University of Massachusetts Amherst ScholarWorks@UMass Amherst
Masters Theses 1911 - February 2014
1982
Quantitative sampling of preimaginal black flies (Diptera: Simuliidae) and drift ecology of Simulium tuberosum Lundstrom complex.
Kenneth Raymond Simmons University of Massachusetts Amherst
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Simmons, Kenneth Raymond, "Quantitative sampling of preimaginal black flies (Diptera: Simuliidae) and drift ecology of Simulium tuberosum Lundstrom complex." (1982). Masters Theses 1911 - February 2014. 3043. Retrieved from https://scholarworks.umass.edu/theses/3043
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QUANTITATIVE SAMPLING OF PREIMAGINAL BLACK FLIES
(DIPTERA:SIMULIIDAE) AND DRIFT ECOLOGY OF
SIMULIUM TUBEROSUM LUNDSTROM COMPLEX
A Thesis Presented
By KENNETH RAYMOND SIMMONS
Submitted to the Graduate School of the University of Massachusetts in partial fulfillment of the requirements for the degree of
MASTER OF SCIENCE
February 1982
Entomology QUANTITATIVE SAMPLING OF PREIMAGINAL BLACK FLIES
(DIPTERA:SIMULIIDAE) AND DRIFT ECOLOGY OF
SIMULIUM TUBEROSUM LUNDSTROM COMPLEX
A Thesis Presented By KENNETH RAYMOND SIMMONS
Approved as to style and content by: //A C /
-f v c John D. Edman, Chairperson of Commrtrto^
C\J. G. JohnU&. Stoffolano,0Member
Michael R. Ross, Member
fr / /! n ' --a John D. Edman Acting Department Head IjCntomology To Anne, who started this project 2 days late hut stuck around for the finish. She knows the rest.
m ACKNOWLEDGEMENTS
This thesis should perhaps he titled "The dead horse that would not die". My most sincere thanks to
Dr. John Edman, my major professor, for never allowing
it to be buried. His patience in allowing me to do my own thing has made me a better scientist -- I hope.
I also would like to thank Drs. John Stoffolano and
Michael Ross for agreeing to serve on my committee
after most of the research had been completed. Dr.
Pedro Barbosa allowed me the liberal use of his truck
during the early phase of this project.
Many special thanks to Steve Bennett, Dennis Lapointe,
Roger Nasci, Ned Walker and Wren Withers for their friend¬
ship and occaisional' jonts to the field to lend a hand.
Thanks also to the rest of the Apiary group for helping
to make graduate school more enjoyable.
I am particularly lucky and thankful for the 52 gang,
whose love and encouragement always helped. My parents
love is dearly appreciated. It got me started and helped
me finish.
This research was supported by Northeast Regional Black
Fly Project NE-118 (Hatch 436) administered by the
Massachusetts Agricultural Experiment Station. That
support was greatly appreciated.
IV TABLE OF CONTENTS
Page DEDICATION. iii
ACKNOWLEDGEMENT . iv
LIST OF TABLES. vii
LIST OF FIGURES . ix
Chapter I. LITERATURE REVIEW . 1
Economic Importance and Control of Black Flies ...... 1 Sampling Preimaginal Black Flies ... 5 Ecology of Preimaginal Black Flies . . 7 Microdistribution . 7 Substratum preference by larvae . . 8 Density of black flies on substratum . 9 Dispersal of black fly larvae ... 10
II. THE SIX LARVAL INSTARS OF THE SIMULIUM TUBEROSUM COMPLEX (DIPTERA: SIMULIIDAE) . 14
Introduction... 1^ Materials and Methods ...... Results. 16 Discussion . 25 Conclusions. 26
III. ANALYSIS OF CERAMIC TILES FOR QUANTITATIVE SAMPLING OF PREIMAGINAL BLACK FLIES (DIPTERA: SIMULIIDAE) IN A ROCKY BOTTOM STREAM. 27
Introduction . 27 Materials and Methods . 28 Field study sites. 28 Description of the tiles . 29 Determining rock surface area ... 29
V Chapter Page
Placement and collection of tiles. 30 Experimental procedures . . c . . . 30 Results. 34 Population dispersion . 34 Tile placement. 35 Density of black flies on rocks compared to tiles. 36 Frequency of instars captured on tiles versus natural substrates. 38 Importance of current velocity and tile angle . 42 Tile colonization. 48 Captures on old versus new tiles . 48 Discussion. 51 Conclusions ...... 55
IV. DRIFTING BEHAVIOR OF SIMULIUM TUBEROSUM AND OTHER BLACK FLIES AND . ITS POSSIBLE CAUSE. 58
Introduction . 58 Materials and Methods ...... 60 Field study area.. • 60 Sampling of S. tuberosum drift . . 60 Artificial stream . 6l Determination of S_. tuberosum ’ s drift pattern. 62 Observation of predator:prey interaction in the artificial stream . 65 Other observations of black fly defensive behavior . . 67 Results. 68 Diel drift pattern of S. tuberosum 68 Foraging behavior of perlid stone- flies . 86 Simulium tuberosum drift and evasive behavior in the artificial stream . 95 Discussion. 102 Conclusions ...... 109
LITERATURE CITED. HO
vi LIST OF TABLES
Table Page
1. Postgenal lengths of Simulium tuberosum instars (micrometers) . 2. Mean body lengths of the six Simulium tuberosum instars (micrometers) . 23 3. Mean number of primary cephalic fan rays of the six S. tuberosum instars. 24 4. Black fly species collected on both tiles and natural substrates in the Saw Mill River, 1977-1980 36 5. Comparison of the number of preimaginal black flies captured on tiles placed in a random versus stratified random design . . 37 6. Comparison of the number of preimaginal black flies collected on tiles and rocks using a random sampling design . 39 7. Comparison of the number of preimaginal black flies captured on rocks and tiles using a stratified design. 40 8. Comparison of the percentage of the total of each black fly species captured on tiles vs. rocks at station 1. 41 9. Comparison of the proportion of the total of each S. tuberosum instar collected on 25 tile samples at station 4. 4l 10. Least squares linear regression of the number of preimaginal black flies (N + 1 log10 transformed) versus current velocity or the data from random and stratified random samples combined .... 42 11. Comparison of the number of S. tuberosum larvae captured on tiles placed at different angles on redwood boards .... 44 12. Comparison of current velocity measured at 4 areas near a ceramic tile on a redwood board at 0, 20, 40, 60, and 90 degrees . . 4? 13. Colonization rate of black flies and other insects on ceramic tiles at station 4 . . 49 14. Comparison of the number of black fly larvae and pupae collected on old and new tiles during winter and summer months ...... 50
