of UMO on plants and the subsequent due to its rapid life cycle and Received: Used motor oil (UMO) is a highly toxic substance that enters the environment as run- URL: https://theiscientist.com/2018/02/07/GeEffects.html Ke tion, rosett has been shown to result in a reduction in fresh weight, in fresh in a reduction shown to result has been and a decline content, in chlorophyll decrease a drastic (Hanano, Almousally and Shaban, germination in seed cal community is a threat to urban biological systems to urban biological is a threat community cal among are plants sessile, Being 2012). al., et (Tchounwou is It effects. environmental these to vulnerable most the and other primarily on roadways found concentrated and enters the envi surfaces impermeable urbanized ronment as runoff, potentiallycausing harmful effects on vegetation and biological diversity (Environment and Climate Change Canada, 2010). Used motor heavy that contains (UMO) is a highly toxic substance oil and diox- hydrocarbons, metals, polycyclic aromatic ecosystem some of the most common ins, which are contaminants (Australian Government, 2010). When metals can heavy organisms, to biological introduced negatively impact cellular organelles (Tchounwou et of exposure The al.,2012). Bioac INTRODUCTION YICHANG GE, DHV YICHANG GE, Int T Arabidopsis thaliana and Arabidopsis persicae and success at plant transformation (Meinke et al., Table 1: Assigned locations of plants for 1998). It is a small plant of the mustard family and is Block A based on a random stratified mod- native to a broad distribution of regions consisting of Europe, Asia, and North America (Meinke et al., 1998). el created using Microsoft Excel software. In 1987, the modern era of A. thaliana research surged 8 6 12 5 into the frontlines (Meinke et al., 1998). This simple an- giosperm has become a pivotal factor to the scientific 11 7 1 9 advances in understanding plant development (Meinke 20 19 2 13 et al., 1998). 17 10 16 18 STUDY SPECIES: HERBIVORE 14 15 4 3 M. persicae is a polyphagous generalist aphid that feeds on over 40 families of plants (Blackman R.L. & Eas- Table 2: Assigned locations of plants for top V.F., 2000). The species is heteroecious holocyclic, Block B based on a random stratified mod- but anholocyclic in tropical areas (Schoonhoven et al., el created using Microsoft Excel software. 2005). A colony is mostly made up of females for the majority of the year, which results in rapid aphid re- 12 15 20 18 production (Blackman R.L. and Eastop V.F., 2000). The 9 1 8 17 high proliferation rate validates the choice of M. per- sicae for this short 11-day study. Wingless aphids were 14 4 5 11 specifically chosen because winged aphids have a lower 7 3 2 6 individual fecundity (Mutti et al., 2008). M. persicae and A. thaliana have been heavily involved in 13 10 16 19 an evolutionary arms race leading to several different The plant locations were determined using the Micro- plant defenses and subsequent herbivore adaptations soft Excel software to produce a random stratified mod- (Schoonhoven et al., 2005). Since A. thaliana is consid- el, as shown in Table 1 and Table 2 (Microsoft, 2017). ered to be a generic plant and aphids are generalist in- The RAND() and RANK() Excel functions were used sect herbivores, the host-attacker relationship can be ology to create a set of unique random numbers from 1 to 20 used as a model in studying plant-animal interactions with each number corresponding to a location from the Ec (Blackman R.L. & Eastop V.F., 2000). There is a plethora top left corner to the bottom right corner in the 4 by 5 of applications to study from this model because of the grid, set up from left to right in each row. This process large majority of plants and animals categorized under was repeated to create a new set of random numbers generalist species. for Block B. This design minimized the effect of biases, causal claims, and accounted for external factors such USED MOTOR OIL as disproportionately distributed sunlight. The plants TheUMO was obtained from Active Green + Ross Tire were placed so the plastic walls of the pots were touch- Automotive Centre located at 1289 Main St. W., Ham- ing, ensuring rosette leaves were not damaged. ilton. The brand and the type of car the oil was extract- The UMO was administered with 10 mL pipette pumps ed from is unknown. on the soil near the base of the plant. UMO contact with A. thaliana was carefully avoided. Each sampling METHODS unit was then inoculated with two adult female M. per- sicae. The aphids were placed at the lower base of the EXPERIMENTAL DESIGN stem, avoiding the trichomes which could have affect- Each sampling unit comprised of A. thaliana in the ro- ed the aphids. The sampling units were then placed on sette life stage. The plants were placed in 5.5cm x 5.5cm plastic trays with a transparent dome-shaped lid. This x 10cm rectangular plastic pots filled with moist soil. enclosed the plants with approximately 30cm of space 40 sampling units were split into two repeated tests, between the top of the plant and the top of the lid. 2 ISCIENTIST | 2018 Block A and Block B. Both blocks consisted of a four There was a mesh screen of about 100cm that allowed by five arrangement of the sampling units. Four UMO for air exchange but prevented aphids from escaping. treatment levels were used: 0mL, 1mL, 3mL, and 5mL. The individual plants were not removed from the de- Each treatment level consisted of five plants, labelled sign during data collection to ensure minimal distur- 1-5, 6-10, 11-15, and 16-20 respectively. bance to other sampling units. At the halfway point (day 5), 3mL of water was added near the corners of the sampling units with an eye dropper. Aphid population 14 count was recorded on days 1, 4, 5, 6, 7, 8, and 11 to ex- Initial and final mean rosette diameters were compared amine the effect of oil contamination on the herbivore with the different treatment groups in order to inves- of the plant. Rosette diameter was measured on days 0 tigate the effect of UMO on plant growth. It was de- and 11 to examine the effect of UMO on plant perfor- termined that the UMO treatment levels had a statis- mance. tically significant effect on the mean rosette diameter per treatment group (ANOVA, F3, 64=3.385, P=0.023). A STATISTICAL ANALYSIS statistically significant difference was determined be- Statistical analysis was completed using R program- tween the rosette diameters of the control and 5mL ming language, on version number 3.4.2. Analysis of treatment level (Tukey, P= 0.048), as denoted with the variance (ANOVA) was used to investigate the inde- star in Figure 1. It is observed in Figure 1 that there is pendent and correlating effects of several variables a suggested difference which supports these statistical (The R Foundation for Statistical Computing, 2016). findings. These include: UMO treatment levels of 0 mL, 1mL, 3mL and 5mL, test days from Day 0 to Day 11 (with UMO EFFECT ON HERBIVORE inconsistent days in between), blocks A or B. The de- pendent variables that were observed are mean aphid population per sampling unit and mean rosette diam- eter. Mean aphid population per day was quantified by taking the total number of aphids on sampling units within a treatment level per day and dividing it by the number of sampling units. Mean rosette diameter was defined as the sum of all rosette diameters divided by the number of sampling units. A Tukey Honestly Sig- nificant Difference post-hoc analysis was used to deter- mine which UMO treatment levels and test days had Ec a statistically significant effect on mean aphid popula- ology tion and mean rosette diameter (The R Foundation for Statistical Computing, 2016). RESULTS Figure 2: Daily mean aphid population per sam- pling unit for both Block A and Block B. Values are UMO EFFECT ON PLANT GROWTH means ±SD. It is suggested through the illustration that there is a significant effect between treatments of 0-1mL and 3-5mL denoted as treatments C and D respectively. Mean aphid population was plotted over the data col- lection period in order to investigate the effect of oil on the herbivore of the plant. The interaction between UMO treatment levels and day had a statistically sig- nificant effect on the mean aphid population per sam- ISCIENTIST | 2018 pling unit of both Block A and Block B (ANOVA, F21, 256 =2.4661, P=0.00052). The interaction between UMO treatment and day demonstrated that the treatment levels had no significant difference within aphid count between the control and 1mL levels as well as the 3mL and 5mL treatment levels (Tukey, P=0.50; P=0.98). These treatment levels can therefore be grouped, denoted by C and D in Figure 2 and the shown graph supports this statistical significant difference. Figure 1: Initial and final mean rosette diameter measured from the farthest tips of each rosette per treatment group for both Block A and Block B. Val- DIFFERENCE IN TREATMENT BLOCKS ues are means ±SD. The star denotes a difference be- In the analysis of the differences between Block A and tween the control and 5mL treatment groups. Block B, the same statistical conclusions were found 15 Figure 3: Mean Myzus persicae population per sampling unit over 11 days after being treated with various amounts of UMO for Block A (left) and B (right). Values are means ±SD. These population means are calculated through the average aphid count at a varying time on each day, roughly from the times 10am to 4pm. as if the data of the two blocks had been run together UMO EFFECT ON PLANT GROWTH (ANOVA, Block A: F = 2.286, P= 0.0013; Block B: 21, 288 The study investigated whether or not UMO was a sig- F21, 288 = 2.286, P= 0.0013). On each both block A and B, there was no statistically significant difference between nificant stress factor in influencing plant performance. the control and 1mL treatment levels on aphid count There was a statistically significant effect (ANOVA, F =3.3853, P=0.023, Tukey, P= 0.048) of oil on ro- (Tukey, P=0.50). Additionally, there was no statistically 3, 64 significant effect on count when comparing the 3mL sette diameter for plants treated with 5mL of UMO, and 5mL treatment levels (Tukey, P=0.98) (Figure 3). as shown in Figure 1. On day 5, the halfway point, 3mL There was a statistically significant difference between of water was added to each plant. A small quantity of the two sub-groups, previously denoted as C and D in an odourless liquid, assumed to be water, seeped out Figure 2. Qualitative observations, taken of Block A onto the paper towel underneath. This may suggest that and Block B at the beginning and end of the experi- the roots of the plants became coated with oil which ment, shown in Figure 4, supported this data. prevented root uptake of water. A study conducted by Abioye, Agamuthu and Abdul Aziz (2012) found that hydrocarbons could coat root surfaces, restricting gas DISCUSSION and water exchange. Without water, plant growth and ology performance are stunted and the photosynthesis pro- cess is left without a vital resource (McElrone et al., Ec 2013). Photosynthesis plays a large role in the growth of A. thaliana, and with the limited presence of water, the plant must actively reprogram its metabolism and growth (Claeys and Inzé, D., 2013). This means A. thali- ana has shown to be flexible toward water limitations. A supporting example of this claim is shown through a study by Skirycz and Inzé (2010), which showed sharp declines in leaf elongation, which was then followed by recovery to a steady growth rate known as acclimation. This could indicate that if given the op- portunity to continue the experiment over time, rosette diameter could restabilize if allowed proper resources. In our plant environment with both biotic (aphids) and abiotic (oil) stress factors, it would be likely that plants focus their resources on surviving. Overall, this suggests that greater concentrated oil treatments damage plant growth in terms of rosette diameter which could be ex- plained by A. thaliana allocating its resources towards survival rather than growth. ISCIENTIST | 2018 UMO EFFECT ON APHID POPULATION Figure 4: Block A (top left) and Block B (bottom left) The ANOVA and Tukey post-hoc results indicate that on day 0 compared to Block A (top right) and Block B (bottom right) on day 11. the 3mL and 5mL treatment levels of UMO had a statis- tically significant effect on M. persicae population (Fig- ure 2). The final aphid populations on the 3mL and 5mL 16 treated plants were much lower than the final aphid small sample size, resulting in an increased influence of populations in the control and 1mL plants. The 1mL outliers on the data trends. A larger sample size of A. treatment had a greater positive effect on aphid pop- thaliana was not possible due to resource restrictions, ulation than all of the treatment levels including the but is necessary in order to further support any prev- control group. This supports the plant stress hypoth- alent patterns identified. Improvement of the experi- esis, which states that environmental stress on plants mental design could potentially include increasing the decreases plant resistance (Joern and Mole, 2005). It number of plants to a sample size of 400. The increase claims that herbivores favour feeding on plants under by a factor of 10 allows for more specific results and in- biotic and abiotic stressors more than unstressed plants creases the accuracy of any trends observed. The exper- (Joern and Mole, 2005). The increased performance of iment suggested a point between 1mL and 3mL where M. persicae on the 1mL treated plants could then poten- significant negative effects occurred on the aphid count. tially be the result of the decrease in A. thaliana defense Therefore increasing the number of treatment levels to mechanisms. Plant defense could have been hindered 10 and using more specific dosage increments would al- in several ways: damaged morphological features, re- low this threshold to be further pinpointed. Resource duction in toxic chemicals present, weakly expressed limitations also resulted in a sample plant population volatiles, and change in foliar chemistry (War et al., with varying initial rosette diameters, meaning there 2012; Joern and Mole, 2005). Trichome deformation was inherent error in rosette diameter change. An eliminates physical barriers to aphid movement and improved experiment would have a controlled initial decreases secretion of poisonous secondary metabolites rosette diameter for all plants used so the change in (War et al., 2012). Plant volatiles are weakly expressed rosette diameter can be increasingly correlated to the resulting in a reduction of harmful signals (Gatehouse, treatment level. 2002). With change in foliar chemistry, herbivores pre- Additionally, the experiment was carried out over a fer to feed on stressed plants with greater nitrogen con- period of 11 days. Compared to the overall lifespan of centrations since it has a central role in all metabolic A. thaliana of approximately six to eight weeks (Koorn- processes (Joern and Mole, 2005; William J. Mattson, neef and Scheres, 2001), the experiment time frame was Ec 1980). considerably shorter, potentially resulting in less signif- icant observed changes in rosette diameter, leaf num- ology BLOCK EFFECT ber and size, and stem length. An improved experiment Initial ANOVA tests determined that there was a statis- would involve measuring results over a larger time tically significant ffecte on aphid count due to a possi- period - potentially the entire lifespan of A. thaliana. Furthermore, contamination effects were studied onA. ble block effect (Fblock(1,308)= 7.7828, P= 0.0056). This idea may be supported by how qualitatively different Block thaliana only in the rosette stage. In order to achieve A was compared to Block B after the testing period, as more accurate results, contamination effects would seen in Figure 4. This raised concerns about the conclu- need to be observed during several different plant life sions made in response to the second research question stages, accounting for factors such as variation in plant since statistics were run on the combined Block A and biomass, defense, and nutrient uptake. B data. As such, individual ANOVA and Tukey post- The effects of UMO on plants and herbivores should hoc tests of Block A and Block B were taken; however, be further examined in greater detail. Possible areas of these tests resulted in the same conclusions and signif- future research include studying the effects of UMO icant differences between UMO treatment levels that contamination on higher trophic levels in an environ- were previously indicated by the tests using the data ment and the results of biomagnification in affected ISCIENTIST | 2018 from both blocks. It is possible that the block effect ecosystems, studying UMO quantity limits before neg- resulted from unequal averages of beginning rosette di- ative impacts on fauna are distinguishable, and study- ameters between the two blocks. It is suggested that ing how plants in different life stages respond to UMO future experiments address this issue by having the two contamination. random stratified blocks begin with equal average ro- sette diameters. Another possible explanation is that CONCLUSION the block effect resulted from the random clustering of The exposure of A. thaliana to 5mL of UMO had a sta- the higher UMO treatment levels within Block A but tistically significant negative effect on plant rosette not in Block B. diameter compared to the control, indicating that oil contamination negatively affects plant growth. The LIMITATIONS AND NEXT STEPS 3mL and 5mL treatments had a statistically significant The study consisted of four oil treatment groups with negative effect on aphid population while the 1mL only 10 individual plants in each group. This is quite a treatment was statistically equivalent to the control, 17 indicating that oil contamination negatively affects the vided by Russ Ellis’s laboratory in the General Science herbivore of the plant once a critical point of contam- Building at McMaster University. Used motor oil was ination is reached. This study indicates that pollutants received from Active Green+ Ross Tire Automotive can influence multiple trophic levels, which should be Centre at 1289 Main St. W. taken into account when looking at future contamina- tion of natural environments. AUTHOR CONTRIBUTIONS C.C., B.C., Y.G., D.J., and S.M., conceived and carried ACKNOWLEDGMENTS out the experiments. S.M. took the lead in running sta- As a collective group, we would like to thank Dr. Chad tistics, with Y.G. collaborating with C.C. to integrate Harvey for his support and always being available as the necessary statistics for figures. B.C. and D.J. had a a resource for the paper. We would also like to thank primary focus experimental design and analysis of re- Sebastian Irazuzta for his help with the experimental sults. All authors contributed to the final writing and design as well as his supervision on data collection days. editing process of the manuscript. Equipment and specimens used for the study were pro- fects in Commonly Used in Vitro Stress Assays. Plant Model Plant for Genome Analysis. Science, 282(5389), REFERENCES Physiology, 165(2), pp.519–527. pp.662–682. Abdel-Shafy, H.I. and Mansour, M.S.M., 2016. A review on polycyclic aromatic hydrocarbons: Source, Gatehouse, J.A., 2002. Plant resistance towards insect Microsoft, 2017. Microsoft Excel. (15.13.3). [computer environmental impact, effect on human health and re- herbivores: a dynamic interaction. New Phytologist, program] Microsoft Office. Available at: 18 Kinesiology ISCIENTIST | 2018 19 rld ear oc- (De y Ret- were stood atures caused After has ll er than st s Wo Chinese gymnast a old med- A er ames, have alendar four c G aft height. e of respectively in asts th sification years. m mpic m), fal and Mary Lou of c ages 30 y c titions such a e gymn Ol ag ast ames. The small the questions, , p s G adia Comaneci, g Games, (144.78 the end 1984 by r the he 4’9” ess than 145 t whether l d age at Olympic on s of Olympic t of ernational compe were ., 2016) or ts cause them to appear young t h l s astics stars N t brought into u 976 Olympic b oubts he 2008 Olympic ORIGINAL RESEARCH ORIGINAL years d Published: 02/07/2018 6 t was 1 medallist 6 t 1 A (147.32 cm) an ontroversies ove ld c o be sting g 10” ’ ca 4 any he petite heig enito Sanz et a aid to be ton, to compete at in Championships of some gymna 16, curred. gymnasts s B allist at the 1 at T m must cm). Past gymn ccepted: 12/03/2017 ccepted: A om/2018/02/07/JayasankarInvestigating.html gymnastics, vertical growth, maturation, HPG maturation, growth, gymnastics, Axis, vertical GnRH, estradiol, artistic Y gymnastics is one of five disciplines within the within disciplines five gymnasticsof one is vestigating the link between Elite-level Elite-level between link the vestigating omen’s artistic gymnastics is one of the most popular spectator sports at the Olym- egrated Science Program, Class of 2018, McMaster University McMaster 2018, Class of Program, Science egrated ARSHA JAYASANKAR SUMMAR W URL: https://theiscientist.c epiphyseal ossification, fractures, short stature fractures, ossification, epiphyseal Keywords: 11/04/2017 Received: pic Games. Artistic gymnasts compete in two streams–elite, regulated at the - interna tional level with ~40 hours/week of training and interclub, regulated at the national level with ~20 hours/ week of training. Noticeable physical features of gymnasts in- clude short stature and short limbs. Height data of gymnasts from the 2012 Olympic Games, 2016 Olympic Games, and 2016 NCAA Gymnastics Championships were - col lected and analyzed to determine whether elite gymnasts were significantly shorter Games Olympic both at gymnasts Elite counterparts. non-gymnast and interclub than were significantly shorterthan (p<0.0001). interclub Additionally, gymnasts from gymnasts each nation an- review were literature a significantly Furthermore, (p<0.05). females shorterfrom height at than average national the the NCAA Championships alyzed physiological and osteological mechanisms behind the differences in stature. Female gymnasts of show reduced levels 17-β-estradiol, luteinizing hormone (LH), and follicle-stimulating hormone (FSH), critical growth hormones in female development. High prevalence of growth plate injuries in the olecranal, patellar and tarsal regions of gymnasts suggest a potential mechanism for shortened limbs in gymnasts. Height data of gymnasts who competed at both the 2012 and and gymnasts growth, displayed mined that more than 50% returning of competitors 2016 Olympic Games - deter growth. break from gymnastics demonstrated (1+ year) a lengthy who took sport of gymnastics recognized by the Fédération Inter- Fédération the by recognized gymnastics of sport nationale de Gymnastique (FIG). It is one of the most popular spectator sports at the Olympic artistic gymnastics, in four athletes compete women’s Games. In different apparatus–vault, uneven bars, balance beam and floor–performing short (30 strength, of combination a to 90s)require routines These each. routines on move- mandatory the complete to agility and flexibility 2004). Girginov, and Parry ments (Girinov, fact about Olympic gymnastsA noticeable their are The most decorated short statures. gymnast at the 2016 Olympic games, Simone Biles, stands at 4’8” (142.24 Artistic INTRODUCTION V Int In gymnastics stature short and investigation, it was determined that all gymnasts were back loop continues until the follicle has fully matured, of their reported age (Macur, 2008). at which point, 17α-hydroxyprogesterone inhibits the Not all gymnasts are of short stature. Two-time Olym- production of estrogens (Mahesh, 2011). pic champion Svetlana Khorkina was 165 cm tall at the Another hormone that plays an important role in peak of her career. Five gymnasts at the 2016 Olympic childhood growth is Insulin-like growth factor-1 (IGF-1), games were Khorkina’s height or taller (Internation- a hormone with similar structural features as insulin. al Olympic Committee, 2016). This study investigates Newborns diagnosed with IGF-1 deficiency are 2-10cm whether elite female artistic gymnasts are significantly smaller than healthy babies, and present growth-relat-ed shorter than their non-elite and non-gymnast counter- defects such as smaller extremities and heads (Lar-on, parts. Furthermore, an analysis of relevant literature 2001). Additionally, the use of IGF-1 as a growth will determine the ecological, hormonal and physio- promoter resulted in an increase of in mean growth logical causes behind short stature in female artistic velocity from 3–4.7 cm/year to 8.2–9.1 cm/year. IGF-1 gymnasts. binds to its specific receptor (IGF1R), which initiates intracellular signaling and activates the AKT signaling BACKGROUND pathway. This pathway is a stimulator of cell growth and proliferation. Specifically, with regards with hu- FEMALE GROWTH AND MATURATION man growth, IGF-1 is responsible for the large growth spurts during puberty. It allows long bones, such as the Puberty is the process by which hormonal changes sig- femur and, and soft muscle tissues to grow (Soliman nal a child’s body to undergo physical transformations et al., 2014). Production of IGF-1 is controlled by hu- to mature into an adult with sexual reproduction ca- man growth hormone (hGH). hGH, also known as so- pabilities. Females begin puberty at approximately age matotropin, is produced in the anterior pituitary gland 10; the process is completed around age 16 (Kail and and released into the bloodstream, reaching the liver Cavanaugh, 2008). Puberty is regulated by hypothalam- for IGF-1 production (Yakar et al., 2002). Stress and ic–pituitary–gonadal axis and is initiated in the hypo- reduced concentrations of sex hormones inhibit hGH, thalamus through the secretion of gonadotropin-releas- which in turn inhibits IGF-1 and reduces skeletal and ing hormone (GnRH) at increasing levels (Millar et al., muscular growth (Caine et al., 2001) especially from an 2004). A possible mechanism from GnRH increase is auxological perspective. The objective of this review is through the increase of leptin, the satiety hormone. to determine if gymnastics training inhibits growth of In addition to the characteristic physical changes such females. as large growth spurts and development of feminine or Kinesiology Adrenarche is period of the growth that precedes pu- masculine body types, puberty is also defined by be- berty, occurring between ages 7 and 10. During this havioural changes–one such change in increased nutri- stage, the adrenal cortex secretes increased levels of an- tional and food intake. Increased intake, especially with drogens such as dehydroepiandrosterone (DHEA) and regards to fats, results in an increase in leptin release dehydroepiandrosterone sulfate (DHEA-S) (Parker, (Clavien, Theintz, Rizzoli and Bonjour, 1996). Leptin 1993). These hormones are responsible for adolescent receptors that initiate the production of GnRH are changes such as public and axillary hair, body odour located in the hypothalamus (Meister and Hakansson, and mild acne. DHEA has also been proven to be a 2001). Via the hypophyseal portal system, a network of partial agonist of estrogen receptors ERα and ERβ, blood vessels, GnRH travels from the hypothalamus to causing similar growth effects as estradiol (Webb et al., the anterior pituitary and binds to receptors on secre- 2006). tory cells, which in turn release luteinizing hormone (LH) and follicle-stimulating hormone (FSH) (Charl- LEVELS OF GYMNASTICS ton, 2008). Women’s artistic gymnastics in the United States, Can- LH and FSH are two critical hormones in female devel- ada, Great Britain and Australia can be divided into opment. LH supports the theca folliculi, a layer of the two different streams–Elite and Interclub. Gymnasts ovarian follicle, which are responsible for the produc- in the Elite stream intend to compete internationally tion of 17- -estradiol precursors. FSH initiates follic- β (including but not limited to world championships and ular growth especially in oocyte precursors (granulosa ISCIENTIST | 2018 Olympic Games), while Interclub gymnasts limit their cells). Both of these actions are part of a positive feed- competition to the national level. The two streams back loop and result in an increase in estrogens. This differ in numerous ways, including but not limited to in turn allows for greater expression of LH, allowing regulatory body, hours of training, scoring systems and for more 17- -estradiol. The development of secondary β routine composition. sex characteristics, vertical growth and changes in body Elite gymnastics is regulated by the FIG. Therefore, shape are driven by the estradiols. This positive feed- 20 data of all gymnasts com- peting in the team event at the 2012 and 2016 Olym- pic Games were collected from the London 2012 and Rio 2016 websites (Inter- national Olympic Com- mittee, 2012; 2016). The heights were separated by National Olympic Com- mittee (NOC, the Interna- tional Olympic Commit- tee designation for nations or territories). Using the roster data from the 2016 NCAA Gymnastics Cham- Figure 1: Heights of female artistic gymnasts in elite (2012 and 2016 Olympics, n=60 for each) and interclub (2016 NCAA Championships, n=66) level competitions. All pionships, height data for three sets of heights are normally distributed, as per the Shapiro-Wilk test. At the collegiate gymnasts were 2012 Olympic Games, the mean was 153.45 cm, with a standard deviation of 6.09. also collected, separated At the 2016 Olympic Games the mean was 153.75 cm, with a standard deviation of by school (National Colle- 7.82. The heights of gymnasts at the 2016 NCAA Gymnastics Championships had giate Athletic Association, a mean of 158.07 cm with standard deviation of 5.24. 2016). NCAA gymnast there is uniformity in the rules, regulations and re- heights are representative quirements that these gymnasts are held to at an inter- of interclub gymnast heights. Statistical analyses were Kinesiology national level. Gymnasts at this level, are scored using completed using GraphPad InStat 3.1 (Motulsky et al., an open-ended system, regulated by the Code of Points. 