Vll "\
Table Page
15. Comparison of the percent of the total larvae and pupae captured and temperature measurements during each sample period of each of the six diel test dates .... 71 16. Mean (N + 1 log base 10 transformed) number of larvae and pupae captured per tile during day and night for each diel period .. 72 17. Comparison of the mean percent of each of the six S. tuberosum instars captured during each of the 5 bay and 3 night sample periods on each diel sample period . 75 18. Comparison of the population age structure of S. tuberosum larvae as determined from random substrate collections and 24 h drift tile samples (1900 - 1900 h). 75 19* Comparison of S. tuberosum larvae captured on drift samplers between the daylight-dark and dark-daylight periods . 79 20. Total number of black fly larvae and pupae collected in eight contiguous 3 h samples over a 24 h period compared to those captured in single 24 h samples on the 2 subsequent days . 87 21. Location of perlid stoneflies on rocks during day and night. 88 22. Digested status of prey items in the guts of perlid stoneflies collected during day and night. 90 23. Drift of S. tuberosum larvae in an outdoor artificial stream with and without stonefly predators . ... 96 24. Prey found in the alimentary canal of most common perlids found at the study site . . 99 25. Electivity coefficients of the predominate prey items in the diet of P. immarginata 100
• • • vi 11 LIST OF FIGURES
Figure Page
1. Plot of the mean postgenal length (log,0) against the respective S. tuberosum instar . 18 2. Linear regression line of body length against postgenal length of individual larvae ... 22 3. Redwood board used to hold tiles at various angles relative to the stream bottom. Tiles were held at desired angles by dowels placed on their underside. The board was placed in the stream parallel to the bottom and perpendicular to the direction of the stream flow. 33 4. Distribution of Simulium tuberosum larvae on the upstream surface of tiles placed at 0, 20, 40, 60, and 90 degree angles relative to the stream bottom on a redwood board. Data is combined from 4 repetitions. Tops of the figures represent the tile edges nearest the water surface. Direction of stream flow is indicated by the arrow ... 46 5. Average drift pattern of S. tuberosum larvae based on the mean percent of the total larvae and pupae captured on tiles for each sample period of each of the six diel test dates. Lines around each mean are standard deviations . 70 6. Diel drift pattern of S. tuberosum instars based on the percentage of each instar captured for each of the 8 sample periods during a diel (1 = 1900 h, 2 = 2200 h, 3 = 0100 h, 4 = 0400 h, 5 = 0700 h, 6 = 1000 h, 7 = 1300 h, 8 = 1600 h; the letters A-E correspond to the sample dates listed in Fig. 7).. 74 7. Drift pattern of S. tuberosum larvae for each of the six diel periods. The solid line is the mean number of larvae per tile and the hatched line is discharge. (A = Aug. 2-3, B = Aug, 6-7, C = Aug. 7-8, D = Aug. 22-23, E = June 26-27, F = July 5-6). 78
IX "X
Figure Page
8. Drift of S, tuberosum larvae during the daylight-to-dark period in mid-July 1980. Lines about the means are standard deviations. 82 9. Drift pattern of 3 genera of Ephemeroptera based on the mean percent per time period of the total number captured for each diel period sampled in 1979 85 10. Number of black fly larvae drifting into the screen of the artificial stream correlated with stonefly activity . 92 11. Number of black fly larvae drifting into the screen of the artificial stream correlated with stonefly activity .... 9^
x ~\
CHAPTER I
LITERATURE REVIEW
Economic Importance and Control of Black Flies
Black flies (Diptera: Simuliidae) are of world¬ wide medical and veterinary importance. They are the only vectors of the human filaria Onchocerca volvulus
Leuckart in Afrotropical and Neotropical regions. This disease infects “between 20-40 million people and causes severe skin reactions and “blindness in the most chronic victims (W. H. 0. 1966, Wenk and Rayhould 1972). In temperate regions general annoyance from hordes of
adults and severe reactions from their “bites make black
flies economically important. Revenue lost by tourist, lumbering and other outdoor-related industries in spring
and early summer can be excessive (Fallis 1964, Jamback
1973, 1976). Sensitized individuals or those with multiple bites may devlop severe reactions leading to
hospitalization (Fredeen 1969, Newson 1977)- Eastern
Equine Encephalitis virus was isolated from both Simulium
meridionale Riley, which feeds on humans during autumn
(DeFoliart and Rao 1965)* and Simulium .johannseni
1 2
Hart that had been fed on infected birds (Anderson et al. 1961). '.•California Encephalitis virus has also been isolated from black flies (Sommerman 1977)*
Monetary loss due to black flies has been better documented for domestic livestock production. Hearle
(1938) reported a 1 week loss of $500,000 from decreased production and death of over 3°0 cattle in Canada.
Outbreaks of Simulium venustum Say and Simulium articum
Malloch have caused drops in milk production of dairy cattle
of up to 50% and severe weight loss of beef cattle
(Fredeen 1958, 1989» Steelman 1978, Shemanchuck 1977)*
Black fly bites can cause severe dermatitis in cattle
requiring treatment (Burghardt et al. 1951)* Simulium
ornatum Meigen vectors Onchocerca gutterosa (Neumann)
to cattle in Europe (Eickler 1977)* A similiar disease
caused by Onchocerca lienalis Raillet and Henry has
recently been discovered in central New York. Simulium
jenningsi Malloch complex, which is common throughout
eastern United States, is the suspected vector (J. B.
Lok and E. W. Cupp person, comm.). Information on the
pathological effects of these two diseases is lacking.
Black flies are also vectors of hematozoan
parasites causing death in birds (Grenier et al. 1975)-
Savage and Isa (1945) reported that black fly-transmitted 3
Leucocytozoan infections killed 5000 turkeys out of a flock of 8000 in Manitoba, Canada. Leucocytozoan simondi
Mathis and Leger was responsible for massive losses of domestic geese in the subarctic (Laird and Bennett
1970). Tarshis (1973) and Herman et al. (1975) reported that black fly transmitted Leucocytozoan sp. caused
71-84^ mortality in Canada Geese goslings preventing
desired population increases at the Seney National
Wildlife Refuge in Michigan. Leucocytozoan smithi
(Lavern and Lucet) affects domestic turkey flocks
throughout southeastern United States (Noblet et al.
1975, Snoddy and Noblet 1976) and recently has been
isolated from turkeys,in New York (E. W. Cupp person,
comm.). A complete review of the diseases vectored
by black flies is included in Crosskey (1973) and
Fredeen (1977)*
The need for black fly control in the northeastern
United States has increased in recent years due to
expanded use of outdoor recreational facilities, parks,
and the improved quality of waterways in which black
flies breed. For example, reduction of pollution in
the Penobscott River in Maine has resulted in a late
summer and early fall problem with the newly described
man-biting species Simulium penobscotensis Snoddy
and Bauer (May et al. 1977)• This species is now 4
found in abundant numbers in late summer in the Miller's
River, Franklin Co., Massachusetts (Simmons unpubl.
data). It is also likely that the restoration of
beaver (Castor canadensis Kuhl) populations has con¬
tributed to black fly problems in Massachusetts.
Outlets to beaver ponds are preferred breeding sites
of Simulium venustum (CC cytotype) which is a severe
pest of humans and horses (Simmons unpubl. data).
Larviciding is the most effective way to control
black flies (Jamback 1973) due to the extensive flight
range of adults (Baldwin et al. 1975). However,
environmental and health risks are associated with
use of conventional chemical insecticides in waterways
used for domestic consumption and recreation. Since no
insecticede is registered for black fly larviciding
in the United States alternative control measures are needed. Approaches that have been taken include
cultural practices (Granett and Boobar 1979), insect growth regulators (Thompson and Adams 1979), and biological agents, which offer the greatest potential
(Laird 1981). Among biological control agents the bacterium Bacillus thuringiensis serotype Hrl4 is most promising. It is effective over a wide range of species, instars, stream conditions, and has low toxicity to most 5
non-target organisms (Lacey et al. 1980, Frommer 1981,
Molloy 1981, Molloy and Gaugler 1981). Control of black flies with B. thuringiensis has been reported 34 kin below the bacteria release site (Lacey et al. 1980).
Sampling Preimaginal Black Flies
Effective implementation of a pest management
program to control black flies will require a thorough
understanding of preimaginal population dynamics.
Central to this problem is accurate estimation of
population densities. The most common method of sampling
preimaginal black flies is placement of substrates
of known area in the stream. Substrates that have
been used include: 1) white tiles (Zahar 1951)»
2) ceramic fireplace tiles (Lewis and Bennett 1974,
Boobar and Granett 1978, Gersabeck and Merritt 1979),
3) boards (Wolfe and Peterson 1958), 4) plastic, metal
or concrete cones (Phillipson 1958, Holbrook 1967,
Benefield et al. 1974), 5) polyethylene tape (Williams
and Obeng 1966, Doby et al. 1967, Boobar and Granett 1978,
Fredeen and Spurr 1978, Ross and Merritt 1978, Gersabeck
and Merritt 1979), 6) cheesecloth (Tarshis 1968),
7) rope (Fredeen and Spurr 1978), 8) vegetation (Disney
1972), 9) bricks (Ali et al. 1974) and 10) styrofoam 6
balls (Walsh et al. 1981). Lewis and Bennett (197*0 suggested ceramic tiles he adopted as standard samplers to allow comparison between studies. However, it has since been shown that tiles are not effective in large, deep rivers where trailing vegetation is the predominant
substratum (Boobar and Granett 1978, Fredeen and Spurr
1978).
Sampling preimaginal black flies with substrates
requires complete knowledge of the ecology and behavior
of the species in question, particularly since populations
are contagiously distributed (Ross and Merritt 1978).