2017). Normality for all three data sets were assessed us- There are two components to this scoring system–an ing the Shapiro-Wilk test, with a 5% significance level. open-ended difficulty score and an execution score out The heights of female artistic gymnasts at the 2012 of 10. Because of this scoring system, elite gymnastics Olympic Games (n=60) was normally distributed with gives preference to difficult skills (Fédération Interna- mean 153.45 cm, and standard deviation of 6.09. The tionale de Gymnastique, 2016a). A recommended train- heights of female artistic gymnasts at the 2016 Olym- ing plan for elite gymnasts consists of two workouts pic Games (n=60) was normally distributed with mean per day (2-3 hours in the morning, 3-4 hours in the af- 153.75 cm and standard deviation of 7.82. Combining ternoon), 6 days week. This results in 30-42 hours/week both Olympic Games (n=120), the mean was 153.6 cm of training (Georgopoulos, 2002). with a standard deviation of 6.95. The heights of gym- Interclub gymnastics typically consists of multi-level nasts at the 2016 NCAA Gymnastics Championships program, regulated by national governing bodies. Scor- (n=66) was normally distributed with a mean of 158.07 ing in this stream is usually out of 10, with the maxi- cm and standard deviation of 5.24. An ANOVA was mum score assigned to a perfect routine. Because of the completed to determine significant difference between difference in scoring in this stream, quality of gymnas- the three groups of gymnasts (Figure 1). tics is emphasized over difficulty. Gymnasts competing The heights of gymnasts from each NOC at both Olym- ISCIENTIST | 2018 at the maximum level of Interclub streams train 18-25 pic Games (n=5 for each NOC, 12 NOCs competed at hours per week, on average (USA Gymnastics, 2017). each Olympic Games) were compared with the national In the United States, gymnasts at this level train with average using one sample t-tests. All NOC means were the goal of athletic scholarships and the possibility to significantly smaller than the national average (Figure compete in post-secondary competitions as a part of 2). The heights of gymnasts at both Olympic Games the National Collegiate Athletic Association (NCAA). were significantly smaller than 2016 NCAA gymnasts At this level, training is regulated to a maximum of 20 (p<0.0001). NCAA gymnasts are significantly smaller hours/week (National Collegiate Athletic Association, than the U.S. national average height (p<0.0001) There 2015). was no significant difference between the two Olympic Games (p>0.05). HEIGHTS OF GYMNASTS To determine if elite female artistic gymnasts are short- SELECTION BIAS er than interclub and non-gymnast counterparts, height It is believed that elite gymnasts are smaller than their 21 Figure 2: Comparison of average Olympic gymnast and national female heights for all nations that participated Kinesiology in the team competition at the 2012 and/or 2016 Olympic Games. Australia, Belgium, Netherlands, and Romania competed at one Olympic Games (n=5), while the other nations were represented at both (n=10). All nations had significant differences between national and Olympic average heights. The asterisk above or below the national flags indicate the level of significance: * (p<0.05), **(p<0.01), ***(p<0.001). interclub counterparts due to a directional selection nasts. A sample of Polish elite gymnasts (n=5) were, on (Malina et al., 2013). In 2009, there were 4932 female average, shorter than their counterparts who dropped gymnasts in the United States competing at the maxi- out (n=4), although the difference was not significant. mum interclub level (level 9 and 10). Only 79 (1.9%) of These height difference continued into late adolescence these gymnasts were classified as elite (USA Gymnas- and full maturation, but were still not significant (Ma- tics, 2009). lina et al., 2013). In a study of Swiss elite gymnasts, Based on the data above, it was determined that there is dropouts (n=12) were significantly taller than their a significant difference between the heights of NCAA elite counterparts (n=12) (Tönz, Stronski and Gmeiner, and Olympic gymnasts, with the latter being smaller. 1990). A similar result was observed in a Belgian and Only 19% of NCAA gymnasts fall below the average Canadian study (Claessens and Lefevre, 1998; Lindner, height of Olympic gymnasts. Of the gymnasts that Caine and Johns, 1991). did fall below the Olympic average height, 50% were classified as and/or competed elite for a minimum of ENDOCRINE AND NUTRITIONAL FAC- one season in their interclub career (USA Gymnastics, TORS ISCIENTIST | 2018 2016). In contrast 77% of Olympic gymnasts fall below In addition to a selection pressure that results in overall the average height of NCAA gymnasts. This data sug- short stature in gymnastics, endocrine and nutritional gests that elite gymnastics, in a sense, selects for shorter factors may also play a role in these physiological dif- gymnasts. ferences. Female athlete triad is disorder characterized Studies in other countries have also determined a po- by the presence of three conditions–reduced caloric in- tential selection towards shorter heights in elite gym- take (negative energy threshold), amenorrhea (absence 22 Kinesiology Figure 3: Endocrine pathways altered in female artistic gymnasts compared to non-athlete counterparts that affect stature and maturation. On the left is the hGH-IGF-1 system which overall reduces skeletal and muscular growth, resulting in reduced stature. The hypothalamic–pituitary–gonadal (HPG) axis is presented on the right; this results in reduced sexual hormones and maturation. The two pathways are interconnected as the reduction in sex hormones in the HPG-axis activates the hGH-IGF-1 system. of menstruation), and low bone mineral density. The ative energy (Loucks, 2006). These trends however, have disorder is most prevalent in sports that emphasize not yet been observed on maturing female athletes. small stature, low body weight, or leanness. When an In a sample of 83 pre-pubescent athletes, 46 gym- athlete is diagnosed with one of the three characteristic nasts and 37 swimmers, there were no differences in conditions, it is highly likely that she is suffering from 17-β-estradiol, DHEAS, LH, and FSH levels. A simi- the other two (Weimann, 2002). The prevalence of this lar result was observed in early pubertal gymnasts. disorder is approximately 31% in female artistic gym- Comparing a sample of early pubertal girls (n=12) nasts–much greater than the general population (<5%) with ectomorphic body types with gymnasts of the (Buck, Bretz and Towns, 2008). These results are based same age (n=12), the latter group had lower LH and on a self-reported survey of gymnasts–actual numbers 17-β-estradiol, but higher FSH levels (Peltenburg et al., could be much higher. Negative energy thresholds can 1984). Female artistic gymnasts typically have very low ISCIENTIST | 2018 delay growth and the onset of puberty, while amenor- body fat levels, which in turn can reduced leptin levels. rhea alters gonadal hormone secretion, altering the lev- A decrease in leptin release reduces the production of els of growth hormones including FSH and LH (Amer- GnRH in the hypothalamus (Meister and Hakansson, ican College of Sports Medicine, 2007). 2001), which in turn reduces the levels of FSH and LH Intensive gymnastics training, correlating to that of in the body. Overall, a decrease in FSH and LH is asso- the elite level, in combination with a negative energy ciated with a lack of growth (Mahesh, 2011). threshold has been determined to be inhibitory on the IGF-1 levels are also affected in female artistic gymnasts. hypothalamic-pituitary-gonadal axis in female artis- In a sample of elite level gymnasts (n=16), measured tic gymnasts (Theintz, 1994). In menstruating adults, serum IGF-1 levels were lower than normal ranges for a negative energy threshold has been associated with females of that age. These gymnasts participated in a decrease and disruption of LH release. Additionally, 3-day period of intensive exercise consisting of ~5 hours an experimental exercise program on post-menarcheal of gymnastics training and ~3 hours of general athlet- females increased LH disruption during periods of neg- ic training. During this training period, the gymnasts 23 showed a decline in IGF-1 levels compared to the basal ges), the feet (metatarsals and phalanges), and the col- values. These measured levels were also lower than the lar bones (clavicles). These bones compromising half of controls (non-gymnasts of the same age) used in this human height–the other half comes from the vertebrae study. Reduced levels of critical hormones in the hy- and the skull. pothalamic-pituitary-gonadal axis suggest that puber- Osteogenesis, or bone growth, occurs through the ad- ty and adolescent development may be delayed female dition of cells and tissue at the epiphyseal (growth) artistic gymnasts (Jahreis et al., 1991). Reduced IGF-1 plates, a layer of hyaline cartilage at the ends or epiph- levels result in reduced skeletal and muscular growth, ysis of long bones. The epiphyseal plates have two which in turn could result in short stature (Caine et al., sides–an epiphyseal side which is closer to the head or 2001) especially from an auxological perspective. The a diaphyseal side which is closer to the main body of objective of this review is to determine if gymnastics diaphysis of the bone. Each plate contains five zones training inhibits growth of females. (Figure 3). relating to growth. The zone of resting cartilage, which is nearest to the epiphyseal side of the plate, contains BONE DEVELOPMENT chondrocytes that do not rapidly divide, but bind to the epiphysis. The zone of proliferation is new cartilage is formed through rapid mitosis of the chondrocytes. These chondrocytes mature and enlarge in the zone of hypertrophy. The zone of calcification, which is near- est to the diaphyseal side contains ossified bones with blood vessels. As bones reach adult size, the epiphyseal plate is ossified and closes, ceasing growth. This occurs anywhere between ages 12 and 25 (Gilbert and Singer, 2003) (Figure 4). EFFECTS ON VERTICAL GROWTH Injuries, especially fractures, to the epiphyseal plate can have a negative effect on bone growth. Epiphyseal frac- tures result in a joint cavity between the epiphysis and diaphysis, preventing osteogenesis from occurring. This can result in shorter than normal limbs. Fractures to Kinesiology epiphyseal plates are referred to as Salter-Harris frac- tures (Salter and Harris, 2001). Sever’s Disease is a con- dition caused by inflammation of the epiphyseal plate of the calcaneus, typically as a result of activity that in- volves walking, running or jumping (Scharfbillig, Jones and Scutter, 2008). Growth plate injuries are common in elite gymnasts. In a study of 349 Australian gymnasts (151 elite, 198 non- elite), over 50% reported injuries to long bone joints. While only 12.3% reported injuries to growth plates, it should be noted that nearly 25% of elite gymnasts re- Figure 4: The five zones of osteogenesis at an ported this type of injury. Only 2.6% of non-elite gym- epiphyseal plate. The zone of resting cartilage is nasts reported a growth plate injury. Growth plate located closest to the head of the bone, while the zone of ossification is located closest to the diaphysis, injuries were the most commonly reported type of in- or midsection of the bone. jury in elite gymnasts in this study (Kolt and Kirkby, 1999). A similar study was completed in the United Endocrine pathways such as the hGH-IGF-1 system de- States, where radiographic images of non-elite gym- scribed above are not the only physiological cause of nasts’ wrists were taken to assess the impact of gym- ISCIENTIST | 2018 short stature–skeletal impacts may also carry an im- nastics training on the distal radial epiphyseal plate. Of portant role. Long bones are one of five types of bones, the 44 participants, 11 showed signs of stress injuries in characterized by the fact that they are more long than the growth plates (DiFiori et al., 1997). wide. In the human body, the long bones are located in Female artistic gymnasts have been identified for the legs (femora, tibiae, and fibulae), the arms (humeri, having short limbs, relatively short legs for height or radii, and ulnae), the hands (metacarpals and phalan- stunted growth of the legs (Buckler and Brodie, 1977). 24 Table 1: Height differences of gymnasts who competed at both the 2012 and 2016 Olympic Games Gymnast Height (2012, cm) Height (2016, cm) Change in Height Age (in 2012) Black (CAN) 155 155 0 17 Bui (GER) 155 155 0 19 Douglas (USA) 150 157 7 17 Fasana (ITA) 152 155 3 16 Ferlito (ITA) 157 160 3 17 Ferrari (ITA) 146 146 0 21 Hypólito (BRA) 147 147 0 28 Mustafina (RUS) 161 162 1 18 Paseka (RUS) 159 161 2 17 Raisman (USA) 157 157 0 18 Seitz (GER) 162 162 0 19 Rogers (CAN) 157 165 8 19 Teramoto (JPN) 136 141 5 17 Average 153.38 155.62 2.23 18.54 Growth plate injuries cause a temporary detachment determined that a lower than normal growth rate could of epiphysis from the diaphysis which prevents bone be associated with gymnastics training. Among with growth and can result in shorter limbs (Gilbert and sample, 4 gymnasts retired from the sport at approx- Singer, 2003). These types of injuries are prevalent in imately age 11. Height velocities in these former gym- elite and non-elite gymnasts. Therefore, it is possible nasts accelerated substantially, which was interpreted Kinesiology that the short limbs and subsequent shorter heights by the authors as catch-up growth caused by the cessa- of female artistic gymnasts can be a caused by growth tion of training (Bass et al., 2000). plate stress and/or injury. Of the 13 gymnasts that competed at both Olym- pic Games, three have publically stated that they had DELAYED GROWTH ceased elite gymnastics training for more than 1 year– Gabrielle Douglas (USA), Alexandra Raisman (USA) Elite gymnastics training may impact female height and Brittany Rogers (CAN). Both Douglas and Rais- through hormonal and physiological changes. How- man took a break from elite competition between the ever, numerous studies indicate that elite gymnastics 2012 Olympic Games and the 2015 competition season, training delays the onset of puberty, and still allow completely ceasing gymnastics training between Au- normal pubertal growth. In a longitudinal study of fe- gust 2012 and January 2015. Rogers did not fully cease male gymnasts, concentrations of 17- -estradiol and β gymnastics training–she trained and competed at the LH were lower than the reference sample for 9-13 year NCAA level with lesser hours than her elite regimen. olds, but showed later increases which were consistent Raisman showed no change in height between the two with delayed maturation (Tönz, Stronski and Gmeiner, Olympic Games. Douglas and Rogers however, showed 1990). an increase of 7 and 8 cm, respectively, the highest of Of the 120 gymnasts that competed in the team event at ISCIENTIST | 2018 the 13 gymnasts. the 2012 and 2016 Olympic Games, 13 competed at both Another case of delayed growth in elite gymnasts can (Table 1). All but one of these gymnasts were past the be seen in Kyla Ross (USA). Ross, then 15 years old, normal age of puberty (16 years), and all but one were competed at the 2012 Olympic Games, measuring 157 below the maximum age of epiphyseal plate ossifica- cm. Ross continued to compete at the elite level un- tion (25 years). Seven of these 13 gymnasts increased in til 2016, when she switched to training at the interclub height between the two Olympic Games, with the av- level in preparation for NCAA competition. During erage increase in height of 4.14 cm. All but 2 gymnasts the time between her retirement from elite gymnas- under the age of 19 showed growth between the two tics in January 2016 to her enrollment as a freshman at Olympic Games. University of California Los Angeles (UCLA) in Sep- Research has also determined that cessation of gymnas- tember 2016, Ross experienced a large growth spurt. tics training for prolonged period of time can trigger According to UCLA’s roster, Ross now stand 5’7” (170.2 the onset of puberty and/or growth spurts. In a longi- cm), a 13-cm growth between the two Olympic Games tudinal study of 21 Australian female gymnasts, it was (National Collegiate Athletic Association, 2016). Over- 25 all, the high prevalence for delayed growth spurts in gymnastics. As per FIG regulations the diagonal dis- elite gymnasts, and larger growth spurts during periods tance between the uneven bars must be between 130 of suspended training, suggest that training at the elite and 180 cm. For a taller gymnast, a 180 cm gap may level may postpone normal pubertal developments in interfere with their ability to complete giant swings, female gymnasts. 360º rotations around the high bar; a smaller gap can exaggerate this disadvantage further. Should their feet ADVANTAGES OF SHORT STATURE hit the low bar during rotation, it can cause injury and the gymnast will receive a mandatory deduction during Overall elite gymnasts are indeed shorter than their competition. On the floor exercise, the diagonal dis- interclub and non-athlete counterparts–this difference tance is 1697 cm; a shorter gymnast can complete more in stature provides them with a biomechanical advan- somersaults and handsprings within this distance, tage. A short stature provides gymnasts with increased while a taller gymnast has a higher chance of landing rotation power. In a longitudinal study of 37 gymnasts out-of-bounds and incurring a deduction. Similarly, over 3.3 years, it was determined that gymnasts with a the length of the balance beam (500 cm) provides the lower growth rate have a higher strength to mass ratio, smaller gymnast the advantage, as she will be able to increasing their potential to perform full-body rota- complete more saltos (Fédération Internationale de tions. Gymnasts with higher growth rates have better Gymnastique, 2016b). performance on back rotation skills, but are lacking in side and front rotational skills, as well as vertical jumps. Skills in gymnastics emphasize rotation around the all CONCLUSION body axes–frontal, longitudinal and sagittal. Elements Across the world, elite female artistic gymnasts are sig- of increasing difficulty involved rotation around mul- nificantly shorter than their non-elite and non-gymnast tiple axes at the same time (Ackland, Elliott and Rich- counterparts. Gymnasts at the 2012 and 2016 Olympic ards, 2003). Games were on average, shorter than gymnasts that A gymnasts’ balance is also impacted by stature. A per- competed at the 2016 NCAA Gymnastics Champion- son’s centre of gravity is approximately below the na- ships. Between interclub and elite gymnasts, a direc- vel. A shorter gymnast, therefore has a centre of gravity tional selection towards smaller heights may be pres- that is closer to the ground. In a 2007 study of collegiate ent, as a larger percentage of elite gymnasts are shorter gymnasts and basketball players, gymnasts had superi- than their interclub counterparts. Endocrine effects or static and dynamic balance abilities than basketball may explain the differences in height of elite gymnasts, players. For reference, the average height of a collegiate as they showed reduced concentrations of hormones Kinesiology basketball player is 6’6” (198 cm), while the average col- in the hypothalamic-pituitary-gonadal axis that legiate gymnast stands at 5’2” (158 cm). While centre of control puberty and growth. Additionally, gravity has some effect on this result, the focus of the epiphyseal plate fractures, injuries and stress may individual athlete’s training can also have an impact. stunt limb growth, affecting a gymnast’s vertical For example, basketball players rarely balance motion- height. While elite gym-nasts at the Olympic Games less on one leg, while this is quite common for gym- are short, some show delayed growth compared to nasts (Bressel et al., 2007). normal puberty standards, or growth during A short stature also helps gymnasts navigate and ma- cessation of training regimens. nipulate the four pieces apparatus in women’s artistic ISCIENTIST | 2018 26 Kinesiology ISCIENTIST | 2018 27 , . ------ - Indian , 18(7), p.53. , 18(7), p.53. Journal of Applied of Applied Journal Clinical Endocrinology Clinical The Endocrinologist, , 110(6), pp.771–781. , 83(11), p.1753. , 38(1-2), pp.89–116. The Journal of Bone and Joint Journal of Bone and The Schweizerische Medizinische Wochen Medizinische Schweizerische , 92(5), pp.2146–2152. , 52(4), pp.385–392. , 120(1), pp.10–20. els of IGF-1 directly regulate bone growth and density. density. and growth bone regulate directly IGF-1 of els Investigation of Clinical Journal 2014. Advances in pubertal growth and factors influ in pubertal growth 2014. Advances pubertal growth? encing it: Can we increase of Endocrinology and Metabolism Journal Theintz, G.E., 1994. Endocrine adaptation to intensive during growth. training physical 41(3), pp.267–272. C., O., S. and Gmeiner, Stronski, 1990. Growth Tönz, gymnasts: female and puberty in 7-to-16-year-old a study. prospective schrift your stats? 1st you know Do Gymnastics, 2009. USA Indianapolis ed. National Program Women’s Gymnastics, 2016. USA Indianapolis. Avail Members (1982 - Current). Team able at: https://theiscientist.com/2018/02/01/LiuMedical.html A brief timeline focusing on the significant events and contributions of the antibiotic era, which era, antibiotic the of contributions and events significant the on focusing timeline brief A 11/04/2017 niitc, eitne bcei, ulc elh arclue Gle Age, Golden agriculture, health, public bacteria, resistance, antibiotics, Accepted: 01/31/2018 - used totreat a variety ofbacterial infections, andcan are Antibiotics 2006). Walsh, and Fischbach (Clardy, isms thatinhibitthegrowth ofothermicroorganisms compounds tobemolecules created bymicroorgan microbiolo gist, coined thetermantibioticsanddescribed these Ukrainian a Waksman, Selman 1941, In has become aglobalthreat tohumanhealth. been apopulartopicinthemedia foroveradecade and sistance may notbenewstotheenvironment, thishas Published: 02/01/2018 ------ISCIENTIST | 2018 Biochemistry 29 ------– Alexander Fleming , in 1939. This them enabled and was intrigued by the idea of idea by the intrigued and was cillin, use enough.” Penicillium chrysogenum Penicillium “There is danger that the ignorant man may “There make them resistant...Moral: if you use peni if you them resistant...Moral: make easily underdose himself by exposing his mi exposing easily underdose himself by crobes to non-lethal quantities of the drug to the drug to non-lethal quantities of crobes to this, society disregarded his warning and the topic of his warning this, society disregarded decades until investigated not was resistance antibiotic later. Chain, a German chemist, were fascinated by Fleming’s Fleming’s by fascinated were chemist, German a Chain, work (Page, 2012). They expanded upon fun a related penicillin from isolated and successfully research his gus, how to demonstrate in vivo testing in mice to conduct penicillin could treat bacterial infections (Page, 2012). Subsequently, they were granted the approval to con duct clinical trials in humans. After multiple failures leading to improvements of the flaws in their experi drug able to show that this miracle ments, they were strep severe had who children treating in effective was tococcal and staphylococcal infections receive (Page, To further funding and patents, 2012). they travelled from England to the United States in 1941 to produce the drug in large quantities, making it available as medication a in the early 1940s (Muniz, 2017). In 1945, in the Nobel Prize and Chain received Fleming, Florey Physiology or Medicine. In Fleming’s acceptance speech, he warned the public of; antibiotic resis about something new and unheard wise antibiotics use to public the cautioned He tance. develop to able were bacteria that understood he as ly Despite 1945). (Fleming, compounds these to resistance halo, halo, indicating an area of no growth (Fleming, 1929). This surprised Fleming as appeared it though as had by the fungus which dissolved been bacteria had the penetrated the broth. He identified the fungus to be Pencillium notatum conduct to him motivated which life, life inhibiting further experiments over the next 10 years (Nicholas and Davies, 2012). Initially, he did not think of using humans, however in infections penicillin to treat reflecting on theresearch he conducted antimicrobial that the proposed lysozyme, Fleming to identify (Bottcher, infections bacterial against be used could wounds heal to ability drug’s the proved he Later, 1959). in rabbits and its effectiveness in issues ethical eliminatingHowever, 2012). (Page, pathogens positive Gram presented lack of biochemistry training and Fleming’s when attempting to isolate the him with challenges in Fleming eventually lost interest active substance. (Page, to test it on humans reluctant penicillin and was findings his on papers published he Nevertheless, 2012). be to strains fungal his allowed and hypotheses and to the public. made available Howard Florey, an Australian pathologist, and Ernst - - Staphylococci (Fleming, 1929). One night, Fleming’s assistant a finelawn grey-greenof mould transparent a and allowing airborne fungal spores to contaminate the to contaminate spores allowing airborne fungal broth. The following day when Fleming observed the was that there dishes, he noticed bacteria in the Petri aureus nutrient dishes containing Petri away putting was the lids properly, to cover however she neglected agar, substance was an enzyme, which he named lysozyme. an enzyme, which he named was substance Hospital while working at St. Mary’s later, Six years in England, he studied the nature of his observations, Fleming performed further studies Fleming performed his observations, in white blood cells to identify the active substance (Bottcher, 1959). In 1922, he discovered that the War War I. During this time, he its through lacerations in healing white blood cells recognized the role of ability to kill bacteria (Bottcher, 1959). Astonished by Dr. Alexander Fleming, a Scottish bacteriologist, Alexander Fleming, a Scottish Dr. during World wound infections on treating worked used appropriately. used by 2050 (Nothias, Knight and Dorrestein, 2016). It is and hospitals organizations, crucial that governments, being that antibiotics are to ensure issue this address resistant resistant bacteria. If used, currently in which they are antibiotics at the rate consumers continue than cancer deaths more will cause bacterial infections to use the misuse of antibacterial products. Scientists are in Scientists are the misuse of antibacterial products. and development the discovery between race constant of new of novel antibiotics, versus the emergence and a treatment no required longer exists, which poses a serious public health threat. It has been determined through to resistance that society has contributed that were previously successful in with a mutant that if an individual is infected means treating it. This multi-resistant be bacteria may the bacterial strain, of new antimicrobial drugs. of new antimicrobial of bacteria when a strain occurs Antibiotic resistance of antibiotics by a class be eliminated no longer can compounds with antibiotic properties from uncultured uncultured from properties with antibiotic compounds bacterial sources yet to be discovered. Thus, for the development area a promising remain sources natural investigate naturally occurring antibiotic compounds compounds antibiotic occurring naturally investigate diffi this Despite 2006). Walsh, and Fischbach (Clardy, of number vast a still is there that estimated is it culty, antibiotics from natural sources (Peláez, 2006). This is partially due to the fact that less than 1% of bacteria can be cultured in the lab, which makes it difficult to and Walsh, 2006). Although most antibiotics current there microbes, from isolated been have ly prescribed of new the discovery in decrease overall an has been products with antibiotic properties or through the through or antibiotic properties with products Fischbach (Clardy, compounds synthetic of creation be created through either the modification of natural BEHOLD THE MOULD ANTIBACTERIAL ACTION tibiotics introduced during this time period. To com- bat the mutant bacterial strains, other forms of antibi- The development of penicillin antibiotics has allowed otics were needed to eliminate the resistant bacteria. physicians to treat life-threatening infections, where However, the microorganisms continued to develop re- patients show noticeable progress within hours after sistance to the new drugs, thus evolving into multi-re- taking the drug. In the 1950s and 1960s, many scien- sistant bacteria, or superbugs. Fortunately, many com- tists were dedicated to understanding the biochemi- panies were on board with antibiotic discovery which cal mode of action of penicillin (Nicholas and Davies, advanced science into a triumphal period in antibiotic 2012). It was determined that beta-lactam antibiotics, history. such as penicillin, inhibit the synthesis of the bacterial cell wall, specifically peptidoglycan. Penicillin binding proteins (PBPs) are a group of proteins found in many GOLDEN AGE types of bacteria that are essential for the cross-linking Most of the antimicrobial classes that are still adminis- of peptidoglycan, which is necessary for the creation tered today, such as tetracyclines and vancomycin, were of a strong cell wall (Saga and Yamaguchi, 2009). The discovered between 1940 and 1960 (Davies and Davies, chemical structure of beta-lactam antibiotics resembles 2010). This time period was known as the Golden Age the C-terminus of peptidoglycan peptide chains; there- of Antibiotics. There are various factors that contrib- fore, penicillin can bind to PBPs to form an enzyme uted to the success as well as the eventual downfall of complex (Page, 2012). Ultimately, this complex inhibits the Golden Age. The discovery of penicillin was the PBPs from performing cross-linking and the cell wall is forefront of medical breakthroughs during this time, not synthesized, which leads to the destruction of the satisfying the high clinical need for these therapeutics bacterial cell. (Mills and Dougherty, 2012). There was a major need Penicillinase is the enzyme responsible for inhibiting for penicillin during World War II as death caused by penicillin, thus contributing to the development of re- bacterial infections were more common than casualties sistant bacteria (Ontario Medical Association, 2017). in battle. As a result, many drug companies, such as Resistant bacterial strains that produce penicillinase Pfizer, invested in antibiotic drug discovery. Addition- decrease the ability of penicillin to kill bacteria. These ally, during this period, there were great decreases in resistant bacterial strains were first discovered in the the death rates caused by infections. Once antibiotics 1940s, but became more prevalent in the 1950s, signi- were made available as treatments, the human life ex- fying the spread of resistance (Saga and Yamaguchi, pectancy increased by 8 years between 1944 and 1972, 2009). In response to this, another penicillin class an- as these common infections were no longer a leading Biochemistry tibiotic, penicillinase-stable methicillin, was developed cause of death (Mills and Dougherty, 2012). in 1959. However, just three years after its introduction, Unfortunately, the 1960s marked the end of the Gold- methicillin-resistant S. aureus (MRSA) was discovered. en Age, where many pharmaceutical companies aban- The antibiotic streptomycin, another novel drug, was doned the search for new antibacterial classes. Some clinically introduced by Waksman in 1944 to treat tu- scientists thought that they had completely cured all berculosis (Davies and Davies, 2010). Similarly, resistant infectious diseases, so there was no need for more an- strains of Mycobacterium tuberculosis emerged rapidly af- tibiotics. Others believed that antibiotic resistance was ter its use to treat the disease (Davies and Davies, 2010). not a concern, and in fact, they considered drug resis- Subsequently, the increased worldwide use of antibi- tance to be rare and the likelihood of bacteria develop- otics after the 1940s acted as a selective pressure that ing resistance to be very low. Furthermore, in the 1980s, led to the development of resistant bacteria (Read and companies that continued antibiotic research shifted Woods, 2014). This became regarded as an issue with their objectives in drug discovery (Page, 2012). Alter- the effectiveness of antibiotics and efforts were made natively, they focused on improving the biochemical to understand the spread of resistance. One mechanism activities and properties of existing antibiotics rather was through spontaneous mutation and natural selec- than developing new classes. Meanwhile, many other tion (Read and Woods, 2014). In 1928, British bacteri- drug businesses abandoned research in antimicrobials ologist Frederick Griffith was conducting experiments and began moving towards other areas of other ther- using Streptococcus pneumoniae when he unknowing- apeutic needs, where they had the potential to make ISCIENTIST | 2018 ly discovered the process of horizontal gene transfer large profits. (HGT) (Arber, 2014). This was confirmed in the 1950s, Overall, the lack of new antibacterial compounds was when it was discovered that bacteria can transfer ge- a factor that contributed to the threat of drug resis- netic material between individuals, including the genes tant bacteria. Additionally, investing in an antibacte- coding for antibiotic resistance (Arber, 2014). rial product was unappealing as it was very expensive All in all, this trend of resistance was seen in all the an- and challenging to get approval by the FDA, and the 30 profits for medications to treat chronic diseases were Report to advise the government and ease anxieties, much greater. Apart from the pharmaceutical industry, which was then distributed to other countries in Eu- antibiotic resistance has also affected other industries. rope and to the United States. Through their research, they concluded that the administration of antibiotics AGRICULTURE in agriculture posed a threat to human health due to the development of antibiotic resistance (Swann et al., Agriculture has been a necessary part of human surviv- 1969). They identified 1523 strains of bacteria that were al and population growth for centuries, providing hu- resistant to certain antibiotics and concluded antibi- mans with food, clothing, heat and employment (Fed- otic use in agriculture was partially to blame (Swann erico, 2005). As the global population has grown, there et al., 1969). This committee recommended that anti- has been stress on the agricultural industry to produce biotics should be separated to be used either in feed or greater amounts of food, and various technological ad- therapeutically and that drugs that are currently free- vancements have allowed for more efficient production ly available should be limited so that they are only be of food products (Federico, 2005). One of the advances available with a prescription. in this sector was the discovery of antibiotics. In the Despite the recommendations of the Swann Report and late 1940s, Thomas Jukes and Robert Stokstad, two further research on this emerging issue, the regulations American scientists, were working at the University of and policies regarding antibiotic use in agriculture California, attempting to find a source of vitamin B-12 remained fairly unchanged. It wasn’t until the 1990s (Wise, 2007). When studying chickens fed with a mix when this issue became the subject of large debates, containing the fermentation products of Streptomyces that more government action was taken (Wise, 2007). aureofaciens, these scientists made an important dis- In 2001, the UK created a committee called the Special- covery for the future of agriculture. They noticed that ist Advisory Committee on Antimicrobial Resistance the poultry experienced a dramatic weight gain, which that had representatives from human and veterinary Jukes attributed this weight gain to the change of intes- Biochemistry medicine, public health, nursing, bacteriology, and vi- tinal flora of the chickens (Stokstad and Jukes, 1950). rology (Wise, 2007). Similarly, a committee was created Although its mechanism of action was not fully under- in Canada called the Canadian Integrated Program for stood, the causative agent was determined to be a prod- Antimicrobial Resistance Surveillance (Government of uct of S. aureofaciens, called chlorotetracycline, which is Canada, 2007). The purpose of these committees is to an antibiotic of the class of tetracyclines (Wise, 2007). monitor the development of resistant bacteria, includ- From this discovery, the benefits of antibiotics on live- ing the contribution of resistance from antibiotic use stock were realized and the worldwide agricultural in- in livestock (Government of Canada, 2007). dustry began to change its practices. There are contrasting perspectives and lifestyles when The main purposes of adding antibiotics into animal it comes to antibiotic use between Western and Eu- feed include the promotion of growth and the pre- ropean countries. Currently in Ontario, there are few vention of infection in livestock populations, as this regulations and little control regarding the types of could have devastating effects on businesses (Khacha- antibiotics and dosage quantities that can be given to tourians, 1998). In the 1950s, the recommended dose of livestock (Ontario Medical Association, 2017). On the antibiotics in animal feed was 10-20 ppm, however in other hand, Sweden banned the administration of an- the 1990s, this dosage had increased to 200-400 ppm tibiotics to livestock as growth promoters in 1986 and (Khachatourians, 1998). With these increasing trends, Denmark followed their lead 9 years later (Ontario the primary use of antibiotics became livestock feed. Medical Association, 2017). Antibiotic bans in Holland, ISCIENTIST | 2018 For instance, of the total production of antibiotics in Denmark, Germany and Sweden were shown to cause the United States in 1998, 50% were used in the agricul- a decrease in resistance rates of one type of bacteria, tural industry with only 5% being used to treat infec- called Enterococcus (Bogaard et al., 2000). As a result tious disease (Khachatourians, 1998). of these bans, the European economy has experienced Although this discovery led to more efficient produc- deficits from the farming industry, which invoked fear tion of animal protein to sustain the ever-growing hu- in other countries and hesitations towards making man population, problems with this new agricultural drastic changes in their policies. practice were soon discovered. The development of Currently, this overuse of antibiotics contributes to the antibiotic resistance gradually became a concern in development of resistance as the bacteria are exposed many different countries, especially in European coun- to the antibiotics more often. The drugs act as a selec- tries. In 1969, the UK parliament created a committee tive pressure for resistance leading to increased devel- to investigate the threat of antibiotic resistance in ag- opment and spread of resistance genes through both riculture and provide recommendations for the future HGT and random mutations, as previously mentioned (Wise, 2007). This committee published the Swann 31 (Arber, 2014). THE FUTURE IS NOW In the history of agriculture, the use of antibiotics is a fairly new development and the consequences of this “Antibiotic resistance is putting the achieve- practice are still being determined. Policies regarding ments of modern medicine at risk.” – World the use of antibiotics as growth promoters are now be- Health Organization ing modified and implemented in many countries, yet agriculture is still considered a major contributor to Given that the consequences of antibiotic resistance antibiotic resistance. were not understood when antimicrobials were intro- duced in the 1940s, regulations regarding the use and HEALTHCARE distribution of antibacterial products did not exist un- til recently. Since antibiotics are used on a global scale, Although there have been large advances in the field the concern of antibiotic resistance is international. of public health care, antimicrobials are still one of the In 2001, the World Health Organization attempted to most important and successful developments in medi- bring policies into action by posting recommendations cine (Aminov, 2010). Evidence of the use of antibiotics and a global strategy on containment of drug resistance has been found from ancient human skeletons dating (WHO, 2017). Furthermore, federal governments have back to approximately 350 CE, but the concerns re- also tried to address the issue. The Canadian govern- garding resistance only began in the antibiotic era, in ment intends on forming relations with animal agri- the late 20th century (Aminov, 2010). Similar to the use culture providers by December 2019 to regulate the of antibiotics in agriculture, the overuse and misuse of approval of veterinary medication and to ensure that antibiotics in the healthcare industry also contribute these products are being used appropriately in animals to resistance. and humans. Through these policies, they anticipate Antibiotics are used in healthcare to control infec- that there will be a decrease in the overall use of antibi- tious diseases that were previously some of the lead- otics. These are only a few examples of efforts that have ing causes of death; however, the misuse of antibiotics been made by different organizations in recent years, can decrease their efficacy (Kardas et al., 2005). Misuse however antibiotic resistance continues to be a large includes the failure of patients to complete antibiotic problem. therapy, skipping doses, reuse of antibiotics and over- Aside from being an international issue, antibiotic re- prescription (Kardas et al., 2005). When antibiotics are sistance also involves affects different fields including used to treat an infection, the drug acts as a selective but not limited to microbiology, healthcare, agricul- pressure which drives the selection of bacteria that are ture, ecology, education, legislative bodies, the phar- Biochemistry resistant and thus not killed by the drug (Kardas et al., maceutical industry and the public (Aminov, 2010). Al- 2005). In previous studies, it was determined that one though efforts have been made to address this problem, third of patients did not comply with the therapy plan more drastic global measures must be implemented, and one quarter had leftover antibiotics for later use in order to prevent widespread resistance and loss of (Kardas et al., 2005). The consequences of this misuse antibiotic efficacy. Since the spread of resistance is a include the accelerated development of resistance. naturally occurring process, it cannot be stopped, but Another practice in healthcare that contributes to the rate of this spread can be reduced. antibiotic resistance occurs when physicians overpre- Some recommendations to address this issue include scribe these drugs. Antibacterial agents are commonly changes in policies in both the agricultural and medic- prescribed for respiratory tract infections which may inal fields, which should be applied to many countries be caused by viruses, instead of bacteria (Wang et al., throughout the world. Prescription antibiotics should 1999). A Canadian study that focused on data from pre- be used to treat bacterial infections and the use of these school children from 1995 estimated that about half of drugs for viral infections should be very limited as this the total cost of antibiotics in Canada was due to over misuse is a major driving force for bacterial evolution prescribing the drugs (Wang et al., 1999). In Canada, it (Kardas et al., 2005). More recently in 2016, the FDA has been estimated that each year, there have been up initiated a change in the marketing status of antibiot- to 26 million prescriptions from physicians that were ics used in livestock (Center for Veterinary Medicine, unnecessary for treatment (Williams and Heymann, ISCIENTIST | 2018 2017). Antibiotics that were previously over-the-count- 1998). This means that the antibiotic acts more often as er are now classified as either prescription or veterinary a selective pressure for bacterial evolution and does not feed directive, which means that they can only be used actually treat the viral infection. under supervision of a licensed veterinarian. Since the majority of antibiotics are used in agriculture, a reduc- tion or even an elimination of antibiotics in this indus- try would lead to a significant decrease in the spread of 32 resistance in bacteria. In addition to limiting use, the further development of resistant bacteria. Compared possibility of novel classes of antimicrobials should be to the billions of years of bacterial evolution, humans explored as new technologies could lead to great dis- have only spent approximately the last 80 years in the coveries (Aminov, 2010). antibiotic era. Although this time is only approximate- From the overview of the history of antibiotic era and ly equivalent to the current human life expectancy, studies regarding the spread of antibiotic resistance, these compounds have revolutionized human medicine we determined that antibiotic resistance first became a with the ability to treat infectious diseases that were major concern in the 1970s (see Figure 1). When looking once considered leading causes of death and to produce at the trends of antibiotic discovery, this time period enough food to support the growing worldwide popu- was considered the end of the Golden Age and after lation. Although resistance is inevitable, there are many this point, there have been few novel classes of anti- measures that may be taken to slow this rate of resis- biotics discovered. Additionally, there were few events tance, some of which are stated in this article, however promoting antibiotic discovery and few policies limit- the fight against resistance requires global cooperation. ing antibiotic use in agriculture and healthcare as well as other products. With the combination of these fac- ACKNOWLEDGMENTS tors as well as the increasing worldwide use of antibi- This review was adapted from the Enrichment Project otics, we propose that this marked the beginning of the of the Integrated Science program. We would like to antibiotic resistance crisis. thank Dr. Sarah Symons for her constant support and Ultimately, though antibiotics are considered relatively guidance. new in terms of human history, this revolutionary med- icine could soon lose its value due to the spread and cipars-picra/index-eng.php> [Accessed 22 Mar. 2017]. Peláez, F., 2006. The historical delivery of antibiotics REFERENCES from microbial natural products—Can history repeat? Biochemistry Aminov, R.I., 2010. A Brief History of the Antibiotic Kardas, P., Devine, S., Golembesky, A. and Roberts, Biochemical Pharmacology, 71(7), pp.981–990. Era: Lessons Learned and Challenges for the Future. C., 2005. A systematic review and meta-analysis of Frontiers in Microbiology, [online] 1(134). http://dx. misuse of antibiotic therapies in the community. Read, A.F. and Woods, R.J., 2014. Antibiotic resis- doi.org/10.3389/fmicb.2010.00134. International Journal of Antimicrobial Agents, 26(2), tance management. Evolution, Medicine, and Public pp.106–113. Health, 2014(1), p.147. Arber, W., 2014. Horizontal Gene Transfer among Bacteria and Its Role in Biological Evolution. Life : Khachatourians, G.G., 1998. Agricultural use of Saga, T. and Yamaguchi, K., 2009. History of antimi- Open Access Journal, 4(2), pp.217–224. antibiotics and the evolution and transfer of antibi- crobial agents and resistant bacteria. Japan Medical otic-resistant bacteria. Canadian Medical Association Association Journal, 52(2), pp.103–108. Bottcher H., 1959. Miracle Drugs. Surrey: Bookprint Journal, 159(9), pp.1129–1136. Limited. Stokstad, E.L.R. and Jukes, T.H., 1950. Further Obser- Mills, S. and Dougherty, T., 2012. Cell-Based Screening vations on the “Animal Protein Factor”. Proceedings Center for Veterinary Medicine, 2017. FDA Reminds in Antibacterial Discovery. In: Antibiotic Discovery of the Society for Experimental Biology and Medi- Retail Establishments of Upcoming Changes to the and Development. New York: Springer Science+Busi- cine, 73(3), pp.523–528. Use of Antibiotics in Food Animals. [WebContent] ness Media, pp.901–925. Available at: Federico, G., 2005. Feeding the World: An Economic Nothias, L.-F., Knight, R. and Dorrestein, P.C., 2016. WHO, 2017. The WHO policy package to combat History of Agriculture, 1800-2000. Princeton Univer- Antibiotic discovery is a walk in the park. Proceed- antimicrobial resistance. [online] Available at: 33 34 ISCIENTIST | 2018 Health Sciences HISTORY OFSCIENCE INTRODUCTION Integrated Science Program, Classof 2019,McMaster University LIZ BALINT,ASHLYN LEUNG Johns Hopkins University (Ome, 2017). The case tar case The 2017). (Ome, University Hopkins Johns against filed was lawsuit a when 2017 of September in light to brought recently been has This poses. it tions ques ethical the and phase trial clinical the within lie research healthcare in involved challenges biggest The ing datafrom humanparticipants. or other related therapies are often achieved by analyz vancements inhealthcare. Newdrugtreatments, diets, toad institutions extendfrom innovative technology each year (Statistics Canada, 2015). Research from these billion $13 approximately for accounting universities, Canadian research and development is performed by of total 40% affiliations. laureate Nobel and facilities leading universities are well known for their research Besides their quality of instruction, many of the world’s review Ethics of clinical research:Ahistorical Research that contributes to significant advancements in healthcare is often achieved Declaration of Helsinki,BelmontReport,Holmesburg Prison,Tuskegee SyphilisStudy Keywords: URL: Received: points thataimto protect therightsof thosewhoparticipate inresearch. These documents carefully considered the issues in their predecessors and include key controversies.ethical modern and regulations current of understanding stronger a to the Nuremberg Code, the Declaration of Helsinki, and the Belmont Report contributes developmentof the Analyzing edited. and shaped havebeen continuouslyguidelines these containing documents the that controversies major various through is It 1940s. the development of ethical guidelinesfor humanparticipationinresearchsince the to led that controversies the examines review historical This rights. participant’s the responsibilities of theinvestigator, andmoreimportantly,ensurestheprotectionkey of the outlines regulations ethical of understanding The per humans. case, on this tests in forming activity, or behaviour a govern that principles moral the are Ethics consideration. ethicalextensive require but science, of progression the for important are participants involvinghuman studies and trials Clinicaltherapies. related other or seek a diverse group of participants to study the effects of new drug treatments, diets, field this in researchers Many participants. human from data of analysis the through SUMMARY https://theiscientist.com/2018/02/08/LeungEthics.html 11/04/2017 tia gieie, ua priiain ciia til, uebr Code, Nuremburg trials, clinical participation, human guidelines, ethical Accepted: 02/08/2018 - - - - that govern one’s actions. Before the 1940s, there were there 1940s, the Before actions. one’s govern that lines and laws. Ethics can be defined as moral principles nize the implications and significance of current guide which modernethicsare based inorder tofullyrecog It is necessary to understand the historical context upon tance inresearch. ment can provide betterunderstandingoftheirimpor establish their to led that events the upon reflecting ical guidelineshave notalways existed inthepastand cases and how they can be prevented in these the future. Eth of reminded be to important is it ago, years 70 over occurred this though Even consent. their without malans withdiseases, includingsyphilisandgonorrhea, Guate infected intentionally who Government, States United the by conducted was study The 1940s. the in Guatemala in study unethical an financed and viewed geted theinvolvementofHopkins’ physicians,who re Published: 02/08/2018 ------ISCIENTIST | 2018 Health Sciences 35 ------1997). The Code particularly focuses onment require of the informed consent, described in the first prin to response direct a was focus This 1997). (Shuster, ciple con experimentation human perverse and tragic the WWII. during scientists and physicians Nazi by ducted After WWII, the victorious Allies held a seriesof mili Nurem The coined Germany, Nuremberg, in trials tary berg (Freyhofer, 2004). Trials These trials were held to World War World II War (WWII) that the first universalcode of formed. was Code, ethics, the Nuremberg blueprint the be to considered is Code Nuremberg The human subjects. involving for studies modern ethics for re of medical subjects that protect standards Current principles in the ten research their roots have search presented in the Nuremberg Code (Table 1) (Shuster, THE NUREMBERG CODE Principle - - - The ten principles that form the Nuremberg Code (Annas and Grodin, 1992). and Grodin, (Annas Code Nuremberg the form that principles ten The Table 1: Table to him to be impossible. him to to During the course of the experiment the scientist in charge must be prepared to terminate the ex periment at any stage, if he has probably cause to in believe, the of exercise the good superior faith, skill judgment and required careful of him that to is result of a the likely in continuation experiment subject. injury, the experimental or death to disability, subject against even remote possibilities of injury, of possibilities remote or death. disability, subject against even experiment The should be conducted only by scientifically qualified The persons. highest degree of skill and care should be required through all stages of the experiment of those who conduct or en gage in the experiment. experiment the bring to liberty at be should subject human the experiment the of course the During seems experiment the of continuation where state mental or physical the reached has he if end an to No experiment should be conducted where there is an a priori reason to believe that death or disab or death that believe to reason priori a an is there where conducted be should experiment No ling injury will perhaps, occur; in except, those experiments where the experimental physicians also serve as subjects. importance humanitarian the by determined that exceed never should taken be to risk of degree The the experiment. by be solved the problem to of Proper preparations should be made and adequate facilities provided to protect the experimental other methods or means of study, and not random and unnecessary and in nature. study, and not random means of other methods or The experiment should be so designed and based on the results of animal experimentation and a knowledge of the natural history of the disease or other problem under study that the anticipated the experiment. of results will justify the performance The experiment should be so conducted as to avoid all unnecessary physical and mental suffering and injury. decision by the experimental subject there should be the and him made to experimental decision the known by nature, duration, inconveniences all conducted; be to is it which by means and method the experiment; the of purpose possibly may which person or health his upon effects the and expected; be to reasonably hazards and qual the ascertaining for responsibility and duty The experiment. the in participation his from come rests upon ity each of the individual consent who directs It initiates, or is engages in the experiment. another with impunity. to be delegated not responsibility which may a personal duty and The experiment should be such as to yield fruitful results for the good of society, unprocurable by The The voluntary consent of the human subject is absolutely essential. This means that the free exercise person to able be to as situated so be should in consent; give to capacity legal have should volved over-reaching, duress, deceit, fraud, force, of element any of intervention the without choice, of power or other ulterior form of constraint or coercion; and should have sufficient knowledge andcompre hension of the elements of the subject as matter to involved enable him to an make understanding and enlightened decision. latter This element requires that before the of acceptance an affirmative 1 # 7 2 3 4 5 9 8 6 10 until the gruesome human experimentation during mischievous” mischievous” (Freyhofer, 2004). These scientists built guidelines ethics to establish ethical upon Hippocratic not but it was involving human subjects, for research follow that system of regimes which, according to my which, according follow that system of regimes ability and judgement, I consider for the benefit of my whatever is deleterious and patients, and abstain from dational theories of modern ethics. Many scientists in ethics. Many modern of theories dational the 1800s were familiar with the Hippocratic 470 and written 360 between BCE, which states, “I will Oath, and left the ethical responsibilities of researchers up to researchers responsibilitiesand ofleft the ethical interpretation. The progression of ethicalcodes prior to the 1940s provides a clear indication of the foun merly referred to merly as referred “human and experimentation” had sub the rights of the research for little consideration protocols, study in discrepancies many to led This jects. no universally accepted ethical guidelines for research guidelines for research ethical no universally accepted for studies were Clinical involving human subjects. prosecute Nazi officials who planned or participated the protection of their rights and welfare, and the re- in war crimes. The first of these was the International sponsibilities of doctors towards research subjects was Military Tribunal, also known as the Major Trial, which overlooked. As a result, a new statement of principles tried the highest-ranking Nazi officials for war crimes for ethical research involving human subjects was put and crimes against humanity (Freyhofer, 2004). This forth: The Declaration of Helsinki. The Declaration of was followed by the Doctor’s Trials, which tried phy- Helsinki was first adopted by the World Medical As- sicians responsible for the abuse of human subjects to sociation (WMA) in 1964 and was largely based on the advance medical sciences. This revealed arguably some fundamentals of the Nuremberg Code, but addressed of the most horrific acts committed during WWII. The clinical research more directly (Goodyear, Krieza-Jeric defendants argued that their acts were legal due to the and Lemmens, 2007). The main principles of the Dec- lack of a universal standard of human research ethics, laration include that medical research can never take and the experiments’ accordance to the law existing at precedence over the rights and interests of the partici- that time (Freyhofer, 2004). The Nuremberg Code was pant, the duty of the physician to protect the integrity developed during the Doctors’ Trial in order to produce and confidentiality of the participant, and that medical a set of medical standards with which to judge the phy- research must only be conducted by those with appro- sicians for their crimes (Annas and Grodin, 1992). priate scientific qualifications (World Medical Associ- While the focus of the Doctor’s Trial was the criminal ation Declaration of Helsinki, 2013). The Declaration nature of the experiments and the standards of human contains refinements and major advances to the prin- experimentation at the time, the broader concerns of ciples outlined in the Nuremberg Code (Stone, 2004). medical research ethics were also examined and debat- The document was carefully examined by international ed. The Code was ultimately based upon past literature, organizations and critiqued by the world’s preeminent events, and philosophies (Annas and Grodin, 1992). physicians, scientists and ethics experts. From their Two primary medical expert witnesses who supported evaluations, the Declaration has been revised seven the prosecution, Andrew Ivy and Leo Alexander, were times since its original publication. The most current largely responsible for the development of the princi- version of the Declaration contains 37 principles, sever- ples in the Code. Both scientists were familiar with the al of which may have been driven by the ethical analysis ethics of medical research proposed by Hippocrates, of controversial clinical studies. which ultimately formed the foundation of the princi- An important outcome following the publication of ples within the Code. It is important to recognize that the Declaration of Helsinki was the protection of vul- the ethical codes depicted by past influential scientists nerable populations in research studies. Vulnerable and physicians significantly impacted the development populations, such as prisoners, are those whose indi- of the ten principles of the Nuremberg Code. vidual freedom and ability to understand risks is affect- Health Sciences The Code has ultimately influenced global hu- ed due to various factors, including decreased freewill man-rights laws and medical ethics. It was not iden- and inability to make informed decisions (Shivayogi, tified as simply a code of ethics, but rather a part of 2013). Principle 25 of the Declaration states that “partic- the final legal judgement in the Doctor’s Trial. The ipation by individuals capable of giving informed con- physicians were judged according to the ten principles sent as subjects in medical research must be voluntary,” outlined in the Nuremberg Code (Annas and Grodin, which was controversial when it was implemented due 1992). Although it originates from the response to the to its ambiguity with regards to prisoners (World Med- tragic experiments at the hands of Nazi physicians ical Association Declaration of Helsinki, 2013). It was and scientists, the