Disney (1972) listed the following factors as determinants
of the number of black fly larvae that will settle on
an artificial substrate: 1) population density, 2)
abundance of other suitable substrata, 3) intensity
and nature of the factors causing larvae to leave
their present substrate, 4) length of time the sub¬
strate is exposed, and 5) acceptibility of the substrate
to the species in the river. Disney (1972), Lewis
and Bennett (1975), Colbo and Moorehouse (1979),
Gersabeck and Merritt (1979) and Boobar and Granett
(1980) showed that various physical factors such as
water velocity, exposure time, and depth influenced the
number of black flies attaching to artificial substrates. 7
Lewis and Bennett (197*0 and Boobar and Granett (1978) reported close correlation of black flies captured on artificial substrates vs. natural substrates but \ allowances for surface area differences were not made.
Studies of colonization rates of artificial substrates suggest maximum densities are reached within an average time of one week (Boobar and Granett 1978, Fredeen and
Spurr 1978, Gersabeck and Merritt 1979)- Further studies on all factors associated with sampling preimaginal black fly populations are needed.
Ecology of Preimaginal Black Flies
Microdistribution. Microdistribution of black flies is defined as the distribution within a specific habitat (Colbo and Wotton 1981). Black flies are often reported as having a contagious distribution but
Ross and Merritt (1978) were the only investigators to describe it statistically as fitting a negative binomial.
The major reason for the contagious distribution of black flies is that larvae predominantly filter feed (Kurtack 1978) and attach to exposed substratum surfaces so they can filter seston from the current with their head fans (Wallace and Merritt 1980).
Thus, larvae congregate on substrates that provide 8
proper physical conditions for feeding.
Several investigators have shown that species prefer a narrow range of current velocities (Harrod
1965, Phillipson 1956, 1957. Lewis and Bennett 1975)-
Kurtack (1978) studied 9 Nearctic species of larvae and
found feeding was more efficient at 30 cm/s than at
50 or 70 cm/s. Chance (1977) reported that larvae
select areas of substratum where current flow is rapid
and the boundary layer (Hynes 1970) is thinnest.
Larvae may actually feed more efficiently in turbulent
microcurrents (Kurtack 1978)> perhaps due to more complete
mixing of seston and increased chance of seston
sticking to mucoprotein secretions of the fan rays
(Ross and Craig 198°).
Substratum Preference by Larvae. Substrate preference
by preimaginal black flies has been noted for a number
of species (e.g. Zahar 1951. Maitland and Penney
1958, Carlsson 1962, Chutter 1968, Colbo and Moorehouse
1979). This may be due to one or more of the following
factors: 1) the nature of the stream substrata and
current flow over it, 2) species preference for specific
habitats, such as riffles, 3) availability of substrates,
4) substrate surface topography and the distribution
of microcurrents and 5) placement of eggs by females 9
(Disney 1972, Colbo and Wotton 1981). Selection of
artificial substrates for sampling black flies must
take these factors into consideration. For example,
-\ Boobar and Granett (1978) found that polyethylene
tapes were more effective than tiles for sampling
species that normally colonize trailing vegetation
while Gersabeck and Merritt (1979) found the reverse
true for species that colonize rocks.
Density of Black Flies on Substratum. Black fly larval
populations can reach tremendous densities. Tarshis
and Neil (1970) reported populations so dense that
larvae created their own substrate by forming "silk
cables and webs" to provide space for attachment.
Personal observations of this phenomenon occurred in
a population consisting primarily of Cnephia omithophilia
Wood, Davies and Peterson at the outlet of Wickett
Pond, Franklin Co., Massachusett. Competition for
space can thus be an important component in the ecology
of black flies (Fredeen and Schemanchuck i960, Kureck
1969).
Gersabeck and Merritt (1978) reported that late
instar Prosimulium mixtum/fuscum Syme and Davies larvae maintain a space around their anchor points causing 10
occasional intraspecific fights. Disney (1972) reported the same phenomenon for Simulium sp. in Africa.
Colonization experiments on artificial substrates suggest
densities reach a maximum and then level off after ca.
one week (Boobar and Granett 1978, Fredeen and Spurr
1978, Gersabeck and Merritt 1979)* Pearson and Franklin
(1968) observed a significant correlation of black fly
population density with levels of drift. Competition
for space with other colonizing insects may also be
a factor. For example, Disney (1972) reported an
increase in Simulium sp. relocation rate when nocturnal
mayflies were added to their troughs.
Dispersal of Black Fly Larvae. Downstream movement
( = drift) of black fly larvae is accomplished by
extrusion of a silk thread, called a "life-line"
(Hynes 1970), from the salivary glands in their mouths
(Craig 1977) and descending in the current until the
thread catches onto another object (Ruhm 1970). The
distance travelled by drifting larvae depends on stream
discharge and nature of the substratum. Based on
captures on ceramic tiles placed at various depths,
Lewis and Bennett (1975) concluded that larvae drift
closer to the water surface where current is fastest. 11
There are 3 major categories of drift hy stream invertebrates: 1) behavioral - drift patterns charac¬ teristic to a species resulting from response to a
stimulus, 2) constant - low level drift resulting
from factors such as accidental loss of substrate
contact, and 3) catastrophic - physical disturbance
of bottom fauna, usually by floods and subsequent
bottom scouring, drought, and anchor ice (Waters 1972).
Separation of behavioral and constant drift is not
possible in field studies (Waters 1972).