Nuremberg Code emerged from the argued that prisoners were not in a legal state to ex- horrors of WWII to contribute to safer, more ethical press choice and therefore, should their consent be giv- practices in medical research involving human subjects. en, it should not be considered informed. One example The Code has influenced the acceptance of informed of the exploitation of inmates is the study that oc- consent in international law, and is used as a basis upon curred at the Holmesburg Prison in Philadelphia in the which many ethical guidelines are built, such as the 1950s. In the studies conducted by dermatologist Dr. Declaration of Helsinki and the current regulation, the Albert Kligman, inmates were given experimental skin International Ethical Guidelines for Biomedical Re- products, including creams, powders, and deodorants ISCIENTIST | 2018 search Involving Human Subjects (Shuster, 1997). (Homblum, 2012). Although the inmates were paid for their participation, many suffered burns and scars as a THE DECLARATION OF HELSINKI result of the medical tests. Those who underwent the patch test, a 30-day trial, had up to 20 different prod- The Nuremberg Code provided important guidelines ucts applied to their back and were exposed to heat for the necessary measures to respect the research sub- from a sunlamp for a period of time (Homblum, 2012). ject’s wishes. However, little emphasis was placed on 36 The patch test sometimes caused skin to peel, itch, and to afford. Thus, researchers sought to test cheaper pre- blister, however, the study was not deemed unethical vention strategies, such as shorter treatment regimens due to the uncertainty of Principle 25. In addition, Dr. or HIV-antibody injections, and performed a study Kligman’s research was not investigated by institution- in several developing countries, involving over 17 000 al personnel, nor was it supervised by the government, pregnant women (Annas and Grodin, 1998). The ques- which allowed Holmesburg Prison to host one of the tion of including placebos in these study groups led to largest medical experimentation centres in the country ethical debate since the original AZT treatment had for two decades. Holmesburg Prison became a key driv- already shown effective long-term results (McIntyre, ing factor for the implementation of ethical guidelines 1998). The fourth revision of the Declaration hoped and the importance of protecting research subjects. Al- to address this situation by specifying that placebos though studies on federal prisoners ended in the mid should only be used where no other therapeutic meth- 1970s, studies on other vulnerable populations contin- od exists. Even though the study was approved by Afri- ued until further revisions of the Declaration were ef- can national ethics committees, thus complying to that fectuated. Among the seven revisions of the Declara- aspect of the Declaration, bioethicists were sceptical tion, the greatest changes occurred in the first, fourth due to the reputations of said committees. Cases have and fifth revisions. been noted where materials for the studies were sent The first revision of the Declaration of Helsinki was from abroad prior to study approval and bribery was published in 1975 after the realization that abuse of re- used to gain ethical approval (McIntyre, 1998). These search subjects remained a common problem. Among reports provide evidence for increasing the stringency the most important aspects of this revision was the of the principles outlined in the Declaration through elaboration of the requirements for informed consent constant revisions. and the rights of human subjects. This was specified Questions surrounding the ethics of placebo-controlled in Principle four which read, “In research on man, the trials arose again in 2000 after the fifth revision of the Health Sciences interest of science and society should never take pre- Declaration, where Principle 29 stated, “The benefits, cedence over considerations related to the well-being risks, burdens and effectiveness of a new method should of the subject” (Williams, 2008). To further protect the be tested against those of the best current prophylac- welfare of participants, it was also added that the re- tic, diagnostic and therapeutic methods. This does not search study must undergo advanced review by an in- exclude the use of placebo, or no treatment, where no dependent committee prior to conducting the study in proven prophylactic, diagnostic or therapeutic method order to publish the results of the research (Williams, exists” (Lewis et al., 2000). Although this principle does 2008). One of the factors that may have driven this re- not appear to differ from that of the fourth revision, vision was the controversy surrounding the Tuskegee it’s importance was demonstrated through the contro- Syphilis Study of 1932 to 1972. versy it generated. It’s been argued that if Principle 29 A fourth revision of the Declaration of Helsinki oc- was taken literally, all clinical trials should be barred curred in 1996 to address placebo-controlled trials. because research subjects receiving the investigational The existing principle stated, “In any medical study, treatment would not be getting the best proven treat- every patient – including those of a control group, if ment (Simon, 2000). Instead, they believe that place- any – should be assured of the best proven diagnostic bo-controlled trials can be conducted ethically so long and therapeutic method,” to which the fourth revision as the omission of the proven treatment would not be added “This does not exclude the use of inert placebo detrimental to the patient’s health and patients are in studies where no proven diagnostic or therapeutic completely informed about the alternative treatments ISCIENTIST | 2018 method exists” (Carlson, Boyd and Webb D, 2004). available. Motivation for this revision was driven by the increas- Over 50 years after its initial development, the Decla- ing concerns regarding the use of placebo controls in ration of Helsinki continues to be an important docu- prenatal HIV transmission in developing countries, ment that sets the ethical standards for modern clin- specifically the trials for azidothymidine (AZT) in ical studies. The Declaration has remained dynamic 1994 (Carlson, Boyd and Webb D, 2004). Prior to these and adapted to changes as ethical problems arose. Due trials, a study of HIV infected pregnant women, con- to this document, physicians are more aware of their ducted by the USA and France, found that intensive responsibilities and research subjects are more aware treatment with AZT reduced maternal transmission of of their rights. It has brought substantial awareness to HIV in 70% of cases (Cohen, 1997). Researchers were the importance of ethics in human research; however, aware that the incidence of HIV/AIDS was greatest in a number of unethical studies are still being conduct- developing countries, but AZT treatment costs $800- ed despite the principles in the Declaration. Although $1000 per person, which is difficult for those women seven revisions have been made in the past, it must con- 37 Table 2: The three fundamental principles of the Belmont Report (Annas and Grodin, 1992). Principle Responsibilities Protecting the autonomy of all people and treating them with courtesy and re- Respect for persons spect; this is applied in the informed consent process. Researchers must be truth- ful and conduct no deception Incorporating the philosophy of “Do no harm” while maximizing benefits for the Beneficence research project and minimizing risks to the research subjects is applied through risk/benefit assessments Ensuring reasonable, non-exploitative, and well-considered procedures are ad- Justice ministered fairly and equally and applied to the selection of research subjects tinue to change in the future to provide ongoing pro- search protection system to uphold the rights of human tection for human research subjects. participants and prompted the development of regula- tions requiring the establishment of Institutional Re- THE BELMONT REPORT view Boards at federally funded institutions (Singer and Levine, 2003). This led to the creation of the National Around the time of the first revision of the Declaration Commission for the Protection of Human Subjects of of Helsinki, the controversial conditions of the Long- Biomedical and Behavioural Research (Kim, 2012). The term Study of Untreated Syphilis in the Negro Male, Commission was required to identify principles that known as the Tuskegee Syphilis Study, were revealed. underlie the ethical conduct of biomedical and be- The study was conducted from 1932 to 1972 and was un- havioural research. Their main product, the Belmont dertaken by the U.S Public Health Service which later Report, was published in 1979 and marked a key point became the Centers for Disease Control and Preven- in history for the development of ethical requirements es tion (CDC). The study sought to document the course involving the use of human subjects in research (Singer of the disease in 400 African Americans to determine and Levine, 2003). The three fundamental principles of racial differences in the manifestation of syphilis (Cor- the Belmont Report (Table 2) influenced the criteria bie-Smith, 1999). When the study first began, there was for the protection of human subjects and continues no effective treatment for the disease, which was the to be an important reference for Institutional Review case until 1943, when researchers found that penicillin Boards. The Belmont Report formed another base upon could be used to treat syphilis. Despite this discovery, which medical research regulations are formed, and has the study was continued and the subjects were never widely influenced the standards that exist today. notified of the existence of a treatment (Singer and Health Scienc The controversies surrounding the Tuskegee Syphilis Levine, 2003). In response to the ethical concerns that Study served not only as a representation of the ex- arose during the study, the Belmont Report was writ- ploitation of vulnerable populations through human ten to establish stricter regulations. experimentation, but also as an inciting incident to Even though the Declaration of Helsinki was formed produce new regulations to protect the rights of future in 1964, no changes were made to the Tuskegee Syphilis human volunteers. It can be argued that the most im- Study (Kim, 2012). In 1969, the CDC formed a panel to portant consequence of the study was the creation of the review the experiment, but they allowed the continua- National Research Act, the National Commission for tion of the study without modification to the protocol, the Protection of Human Subjects of Biomedical and for unknown reasons (Shovers, Lynch and Burmeister, Behavioural Research, and ultimately, the formation of 2000). In 1972, a story published in the New York Times the Belmont Report. The lack of ethics surrounding the and the Washington Star revealed the ethical concerns study have evidently led to the development of more of the study, and the resulting public outcry led to hear- protective guidelines to prevent these events from re- ings directed by Senator Edward Kennedy and closure appearing in the future, and have powerfully impacted of the study (Kim, 2012; Shovers, Lynch and Burmeister, the ethics of current medical research. 2000). At this time, participants were finally given the appropriate treatment, but it was too late for the ap- MODERN CONTROVERSIES ISCIENTIST | 2018 proximately 100 men who died due to untreated syph- ilis or syphilis-related complications (Corbie-Smith, Although the regulations outlined in official docu- 1999). ments such as the Nuremberg Code, the Declaration of Within two years of the termination of the Tuskegee Helsinki, and the Belmont Report drastically improved Syphilis Study, the United States Congress passed the the ethical requirements surrounding clinical trials, National Research Act. This established a human re- there are still many controversies surrounding modern 38 clinical research. result, patient risks increase, and consequences may One example of this is known as “pay to play” clini- remain unknown. Drug companies are mainly driven cal trials. Most studies offer monetary compensation by commercial interests and may prioritize them over for their research participants, but a number of recent the interest of the patient. To accommodate this grow- studies require payment for participation. Some re- ing issue, bioethicists recommend that new policies be searchers charge patients to be enrolled in their study designed specifically for these trials (Wenner, Kimmel- as both a way to fund their research and for personal man and London, 2015). Such policies should promote profits (Emanuel et al., 2015). Patient funded studies research methods that minimize bias when collecting have been conducted for conditions including Parkin- data and ensure that the paying patient is aware of all son’s disease, and Multiple Sclerosis. One particular risks surrounding the trial. Should this type of trial case surrounds Novastem, a Mexican stem cell product continue in future studies, it is important to form new distributor for the U.S company Stemedica. Novastem accommodating guidelines to ensure the safety of the reportedly charged their participants $30 000 USD for participant. enrollment in their trial which uses neural stem cells to treat stroke-related brain damage (Wenner, Kimmel- CONCLUSION man and London, 2015). This raised concerns regarding In the past century, various documents have been estab- patient participation and regulation of the study. lished which state that certain ethical principles must First, selection of subjects for Novastem’s study were be incorporated in clinical studies to protect the re- based on one’s ability to pay, thus prioritizing the needs search subject’s rights. It is necessary for researchers to of the wealthy. Not only does this pose an ethical di- understand how ethical regulations have been formed lemma, but this can affect the validity of the study, in order to understand their significance and potential as the selection of participants should be primarily for improvement. Knowledge of the biases, opinions, Health Sciences based on the goals of the research study. Issues then and controversies that have produced these regulations arise when considering the need for placebos because provides a better perspective with which these ethical paying participants will be less willing to accept ran- codes can be understood. Since the 1940s, controversies domization when there is a possibility that they will surrounding experiments with human subjects have not receive the treatment. Also, patient funded studies led to new guidelines that shape research ethics. The can cost thousands of dollars, causing patients to spend Nuremberg Code, the Declaration of Helsinki, and the large amounts of their savings. Severe illnesses can com- Belmont Report were guidelines that formed as a result promise decision making when deciding to spend large of controversial studies responsible for the improper funds on a potential treatment (Emanuel et al., 2015). treatment of human subjects. These regulatory ethical This could allow researchers to exploit desperate pa- research codes have helped to shape the development of tients, even though the majority of experimental agents our current ethical standards, and will continue to be u used in early clinical trials fail. used as a basis for future ethical guidelines. Another concern regarding patient funded trials is the production of reliable medical evidence to ensure the safety of the study. New forms of treatment can be ACKNOWLEDGMENTS ineffective, and even dangerous, thus, several factors We thank Dr. Sarah Symons for her assistance, guid- to oversee the process are typically required. Studies ance, and support throughout this project. must produce adequate evidence for the safety, tox- icity, and efficacy of their treatment prior to submit- AUTHOR CONTRIBUTIONS ISCIENTIST | 2018 ting a clinical trial application (Wenner, Kimmelman L.B. wrote the Nuremburg Code and Belmont Report and London, 2015). Since trials are expensive, private sections and A.L wrote about the Declaration of Hel- sponsors may minimize the sample size and duration sinki and modern controversies. Both authors contrib- of the study to generate the required evidence. As a uted equally to the introduction and conclusion. 39 40 ISCIENTIST | 2018 Health Sciences Investigation. Tuskegee SyphilisStudy:Considerations forClinical 1999.TheContinuingLegacy ofthe Corbie-Smith, G., er CountriesAttacked. 276(5315),p.1022. 1997. AIDS:EthicsofAZTStudiesinPoorCohen, J., future. 57(4),pp.695–713. of theDeclaration ofHelsinki:past,present and Boyd,K.andWebbCarlson, R., D, 2004.The revision it=0>. action?objId=88-221-X&objType=2&lang=en&lim Available at: Published: by showing that participants reported experiencing ad by showing that participants reported verse effects when theyconsumed test food itemscon taining MSG, and not when they consumed the food item without MSG. both However, studies had a small solely on participants who conducted sample size, were to the symptoms, and as being susceptible self-reported or detailed protocols well-described neither included results. the responsible agent, but rather mentioned it in a list mentioned but rather agent, the responsible wine, cooking including soy sauce, causes of speculative 1968). and sodium (Kwok, many further anecdotal publication, Kwok’s Following of investigations and a variety published, letters were of of this constellation into the cause conducted were symptoms (Anon., 1968). This included a double-blind wonton soup participants consuming study with three cross single-blind a and 1968), Byck, and (Schaumburg over study with thirty-fivetomato participants juice (Ambos, et al., consuming 1968). These small initial studies concluded that MSG was the responsible agent LETTER TO THE ISCIENTIST THE TO LETTER ------03/12/2018 Accepted: Accepted: , Kwok coined the term coined , Kwok monosodium glutamate, adverse effects, monosodium glutamate symp monosodium glutamate effects, adverse monosodium glutamate, 01/22/2018 https://theiscientist.com/2018/03/12/LaiAdverse.html SUMMARY Keywords: additive food syndrome, Chinese restaurant complex, tom Received: URL: often dismissed, some evidence points to possible effects of MSG on human physiol further controlled study, such as better warrant persist that ogy; gaps in the literature with food. MSG consumption studies involving that although the constellation of symptoms, now referred to as MSG symptom com and are thus study conditions in controlled reliably been reproduced not have plex, biologic effects of glutamate, the active compound in MSG, are first briefly discussed, human consump of effects on adverse literature a summary existing by of followed tion of MSG. Overall, based on the current state of evidence, it is argued in this work food additive in 1908. Although the compound has been recognized as safe for human human for safe as recognized been has compound the Although 1908. in additive food be reports and and anecdotal medical regulatoryby exist bodies, there consumption known the work, this In effects. adverse of variety a elicit can MSG consuming that liefs Monosodium glutamate (MSG) is a widely used flavour enhancer,first patented as a Notably, Notably, Kwok did not specifically implicate MSG as neck, generalized weakness, and palpitations. He fur weakness, generalized neck, of Chi friends from experiences similar described ther nese descent aftereating at some Chinese restaurants. tion of symptoms he had repeatedly experienced after at including numbness a Chinese restaurant, at eating to the arms and with radiation the back of the neck ported by Kwok in 1968. In a letter to ported by the Kwok editor in the New England Journal of Medicine “Chinese restaurant syndrome” to describe the collec ponent of MSG is glutamate, which produces the taste known as umami (Sano, 2009). Adverse effects ofcon suming monosodium glutamate were first notablyre Monosodium glutamate (MSG) is a food additive first com active The 1908. in enhancer flavour a as patented INITIAL REPORTS OF ADVERSE INITIAL REPORTS OF ADVERSE EFFECTS OF MSG REVIEW OF EXISTING LITERATURE JAMES LAI JAMES University 2019, McMaster of Medicine, Class School of Michael G. DeGroote brief review brief monosodium glutamate consumption: A A consumption: glutamate monosodium Adverse effects associated with associated effects Adverse STUDIES BY REGULATORY AGENCIES suggested that statistically significant increases in head- In the following years, several further studies were con- ache (p < 0.023), muscle tightness (p < 0.004), numbness/ ducted. In particular, the issue was examined by various tingling (p < 0.007), general weakness (p < 0.040), and food regulatory groups, which conducted literature re- flushing p( < 0.016) could be elicited when participants views to reach their conclusions. In 1980, a committee consumed 5 g MSG disguised in in a citrus-flavoured of the Federation of American Societies for Experimen- beverage (Yang, et al. 1997). A subsequent multicentre tal Biology (FASEB) concluded that MSG may cause study with 130 participants used a similar protocol, but adverse reactions in a portion of the population that included further phases of testing on susceptible indi- is susceptible to its effects, but that evidence was not viduals, one of which included consumption of MSG strong because some studies showed that symptoms with food (Geha, et al., 2000). As in the study by Yang could be elicited without MSG. It further noted that and his colleagues, it was noted that subjects responded MSG levels in use in processed foods were less than to higher doses of MSG; however, with later rounds of those used in the experiments that elicited symptoms. testing, it was found that subjects did not respond con- The average daily MSG intake was estimated at the sistently to MSG challenge. More recently, Baad-Han- time to be around 550 mg/day (He, et al., 2011), while sen, et al. (2010) found increases in headache, pericrani- experiments typically used amounts in the range of 1 to al muscle tenderness, and systolic blood pressure with 12 grams in a single dose. Therefore, it was concluded MSG intake in sugar-free soda versus placebo. Studies that levels in use in processed foods were acceptable using a similar oral MSG-soda challenge have also con- and posed no risk of harm. No comment was made re- cluded that MSG intake leads to sensitization of the garding MSG levels in home- or restaurant-prepared masseter muscle and increased blood pressure (Shima- foods, as this fell outside of the committee’s scope da, et al., 2013; Shimada, et al. 2015), as well as increased (Federation of American Societies for Experimental susceptibility in individuals with temporomandibular Biology, 1980). A joint Food and Agriculture Organi- disorder (Shimada, et al., 2016). However, these studies zation and World Health Organization report in 1987 used a fairly high dose of 150 mg MSG/kg body weight. similarly concluded that levels required for the intend- ed effect of flavour enhancement did not pose a risk, DISCUSSION but suggested caution regarding taking large amounts It is currently widely believed by the public that MSG in a single dose; it did not specify what constituted a does not cause adverse effects, and that studies have “large” dose (World Health Organization, 1987). A fur- definitively shown that the MSG symptom complex is ther review of MSG by the FASEB in 1995 suggested not reliably reproduced in laboratory settings. Based the less pejorative name “MSG symptom complex” and on the review of the current literature, it is argued in Health Sciences concluded that evidence suggested that causality for this work that a more nuanced and balanced position MSG and reported side effects had been established for would be more appropriate. It is argued here that: a subset of the general population, noting reactions as 1. MSG consumption in large amounts without described by previous reports to oral MSG boluses ≥ 3 g food have been shown to elicit uncomfortable without food, and more frequently if taken on an emp- but non-life-threatening symptoms ty stomach. However, despite the abundance of reports 2. One well-conducted double-blinded place- on these symptoms, the quality of much of the litera- bo-controlled trial (Geha, et al., 2000) showed ture available at the time was deemed to be poor (Fed- that the symptoms elicited were not consis- eration of American Societies for Experimental Biolo- tently reproduced gy, 1995). Food Standards Australia and New Zealand reviewed the safety of MSG in 2003 and concluded that 3. There is a paucity of well-designed studies ad- many earlier studies had methodological flaws. How- dressing MSG consumption in large amounts ever, they agreed that the ingestion of large amounts with food, and the fact that both food and the of MSG (≥ 3 g) without food did provoke symptoms pattern of MSG consumption during a meal in a small population of people. It was also noted that is expected to affect the dynamics glutamate the literature did not adequately address whether con- absorption sumption of MSG with food would alter the effect on 4. Arguments asserting that the average daily ISCIENTIST | 2018 participants (Food Standards Australia New Zealand, consumption of MSG do not exceed recom- 2003). mended maximum acceptable doses have only addressed MSG consumption from processed RECENT CLINICAL STUDIES food and home cooking, and not the MSG Various double-blind placebo-controlled (DBPC) trials content of restaurant food; thus, further re- have been conducted. One trial with 61 participants search should be conducted in this regard 42 Figure 1: Comparison of free glutamate levels in various food items and restaurant meals. Values obtained from Yamaguchi and Ninomiya (1998, cited in Food Standards Australia New Zealand, 2003) and Nicholas and Jones (1991, cited in Food Standards Australia New Zealand, 2003); maximum values were selected for those with ranges provided. Of particular note, food regulatory agency reports have MSG symptom complex was originally associated with Health Sciences acknowledged that large doses of MSG consumed with- eating restaurant food containing MSG, it is unclear out food appear to elicit temporary adverse effects that whether MSG consumption amounts as previously es- are not life-threatening. Double-blinded placebo-con- timated from processed food and home cooking, rang- trolled studies agree with this conclusion, but have ing from 0.6 to 1.5 g/day (Henry-Unaeze, 2017), apply shown that symptoms are not elicited consistently. The to restaurant food. One estimate of MSG content in literature review conducted for this work did not find restaurant food reported up to 1500 mg free gluta- any major DBPC since that of Geha, et al. (2000); we mate/100 g food (Nicholas and Jones, 1991 cited in Food argue it would be prudent to replicate this study giv- Standards Australia New Zealand, 2003) (Figure 1). It en the relatively low number of participants that were has also been estimated (although, without justification tested in the later phases of their study, which pre- or reference) that a highly seasoned restaurant meal cluded statistical analysis (12 participants in Protocol may contain as much as 5 g MSG (Yang, et al., 1997), C, and 2 for Protocol D). Furthermore, following the which falls within the range of MSG levels consumed work of Geha, et al. (2000), in which MSG consump- in experiments. It may be reasonable to systematically tion with food was assessed in Protocol D, no further survey restaurant food for its MSG content to more ac- DBPC with MSG with food were located. It has previ- curately inform MSG with food studies; new protocols ously been asserted that consumption with food may have been developed that may facilitate this, such as affect absorption of glutamate; it has also been noted the liquid chromatography-tandem mass spectrometry that consumption of MSG in food may occur in a va- used by Cebi, et al. (2018). ISCIENTIST | 2018 riety of temporal patterns over the course of the meal. In conclusion, evidence currently suggests that inges- Studies should therefore evaluate MSG consumption tion of large amounts of MSG without food may trig- with food to better elucidate possible adverse effects of ger symptoms consistent with MSG symptom complex consuming MSG in real-life situations. in susceptible individuals. However, there is currently The basis of several assertions of MSG’s safety includes a lack of studies regarding the adverse effects of MSG the argument that MSG is not consumed in amounts as consumption with food, and estimates of MSG con- high as those used in studies. However, these assertions sumption are currently based on processed and home- are either based on the claim that high consumption of cooked foods, not restaurant foods. Therefore, it is MSG does not occur because it is not required for its argued that further DBPCs challenging participants intended flavour-enhancing effect (World Health Or- with MSG in food, informed by better quantification ganization, 1987), or based on calculations and surveys of MSG in restaurant foods, should be conducted be- of processed foods and home cooking (Rhodes, et al., fore dismissing the possibility that MSG may produce 1991; He, et al., 2011; Henry-Unaeze, 2017). Given that adverse effects when consumed. 43 44 ISCIENTIST | 2018 Health Sciences Standards Australia NewZealand. assessment safety a glutamate: sodium 2003. Zealand, New Australia Standards Food Societies forExperimentalBiology. (MSG) tamate ology,1995. Federation of American Societies for Experimental Bi of American SocietiesforExperimentalBiology. glutamates as food ingredients 1980. Experimental Biology, for Societies American of Federation bust LC-MS/MSmethod. 2018. A survey of freeO., glutamic acid Sagdic, in foods using and a roO. E. Olgun, E., C. Dogan, N., Cebi, Cephalalgia pericranialheadacheand on (MSG) sensitivity.muscle son, P., 2010. Effect of systemic monosodiumSvens glutamate and M. Ernberg, B., Cairns, L., Baad-Hansen, Medical Association Journal syndrome. restaurant Chinese 1968. Anon., England Journal ofMedicine glutamate. on accent cib-syn: Sin 1968. B., S. na, Marmorek, L.andWolschi Leavitt, N.R., Ambos, M., REFERENCES , 30(1),pp.68-76. Analysis of adverse effects to monosodium glu monosodium effectsadverse of to Analysis . Bethesda, MD: Federation of American of Federation MD: Bethesda, . Evaluation of the health aspects of certain of aspects health the of Evaluation , 99(24),pp.1206-1207. , 279(2),p.105. Food Chemistry . Bethesda, MD: Federation Cner: Food Canberra: . , 248,pp.8-13. Canadian Mono New ------ican Journal ofClinical Nutrition Sano, 2009. 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Cairns, A., Shimada, 279(2), p.105. cib-syn: accent onglutamate. Sin 1968. R., Byck, and H. H. Schaumburg, , 99(6),pp.757-762. New England Journal of Medicine of Journal England New Nutrition European Journal of Pain of Journal European Journal of Headache PainHeadache of Journal Journal of Allergy and Clinical . Geneva: WorldHealth Geneva: . , 31(2),pp.315-323. Evaluation of certain of Evaluation , 20(9), , , 14(1), , - - - - - , ISCIENTIST | 2018 Ecology 45 ------03/15/2018 ORIGINAL RESEARCH ORIGINAL Published: Many definitions of invasive species hinge on the idea of such harmful. An example necessarily that they are a definitioncomes fromRussell and Blackburn (2016), only invasive becomes that an alien species who argue Another threshold. a given damage when it exceeds similar definitioncan be found in Richardson et al.’s should be used. For a comprehensive comparison of the comparison a comprehensive For should be used. the for and literature the in found definitions following 1. Table public, see general DEFINITIONS BASED ON NEGATIVE IM ON NEGATIVE DEFINITIONS BASED PACTS 03/15/2018 Accepted: Accepted: invasion ecology, invasive species, stage-based model, invasive ecology, propagule invasion pressure, 11/04/2017 https://theiscientist.com/2018/03/15/BiderRedefining.html SUMMARY Keywords: capability reproductive energy consumption, availability, resource URL: mented at multiple stages for full at multiple stages for effectiveness. mented Received: invasive species will allow for better communication within invasion ecology research ecology invasion within communication better for allow will species invasive and throughout the scientificcommunity. Ultimately, our definition helps to simpli be imple that can policy prevention invasion a practical fy developing of the process simulates simulates the influence of multiple ecological factors (propagule pressure, resource availability, energy consumption, reproductive capability), which can either inhibit or assist the invasion of a species. Adopting a more comprehensive method for defining tion, and parasitism in addition to contingent physical forces such To as definition. flooding, this into freez fit to able be will species of variety wide a Therefore, fire. and ing, demonstrate its effectiveness, we test our definition with an agent-based model that nition is to classify a given species by its progression through the invasion process. By process stage-based the of version novel a propose we definitions, existing integrating of invasion. This approach incorporates extrinsic factors such as competition, preda characteristics characteristics of the (potentially) invasive species. This process encompasses three main stages: establishment, transport, purpose and of spread. a The stage-based defi based on human perception. The objective of this objective project to was based better define species on human The perception. using invasion a mechanistic, authors stage-based attempt have perspective. Several ed to develop a definition thatfocuses on the processof invasion,rather than on the Invasion ecology explores the impacts of new species entering established ecological ecological established entering species new of impacts the explores ecology Invasion discussed frequently are species invasive of effects the While communities. and niches and arbitrary, vague, remains invasion of definition the academia, industryand both in to be a consensus regarding how and when these terms regarding to be a consensus use some or all of these terms, but each with their own with their each these terms, but of or all some use awareness ever-growing Despite personal connotation. has yet there ecology, and invasion species of invasive “imported”, “imported”, “naturalized”, and “non-native” frequently poorly remain generally but in the literature, appear will al., Authors et 2008). Valéry 2016; (Pereyra, defined there there have been many different wordsused out of its native range. that has travelled to a species refer to such “exotic”, “introduced”, as Terms “invasive”, “alien”, Since Since the emergence of invasion ecology as a field, INTRODUCTION PASCALE BIDER, TANYA DANIEL, ADAM TWEEDLE ADAM DANIEL, TANYA BIDER, PASCALE University 2019, McMaster Class of Program, Science Integrated based process based Redefining invasive species using a stage- using species invasive Redefining Table 1: Comparative table highlighting the various definitions of “invasive” used in invasion ecology (based on Pereyra’s 2016 methodology). Both academic and public sources were examined. Citation Terms Specific characteristics that must be met in order to be con- used sidered invasive Negative Human Other Impact Mediated Valery et al., 2008 Invasive No No Must become dominant in new ecosystem Russell and Blackburn, Invasive Yes No - 2017 Tassin et al., 2017 Invasive No No - Richardson et al., 2000 Invasive No Yes Must be relocated beyond a specific distance Hulme et al., 2016 Alien No No - Pycek et al., 2004 Invasive No Yes Depends on specific term used Heger and Trepl, 2003 Invasive No No - Pycek and Richardson, Invasive No Yes Must produce reproductive off- 2010 spring at locations far from the site of introduction Powell, Chase, and Invasive No No Must have high rates of popula- Knight, 2013 tion growth and become domin- ant in new ecosystem Wikipedia, 2017 Invasive Yes Yes Must directly threaten agricul- ture or biodiversity USDA, 2016 Invasive Yes No Must cause environmental harm or harm to human health Ecology Ontario's Invasive Spe- Alien, In- Yes Yes - cies Awareness Pro- vasive Alien gram, n.d. Government of Canada, Alien, In- Yes No - 2016 vasive Alien (2000) paper, who claim that alien species makes the confusing language and misuse of terms (Tassin et al., shift from “naturalized” to “invasive” when it begins to 2017). Additionally, defining “invasive species” based on have detectable consequences. This paper also proposes their negative impacts makes it difficult to categorize to label those alien species, once they are regarded to be species who have the potential for negative impacts in harmful, as “pests” and “weeds”. the future (Russell and Blackburn, 2016). The method of classifying species based on how harm- ful they are to their surroundings has some obvious DEFINITIONS BASED ON HUMAN issues. To use such a definition, we must define what FACILITATION constitutes “invasive”, as well as what constitutes a Another problem that is presented by many current “negative impact”. Our perception of what is “negative” definitions of the term “invasive” is that, by some stan- is heavily based on social values (Tassin et al., 2017). dards, an invasive species must be introduced through Invasive species have a variety of consequences, rang- ISCIENTIST | 2018 human means. Gilroy et al. (2016) argue that, since we ing across ecological, biological, economic, or social need different management practices for species intro- impacts. The introduction of a new species may result duced with human aid and species introduced without, in negative consequences within certain areas, but not any definition used should clearly differentiate the in others. When using these types of definitions, some two. It has been proposed that terms such as “invasion” may choose to label a given species as invasive, while be reserved for the cases where species distribution is others will not. Differences in values could result in changed by human activity (Gilroy et al., 2016; Pysek 46 et al, 2004). Several legislative bodies have incorporat- are used, it may seem unrealistic to remove them from ed this school of thought in their definitions (Gilroy et our language altogether. al., 2016). For example, the European parliament stated in 2014 that an “alien” species is identified as “any live DEFINITIONS THAT ARE EXCESSIVELY specimen introduced outside its natural past or present SPECIFIC distribution” where introduction signifies “the move- Throughout the literature, several definitions of “inva- ment by human intervention” (Gilroy et al., 2016). Ad- sive” require the species to fit within highly specific, ditionally, it has been said that since non-human-medi- and relatively arbitrary, criteria. For example, Richard- ated introductions are so rare, they should be classified son et al. (2000) propose that, in order to be considered as natural rather than invasive (Richardson et al., 2000). invasive, a plant must be carried further than 100 km This type of definition is problematic for many reasons. from their native range. Then, plants spreading by seed First, organisms can travel to new regions without the must continue to expand in their new range at a rate aid of humans. For example, in one of the first works to of over 100 m in under 50 years (or over 6 m in under ever describe invasive species, Charles Elton explains 3 years for plant spreading by root or rhizome). These that since many migratory birds travel across the globe, minimum distances must be met in order for the plant they can easily transport seeds, small eggs, or even mi- to be considered invasive (Richardson et al., 2000). An- croscopic organisms far from their usual range (Elton, other example can be found in the 2004 article by Pysek 1958). Secondly, classification is difficult when a species et al. where it is stated that a “naturalised alien” is a uses human-mediated processes only temporarily, or plant that has sustained a self-replicating population for only a part of their journey to a new habitat. For for at least 10 years without aid from humans. These example, bats may temporarily rest on boats when trav- overly precise definitions can be difficult to apply to a elling over water from one range to the next (Gilroy et wide range of species. In addition, some feel that overly al., 2016). There is ambiguity when a species travels to a specific, academic language can complicate - commu new area by more than one pathway, some human-me- nication between invasion ecologists and the general diated and some not (Hulme et al., 2016). This approach public (Pereyra et al, 2016). Ecology can also complicate the labelling of a newly-arrived species whose vector of transport is still unknown. THE IMPORTANCE OF A STANDARD- Some argue that once a species is introduced, it makes IZED DEFINITION no difference from an ecological perspective whether This inconsistency present in invasion ecology vocabu- the introduction was human-mediated or not (Heger lary has arisen as a result of many factors. First, it has and Trepl, 2003). Therefore, no distinction needs to be been speculated that since invasion biology has not made between the two. Many texts have now adopted been studied as a holistic field until recently, the sepa- this point of view (Valéry et al, 2008; Colautti and Ma- rate disciplines previously concerned with species inva- cIsaac, 2004; Vermeij, 1996). sion have each established their own definitions (Pysek et al., 2004). Consequently, we have definitions that DEFINITIONS USING PEJORATIVE come from ecological, biogeographical, and anthropo- LANGUAGE centric perspectives, to name only a few (Pysek et al., Texts describing species establishing themselves outside 2004). Furthermore, invasion ecologists concerned with of their native range will often do so using unnecessar- different taxonomic groups have generated definitions ily pejorative or value-laden terms. The term “invasive” that are applicable only to their taxa and not others in itself can be associated with concepts such as “attack” (Blackburn et al, 2011). In particular, there tends to be a ISCIENTIST | 2018 or “aggression” (Richardson et al., 2000). However, this divide between ecologists studying plants and animals term has been used more often than any other in recent (Blackburn et al. 2011). Some plant ecologists might use literature (Pereyra, 2016). Additionally, some papers the term “invasion” to define the local establishment of may choose to use other pejorative terms such as “pest” new populations, whereas outside of this field, the same and “weed” (Richardson et al., 2000; Pysek et al., 2004). process might be defined as “colonisation” (Richardson This type of language is problematic as it may lead to et al., 2000). For example, Taxillus tomentosus (mistle- biased attitudes towards non-native species (Pereyra, toe) has been called invasive to certain forested areas 2016). Viewing all invasive species as harmful attackers in South India, even though the plant is native in this can cause us to overlook their potential uses and ben- general region (Rist et al., 2010). efits (Geniusz, 2015). Some have proposed the replace- Another potential reason for misunderstanding regard- ment of biased terms by neutral ones, such as “alien”, ing current terminology could be the result of different “introduced” or “neobiota” (Pereyra, 2016). However, languages in which invasion ecology has been studied given how commonly some of these pejorative words and written. Researchers from all over, particularly in 47 Central Europe, have contributed to the field of inva- tected by legislation (Gilroy et al., 2011). The values we sion ecology. Richardson et al. (2000) suggest the nu- attach to the terms we use then inform environmental ances with which terms are used in French, German, agendas (Pereyra, 2016). Evidently, it is vital to establish or other languages are not fully translated into English a clear-cut definition of “invasiveness” that can be ap- (Richardson et al., 2000). Furthermore, changes within plied to all species, while being separated from human the English language itself could also be responsible for values, and accounts for the entire invasion process. this confusion (Richardson et al., 2000). Many of the terms used to talk about invasive species were defined THE STAGE-BASED APPROACH before the complexities of the invasion process were To properly distinguish a native species from a species fully understood. For example, the term “naturalized” currently undergoing the process of invasion, an al- appeared in biological literature in the mid-19th centu- ternative definition must be proposed. This improved ry, but invasion ecology as a field only began to emerge definition must consider the process of invasion, rather a century later (Richardson et al, 2000; Elton, 1958). than focusing solely on a species’ capacity to spread and Overall, this has led to terms such as “invasive”, “na- reproduce. Thus, a stage-based definition studies the tive”, “alien”, and “naturalised” being used throughout progression of the potentially invasive species’ ability to the literature with a variety of definitions and conno- occupy a new environment. There are three fundamen- tations. Imprecise language and definitions cause many tal stages within a stage-based definition: transport, es- problems for studying and writing about invasive spe- tablishment, and spread. Within each stage, there are cies. Ultimately, it hampers comparison between stud- several factors that either inhibit or assist the species ies (Blackburn et al., 2011). By its very nature, invasion when moving from one stage to the next. In our pro- ecology requires comparing data sets from geographi- posed definition of the stage-based process of invasion, cally separate regions (Pysek et al., 2004). In order to these extrinsic and contingent factors are included. detect trends, it is important to know what species are A stage-based perspective is more beneficial as a defi- native to what region, and what species are invasive nition because it can encompass multiple pathways elsewhere (Pysek et al., 2004). If researchers do not use by which a species can invade. Due to the definition’s the same criterion when labelling their species under broader perspective, the focus is now on the ecological investigation, comparison of data becomes impossible logistics - of both the environment and the species - (Hulme and Weser, 2011). Secondly, inconsistent la- which allow for invasion to occur. Since the stage-based belling complicates communication with the general perspective looks at the process overall, the develop- Ecology public. The use of multiple different terms to describe ment of prevention techniques and policies can be cre- invasive species can lead to both the public and the ated to stop the invasion of a species, if necessary. As media misinterpreting information (Russell and Black- Jeschke et al. (2014) state, impact can occur at any stage. burn, 2017). Difficulty in interpreting data surrounding Our definition is broad enough to show how changes to invasive species can then lead to the implementation of the environment can happen, but these effects are not inadequate policy and legislation (Gilroy et al., 2011). implied to be decidedly positive or negative. Thus, it is Since the term “invasive species” is connoted with neg- not assumed that all invasive species are detrimental to ative impacts, the species thus labelled are more likely an environment. to be targets for eradication policy. Meanwhile, species Much of a species’ ability to invade is based on circum- labelled as “native” or a similar term might become pro- stance and random factors (Heger and Trepl, 2003). Our ISCIENTIST | 2018 Figure 1: Our proposed model to demonstrate the process of invasion. 48 definition looks to include contingency and variability gards transport of invasive species as any movement among species and the environment so that invasion of a species, regardless if it was accomplished through can be generalized to a process. Furthermore, looking a human-mediated vector or a non-human-mediated at invasion as a process allows investigation into the re- vector. Lockwood et al. (2007) and Sakai et al. (2001) lationship and properties of both the invading species distinguish several pathways of transport caused by hu- as well as the ecosystem itself. Invasion could be regard- mans which include but are not limited to: farming and ed as a temporally-dependent process rather than spa- hunting, environmental enhancement, biocontrol, con- tially-dependent. Overall, our proposed model builds servation, and scientific pursuits. Examples of uninten- on a more holistic perspective. tional human-mediated transport include movement of species from airplane, automobile, and boat travel THE PROPOSED INVASION PROCESS (Lockwood et al., 2007; Sakai et al., 2001). Although the Expanding from an invasion model propsed by Black- transport of invasive species is usually due to human ac- burn et al. (2011), we integrate previous versions of the tivity, natural transport can also occur, such as by wind invasion model to develop a broader stage-based defi- and bird migrations (Lockwood et al., 2007). It should nition (Figure 1). be noted that the success rate of a particular transport vector can change over time. For instance, the ability Stage 0: Origin– Most models of invasion are funda- to travel across greater distances is increasingly becom- mentally built around three main stages: transport, es- ing faster and more accessible. Additionally, transport tablishment, and spread (Blackburn et al., 2011; Lock- is its own stage because it may take multiple trials and wood et al., 2007; Colautti and MacIsaac, 2004; Heger various new habitats before an invasive population can and Trepl, 2003; Sakai et al., 2001). We have decided to settle. keep these three stages as well as add a Stage 0: Or- For survival through Stage I, there are multiple factors igin. This stage represents a species in its natural hab- to take into account in order to overcome Barrier a: Ge- itat. Although several papers have varying definitions ography. The vector itself must be sustainable to support for the meaning of a native species and its home region the invasive population. For example, a greater distance Ecology (Lockwood et al., 2007; Colautti and MacIsaac, 2004; to travel will likely result in fewer viable healthy in- Heger and Trepl, 2003; Sakai et al., 2001), the general dividuals to establish in the new region. To counter consensus is that a native habitat is wherever the in- this effect, a greater number of individuals could be vading species has travelled from. For our purposes, transported to increase the chance of survivors. This is we will regard Stage 0 as a relative region so that a referred to as propagule pressure, which is defined as change in habitat is evident. Thus, this could be the size of the invading population as it enters the new regarded as the donor region. Species survive and habitat (Blackburn et al., 2011; Lockwood et al., 2007; thrive in this first region before entering Stage I of the Colautti and MacIsaac, 2004; Sakai et al., 2001). An- invasion process. The Region numbers of our model are other factor associated with propagule pressure is the intended to quan- tify species movement as it frequency of the transport, and whether or not the in- progresses through the stag- es of invasion. To classify vasive species population is being regularly replenished a species as being in Stage 0, the species must not with healthy individuals to assist in mating, dealing inhabit more areas than those it is native to. with environmental stochasticity (unpredictable and However, once the species has commenced the stressful weather patterns), and maintaining competi- invasion process, it will move to occupy increasing- ly tive interactions with species in the novel region (Lock- more regions over time. wood et al., 2007). As the density of the invasive popu- ISCIENTIST | 2018 lation develops, it is more likely to establish. Stage I: Transport–In current literature, the first stage Overall, the greater the propagule pressure, the more of transport (Stage I) is presented in a manner that is likely a species will succeed in entering Stage II. Our saturated with human bias. One source of this issue is definition of Stage II resembles the description by that invasive species are believed to necessarily use hu- Blackburn et al. (2011), which is that establishment re- man-mediated vectors. Our version of Stage I is broad fers to a single self-sustaining population over multiple enough to include human-facilitated transport as well generations in a localized region. Interestingly, there as any other contributor to species movement that can be an intermediate period of time before an inva- might arise. We primarily utilise Vermeij’s (1996) defi- sive species becomes self-sustaining in Region 2, which nition of biotic interchange, which refers to the expan- is denoted as The Lag Period (Lockwood et al., 2007; sion of a species’ range, as our definition for transport. Sakai et al., 2001). This has been suggested to be the More simply, transport is a broad term for a portion result of exponential population growth trends or a of a species’ population leaving its original habitat temporary timeframe used to increase propagule pres- and entering a novel environment. Our definition re- 49 sure (Sakai et al., 2001). Since the time frame varies per et al., 2012; Lockwood et al., 2007; Sakai et al., 2001). species, has not been well-recorded, and would not be Without genetic diversity, a population may not have considered a barrier to overcome, it is not included in the suitable characteristics to survive in the new hab- our model as a separate section, but rather as a factor itat. This phenomenon is known as the founder effect, within Barrier b: Survival. resulting in a bottleneck population. Limited genetic variation within a population leads to inbreeding de- Stage II: Establishment–Colautti and MacIsaac (2004) pression and can limit population growth, and thus separate establishment as 1) widespread but rare or 2) decreases establishment potential (Tsutsui et al., 2000; localized but dominant. We chose not to separate es- Sakai et al., 2001). However, the funneling of genetic tablishment into two distinct types but rather focus on diversity could also be beneficial to allow for adaptive the process of establishment within invasion. Stage II is evolution. The invading species may already have rela- broken down into two barriers because success in sur- tively pre-adapted traits that allow the species to grow viving is separate from success in reproducing (Black- in the new environment (Sakai et al., 2001). In summa- burn et al., 2011; Heger and Trepl, 2003). Within Barrier ry, if the alleles that express the greatest fitness remain, b: Survival, the species must be able to survive against the invasive species is more likely to establish. If the predators as well as compete for resources among nat- alleles express unsuitable traits, then the invasive spe- ural inhabitants. Essentially, basic physiological needs cies is more likely to fail. Furthermore, hybridization of must be met. This ability to survive in the new environ- native species with invasive species can be both benefi- ment stems from the interactions between the natural cial or inhibiting (Sakai et al., 2001). The introduction region’s species and the invasive species. This includes of these new genes may lead to an increase in fitness or competitive exclusion, predation (predator-prey), para- it can lead to genetic assimilation. In all of these cases, sitism (parasite-host), herbivory (herbivore-plant), and natural selection may favour the native species instead mutualism (Lockwood et al., 2007; Sakai et al., 2001). of the invasive species and vice versa. Evidently, genetic Furthermore, r-selected species would have drastical- diversity and propagule pressure play an essential role ly different establishment and reproductive strategies in conjunction with species-species interactions to al- than k-selected species. Since factors like fecundity, low for an invasive species to establish. generation growth time, individual growth rates, and specialist versus generalist qualities depend on the par- Stage III: Spread–Once capacity of Region 2 has been ticular species, they are not the sole reason for a spe- reached or enough individuals are present in the set- Ecology cies’ ability to invade. As a result, it is more correct to tled population, the invasive population can move onto look at ecological interactions. All species-species in- Stage III with the proper transport. Barrier d: Dispersal is teractions may work in favour or against an invading similar to Stage I, but the colonization outwards from species. For instance, the invasive species may find that Region 2 tends to be more localized (Chapple et al., 2012; access to resources is easier in the new environment Sakai et al., 2001). In these newly introduced regions, due to lack of predators and similar niche-sharing spe- the previously single, self-sustaining invasive popula- cies. On the other hand, the invasive species may not tion has now increased to two or more established com- have the fitness to exploit or compete for nutritional munities. For Barrier e: Survival, this is similar to Stage II resources with current inhabitants. Moreover, mutual- in terms of the factors that allow an invading species to ism can have both positive or negative effects. A lack live in the newest habitats. Genetic diversity, morpho- of mutualism can inhibit a species’ population growth, logical traits, and reproductive rates are all key compo- while on the other hand, a newly established mutualism nents in invasion. Stage III can theoretically occur in an interaction can allow the species to thrive (Sakai et al., infinite number of new regions for an infinite amount 2001). In addition to species-species interactions, Barri- of time. In simple terms, Stage III is considered to be a er b also includes the theory of disturbance and its role range expansion of the initial invading population. in allowing a species to invade (Lockwood et al., 2007; Sakai et al., 2001). This theory explains that environ- SUMMARY mental changes, as a result of human activity or natural At any phase of the invasion process, a species can in- activity, can lead to niche modifications which allow definitely stay at a particular stage or the species can ISCIENTIST | 2018 for invading species to enter (Lockwood et al., 2007; fail to invade. The former occurs when the independent Sakai et al., 2001). species population is able to remain self-sustaining. As Within Barrier c: Reproduction, the invasive species must discussed, there are many factors that could result in be near other viable mates. This proximity impacts an the latter, which include, but are not limited to, prop- invading species’ genetic variation, which can be both agule pressure, species-species interactions, and genetic advantageous or detrimental to the population (Dick diversity. Thus, pressure from the new environment can 50 result in a lack of potential mates or inability to at- tain basic survival needs. As for the nomenclature, both stage and barrier are incorporated in a numeral-letter system. For example, a species that has established an invasive population in Region 2, but is not reproducing, would be in Stage II-b of the invasion process. NETLOGO AGENT-BASED MODEL To visualize and observe trends in the stage-based mod- el, an agent-based model was developed. The model was adapted and modified from the “Life Strategy and Inva- Figure 2: A new specialist species is introduced to sion Biology Model”, designed by Kyra Simone (2016), the environment . The amount of native species in- in NetLogo (Wilensky, 1999). Parameters based on the dividuals is shown in dark green , the amount of interaction between a native ash borer species and an invasive species individuals is shown in light green, invasive ash borer species were set to demonstrate their and the total energy of all ash trees , that both roles within the invasion process. species eat, is displayed in medium green. The initial population size of the native and invasive Next, an environment was created where the new spe- specialist demonstrates how the population size of the cialist and the native specialist were similarly matched native species will affect the fitness of the invasive spe- in terms of fitness (Figure 3). In this case, neither of cies to progress through the model, as well as the effect the species died out and both appear to reach relatively of propagule pressure upon the native species. stable population sizes. The energy consumption of both species can be altered, helping to demonstrate how an organism’s ability to consume at a more efficient rate relative to another can provide a distinct advantage for population establish- Ec ment and reproduction. ology The minimum reproductive energy required of both the native and invasive borer species can be adjusted to demonstrate how high fecundity levels have signifi- cant impacts on propagation towards the surrounding region and throughout an environment. The maximum number of offspring can be altered to simulate a species with a high fecundity rate and to aid in understanding its implications on the population Figure 3: Invasion where the native species and the