Reviews of the literature on black fly drift are
included in Hynes (1970), Waters (1972), Muller (197*0.
and Colbo and Wotton (1981)- Drift patterns with
night peaks are most common (Anderson 1966, Muller
1966, Chaston 1969, Bishop and Hynes 1969» Elliot
1969, Hynes 1970, Reisen 1977) But diurnal peaks
also have been reported (Clifford 1972, Colbo and
Moorehouse 1979» Wotton et al. 1979)*
The cause of black fly behavioral drift as well
as other aquatic drift has been the subject of much
speculation. Aside from documented cases of drifting
by newly eclosed first instar larvae to feeding sites
(Rubztov 1964, Ruhm 1970, Colbo and Moorehouse 1979)
there is little experimental evidence to explain black 12
fly drift. Chaston (1969) demonstrated in artificial stream and field experiments that it was triggered when light dropped below a certain critical threshold, which was found to be a mean of 1.584 lux. However, none of the experiments in Chaston’s study were run with black flies only and thus interaction with other insect species cannot be excluded. Nonetheless, drift
is somehow related to light intensity as drift suppression during full moonlight has been observed (Anderson 1966,
Bishop and Hynes 1969).
Drift of other aquatic insects has been associated with increased chance of being swept off rock surfaces
during nocturnal feeding periods (Waters 1972),
but black flies have no diel feeding pattern (Kureck
1969) and remain on the photopositive zone of the
substrate continuously. A consequence of this behavioral
trait is that black flies, which are relatively
sessile (Colbo and Wotton 1981), would come into
contact with foraging nocturnal insects (Hynes 1970,
Muller 1974). This has led to the suggestion that
drift may be caused when larvae are simply knocked off
the substrate by other insects (Minshall and Winger
1968, Elliot 1969, Kurck 1969* Disney 1972, Colbo and
Wotton I98I, Wotton et al. 1979)- Disney (1972)
provided the only experimental evidence for this
possibility by demonstrating an increase in relocation 13
of Simulium sp. in troughs when nocturnal mayflies were added.
Black fly larvae have a number of natural predators including Trichoptera (Burton and McRae
1972), Chironomidae (Simmons unpubl. data), Plecoptera
(Siegfried and Knight 1976 a, b) and fish (Hynes 197°) •
Therefore, it is logical to hypothesize that a major portion of black fly drift might be caused by
predator escape response. Predator: prey interactions
have only recently been considered an important factor
in drift when it was shown that mayflies respond
defensively to stonefly predators by drifting (Peck-
arsky 198O, Walton 198I). Peckarsky (1980) has
proposed that stoneflies may be a major factor con¬
trolling the distribution of lotic dwelling invertebrates. CHAPTER II
THE SIX LARVAL INSTARS OF THE SIMULIUM TUBEROSUM
COMPLEX (DIPTERA: SIMULIIDAE)
Introduction
Simulium tuberosum Lundstrom complex is abundant throughout North America in a wide range of stream types (Stone and Snoddy 1969) and is reported as a biter of man and domestic animals (Stone and Snoddy
1969, Magnarelli and Cupp 1977)* Little information exists on the larval biology of this species, including data on the number of instars. This lack of diagnostic information hampers quantitative life history studies.
To facilitate my studies on sampling and drift of
S. tuberosum in the Saw Mill River, the number of larval instars was determined.
Materials and Methods
Simulium tuberosum larvae were collected weekly during August 1979 and from late June to mid-July 1980 from the Saw Mill River, Franklin Co., Massachusetts,
9 km below Lake Wyola. Each collection period encompassed
14 15
approximately an entire generation starting with a peak in first instars. Larvae were preserved in 95$ ethanol in the field and examined within 1 week.
Postgenal lengths (PGL) were measured following the procedures of Fredeen (1976) and Ross and Craig
(1979)* Measurements were made to the nearest 0.5 division of an ocular micrometer with 120 divisions per mm (Craig 1975) at 25 or 50X using a binocular microscope.
A size frequency distribution (Fredeen 1976) of
the PGL suggested six larval instars. Instar groups
constructed from this distribution were tested for
significance with a Student's t-test (Elliot 1977)•
To test whether any instar had been missed the log^Q
of the mean PGL of each instar was plotted against
the representative instar number. A straight line
indicates that no instars have been overlooked
(Wigglesworth 1972). Additionally, both PGL and
body length of 80 individual larvae were measured
(Resh 1976) and the former regressed on the latter
to determine the goodness of fit of the data to a
straight line.
Body lengths and the number of primary cephalic
fan rays were also determined for each instar. Larvae 16
were selected at random t>y placing them in an 11 dram vial filled with 95^ ethanol, shaking the vial, and
removing them with a pipet with a 3 mm opening. Each
larva was then sorted to instar “based on PGL and total
body length was measured (Fredeen 1976, Ross and Craig
1979)• Primary cephalic fan rays were counted at
100X on a compound microscope by removing a fan and
placing it on a microscope slide with a cover slip.
A Student's t-test was used to test for significant
differences in body length and number of fan rays
between instars.
Results
The S. tuberosum larvae examined in this study
have 6 instars, as determined by significant differences
(P^ 0.001) between the means of the PGL (Table 1).
Brook's Ratios were all in the vicinity of 1.4 (Table
1) supporting Brook’s (Dyar's) Rule (Crosby 1973)*
Crosby's Ratio of the difference of Brook's Ratio
between instars did not exceed 10% (Table 1), indicating
that no instar was overlooked (Crosby 1974, in Craig
1975). The plot of the log1Q of the mean PGL versus
instar number is a straight line (Fig. l) indicating that no
instar was missed (Wigglesworth 1972). The significant Table 1. Postgenal lengths of Simulium tuberosum instars (micrometers). * * * -P - 00 -P rQ +3 - o O - O X -H £ CQ 0 ft £ PQ >vH cq £ O ft CQ cd CQ O 0 o -p £ cc; 00 Q 00 w § s ft -ft M -p £ 0 cd £ ttf) 0 cd £ cq cd £ • • • • -3- VO 00 VO VO 00 ft- CO x-H 0s- ft- o O CM -ft t—1 * * * 1 • • vO CN- ON vo X—1 CM • • VO VO VO o -=3- + x“H CM ft- o ON VO (M X—1 T—1 av \—1
P <0.001 18
Figure 1. Plot of the mean postgenal length (log1Q) against the respective S. tuberosum instar. LOG OF MEAN POSTGENAL LENGTH 19 20
correlation coeficient of the regression of PGL on body length (Fig. 2, R2 = 0.94, Y = -424.57 + 11.8?,
P>< 0.001) also indicates that no instar was overlooked
(Resh 1976).
Individual larvae selected at random and sorted to instar by their PGL had significantly different body lengths with no overlap between instars (Table 2).
The number of rays on the primary cephalic fan is not as diagnostic for separating instars as PGL or body length since the difference between the mean number of fan rays for first and second instars is not significant
(Table 3)* The number of fan rays is significantly different between the other instars, but there is overlap between instars 4 and 5> and 5 and 6 (Table 3)*
However, where overlap exists developmental traits can be used to insure instar separation. First instars can be distinguished from all others by the egg burster on their cephalic apotome. Fourth instars are dis¬ tinguishable from fifth instars by the degree of development of the respiratory histoblast. In fourth instars the respiratory histoblast is developed only slightly and there are no filaments present. In fifth instars, filaments are starting to develop but individual 21
Figure 2. Linear regression line of body length against postgenal length of individual larvae. BODY LENGTH (JLj M ) 22 Table 2. Mean body lengths of the six Simulium tuberosum instars (micrometers) 23 Table 3* Mean number of primary cephalic fan rays of the six S. tuberosum instars TJ -p -P 00 -p £ CQ 2 0) GO Q w GQ S -It PC H £ 0 ctf £ CD fctO ctf £ • • • • NO NO UN CM O \-1 tH CM o o CM T—1 ON -It • 1 S GQ • • • • UN O IN- • NO it o ON o CM CM t—1 o CM t—1 UN CM vH -It cm n'A ^ • 1 * 9 • T—1 ON O • CM -cH NO CO ON IN- O ^—1 O IN- o vH -It CM CM • 1 * • • -t CO ON • tH on IN- UN UN 00 UN CM ON O o CM ON CM -It 1 • * * • • NO CN CM • NO -3" CM ON o UN IN- CM ON o O -It ON CM IN- 9 * * 1 • • CO UN on • NO 00 rH IN- ON o ON CM ON IN- -It ON IN- ON O • 1 • ■ o o Ph 9/ * * V/ o pH o 24 25
filaments cannot be discerned. Sixth instars are easily separated from the others by their fully developed respiratory filaments, which are white in recently molted larvae and black in fully mature individuals.
The cervical sclerites are separated from the post¬ occiput only in sixth instar larvae.
Discussion
Six larval instars of S. tuberosum were clearly
differentiated using 2 continuous characters, postgenal
length and body length. A discrete character, the number of primary cephalic fan rays, was diagnostic
for all instars except the first and second. Un¬
doubtedly a more comprehensive morphometric analysis
would reveal other diagnostic characters, as has been
the case with other black fly species (e.g. Craig 1975*
Ross and Craig 1979. Ross 1979). The characters used
in this study apear to be sufficient for S. tuberosum.
The number of larval instars reported for other
black fly species ranges between 4 and 9 (see Ross
and Merritt 1978 and Colbo and Wotton 1981 for reviews)
with 6-7 being the most common. Local environmental
conditions may influence instar number as shown for
Prosimulium mixtum/fuscum complex Syme and Davies by
Ross and Merritt (1978). However, the variation in 26
instars observed by these authors may also have been related to the species complex posed by P. mixtum/
fuscum (see Rothfels and Freeman 1977)- The fact that S. tuberosum also is a complex of at least 5
sibling species (Rothfels 1979). is widely distributed,
and is found in a variety of lotic habitats also
raises the question of whether 6 instars is the rule
for S. tuberosum. Ideally, laboratory rearing of
different siblings under varying environmental con¬
ditions is the best method for absolute instar det¬
ermination, but obtaining fertile eggs from individual
S. tuberosum females poses problems. Nevertheless,
studies based on field populations provide an essential
framework for much needed quantitative ecological
studies of black fly larval populations.
Conclusions
1. The S. tuberosum population 9 km below Lake Wyola
on the Saw Mill River had 6 larval instars.
2. Postgenal length and body length were the most
accurate characters for instar separation. CHAPTER III
ANALYSIS OF CERAMIC TILES FOR QUATITATIVE
SAMPLING OF PREIMAGINAL BLACK FLIES (DIPTERA:
SIMULIIDAE) IN A ROCKY BOTTOM STREAM
Introduction
Accurate sampling of preimaginal black fly pop¬ ulations is critical to properly assess population density for bionomic and control studies. The most common sampling technique is to place various artificial substrates of known surface area in the stream (Zahar 1951»
Wolfe and Peterson 1958, Williams and Obeng 1962,
Doby et al. 196?, Johnson and Pengelly 1966, Tarshis
1968, Disney 1972, Ali et al. 197*1, Lewis and Bennett-
197^, Boobar and Granett 1978, Merritt 1979, Walsh et al. 1981). Lewis and Bennett (197*0 suggested that ceramic fireplace tiles be adopted to allow comparison between studies. Subsequent studies in large rivers have shown that tiles are less effective than trailing vegetation mimics such as polyethylene tape or rope
(Boobar and Granett 1978, Fredeen and Spurr 1978).