invasive propagules are equal. The amount of native dynamic in both the native and invasive borer species. species individuals is shown in dark green , the The final alterable parameter is the density of ash trees amount of invasive species individuals is shown in present within the environment. This demonstrates light green, and the total energy of all ash trees, that how a larger availability of hosts can significantly de- both species eat, is displayed in medium green. termine the ability of an ecosystem to support a native species as well as an invasive species simultaneously, or A third simulation was performed where the propa- if the resources available will only support one species. gule pressure of the new specialist was low (Figure 4). ISCIENTIST | 2018 Using these parameters, we hope to demonstrate the ef- In this scenario, the new specialist died out while the fect that these factors will have on the invasion process native specialist population size remained relatively and how they will affect the progression of an invasive unchanged. species through the stage-based model. Finally, an invasion scenario was created where the new specialist was not as well adapted to its new environ- RESULTS ment as it has a reduced ability to extract energy from Using the NetLogo code, various invasion situations its food source (Figure 5). In this situation, the size of were simulated. First, the code was used to re-create a the new specialist population initially increases, then situation with an abundance of food for the invading drops back down to disappearance. species, low competition, high reproductive ability, and high propagule pressure (Figure 2). When the new spe- DISCUSSION cialist was introduced, its frequency increased rapidly With variation in starting conditions, it is possible to while the native specialist died out. demonstrate how species in different situations will fit 51 the definition of our stage-based model with diversecresults. In a situation where an invasion is favoured, the invad- ing species has a high fecundity rate, an abundance of food, few competitors and a large number of invading propagules that reach the new environment. As seen in Figure 2, the initial spike in invasive species individ- uals marks the introduction and beginning of estab- lishment. Consequently, the invasive species will thrive and its numbers will continually rise, characterising it as a species in Stage III-e. In a scenario where the native species and the invasive Figure 5: Failed invasion where the invading species propagules are evenly matched in numbers, ability to is not well-adapted for the new region. The amount of native species individuals is shown in darl green, acquire nutrition, and reproductive ability, the arrival the amount of invasive species individuals is shown of the invader modifies the environmental steady-state in light green , and the total energy of all ash trees , conditions and a new equilibrium is reached (Figure 3). that both species eat, is displayed in medium green. As evidenced by the relative steady population size in any nutrition from their environment. The species is both native and invasive specialists, it appears that the not able to cross the barrier of survival and the invasion environment is able to sustain this equilibrium. The in- fails. vader is able to establish itself and sustain its popula- Due to the number of combinations that can be created tion, however its numbers do not increase. As such, it and simulated using the slider functions of the Netlogo would be labelled as a Stage II-c species. model, all results were not acknowledged. The combi- nations that were highlighted include two instances of a successful invasion and two examples in which in- vasion fails. Successful invasion is apparent when the invader is able to integrate a population into the new environment, whether or not the native borer species is affected. When the invasion process fails and the invad- ing borer species is not able to establish a population, Ecology this is characteristic of invasion failure. CONCLUSION Ultimately, it is suggested that previous invasion vo- Figure 4: Failed invasion where the initial propagule pressure is relatively low. The amount of native spe- cabulary should become umbrella terms, and that our cies individuals is shown in dark green, the amount stage-based, numeral and letter system be used to spe- of invasive species individuals is shown in light cifically refer to species in the invasion process. Our green and the total energy of all ash trees, that both proposed novel definition is an improvement over ex- species eat, is displayed in medium green. isting definitions in several ways. First, it demonstrates minimal human bias, as it does not rely on human A third situation that may occur is when the potential perception of supposedly damaging or harmful im- invader fails (Figure 4). In this case, the initial number pacts. Further, it does not depend on arbitrary values of propagules that reach Region 2 is too low, and even of distance or spread. Third, our model focuses on the though both species are equally able to feed themselves plant-animal-environment interactions, allowing for a and reproduce, the invading species is not able to cross holistic approach to defining the invasion process. Fi- the barriers of survival and reproduction. nally, it can be can be simply applied to a wide range of Similarly, invasion failure will also be induced if the plant and animal species. Our NetLogo model provides invading species is ill-adapted to its new environment a user-friendly reference point for understanding how ISCIENTIST | 2018 (Figure 5). In this scenario, the invasive species is not to apply our definition. If this stage-based perspective able to get sufficient nutrition once it is transported is adopted into the literature, communication within to the second region. Although the initial number of scientific communities as well as conversation with the invasive propagules is almost double the initial amount general public will be more readily understandable. of native specialists and they have equivalent reproduc- Standardizing the language used when communicating tive capabilities, they have a lower likelihood to obtain with and across both groups will minimize confusion 52 and reduce the risk of mislabelling species. Clear lan- AUTHOR CONTRIBUTIONS guage will allow for better legislation and prevention P.B. conducted a literature review on the existing lan- policies to be developed, as well as more effective out- guage used in invasion ecology for the introduction and reach and education campaigns. Ideally, this will lead assisted in observing and analyzing the results from the to greater ecological awareness in the community and a stage-based invasion simulation. T.D. conducted the greater effort towards environmental stewardship. research for several proposed stage-based approaches for invasive species. Afterwards, they were the main ACKNOWLEDGMENTS author to integrate these approaches to be used for our We would like to thank Dr. Chad Harvey for his guid- novel definition. A.T. developed the NetLogo modelling ance and support during this project. We would also code for the stage-based invasion simulation as well as like to thank Andrew Colgoni for his insight and edit- helped in writing the overall analysis and discussion of ing, and Dr. Sarah Symons for her NetLogo expertise. the outputs with P.B. All authors contributed in devel- oping the proposed definition and were responsible for editing and revising one another’s work. Pyšek, P., Roques, A. and Vilà, M., 2016. Blurring alien Sakai, A.K., Allendorf, F.W., Holt, J.S., Lodge, D.M., REFERENCES introduction pathways risks losing the focus on in- Molofsky, J., With, K.A., Baughman, S., Cabin, R.J., Blackburn, T., Pyšek, P., Bacher, S., Carlton, J., vasive species policy. Conservation Letters. Cohen, J.E. and Ellstrand, N.C., 2001. The population Duncan, R., Jarošík, V., Wilson, J. and Richardson, D., biology of invasive species. Annual review of ecology and Hulme, P. and Weser, C., 2011. Mixed messages from 2011. 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Available at: minology to define ‘invasive’ species. Diversity and Dis- Ecology invadingspecies.com/invaders/> [Accessed 5 Apr. 2017]. T.J., 2000. Reduced genetic variation and the success of tributions, 10(2), pp.135–141. an invasive species. Proceedings of the National Academy NOBANIS, n.d. Definitions used by NOBANIS. [online] Dick, J.T.A., Alexander, M.E., Ricciardi, A., Laverty, of Sciences, 97(11), pp.5948–5953. NOBANIS. Available at: 53 54 ISCIENTIST | 2018 Ecology ORIGINAL RESEARCH INTRODUCTION Integrated Science Program, Classof 2019,McMaster University ADELMAN, AUDREY TAM MONICA TAKAHASHI, PASCALE BIDER,ARIANAFRASCHETTI, CAMILLA GILLIS- crease insodiumconcentration hasled to groundwater in the that shown have studies Moreover, and 2003). Buff, Hafner (Godwin, America North in freshwater ly increases theconcentrations ofsodiumin bodiesof when used to de-ice roads and sidewalks, it significant Road saltisusuallymadeupofsodiumchloride,and way tomeltsnow andice duringthewintermonths. efficient and cheap a is it since Canada in pollutants Road saltisoneofthemostwidely-used environmental persicae The effects of water salinityon ogy Keywords URL: Received: salinization, plantswillhave to face thedoubleburdenof saltstressandherbivory. byaffected be to appear not do aphids Sincethe solution. saline bythe affected been have not may aphids the by preferred plant the of regions the Secondly, aphids. the plant may have allocated itsresources towards rather survival thandefence against the Firstly, reasons. main two to due be could aphids the on effect of lack fitness. The the low and high salt treatments, suggesting that salt did have some effect on plant between found was significance However,statistical treatments. salt low and control the or treatments salt high the and differencesignificant control the between no was on the aphid population growth, but did significantly affect the leaf surface area. There on the course of the experiment andanalyzed to determine theeffect of salinized water tion. Rosette leaf surface area and aphid population were recorded for each plant over watered with either regular tap water, a 20 mM saline solution, or a 60 mM saline solu sicae saline solutionswould affect thehealthandgrowth of theplantsaswell asthe of concentrations different with plants the watering that hypothesized was It ment. and type Wild them. on feed that herbivores the and plants ecosystem.rounding The experimentaffectshowthis ofsalinity study was topurpose detri has mental effects onnotonlytheorganismsthatfeedThis ontheseplants,butalsothesur health. plant impact can which salinization, soil to lead containing can salt Runoff road issues. environmental serious some poses also it However, snow. efficient and way melt cheap to a as months winter the during used often is salt Road SUMMARY A. thaliana , Myzus persicae plant https://theiscientist.com/2018/03/15/TakahashiEffects.html populations. Wild type Wild populations. - 11/04/2017 : animal s a l inity and and interactions , Arabidopsis M. persicae (green peach aphid) were the model organisms used in the experi Accepted: Arabidopsis thaliana A. thaliana . The saline treatments did not havesignificant not Theeffecta did . treatments saline thaliana 03/15/2018 , Myzus plants were inoculated with - - be less apparent. ence ontherest oftheecosystem and otherspecies may may cause noticeable changes in plant health, any influ it. While the negative effects of sodium on plant species plants, butalsoontheecosystem thattheplants inhab the impacts high salinity would have on not only the study to important is it Consequently, 2002). Barker, ity touptake water and photosynthesize (Bryson and mental effects to plant health, affecting the plant’s abil Alexander, 1998). Increasing soil salinity also has detri contamination and plant toxicity issues (Forman and persicae Published: , soil Arabidopsis thaliana , road Myzus 03/15/2018 salt , de M. persicae - icing , ecol M. per plants and ------ISCIENTIST | 2018 Ecology 55 - - - ro- the random random vel Low Low leaves. Ran leaves. High High None None None in Hamilton. on their lack A. thaliana thaliana A. (Table 1). The Salt Le based done using a fficient, four-number a e Greenhouse healthy (green peach aphid) that had that aphid) peach (green more 1-4 5-8 9-12 f Numbers and Sabin, 2009). o make 13-16 21-24 e used to facilitate the watering to facilitate the watering e used 17-20 25-28 or tobacco thaliana or tobacco A. same manner. out of a selection of plants grown of plants grown selection out of a TMENT SET-UP deemed process Range o Arabidopsis thaliana plants and their M. persicae chosen on either were were F E C B G A D was chosen for each plant. For example, numbers example, numbers plant. For for each chosen was had not begun the bolting stage or had bolts that or had bolts the bolting stage had not begun opipette tips wer random number table was used again to determine to again used was table number random Plant discolouration or torn leaves. All chosen plants ei or torn leaves. discolouration ese plants were corresponding random table number ranges and ranges table number random corresponding The 1: The Table or the experiment, 24 healthy wild type type wild healthy 24 experiment, the or treatments. Although only plants A-G are shown in are shown treatments. Although only plants A-G the table, the remainder of the plants were assigned the plants were the table, the remainder of a letter, a four-number range, and a treatment in the range, a four-number a letter, one to four corresponded to plant A, numbers five to eight corresponded to plant B, etc. settes in the McMaster University Th of ther had been previously Theremoved. study used 96 vivip- female arous reared been was treatments domization of the number table (Petrie range treat to assign then used was number table random ments to plants in the systematic pattern of “no salt”, “low salt”, and “high salt”, resulting in eight plants per treatment. randomization Micr SALINE TREA SALINE TERIALS AND METHODS AND TERIALS MA DESIGN EXPERIMENTAL F of the plants. 96 tips were retrieved and cut at the - sec ond line from the bottom so that each tip was 4.5 cm long. This allowed water travel to through at a fast- This the placement of the plants in the two trays. The se- quence of two-digit numbers in the table dictated the Plants in the tray. placed in which plants were order were placed one at a time 1, into firstTray filling up a there until the next to on going then and of three row were four rows of three plants each. 2 Tray was filled in the same manner (Figure 1). in Plants were arranged a staggered fashion to minimize thecontact between of aphids migrating the chance plants and to reduce plants. between . - - - Myzus per- A. thaliana A. thaliana and d through both d through egulate and one of its predators, and one of its predators, A. water thaliana plants with salinized , more commonly known as the green peach aphid peach known as the green commonly , more sicae - follow the answer to order in conducted was study The ing set of questions: of (1) is rate the growth does how (2) and salinity, soil of increase an by affected the size and growth of salinity affect the concentration on A. thaliana? populations feeding of the M. persicae the Watering and a plant growth decreased to cause predicted was reasons. in aphid population size for several reduction the cessa in Continual high salinity in plants results in the expansion, a decrease surface tion of plant leaf (Parida of photosynthesis, and plant death productivity and Das, 2005). Thiscauses aphid populations, which rely on the phloem sap of plants for nutrition (Gior- danengo, et al., 2010), to be left without a food source. plant me high salinity in plants increases Additionally, (Boughalleb, Denden sophyll and epidermal thickness and Tiba, 2009). This may make it more difficult for the aphids to pierce the leaves for food (Giordanengo, et al., 2010). By observing the effect of saline soil on M.understand better to aims study present the , persicae the effects ofroad de-icing on primary producers and the organisms that feed on them. The the future for important implications have resultsstudy may of the and around Cootes Paradise both around of ecosystems the world. tions would be sure to have repercussions elsewhere in elsewhere repercussions to have tions would be sure the food it chain. isTherefore, important to determine whether road salt influences plant growth and the ef- this has on herbivores. fect designed was experiment 13-day a issue, this address To the plant species between to observe the interactions thaliana Arabidopsis top-down top-down and bottom-up interactions, the influence in the A discussion population. in each varies of either structure that community argues Naturalist American ev- meaning bottom-up interactions, by is controlled ery trophic level is “food-limited” and (Hairston, Slobodkin, Smith 1960). It is also argued that interactions, by top-down is regulated nity structure - commu size level limits the population trophic each whereby of the trophic level below it (Hillebrand, et al., 2007). In a community that operates via bottom-up - interac negatively likely in soil salinity would tions, an increase which depends upon of the ecosystem, impact the rest the affected plant species for energy. The influence of the salt would be seen first within the populations such as herbivore and then within primaryducers pro- popula Any impact on the size of these insect insects. Although population size is r population Although Tray 1 G W K Q O B S I H U R M Tray 2 A X D L E V T J C F N P Figure 1: Plant treatments (red - no salt, green - low salt, blue - high salt) and tray placement. er rate. Four of the modified micropipette tips were placed near the edge of the container in the middle of each side. They were then pushed in at an angle approx- imately 65° from the horizontal into the soil until their tops aligned with the surface (Figure 2). Figure 2: A top-view image of a plant that was taken The plants were watered with a 20 mM salt solution, a using the quadripod. The four circles at the north, 60 mM salt solution, or regular tap water depending east, south, and west sides of the plant pot are the micropipette tips that were inserted at an angle of on their treatment group. Two 500 mL solutions were approximately 65° and used to water the plant. created for each concentration so that plants could be watered with the same solution each time. Tap water of aphids for all 24 plants. Each plant was put on the Ecology was used instead of distilled water to ensure that the lab bench after inoculation to avoid mixing up inoc- plants still received some of the minerals that are found ulated and uninoculated plants. After every plant had in tap water. received two aphids, a second round of inoculation oc- The plants were watered one tray at a time. A micro- curred, whereby each plant was populated in the same pipette was used to fill each of the four modified -mi way with two more aphids. In the end, each plant had a cropipette tips with tap water in all the plants in the total of four aphids. “no salt” treatment group (i.e. the control plants) on the first tray. Each of the 4 micropipette tips were first DATA COLLECTION filled with 1 mL of water, adding up to a total of 4 mL Front-view and top-view photos were taken of each per plant. Immediately after, the same plants were wa- plant for comparison throughout the study. A quadri- tered with another 4 mL of tap water, resulting in a pod was built out of a cardboard box in order to ensure final volume of 8 mL of tap water in each control plant. that the top-view photo of every plant was taken from The water was allowed to sink into the soil through the the exact same position, height and angle each time micropipette tips while the plants in Tray 2 were being (Figure 3). Several holes were cut out of the box to allow watered in the same manner. This process was repeated the maximum amount of light to reach the plant. An one more time, so that in the end, each control plant outline was drawn on the top of the box to ensure that had received a total of 16 mL of tap water. The process the photography device (an iPhone 6) was placed in was repeated in its entirety using the 20 mM and the 60 the same location for each photo. A sheet of blank pa- mM salt solutions for the plants in the “low salt” and per with an outline of the bottom of the plant pot was ISCIENTIST | 2018 “high salt” treatments, respectively. attached to the legs of the quadripod to aid with the placement of the plant. A small piece of cardboard was INOCULATION OF M. PERSICAE used to make a frame with a white background to give Every plant was first inoculated with two aphids using sufficient contrast for the pictures. A ruler was taped a metallic probe and a magnifying glass. This number to the cardboard to provide a scale for pictures. The was chosen to ensure that there was a sufficient number photos of all the plants were taken in the exact same 56 location each time to ensure that there were similar on the A. thaliana. amounts of lighting. Any significant results (p < 0.05) from the two-way The two trays of plants were placed by the window so ANOVA analysis were further explored using a post- that each tray received equal amounts of light. They hoc Tukey’s Honest Significant Difference (HSD) test were put beside each other to ensure that all plants to compare the means of the different treatment groups. were growing in the same conditions. The trays were covered with a plastic lid to protect the plants from disturbance or contamination and to prevent aphids from escaping. The number of aphids on each plant was counted and recorded in a master data sheet six times throughout the experiment. Each time, top- and front-view photos of every plant were taken and observations regarding plant health were recorded. The plants were watered every four days, with the exception of the seventh day, when the plants were watered due to limitations with the laboratory. The plants were watered with their re- spective solutions using the same technique that was used for the first round of watering. STATISTICAL ANALYSIS The data collected from the study was analyzed using R. Figure 4: Aphid population change throughout A lattice add-on package was installed to enable better time on plants watered with normal water, with a visualization of the data. A scatter plot of aphid count low-concentration salt solution, and with a high-con- and date with treatment as a conditioning variable was centration salt solution. Ecology created using the lattice add-on. This allowed for better RESULTS Figure 3: The cardboard quadripod for taking top- view images of the plants (the ruler is not present in the photo). The square frame was used to provide a APHID POPULATION CHANGE white background to enable better visualization of Within the different treatments, there were no notice- the rosette leaves. The plant pot was placed in the outlined area on the sheet of paper and the iPhone able trends with respect to aphid population growth. 6 was positioned using the outline on the top of the Both of the treatments and the control saw the aphid quadripod. population increase with time, and there was no no- ticeable difference regarding the manner in which the visualization and analysis of the data as the effect of populations increased (Figure 4). This suggests that the treatment was separated from date. A two-way analysis salt treatments did not have an impact on the aphid of variance (ANOVA) test was conducted to see wheth- population growth. This is also evident when the aver- er treatment and date had a significant effect on aphid age aphid population count is compared within each of population growth and whether there was a significant interaction effect from the two variables. This interac- tion effect would indicate that variance in aphid pop- ISCIENTIST | 2018 ulations between treatments depended on the number of days elapsed. The aerial photos of the plants were analyzed using an image processing software, ImageJ. Using the software, the surface area of the exposed rosette leaves (in cm2) of each plant was calculated for every observation day. This value was used as an estimate of total plant leaf surface area. The scale of the program was calibrated us- ing the ruler in the plant image. A boundary was drawn by hand around the plant leaves to accurately select the Figure 5: The average population of aphids on plants area for measurement. This data was analyzed using the watered with tap water, a low-concentration salt same statistical method (a two-way ANOVA) as the solution and a high-concentration salt solution. The error bars represent ±SE. aphid population data to see if treatments had an effect 57 Figure 6: Average plant surface area change through- out time of plants watered with a high-concentration saline solution (60 mM), a low-concentration saline solution (20 mM), and tap water. (A) shows the rela- tionship between the “high salt” treatment and the control; (B) shows the relationship between the “low salt” treatment and the control. (C) shows the rela- tionship between the “high salt” treatment and the “low salt” treatment. The Tukey HSD test indicated that there was a significant difference between the high and the low treatments (p = 5.276 x 10-4). All the error bars were calculated by measuring the stan- dard error of the data points. the three treatments (Figure 5). significant (p = 3.27 x 10-4). It should be noted that time A two-way ANOVA test was conducted on the rela- also had a significant effect on the plant surface areap ( tionship between the aphid population growth and = 2.45 x 10-5). However, time and treatment affected the both the treatment and the date. The relationship be- plants independently of each other (p = 0.856). Thus, tween aphid population growth and the treatment was the manner in which the date affected the surface area deemed insignificant p( = 0.76). The only variable that change did not impact the way in which the treatment was significant in relation to aphid population growth Ecology affected the surface area change. was the date (p < 0.001). In other words, the different To determine which treatment had an effect on the salt solution used to water the plants did not have any plant surface area change, a Tukey HSD test was run impact on the growth of aphid populations. (Table 2). The test revealed that there was a significant difference between the high and the low treatments p( PLANT SURFACE AREA CHANGE = 5.276 x 10-4). Interestingly, it also revealed that there Using ImageJ, the surface area of the exposed rosette was no significant difference between the control and leaves of each plant was measured for each of the days the high salt treatment or the control and the low salt that they were surveyed. Each plant began with a treatment (p = 0.1884 and p = 0.09536, respectively). unique and distinct surface area. In order to account To determine how the distinct treatments affected the for this difference, the surface area change of each plant plant surface area growth with respect to time, the through time was compared. Initially, it was difficult to change in surface area of all plants for each treatment notice any trends in the data collected. As well, each was averaged for each day that data was collected. The plant’s growth and shrinkage pattern was unique, mak- averages of the variable interactions in Table 2 were ing visual analysis of the data impossible. graphed against each other (Figure 6). The plant surface Due to the three distinct variables (high salt, low salt, area of the plants in the high salt treatment group de- and no salt), a two-way ANOVA was used. The relation- creased the most throughout time; however, all of the ship between surface area change and treatment was treatments had a net negative impact on plant growth Table 2: Result of a Tukey Honest Significant Difference test with a 95% family-wise confidence ISCIENTIST | 2018 level analyzing the differences between the impact of the salt treatments on Arabidopsis thaliana plants. Variable Interactions Difference Lower bound Upper Bound Adjusted p-value Low - High 2.497 0.9595 4.035 0.0005276 No - High 1.140 -0.3980 2.678 0.1884 No - Low -1.358 -2.895 0.1804 0.09536 58 over time. There was no significant difference between ent ways. Its constitutive defences include trichomes the control plants and the low salt treatment group or and toxic antibiotics in its phloem (Louis and Shah, between the control and the high salt treatment (Figure 2013). The plant also makes use of inductive defences 6A and 6B). The significant difference between the low such as increasing starch and glucosinolate concen- and high treatments is visible since the error bars of tration when attacked, or inducing premature death the graph do not overlap (Figure 6C). In other words, of leaves on which M. persicae colonies are established the three graphs support the results obtained using the (Louis and Shah, 2013). While effective, these induc- Tukey HSD test. tive mechanisms of defence are costly to the plant. It is possible that, when under stress from increased salt DISCUSSION concentration, the A. thaliana would preferentially al- locate resources to survival rather than defence. If this These results give some insight into the impact of win- were the case, M. persicae would not be impeded in its ter road salting on both plant and aphid populations. feeding and the colonies would thrive. Another possi- First, this study suggests that varying saline concentra- bility is that although the leaf surface area decreased in tions in the soil have an impact on A. thaliana growth. the high-salinity treatment, this did not have a popu- The results of this experiment did not entirely confirm lation-limiting effect on the aphids. During the study, the initial hypothesis because the control group did not the aphids seemed to aggregate preferentially on the differ from either treatment groups. Had the hypothe- stems and bolts, so perhaps a slight reduction in leaf sis been entirely accurate, the decrease in plant surface area would not have caused a measurable impact on area would have been significantly greater in the high aphid habitat. salinity plants than in the control, which was not the These findings carry some heavy implications with re- case. However, the results of this study did indicate gards to the use of salts for road de-icing. Naturally that increasing the salt concentration from 20 mM to high levels of soil salinity already impede plant growth 60 mM negatively impacted plant leaf area. Salted soils in several locations around the world, and further salt are known to impede the growth of many plant spe- addition through road de-icing only exacerbates the Ecology cies (Qados, 2011). Increasing salinity has been shown problem (Qados, 2011). By some estimates, salinization to decrease water uptake by the roots, alter leaf osmotic of arable land will have caused a 50% loss of agricultural potential, modify protein content, increase sodium and space by 2050 (Heidari, et al., 2011). Furthermore, stud- chloride ions in the chloroplasts, reduce the quantity ies have shown that some plants are even more vulner- of photosynthetic pigments present in the leaves, and able to high salt concentrations when under drought inhibit photosynthesis (Ueda, Tsutsumi and Fujimo- conditions (Ha, et al., 2008). Given the water shortages to, 2016; Heidari, et al., 2011; Qados, 2011; Gama, et al., currently devastating several regions of the world, the 2009). This, in turn, can have a negative effect on leaf safekeeping of our agricultural lands is crucial. Addi- number, plant dry mass, and leaf area (Qados, 2011). tionally, if herbivores such as M. persicae are unaffect- Additionally, exposure to salt stiffens the epidermal ed by the increase in salinity, plants will have to face cell wall of the plant (Zörb, et al., 2015). The present the double burden of salinization and predation. This study indicates that A. thaliana may be susceptible to could have dire consequences on food production as salt stress and that exposure to salted soils might nega- well as on fragile ecosystems such as Hamilton’s Cootes tively impact its fitness. Paradise. Contrary to A. thaliana, however, the M. persicae popu- In light of these potential consequences, it seems vi- lation was in no way affected by increased salinity. This tal to continue research in this direction. The present ISCIENTIST | 2018 rejects the hypothesis that was initially postulated. M. study had some limitations that future research could persicae feeds by sucking the phloem of the plant (Dut- address. In this experiment, data was collected by man- ton, et al., 2002). If the salinity of the soil had caused a ually counting the live aphids, which may have allowed change in phloem chemical composition or availabili- for some errors during the count. In future studies, a ty, it should have negatively impacted the population more accurate way of assessing aphid population growth growth of M. persicae. Additionally, since salt causes should be employed. This could include the use of mi- plants to thicken and stiffen their epidermis (Zörb, et croscopes as well as a greater number of independent al., 2015), it could have been possible that the aphids counts of each plant. As well, ImageJ, the program used were unable to pierce it with their stylets and therefore to calculate rosette leaf surface area, has limitations as died from lack of food. However, no such effect was it can only analyze images two-dimensionally. This did observed. not account for the fact that some leaves naturally lie There are a few potential reasons why the aphid pop- more flat while others grow in an upward direction. In ulation was not affected by the increase in salinity. A. future studies, a more accurate technique should be em- thaliana defends itself from aphids in multiple differ- 59 ployed. Furthermore, the potted plants were kept very It should be noted that, although no effect on herbi- close to each other over the course of this experiment, vores was observed, future research should be done to which may have allowed for migration of the aphids determine whether this is true for other herbivores and from one plant to another. It is also worth noting that plant types. Overall, the negative impact that salt had the plants did not all start off at the same stage of on A. thaliana in this experiment provides further jus- growth. Some were already bolting, while others were tification as to why an alternative method for de-icing still in the rosette stage. In an effort to normalize the should be researched, as protection from salinization is growth stage, the bolts were cut off of the more mature crucial for ecosystem preservation. plants. However, this may have further impacted their growth. Additionally, the saline solutions used to water ACKNOWLEDGMENTS the plants were of relatively low salt concentrations. A. The researchers would like to thank Dr. Susan Dudley thaliana has been known to survive at saline concentra- (McMaster University), Sebastian Irazuzta (McMaster tions of 100 mM (Ueda, Tsutsumi and Fujimoto, 2016). University), and Russ Ellis (McMaster University) for It is possible that repeating this experiment with high- their guidance and assistance. McMaster University er salt levels would have a greater impact on the plants, Greenhouse is acknowledged for providing the A. thali- thus yielding clearer results. Finally, and perhaps most ana and M. persicae. importantly, the experiment only lasted for a duration of 13 days. While this allows us to gain some informa- tion on the effect of salinization on aphid populations, AUTHOR CONTRIBUTIONS the salt levels that accumulated in the soil over 13 days All authors contributed equally towards the gathering are not entirely comparable to a season of road salt of background research, designing of the experiment, build-up. Future studies could investigate this system collection of data, and editing of the final manuscript. over a greater time period. In terms of composing the manuscript, M.T. and A.T. wrote the abstract and A.F. focused on the introduc- CONCLUSION tion. M.G-A. and A.T. wrote the methods, while M.T. performed the statistical analyses, wrote the results, This experiment demonstrated the negative effects and produced the figures. P.B. focused on the discussion salt has on plant growth and health. The aphid popu- and A.F. wrote the conclusion. lation sizes were not significantly correlated with in- creased salinity levels, but the leaf surface area of the Ecology A. thaliana decreased with increased saline treatments. interactions: How aphids manipulate plant responses. pp.10904-10909. REFERENCES Comptes Rendus Biologies, 333(6-7), pp.516-523. Louis, J. and Shah, J., 2013. Arabidopsis thaliana— Bryson, G. and Barker, A., 2002. Sodium accumula- Godwin, K.S., Hafner, S.D. and Buff, M.F., 2003. Long- Myzus persicae interaction: shaping the understanding tion in soils and plants along Massachusetts roadsides. term trends in sodium and chloride in the Mohawk of plant defense against phloem-feeding aphids. 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M. and Bonnemain, J., 2010. Compatible plant-aphid Proceedings of the National Academy of Sciences, 104(26), 60 Ecology ISCIENTIST | 2018 61 (Lou- A. thaliana resource allocation, allocation, resource fertilizer, fertilizer, ORIGINAL RESEARCH ORIGINAL Published: 03/22/2018 Y SYSTEM abidopsis is a weed thaliana, known as thale cress, stronger defenses against herbivory (Sohlenkamp et al., defenses against stronger 2002). Thus, it iscommonly used in fertilizers.Despite its benefits, nitrogen fertilizercan alsocause - environ - surround to crops from runoff through damage mental ing biota (Liu et al., 2014). This affects plantgrowth, alter plant-animal dynamics. eventually and can Ar STUD commonly used as a model organism due to its small as a model organism used commonly short life span (Koornneef, and relatively genome Alonso-Blanco and 2004). Vreugdenhil, Myzus , persicae green as the known commonly organism model another on the phloem that feeds is a generalist aphid, peach of plants, including of a wide variety is, Singh, and Shah, 2012). In general, plants employ a defenses to deter and mechanical number of chemical , aphid, nitrogen, soil, ccepted: 03/22/2018 ccepted: A thaliana, M. persicae A. Y al elements serve as the building blocks of ecosystems, and cycle through the BalintTesting.html https://theiscientist.com/2018/03/22/ are the basis of ecosystems, and changes in plant in changes and ecosystems, of basis the are egrated Science Program, Class of 2019, McMaster University McMaster 2019, Class of Program, Science egrated esting the effects of nitrogen on the on the nitrogen of effects the esting SUMMAR Keywords: plant-animal interactions URL: Natur 01/21/2018 Received: biosphere. Nitrogen is one of the most important elements for plant growth. To further To growth. plant for elements important most the of one is Nitrogen biosphere. increase nitrogen plant is fertilizer,growth, as though artificially addedto ecosystems it may put nearby organisms at risk. The impact of fertilizer runoff affects many - envi ronments and the organisms that inhabit them. For these reasons, it is important interactions. plant-animal on nitrogen to of amounts increased of effects the understand To do so, we studied the effect thali- of Arabidopsis varyingbetween interaction ammonium the on nitrate fertilizer, in (AN) found commonly concentrations, compound a with water, treated group A. thaliana plants were control The ana and Myzus persicae. while low and high dose groups were treated with varying concentrations of AN. We counted the number of M. persicae present on each A. thaliana plant throughout the study period. low dose The group begins to plateau after the sixth day, while the - con affects content nitrogen soil that suggest results These grew. groups dose high and trol optimal trophic plants between treatment interactions and was a The herbivores. low dose of AN, as population of growth M. plateaued, persicae limiting herbivory and po- A. thaliana. benefiting tentially growth growth have a substantial effect on (Maron saidand Crone, ecosystems 2006). Therapidly increasing hu- demand for to an increased man population has led plants used for food, oil, and energy (PRB, 2016). heavily rely fuel this demand, agriculturalists to order In or ammonium nitrogen, on fertilizers such as inorganic by plants, as it absorbed (AN). AN is readily nitrate abundant be held by the can that charge has a positive ions in charged negatively soil (Liu et al., 2014; UCAR, 2016). Nitrogen is an essential nutrient for plants due and plant structure. to its use in metabolic processes give plants levels of nitrogen By extension, increased Plants NITROGEN AND PLANTS NITROGEN INTRODUCTION LIZ BALINT, LEENA ALSHENAIBER, NOEL KIM, ISOBEL SHARPE, MEGAN SWING KIM, ISOBEL SHARPE, MEGAN ALSHENAIBER, NOEL LIZ BALINT, LEENA Int T and A. thaliana and M. persicae of interaction this sort of herbivory. In our study system, A. thaliana ulation of M. persicae on A. thaliana to infer the poten- has several defenses against the phloem-sucking herbiv- tial effects of runoff water contaminated with nitrogen ory of M. persicae (Kuśnierczyk et al., 2008). fertilizer on plant growth. We hypothesize that the M. persicae population will initially increase with higher PLANT DEFENSE concentrations of nitrogenous compounds available in the soil. Over time, we predict that A. thaliana will use A. thaliana has a variety of characteristics used to defend the added nitrogen in the soil to strengthen its defenses itself against the herbivory of M. persicae. Defensive re- against the increasing rates of herbivory. sponses in the plant are elicited by the saliva secreted by M. persicae when sucking on the phloem of A. thaliana, MATERIALS AND METHODS as well as the penetration of the phloem cell wall itself (Miles, 1999; Kuśnierczyk et al., 2008). Components of EXPERIMENTAL SETUP M. persicae saliva lead to the accumulation of jasmon- ic acid in the plant, a compound known to transduce This experiment was conducted over a 13-day period in signals of attack throughout the plant (Kuśnierczyk et a laboratory at McMaster University in Hamilton, On- al., 2008). Salicylic acid signalling pathways are also in- tario during early autumn. 24 wild type (Col) A. thali- duced during herbivory, although jasmonic acid path- ana plants were obtained in the rosette stage of growth. ways are more efficient. Due to antagonistic crosstalk The plants received sunlight from a north-facing win- between the two signalling pathways, M. persicae can dow and were kept at room temperature. induce salicylic acid signalling responses, thereby sup- pressing jasmonic acid signalling (Wees et al., 2000; De TREATMENT AND INOCULATION Vos, Kim and Jander, 2007). During herbivore feeding, Ca2+ membrane channels also open, allowing areas of TheA. thaliana plants were randomly divided into three the sieve element membrane that were broken by M. groups of eight: control, low dose, and high dose. Two persicae to be plugged by proteins (Will and van Bell, concentrations of AN were created by dissolving solid 2006; Kuśnierczyk et al., 2008). M. persicae then attempt crystal AN in water, with a concentration of 60 ppm to prevent these defensive responses by injecting Ca2+ for the low dose group and 300 ppm for the high dose binding proteins into the sieve element membranes, group, which are realistic fertilizer runoff concentra- ology which stops A. thaliana’s proteins from repairing mem- tions (Kim et al., 1986). Next, 12 mL of distilled water Ec brane damage. was added to the control plants, and 12 mL of the low and high dose AN solutions were added to the respec- OUR STUDY tive plants, using a pie pump. The AN was dissolved in water to allow for a faster uptake of nitrogen from soil The interaction between nitrogen fertilizers and vari- to plant. The solution was added to the control group ous plants has been studied, demonstrating that nitro- first, then to the low group, and lastly to the high group gen addition to the soil affects some aspects of plant in order to avoid contamination of solutions by the pie growth and development, but not others (Chechin pump. Using a teaser needle, each A. thaliana was inoc- and Fumis, 2004; Houle and Moore, 2008). Addition- ulated with four M. persicae. This is a suitable number ally, the interactions between M. persicae and A. thali- of M. persicae; even if one or two insects do not survive, ana have been studied extensively (De Vos and Jander, there is still opportunity for population growth. The 2009; Kim and Jander, 2007). ). Beyond this, the impact plants were labelled and placed into three large plastic of M. persicae on nitrogen metabolism in plants such as trays which were covered with clear plastic lids to avoid alfalfa has been explored by Girousse et al., 2005, who external contamination and the transfer of M. persicae. found that M. persicae infestation led to reduced nitro- gen content and associated negative nitrogen deposi- DATA COLLECTION tion rates. Altered nitrogen availability influences the magnitude of response to herbivory by altering the Data was collected on days 4, 5, 6, 7, 11, and 12 after levels of compounds such as jasmonic acid, which alert inoculation. The number of M. persicae was counted for ISCIENTIST | 2018 plants to biotic and abiotic stressors (Schmelz et al., each plant by removing one pot at a time from the lid- 2003). However, the effects of nitrogen fertilizer on the ded containers to prevent any alates from escaping. To interactions between M. persicae and A. thaliana have reduce human error, each plant was counted by two re- not been demonstrated. Subsequently, in this study we searchers, and the mean of the two counts was used for propose to investigate how varying amounts of nitrog- analysis. On each of these days, we also noted whether enous compounds in the soil of A. thaliana affect the the plant was bolting and the number of leaves begin- per capita growth rate of M. persicae. We used the pop- ning to yellow per plant. Photographs of each of the 62 plants were also taken. DISCUSSION STATISTICAL ANALYSIS Our results suggest that the effectiveness of plant de- fenses may change based on the concentration of AN All analyses were done using the statistical software, added to the soil. Based on our findings, the addition R (R, 2015). An analysis of covariance (ANCOVA) was of low doses of AN to the soil of A. thaliana can be ben- performed on the mean M. persicae count, the treatment eficial when limiting herbivore populations. However, group (control, low dose, high dose), with a continuous adding higher doses of AN can increase herbivore pop- predictor of time (measured in days). An interaction ulations. During the first four days of the study period, term between treatment and day on the mean M. persi- the control, low dose, and high dose groups all experi- cae count was also analyzed in the ANCOVA. Plant H6 enced an increase in mean M. persicae count over time from day 12 was an outlier and removed from analysis. (Figure 1). These trends suggest the beginning of an exponential population growth due to back-and-forth Table 1: Summary of ANCOVA for the effect of competition between the herbivore and plant (Ayala et treatment, day, and the interaction between al., 1973). treatment and day on mean M. persicae count over a On the sixth day, the mean count for the low dose 13-day period. group began to plateau, which demonstrates the po- Mean M. persicae count tential benefit of a low dose of AN (Figure 1). Uptake Df Mean Sq F value Pr(>F) of nitrates occurs in the roots of the plant. Here, var- Treatment 2 9.5 0.1160 0.89060 ious transporters move the nitrates from the apoplast Day 1 6653.2 84.4438 1.454e-15 of root epidermal and cortical cells, located outside of Treatment:Day 2 430.8 5.2731 0.006214 the plasma membrane of plant cells, into the symplast, Residuals 11191.6 81.7 located inside the plasma membrane (Sohlenkamp et RESULTS al., 2002). Nitrogen is an essential nutrient to plants, as it is an important structural component and is essential The effect of time on mean M. persicae count was sta- Ec for metabolic processes, including photosynthesis. By tistically significant (p<0.05; Table 1). Furthermore, the extension, increased levels of nitrogen allow A. thaliana ology interaction term between treatment and day was sta- to have stronger defenses against herbivory. Our re- tistically significant (Table 1). The treatment as the sole sults suggest that the low dose of AN gave A. thaliana factor did not have a statistically significant influence the extra resources needed to strengthen its defenses, on the mean M. persicae count (Table 1). preventing the population of M. persicae from growing For the first four days of the experiment, the number of larger (Kuśnierczyk et al., 2008). Stout, Brovont and M. persicae increased over time (Figure 1). The low dose Duffey (1998) found a similar result. They reported that increased at the highest rate, then the high dose, then low levels of AN addition to the soil produced larger the control. At 5 days, there was a small peak in the increases in phenolic levels in tomatoes compared to mean M. persicae count for all treatment groups. After 6 plants with high levels of AN addition. Phenolics are a days, the mean M. persicae count plateaued for the low secondary metabolite that plants release in response to dose treatment group, while the control and high dose stress. Overall, the plateau in our low dose group rep- group continued to increase. Figure 1: Mean M. persicae count over the course of 12 days ISCIENTIST | 2018 for plants with no nitrogen added to the soil (Control), plants with 60 ppm ammonium nitrate (AN) added to the soil (Low), and plants with 300 ppm AN added to the soil (High). Error bars represent standard error. 63 resents the ability of A. thaliana to strengthen its de- foliar biomass resulting from an increase in nitrogen fenses against herbivores using AN from the soil. concentrations. The benefits of a low dose of AN to the plant also aligns Despite our promising results, there are several fac- with the theory of resource allocation. All plants must tors that may have altered our data. The A. thaliana use their resources, including nitrogen, for three main plants available for this experiment were not of con- functions: reproduction, defense, and growth, which sistent size, age or health, representing a confounding all interact and compete for resources (Bazzaz et al., variable. For example, some of the plants entered the 1987). These three functions interact and compete for bolting stage earlier than others, and based on our ob- resources. Allocation to these functions varies by in- servations, there were higher numbers of M. persicae on dividual and over time, as well as among populations bolting plants than on rosettes. Additionally, since all and species (Bazzaz et al., 1987). In the case of our study plants in one treatment group were kept in the same system, we may see the plateau for the low dose group lidded container, there was a possibility of winged after the sixth day because the plant has had the chance alates traveling between plants. Alates may have devel- to react to the herbivory of M. persicae and allocate oped due to lack of space and changes in quality of the more resources to defense. To confirm this theory, fu- host plant (Dixon, 1977). To reduce this error, all plants ture studies should measure plant biomass and repro- could have been kept in separate containers. Another ductive rates. As demonstrated by Pratt et al. (2005), source of error is the fact that for the low and high dose plants damaged by herbivory will have lower rates of groups, we added AN to the soil but did not measure reproduction than undamaged plants. In the case of M. concentrations of nitrogen in the soil itself. These val- quinquenervia, the paperbark tea tree, the undamaged ues can be different from one another, as Mulvaney, plants were 36 times more likely to reproduce than the Khan and Ellsworth (2008) found that addition of AN plants damaged by herbivory. This points towards re- fertilizer to the soil decreases amounts of natural soil source allocation for defenses, taking away energy from nitrogen. This is because when nitrogen is added to the reproduction and growth. soil, it can undergo changes in its availability to plants. The control group population continued to increase From this study, we can apply our findings on the effect for the duration of the experiment, which suggests the of nitrogen on aphid population growth to surrounding beginning of exponential population growth over our ecosystem. For example, trophic levels may be affected short experimental time period (Figure 1). The high by nitrogen fertilizers through tri-trophic interactions. dose group also followed this trend, suggesting that This involves the effects of plants, herbivores, and her- Ecology high amounts of AN are not beneficial in promoting bivore predators on one another. Nitrogen can affect the defenses of A. thaliana against herbivores. In accor- nutrient availability to plants, which is a bottom-up dance with this result, Minkenberg and Fredrix (1989) force that involves the removal of a primary consumer show that leafminers, namely Liriomyza trifolii, prefer or producer. This bottom-up force can either increase to feed on plants that were fertilized with higher levels or decrease plant populations (Capinera, 2008). Addi- of nitrogen, suggesting that the plant is more attractive tionally, this can alter animal community composition, to herbivores like M. persicae. Other studies have found influencing herbivores. This was observed in our study similar results (Chen, Olson and Ruberson, 2010). In through the interaction between the nitrogen fertilizer the context of our study system, the increase in attrac- treatment and the change in the population of herbi- tiveness along with nitrogen concentrations may ac- vores. Herbivores are also influenced by a top-down count for our observed results in the high dose group. force by their predators, carnivores (Capinera, 2008). Although the A. thaliana in the high dose group may Carnivores are affected by the populations of their have been more attractive to the herbivores, the pla- prey, which we know from our study are influenced teau in the population of M. persicae suggests that A. by the interaction between nitrogen availability for thaliana may have exhibited increased defenses. These plants, and time (Capinera, 2008). Therefore, all three counteracting forces provide an explanation for the trophic levels are affected by the addition of nitrogen trends observed in our results for the high dose group, fertilizer in plants showing the ecological impact of our which were similar to the control group. Additionally, study and further studies involving nitrogen fertilizers high levels of nitrogen may have allowed A. thaliana to in plants. ISCIENTIST | 2018 grow larger and thus be able to support more M. persi- Overall, this study provides greater insight into the in- cae. However, in our study we did not directly measure teractions between plants and animals. Through a con- plant growth. Further studies should measure growth trolled laboratory experiment, we found that the addi- rate and foliar biomass of A. thaliana to determine if tion of ammonium nitrate to the soil of A. thaliana has a increased attractiveness is directly due to increased ni- significant effect onM. persicae population growth over trogen concentrations, or indirectly due to increased time. These findings can be used to better understand 64 the potential impacts of nitrogen fertilizer runoff on analysis. We would also like to thank Dr. Chad Harvey surrounding ecosystems. In addition, our research can for reviewing our manuscript and providing additional be used by future studies as an analogous experiment comments and critiques. to apply over a longer time period to analyze the long- term effects of nitrogen on interactions between plants AUTHOR CONTRIBUTIONS and animals. Further studies must be done to under- L.A. contributed to the introduction and discussion. stand all effects of nitrogen fertilizer on the interaction N.K. contributed to the methods and results and per- of A. thaliana and M. persicae, as well as interactions be- formed the statistical analysis. L.B helped write the tween other plants and herbivores in susceptible envi- introduction, methods, and results and performed the ronments. statistical analysis. I.S. wrote the discussion section and the methods for treatment and inoculation. M.S. ACKNOWLEDGMENTS contributed to the introduction, abstract, and methods We would like to thank Dr. Susan Dudley for their sections. All authors contributed equally to experimen- guidance through this project and help with statistical tal design, data collection, and editing.sdfdf Kim, J. and Jander, G., 2007. Myzus persicae (green Global Dilemma for Sustainable Cereal Production. 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One major concern that hasbecome prevalent in humidity, salinity, andnutrientavailability insoiland ecosystems byalteringpHlevels,temperature, climate, climate change have directly and indirectly impacted to contributions anthropogenic The ways. ir- reversible, often and detrimental, in environment rounding INTRODUCTION Integr NADIA ALHASHEMI interactions acid contamination onec Eff For centuries, humansha ANGELA DITTRICH,JOSEPHINEAGUECI,TAYLORLUU,MEGHRATHOD, URL: https://theiscientist.com/2018/04/10/DittrichEffects.html Received: 02/25/2018 minimizing thiscontamination. provides acid evidencefurther acetylsalicylic for theecological impactof pharmaceuticals, andtheimportanceof of presence the in thaliana A. and persicae M. between interactions the investigating Overall, growth. population persicae M. on height bolt tween ASA concentration and bolt height growth. There were effects no observable of tion was increasedfor bothgenotypes,while an inverse relationshipwas be observed concentra- ASA as observed was population aphid in reduction greater A responses. concentrations were found to be significant for both aphid population and bolt height pact on the population growth of M. persicae. Interactions im- between significant genotype and a ASA had concentration ASA nor genotype neither that concluded was It plants. Col the to relativerategrowth population aphid mean negativeexperienced a with plants Overall,sid2 0.2mM). 11nM, was treated 1.1nM, concentrations(0M, ASA aqueous four of one plant Each sid2. induction-deficient acid salicylic mutant the and line Col wild-type experiment:the this in wereused Twothaliana A. of ascensionlines thaliana. A. growthof concentrationsplant soil the ASA impact howvarying analyzed study this Additionally, thaliana. Arabidopsis of strains different genotypically on cae persi- Myzus of growth population the on concentrations soil ASA varying the of determine impact to aimed study This pathways. immunity acid salicylic enhancing by a commonly used medication that has been found to have an impact onplant defence is ecologicalAspirin, on as interactions.knownimpacts Acetylsalicylic also (ASA), acid Pharmaceutical Keywords: Myzuspersicae, immune pathways anthropogenic impacts,soilcontamination, crosstalk,ecologicalinteractions, SUMMARY ated Science Program, Classof 2020,McMaster University ects of anthropogenicacetylsalicylic soil ve be contamination has become a rising concern based on its potential en alteringtheirsur Acc Arabidopsis thaliana,acetylsalicylicacid, ept ed: 04/10/2018 - - 02 ad otaetvs n h edcie systems endocrine the on contraceptives al., and et Lajeunesse2012) 2010; al., et Schultz 2003; Steuger, fects ofantidepressants (Metcalfe, Miao,Koenig and variety ofstudieshave observed andanalyzed theef throughwide excretiona human While (Crowe, 2014). or runoff direct via rivers and lakes of contamination posal ofmedication, results inincreasing chemical combined withover-prescription andimproper dis in North America (Health Canada, 2012). This increase, annually percent 15 to 10 rise to continues use ceutical tan Water Agencies, 2008). It is estimated that pharma- ological Published: 04/10/2018 - - - ISCIENTIST | 2018 Ecology 67 - - - ), were sid2), were M. on persicae - possess ascension with four different different four with Col and 16 sid2 sid2), or Col A. thaliana? The corresponding is 60 days (Serino and Gusmaroli, 2011), across across different plant genotypes? (2) How experiment was conducted over a span of 12 days TERIALS AND METHODS A. thaliana so selecting plants in these stages minimized flowering of the exper prior to the completion and plant death 1.1nM, (0M, line ascension each within treatments ASA - concentra four the for justification The 0.2mM). 