Tiles are effective samplers for black flies in shallow, rocky bottom streams (Lewis and Bennett 197*1 a, b, 1975*
27 28
Gersabeck and Merritt 1978, Simmons unpubl. data).
During a 2-year study on the bionomics of black flies on the Saw Mill River (Simmons unpubl. data) it was observed that more detailed information on sampling techniques was needed in order to properly interpret data from ceramic tiles. Most sampling studies with artificial substrates have compared capture rates on different types of substrate but lack detailed data on other aspects of sampling.
Five specific aspects were selected for detailed investigation: 1) spatial dispersion parameters of Simulium tuberosum Lundstrom, 2) tile placement,
3) comparison of densities on natural substrates versus tiles, 4) comparison of S. tuberosum stages captured on tiles versus natural substrates, and
5) tile colonization rates.
Materials and Methods
Field study sites. Sampling was done in the Saw Mill
River, Franklin Co., Massachusetts. Sample stations were: 1) Lake Wyola outlet from just below the dam to 75 M downstream, 2) three km below Lake Wyola,
3) four km below Lake Wyola, and 4) nine km below
Lake Wyola. Each station had predominantly large 29
cobble, an average depth of 10-30 cm and a width ranging from ca. 7 M at station 1 to 10 m at station
4. The only vegetation was sparse Frontinalis sp.
Description of the tiles. Tiles were rust-colored
(American Oleon, Canyon Red color), which was the color Lewis and Bennett (1974) found most attractive to larvae and were 15 X 15 X 1 cm with 2X2 mm
contours on one side. Contoured tiles were used on the suppostion that they would provide microhabitats
similar to rocks. Total surface area and weight were p 625.5 cm and 640 g, respectively. Tiles were larger than those used by Lewis and Bennett (1974) because preliminary studies indicated they were more stable
in riffles and during spates.
Determining rock surface area. The approximate surface
area of a rock exposed above the substratum was
determined by firmly pressing light-weight aluminum
foil over the rock until a smooth film covered all
contours. The foil covering the section of rock that
was buried in the substratum (buried and exposed rock
regions can be differentiated on the basis of color)
was trimmed and the remaining foil cut so it could be
evenly flattened without altering surface area. The
foil outline was traced with a compensating polar 30
planimeter to determine area. Accuracy of this technique was established by determining surface area
of several odd-shaped objects of known area. Det¬
erminations were within 95^ of the actual area.
Placement and collection of tiles. Tiles were placed
among the cobbles in the stream at about a 20° angle
with the contoured side up and perpendicular to the
flow. In preliminary tests there was no significant
difference among tiles placed with contoured or smooth
side up„ Unless otherwise noted tiles were left in
the stream for 7days prior to counting. To count
the black flies on a tile, the tile was approached
from downstream and slid from the water as smoothly as
possible. Drift nets placed below the tile indicated
that few larvae detached with this method. While
holding the tile over an enamel pan, the larvae and
pupae were counted and then transferred to 95^ ethanol.
The tile was then returned to its original position
in the stream and depth and current velocity (Model
No. 2030 General Oceanics current meter) were measured.
Experimental procedures. Population dispersion
parameters of S. tuberosum were determined from counts
on 75 tiles randomly placed in a riffle at station 4.
A comparison of captures on tiles placed in a random 31 versus stratified random (subhabitats in riffles where larvae congregate) design was made by placing 10 tiles in each treatment at stations 1-4. Preimaginal densities on rocks and tiles were initially compared by positioning 25 tiles in a random design at stations
1-4. On the day tiles were sampled an equal number
of rock samples were randomly collected in a similiar manner. Larvae were counted by washing them off the
rock with ethanol into an enamel pan. Larvae and pupae remaining on the rock were counted with the
aid of a flashlight and magnifying lens. Current
velocity and depth were recorded for each rock sample.
In an effort to improve the fit of tile versus rock
density estimates, 10 additional tile and rock
samples were taken at each station using a stratified
random design.
The effect of tile angle on capture rate was
determined by comparing the number and position-.-Df
larvae collected on tiles placed at 0, 20, 40, 60
and 90° angles relative to an even channel of stream
bed at station 4. Tiles were positioned on a redwood
board anchored parallel to the bottom by steel posts
(Fig. 3) and were held at the correct angle by dowels
positioned on their underside. Tile positions on the 32
Figure 3* Redwood board used to hold tiles at various angles relative to the stream bottom. Tiles were held at desired angles by dowels placed on their underside. The board was placed in the stream parallel to the bottom and perpendicular to the direction of stream flow. 33 34 board were randomized. Following inspection, each tile was returned to its original position on the board and current velocity was measured directly over it, on each side and 40 cm upstream.
Four repetitions lasting 48 h each were conducted.
The samplers collected only drifting larvae.
Simulium tuberosum was the only species at station
4 during the tests.
Tile colonization rates were determined by comparing S. tuberosum captures on tiles left in the for 24, 48, 96 and 144 h. Twenty tiles placed in a stratified random design were used for each treatment.
All other insects on the tiles were counted and identified to family.