11nM, - vari response observe to control the was 0M were: tions ables without exposure to ASA, 1.1nM and 11nM were concentrations of ASA bounds upper lower and the (Cleuvers, in the environment recorded been that have 2004), and 0.2mM was a physiologically active - concen plant immune responses to induce observed tration (Senaratna et al., 2000). A double tray apparatus was used to minimize cross contamination of wa- different ter treatments between neighbouring plants. The trays on the bot with a solid plastic tray structured were in rosette or early bolting stage. Theaverage lifespan of groups into two separate divided iment. Plants were line ( on ascension based for allow to overtop layered tray permeable a and tom, A gap contain. and the solute it may of water drainage by folding paper towels created was the trays between trays two of the perimeter the along them wedging and creating separation. Each corner tray was labelled, - cre ating a grid standardized system to define the location trays the of orientations varying the and plant each of ra ra and Dangl, 2011). In the present investigation, the functional possesses (wild-type) ascension Columbia SA and JA pathways, while the es a mutation to the SA-induction deficient 2gene in Arabidopsis . thaliana Thisgene codes for isochorismate SA in a crucial role plays an enzyme which synthase, this al.,of et Mutation (Ghazijahani 2014). biosynthesis ability to accumu in plants with a reduced results gene late SA, and thus enhanced susceptibility to pathogens to pathogens susceptibility late SA, and thus enhanced acid (SA) is one Salicylic stresses. and environmental regulating plant growth available of the most readily materials, and is also effective in other forms such as et al.,methyl salicylate (Ghazijahani 2014). following the address to was study this of objective The - concentra soil ASA varying do How (1) questions: two of growth tions impact the population A. thaliana the growth impact ASA soil concentrations do varying and overall quality of null hypotheses were the following: (1) ASA - concen tration and genotype do not affect aphid population. and (2) growth the impact not does concentration ASA quality of A. thaliana. overall This MA in an indoor facility at McMaster University in Ham- in University McMaster at facility indoor an in ilton, Ontario, from September 21st to October 3rd, 2017. All 32 A. thaliana plants (16 - - - - aquatic aquatic organisms such as fish and frogs (Park and Myzus , persicae in order to upregulate the less - effec ling molecule that plays an important role in the plant an important role that plays ling molecule pathways defence via SA-inducible immune system (Clissold, 1996). Salicylic acid acts as a stress - messen biotrophic against to resistance linked and has been ger pathogens and promotion of - Systemic Resis Acquired attack, an following plants ‘primes’ which (SAR), tance ensuring faster and more effective defence responses against a second attack. (Conrath, 2006, Mai and Gabrys, 2011). Another major plant defence is Morkunas; the jasmonic acid (JA)-inducible pathways, which also involve the use of JA molecules as messengers that ac- un However, predation. to address genes certain tivate been observed have the JA SA pathways like pathways, to be an effectiveresponse to stressors such as- necro (Conrath, insects and herbivorous pathogens trophic 2006). Thepresence and activation of pathways both in an at in crosstalk within an individual plant results the and employ allocation resource to optimize tempt - preda the to respective response defence effective most in the following studies that it is evident However, tor. it has been observed that SA and JA pathways share by cer when confronted relationship an antagonistic tain biotic stresses. Although JA-induced mechanisms are more effective against predators such as generalist observed has been aphids, the SA signalling pathway to suppress the JA pathway by means that are still un- - SA-medi that concluded (2010) al. et Leon-Reyes clear. ated suppression of the JA signalling targeted pathway regions downstream of the JA biosynthesis pathway. - SA-me towards provided also been have Observations diated suppression of JA-inducible genes (Kourneef et al., 2008). Nevertheless, suppression of the JA signal- of for the secretion evidence has provided ling pathway aphids such peach by green SA-inducible compounds as (Louis and Shah, 2013). mechanism tive plant defence Interest in crosstalk between the SA and JA signaling to the experimental use of ascension has led pathways lines deficient in one of mechanismsthe (Nishimu- Kidd, 2005; Kloas et al., et al., 2009; Runnalis 2013; - Or lando and Ellestad, 2014), the impacts of - pharmaceuti Many understudied. severely plants is absorption by cal defence a as pathways chemical inducible on rely plants or concentration so altering the presence mechanism, major have slightly could even compounds of various consequences et (War al., 2012). Thus, it is crucial that inter- ecological on medications common of effects the to quantify the actions magnitude is of examined these consequences. An investigation was conducted on the acid acetylsalicylic with contaminated soil of impacts the by known Commonly interactions. plant-animal on brand name Aspirin, this nonsteroidal anti-inflamma- of salicylic acid (SA), a signal tory drug is a derivative of Figure 1: Initial experimental setup. The Col tray contained plants A1- C6, while the sid2 tray contained plants D1- F6. The initial setup on Day 0 (A) had the Col tray to the west of the sid2 tray, with the “1” side of the tray against the north window ledge as illustrated by the striped area. The trays were switched on Day 5 (B) with the “1” side of the tray against the north window ledge but the sid2 tray to the west of the Col tray. The trays were switched on Day 5 (B) with the “1” side of the tray against the north window ledge but the sid2 tray was placed to the west of the Col tray. On day 8 (C), the “6” side of the tray was placed against the north window ledge with the Col tray to the west of the sid2 tray. On day 11 (D), the “6” side of the tray was placed against the north window ledge with the sid2 tray to the west of the Col tray. throughout the experiment. Trays were re-oriented on measured in days, was interpreted as a continuous in- days 5, 8, and 11, to eliminate any environmental bias dependent variable. The effects of genotype and ASA (see Figure 1). A number generating software (Haahr, concentration on bolt height growth were also assessed 1998) was implemented to randomly assign a location with an ANCOVA followed by Holm’s test. Possible and ASA concentration to each plant. block effects arising from growth of Col and sid2 plants in different trays were included in the above analyses. PREPARING TREATMENT SOLUTIONS The potential impacts of bolt height on aphid popu- AND INOCULATION WITH APHIDS lation growth were assessed using a linear regression The ASA solutions were prepared from 100 g of 99% model. All analyses were carried out using R version ology powdered acetylsalicylic acid and distilled water on a 3.3.2 (R Development Core Team, 2016). VMR stirrer. 10 drops of MeOH were added to ensure Ec solubility in water. 15 millilitres of the corresponding RESULTS SA solutions were exogenously delivered to the plant via soil drenching. The plants were first watered on Day APHID POPULATION GROWTH 0 with respective solutions and twice more throughout the duration of the experiment (Day 4 and Day 11). On Day 4, three aphids were inoculated from pre-ex- isting A. thaliana plants onto the rosette leaves of each of the 32 test plants. The aphids were all in an inter- mediate development instar stage and were reared on a wild type (Col) Arabidopsis thaliana. Rearing them on A. thaliana ensures the aphids were not starved prior to inoculation, avoiding any malnourishment. As well, during the investigation, the aphids were subjected to the same conditions by maintaining spatial and tempo- ral proximity of the population. Following the inocula- tion, aphid numbers were counted for the duration of the experiment by inspection with a probe and magni- Figure 2: Mean Aphid Count of Col for Four fying glass. Bolt height was measured on days 0, 4, 8, 11, Concentrations over Eight Day Period. Control = 0M, and 12, as a quantitative factor coupled with qualitative Low = 1.1nM, Intermediate = 11nM, High = 0.2mM. ISCIENTIST | 2018 observations, serving as a proxy for plant quality. Error bars show standard deviation (<0.95 interval). STATISTICAL ANALYSIS Each A. thaliana plant used in this experiment was ex- posed to one of four ASA concentrations: 0M (control), The effects of genotype and ASA concentration on 1.1nM (low), 11nM (intermediate) or 0.2mM (high). aphid population growth were assessed with an AN- Among all eight combinations of genotype and treat- COVA, followed by a post hoc Holm’s test. Time, 68 Table 1: Three-Way Analysis of Covariance of the Effect of Genotype and ASA Concentration Difference on Myzus persicae Population Growth Across Eight Day Test Period. * indicates a significant p-value. Source d.f. Mean Square F Value Pr(>F) Plant x 3 20.7156 4.7946 0.003255* Treatment x Day Treatment 3 13.2290 3.0618 0.030168* x Day Plant x Day 1 12.4256 2.8759 0.092078 Plant x 3 23.7750 5.5026 0.001318* Treatment 1 8.2656 1.9130 0.168766 Day Figure 4: Mean Change in Aphid Count of Col and Treatment 3 16.0417 3.7128 0.013064* sid2 Plants at Various ASA Levels over Eight Day Test Plant 1 30.6250 7.0880 0.008643* Period. ‘1’, ‘2’, ‘3’, and ‘4’ represent 0M, 1.1nM, 11nM and Residuals 144 4.3207 0.2mM. Error bars show standard deviation (<0.95 ment, the control Col plants experienced the only net interval). increase in mean aphid count over the test period, peak- pressed a similar overall decrease in mean aphid count ing at a mean of 6.50±6.22 aphids on Day 11 (Figure 2). in both the Col and sid2 plants of -2.25±0.83 and -2±0.71, This peak is significantly different (P<0.05) from only respectively. The intermediate concentration group ex- two other treatment combinations (low Col on Day 11, hibited 0±3.00 in mean aphid population for Col plants d.f. = 3, p = 0.039; and intermediate sid2 on Day 12, d.f. and a -3±0.00 change in aphid population count in the = 3, p = 0.039) (Table 1). These were the only signifi- sid2 plants, the largest decrease observed among the cant differences found with interaction between geno- genotypes and treatments. The high Col and high sid2 type, treatment, and day. The high sid2 plants also had plants expressed a change in aphid count of -2.25±0.83 a similar trend, exhibiting the largest increase among and -1.25±3.03, respectively. No significant differences all sid2 treatments and peaking at 3.50±2.29 aphids on were found in the overall change in mean aphid count Ecology Day 11 (Figure 3). The high and lowCol plants exhibited between genotypes for any treatment (d.f. = 3, p = 1.000 a gradual decrease in mean aphid count, while the in- for almost all of differences in mean) (Table 2). termediate Col plants gradually increased over the test Table 2: Significant interactions (P < 0.05) between period, but had no overall change. However, for sid2, genotype and ASA concentration on aphid mean aphid count decreased from Day 4 to Day 12 for population count across eight day period since ASA all concentrations without a significant difference be- inoculation Genotype. Sid. Col. Sid. Col. Sid. Col. tween one another, suggesting that they were similarly Conc.Day high.8 control.11 control.11 low.11 low.11 int.11 negative during this time period. Col.low.11 1.000 0.039 1.000 - - - Sid.int.12 1.000 0.039 1.000 1.000 1.000 1.000 BOLT HEIGHT ISCIENTIST | 2018 Figure 3: Mean aphid count of sid2 for four ASA Figure 5: Mean bolt height of Col plants for four concentrations of eight day period. Error bars show treatments for 12 days since initial inoculation standard deviation (<0.95 interval). with ASA. T0 = 0M; T1 = 1.1x10-9M; T2 = 1.1x10-8M; T3 = 2.0x10-4M. Error bars show standard deviation (<0.95 Additionally, observations were made regarding overall interval). change in aphid count from Day 4 to Day 12 (Figure 4). The control Col experienced the only increase observed Interactive effects between genotype and concentration among all treatments and genotypes with a mean in- on mean bolt heights of Col (Figure 5) and sid2 (Fig- crease of 3.25±6.42 aphids, approximately double that ure 6) plants were observed on specific days over the of the sid2 decrease. The low concentration group ex- 12-day test period (Figure 7). All Col ASA concentra- 69 mean growth is approximately three times greater. At high ASA concentration, the only overall decrease in bolt height is observed in sid2 plants, while Col bolt growth is similar to low ASA concentration. The control and intermediate sid2 plants demonstrat- ed increasing bolt height growth rates, with that of the intermediate plants being at a steeper incline. Almost no change in mean bolt height was observed at low sid2. The notably small standard deviation (0.125±0.083cm) suggests the presence of a threshold of effect of ASA concentration on bolt height. Interestingly, all sid2 Figure 6: Mean bolt height of sid2 plants for 4 treatments over 12 days from initial ASA inoculation. ASA concentrations had very similar growth rates from Day 0 and Day 4, after which the different treatments Error bars show standard deviation (<0.95 interval). diverged with rates that were not significantly differ- ent (d.f. = 3, p = 1.0000). Most importantly, bolt height growth appeared to share an inverse relationship with ASA concentration among both Col and sid2 plants. HEIGHT ON APHID POPULATION GROWTH The intercept of the linear regression plot of bolt height against aphid population was significant (Table 4; p = 5.82e-15) while the slope was not (d.f. = 1, p = 0.828) (Figure 8). Figure 7: Mean change in plant bolt height of col Table 4: Linear regression model of the effect of and sid2 plants at various ASA levels over 12-day test height on aphid population for all genotypes and period. ‘1’, ‘2’, ‘3’, and ‘4’ represent 0M, 1.1nm, 11nm treatments. * indicates a significant p-value. and 2.0μm. Error bars show standard deviation Source d.f. Estimate Pr(>|t|) F Adj. R2 (<0.95 interval). Intercept 2.07448 5.82e-15* Height 1 -0.01722 0.828 0.04741 -0.007679 Ecology tions exhibited positive bolt height growth, while sid2 Residuals 124 exhibited varying trends across different ASA concen- trations. Control Col plants demonstrated the greatest DISCUSSION mean bolt height growth compared to the three other ASA concentrations, with a steady increase ending at APHID POPULATION 6.43±1.64cm. Bolt height means on Day 11 and Day 12 Control Col exhibited the greatest overall increase in for control Col were significantly different from Day M. persicae populations, peaking on Day 11 (Figure 2 0 means for all treatment groups (Table 3; d.f. = 3, p and Figure 3). This was contrary to the expectation that = 0.0109, p = 0.0094, p = 0.0159, and p = 0.0170, from control Col would experience the greatest decrease in control to high respectively). Control Col on Day 12 aphid population compared to all other Col treatments, was also significantly different compared to high sid2 as the plants had the smallest degree of crosstalk be- on Day 8 (P<0.05, d.f. = 3, p = 0.0283) and Day 11 (d.f. tween SA and JA, due to the absence of upregulation of = 3, p = 0.0304). Bolt height growth for low and high SA by ASA. Evidence from other studies suggests that Col were similar in both progression and final heights SA-inducible pathways do not play an important role of 2.8±2.60cm and 2.43±2.56cm, respectively, while in- in addressing aphid feeding stress. For example, mutant termediate Col demonstrated the smallest bolt height A. thaliana such as sid2 and npr1, which have a loss of growth (1.20±1.27cm). In addition, Col exhibits greater key SA regulator and receptor NPR1 (non-expressor of bolt growth at all ASA concentrations compared to PR genes 1), have unchanged aphid population growth sid2, except at intermediate concentration, where sid2 compared to wild-type (Moran and Thompson, 2001; ISCIENTIST | 2018 Table 3: Significant interactions (p < 0.05) between genotype and ASA concentration on bolt height across eight-day period Genotype. Col. Col. Col. Col. Col. Sid. Col. Sid. Sid. Conc.Day cont.0 high.0 mod.0 low.0 mod.4 high.8 mod.8 high.11 high.12 Col.control.11 0.0212 0.0183 0.0304 0.0325 0.0304 - - - - Col.control.12 0.0109 0.0094 0.0159 0.0170 0.0159 0.0283 0.0498 0.0304 0.0375 70 Mewis et al., 2005). In fact, observed crosstalk between that addition of ASA via soil drenching may be an ef- JA and SA-inducible pathways often appears to be an- fective substitute for SA in the absence of endogenous tagonistic, with suppression of JA pathways promot- SA. ing susceptibility to aphid feeding (Mewis et al., 2005; Support for a greater role for SA-inducible pathways is Louis and Shah, 2013). The large amount of evidence contradicted by observations made with high sid2. Ex- led Louis and Shah (2013) to conclude that SA-induc- posure to the highest ASA concentration in sid2 was ex- ible pathways are not important in addressing aphid pected to yield the highest aphid population numbers feeding stress, and may in fact negatively impact plant among all sid2 plants due to greatest suppression of JA. defence. Thus, the observations made regarding the no- However, the small mean aphid population on high sid2 might be explainable by error. From Day 11 to Day 12, Figure 8: Relationship between mean change in all genotype and treatment combinations experienced bolt height and mean change in aphid population a decrease in aphid population, excluding a small in- across all genotypes and ASA concentrations crease observed among low Col that is likely a conse- quence of aphid migration. This shared decrease may be ticeable increase in aphid population numbers on con- due to contamination of the ASA solutions produced trol Col conflict with the above evidence. The absence on Day 0 and used throughout the experiment for soil of ASA in control Col should favour SA pathways to drenching. A contaminant must have been introduced, the smallest degree among all Col treatments and thus as the development of an unidentified biological con- result in reduced aphid populations. Although signifi- taminant was observed in the high ASA concentration cant differences in aphid population count were only beaker on Day 11, thus contamination of the other solu- found between control Col on Day 11, and low Col and tions may have been unnoticeable to the naked eye. The intermediate sid2 on Day 11 and Day 12 (d.f. = 3, p = contaminant was observed to be cloudy yellow-white 0.039; d.f. = 3, p = 0.039), evidence for interaction be- in colour, irregular in shape, and filamentous (refer to tween genotype and ASA concentration demonstrated Figure 9). This common decrease may have also arisen by the trend of control Col may suggest a greater degree from the introduction of predators into the environ- Ecology of effectiveness of SA-inducible pathways against aphid ment. Aphid predators were first noticed on Day 8, and feeding than previously believed. Conflicting observa- could have potentially been responsible for any drops tions have been made on the exogenous application of in aphid count observed throughout the experiment. SA derivatives on aphid settling and growth; applica- Thus, had the experiment ended on Day 11 rather than tion of synthetic analogs of SA has no impact (Moran 12 due to significant impact by error, the trend of mean and Thompson, 2001), while the SA analog benzothia- aphid count on high sid2 would have become an overall diazole has resulted in decreases in aphid reproduction increase, suggesting that upregulation of SA-inducible (Morkunas, Mai, and Gabriś, 2011). Furthermore, sever- pathways adversely impacts plant defence. However, al SA-related pathways have been identified as effective this appears to be an outlier trend and may suggest the defences against aphid feeding. For example, the reg- presence of a threshold ASA concentration at which ulatory gene PAD4 (phytoalexin deficient4) is one of ASA exposure changes from positively to negatively the genes responsible for inducing SA synthesis, as well impacting plant defence. as activating a defence pathway that adversely impacts aphid settling and fertility (Wiermer, Feys, and Barker, 2005). In turn, SA molecules enhance PAD4 expression, resulting in a positive feedback loop. Thus, it is suggest- ISCIENTIST | 2018 ed that SA-inducible pathways, while possibly less ef- fective than counterpart JA pathways, may still play an important role in reducing aphid population numbers. This is supported by the low, intermediate and high treatments of both genotypes - excluding high sid2, which exhibited only decreases in aphid population - which suggest a more effective plant defence response to aphid feeding with greater exposure to ASA (Figure 3). The lack of significant differences between Col and sid2 suggests that they were similarly effective in plant defence, and that exogenous exposure versus no expo- sure to ASA impacts plant response more greatly than Figure 9: Unknown contamination observed in the degrees of ASA concentration. This may also suggest high ASA concentration beaker on Day 11. 71 Consideration of more ASA concentrations within the intermediate, and high treatments further supports the experimental setup of a future study may make these possibility that changes in bolt height are most greatly trends more discernable, and provide more accurate influenced by the exposure or lack of exposure to ASA. observations regarding the importance of the SA sig- Interestingly, trends for M. persicae populations and nalling pathway. This would be aided by the inclusion bolt growth for low and high Col plants appeared to be of an ascension line possessing a non-functioning JA quite similar, both demonstrating gradual decreases in signalling pathway, in order to further investigate the both response variables. However, the aforementioned impacts of JA-SA crosstalk. relationship is only speculated based off of the minimal Interestingly, mean aphid population decreased on all data in this study and should be further investigated plants between the first and second day since aphid with more specific future experiments. inoculation, irrespective of genotype or ASA concen- tration. This was believed to arise from stress placed BOLT HEIGHT ON APHID POPULATION on the aphids as they were introduced to a new envi- The intercept of the linear regression plot investigating ronment and each of the plants were watered with the possible effects of bolt height upon aphid population appropriate ASA solution. indicated that at changes in mean bolt height of 0cm, there was an expected significant mean aphid popula- BOLT HEIGHT tion increase of 2.07. Interpreting change in bolt height There is an apparent inverse relationship between ASA as a proxy for plant quality, this may suggest that A. concentration and change in bolt height for both geno- thaliana in poor health experience an increase in aphid types, supported by all treatment combinations except population. However, the absence of a significant slope intermediate Col and sid2. At higher ASA concentra- suggests that there does not exist a relationship be- tions, reduced bolt height growth was observed. While tween the two variables, and it must be acknowledged significant differences were only found between con- that bolt height is an unreliable indicator of plant qual- trol Col on Day 11 and Day 12, and the other three ASA ity that can be affected by factors such as ascension line concentrations of Col and high sid2 (Table 4), this trend and and significant variation in the development rate was expected. SA plays an integral role in plant senes- of A. thaliana (Gnan, Marsh, and Kover, 2017) (Figure 7). cence and cell death, and exogenous application of SA Nevertheless, the significant intercept warrants further causes programmed cell death (PCD) and early senes- investigation, perhaps through consideration of the cence in association with SAR (Vogelmann et al., 2012) effects of interaction between genotype and height on Ecology (Brodersen, 2005). Furthermore, SA is involved in reg- aphid population. ulation of flowering time, and in response to both bi- otic and abiotic stresses, high SA endogenous levels are CONCLUSION observed in plants that transition earlier from bolting Overall, interactions were found between both ASA to flowering to increase chances of population survival concentration, genotype and time. These results led (Martinez et al., 2003) (Lyons et al., 2015). This furthers to the following conclusions: the null hypothesis that the notion that SA promotes plant resource realloca- ASA concentration and genotype do not affect aphid tion, diverting resources from increasing biomass to population was rejected. Additionally, as a significant processes involved in reproduction, thus increasing the relationship was found between ASA concentration overall survival of the population rather than the in- and bolt height, the null hypothesis that ASA concen- dividual. The outlier bolt height trend of intermediate tration does not affect bolt height was rejected. These sid2 can be explained by the large standard deviation of findings present further evidence for the impact of an- 2.575±2.712cm. The sole significant differences between thropogenic contaminants on not only the quality of control Col and four other treatment groups, ASA con- A. thaliana and its associated immune response, but on centration combinations may suggest that exposure or the population growth of insects such as M. persicae. non-exposure to ASA has a greater impact on Col bolt The irresponsible disposal of pharmaceuticals is having height growth than different degrees of ASA exposure. a significant impact on the environment. Thus, efforts Furthermore, all four ASA concentrations of Col expe- should be made to implement new policies and proce- rienced a net increase in mean bolt height and sid2 ex- dures to limit pharmaceuticals such as ASA from en- ISCIENTIST | 2018 hibited various trends, suggesting a possible influence tering the soil and water, and decrease anthropogenic of genotype upon bolt height. However, standard devi- ecological impacts. ation for the low, intermediate, and high Col were very large, and shared large amounts of overlap with the means and standard deviations of the sid2. This observa- ACKNOWLEDGEMENTS tion, and the lack of significant differences among low, We would like to thank Dr. Chad Harvey and Sebastian 72 Irazuzta for their supervision, guidance, support, and uscript. feedback. We would also like to thank the Integrated Al Hashemi: Research contributions included: re- Science program for providing us with the resources searched proper care and conditions for A. thaliana. and opportunity to conduct this study. We would like Authorship contributions included: wrote the study to acknowledge Dr. Robin Cameron for her insightful system portion of the report, general editing and con- guest lecture on crosstalk which furthered our interest tributed to the discussion and conclusion. in investigating the salicylic acid inducible pathway. Agueci: Research contributions included: Researched Finally, we would like to thank Russ Ellis for coordi- background information on M. persicae and A. thali- nating the lab and providing us with the resources and ana (general traits, aphid and plant relationships with space to conduct this experiment. jasmonic/salicylic acid pathways, plant defence mech- anisms against aphid feeding, the effects of acetylsali- AUTHOR CONTRIBUTIONS cylic acid on plants and aphids), experimental design. Authorship contributions included: formulation of Rathod: Research contributions included: researched appropriate research questions and hypotheses, created the study system of A. thaliana and M. persicae and Excel graphs and analyzed their trends for the Results experimental methods on exogenous delivery of sub- and Discussion sections. Contributed to edits and for- stances to plants, and experimental design. Conducted mulation of Abstract and Conclusion. research on SA and JA defense pathways. Authorship Dittrich: Research contributions included: investigated contributions included: wrote materials and methods various anthropogenic ecological impacts to shape the section, and co-authored study system and results. research topic, followed by researching pharmaceuti- Contributed to general editing. cal and ASA impacts on water, soil, and plants once Luu: Research contributions included: researched the the topic was narrowed down, researched JA and SA SA and JA pathways, including mechanisms, genes in- pathways to determine which strains were appropriate duced, SAR, and local responses. Also researched cross- for the experiment, and background research for the talk between SA and JA, and the impact on plant health Introduction section of the manuscript. Authorship Ecology and aphids. Conducted statistical analyses (ANOVA, contributions included: compiled most of the abstract, ANCOVA, post hoc) for all data, created graphs along- wrote the Introduction section and part of the Materi- side another group member to display trends of data. als/Methods section, wrote the references, and edited Authorship contributions included: contributed to re- the rest of the manuscript. sults and discussion sections of the manuscript. Con- tributed to general edits and formatting for final man- ISCIENTIST | 2018 73 74 ISCIENTIST | 2018 Ecology Signaling &Behaviour jasmonate signalingpathways inArabidopsis. identify regulators to ofcross-talk between salicylateand system reporter a Towards 2008. al., et Kourneef 1163, pp.187-200. vertebrates. Kloas, W. ography of theaphid, and land use influences on the and phenology numbers M. Hullé, Environment Study (2012). Canada. Health tegrity Services, Dublin. 1998. M., Haahr, early flowering. on constraints trade-off from plants thaliana dopsis Arabi releases fitness to contribution photosynthetic 2017.Kover,Inflorescence & T., P.X., Marsh, S., Gnan, basil sweet ( in minerals some of acquisition root of sprays ofcitricacidandsalicylicalterthepattern Ghazijahani, Hadavi, & N., E., Jeong, B. 2014. R., Foliar Resistance. Acquired Plant Signal&Behaviour Systemic 2006. U., Conrath, salicylic acid. of derivatives related and Aspirin 1996. S.P., Clissold, Safety naproxen, and acetylsalicylic acid. ibuprofen, diclofenac, drugs anti-in the flammatory of toxicity Mixture 2004. M., Cleuvers, ology Arabidopsiscelldeathin of duction acd11. in the in acid salicylic of The role 2005. Brodersen,P., sociation ofClean Water Agencies. WaterEnvironment the in Pharmaceuticals (2008). Agencies. 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RFoundation for , 47(4), pp.2077- Environ 30, 7, 7, ------, THE EDITORIAL BOARD Laura Green & Varsha Jayasankar Senior Editors Tanya Daniel & Sabrina Macklai Level III Editors Yajur Iyengar & Sam Marchetti Level II Editors Andrew Colgoni & Dr. Sarah Symons Editors-in-Chief Jonathan Panuelos Webmaster and Promotions Isobel Sharpe Workshop Coordinator James Lai Senior Advisor CONTACT US journals.mcmaster.ca/iScientist COVER PHOTO TheiScientist.com An aerial view of the coast of Hawai’i (the Big Island) with mountains in the distance. [email protected] Photograph by Abby Lindzon H.G. Thode Library, Room 306 1280 Main Street West ISSN: 2560-9939 (Print) Hamilton, ON L8S 4P5 ISSN: 2560-9947 (Online) THEi SCIENTIST TheiScientist.com