Tiles became darkly stained after prolonged use in the stream. The effect of this staining on capture was tested in February-early March and
August by comparing black flies captured on new and old tiles placed in a stratified random design at stations 1 and 2. New tiles had never been used while old tiles had been used for at least 9 months but were cleaned weekly with a nylon brush.
Results
Population dispersion. Based on the Chi-square test 35
of the variance to mean ratio (Elliot 1977) > "the
S. tuberosum population at station 4 was contagiously
i ^ distributed (mean and standard deviation = 33*43 -
39.28, normal variable = 365*32). The population fits a negative binomial distribution as determined by the statistic U method (Elliot 1977; k = 0.42, determined by the proportion of zeros method, U = -1150.71» standard error of U = 2.31)* Since U is less than its standard error, agreement with the negative bi¬ nomial is accepted at the 5$ level (Elliot 1977) •
The low k value is indicative of pronounced clumping
(Pielou 1977). The 15 other black fLy species
in the Saw Mill River (Table 4) were not examined
for spatial dispersion parameters but weekly natural
substrate and tile samples at 14 stations in 1977
indicated they were also contagiously distributed
(Simmons unpubl. data).
Tile -placement. Significantly more larvae and pupae
were collected on tiles from the stratified random
design at each station (Table 5). The average difference
in the number captured per tile ranged between 7-100
more on the stratified samplers than on randomly
placed samplers (Table 5). The variation between
tiles was large for both designs but it was noticably 36
Table 4. Black fly species collected on both tiles and natural substrates in the Saw Mill River, 1977-1980.
Prosimulium fuscum Syme and Davies P. magnum Dyar and Shannon P. mixtum Syme and Davies Cnephia dacotensis Dyar and Shannon Cn. ornithophilla Davies, Peterson and Wood Stegopterna mutata (Malloch) Simulium aureum Fries S. croxtoni Nicholson and Mickel S. decorum Walker S. fibrinflatum Twinn S. jenningsi Malloch S. quebecense Twinn S. tuberosum Lundstrom S. venustum Say S. verecundum Stone and Jamback S. vittatum Zetterstedt
less for the stratified design (Table 5). The percentage
of the total of each species collected on the tiles
during the test was: STATION 1 - S. decorum (17)»
S. tuberosum (47), S. verecundum (12), and S. vittatum
(24), and STATIONS 2-4 - S. tuberosum (99) and
S. verecundum (1).
Density of black flies on rocks compared to tiles.
Tile versus rock samples indicated tiles provide a
reasonable estimates of natural density but sample
design is important. On the random samples there
was an average of 3*9 times more black flies collected Table 5. Comparison of the number of preimaginal black flies captured on tiles placed in a random versus stratified random design. i o i £ I ■ 1£ ; to | I 1 1 I I I 0 ] |TO 1 |CH I |«H ? icd i 1-p \ *H i GO ! I-P 1 1 ■ cd Oh * * * * \ •H \ O i—1 EH Q GO •H S + 1 O PO EH 1-1 05 Oh Q 0 GO •H S + 1 ex £ s 0 cd £ UD 0 -P GO -p cd 05 £ 0 £ 0 cd UD 0 S • £ o cd • • • • •3* NO NO O NO On \—I -t- co £>- o -b- O O- o- ^—! CNJ + 1 CNJ O- O o CNJ ntn CNJ ON o 0~N o tH tH ON o + 1 ON ^—1 ° T—1 • 1 • • 1 • • it NO NO CNJ O NO T—1 r—1 O \-1 -f CNJ \-1 CNJ -=t o NO + 1 T-1 o rH o ON ON ON O tH rH o + 1 o CNJ m 1 • • 1 1 * • it* NO On o o On rH CNJ 00 00 00 o CNJ o NO T—1 o + 1, T—1 o tH o ON ON tH + 1 o vH o • 1 • • 1 1 • • -cf NO NO NO -t- -Tt -t \—1 ON CNJ o o NO oo CNJ o tH CNJ -t CNJ + 1 o ON ON o o T—1 CNJ ^—1 T—1 o o + 1 1 • • • 1 1 • • * •H TO •H I—I -P -P i—I -P S 0 0 m o O £ 0 cd >5 > 0 O O 05 u u 0 {0 $0 O 0 cd VO o •H 00 • -P -P -p -p Q «H rH Ch On V/ £ 0 0 0 Oh 0 cd 0 £ m 0 0 £ 0 S O £ Oh o ro £ cd • o v—' Ph O 37 38
2 on tiles than rocks per 100 cm at stations 1-3
(Table 6). Station 4 was excluded because of the zero estimate/100 cm2 for the rock samples. A Mann-
Whitney U-test of the raw data indicated significantly more black flies were collected on tiles than rocks at each station (Table 6). The average tile/rock ratio
o was 0.79 black flies per 100 cm of the stratified random samples at stations 1-3 (Table 7)• Significantly more larvae and pupae were collected on tiles at stations
1 and 4 based on the raw data (Table 7)* The per¬
centage of each species collected on tiles and rocks
at station 1 was similar (Table 8). Stations 2-4
were dominated by S. tuberosum.
Frequency of instars captured on tiles versus natural
substrates. The percentage of each of the 6 S. tuberosum
instars (Chapter II) and pupae collected on tiles
versus natural substrates was not significantly
different though 6 times as many were collected on
tiles (Table 9). Natural substrate and tile samples
during 1977-1978 also indicated that tile collections
provided accurate estimators of natural population age
structure of the other black fly species in the
Saw Mill River. The exception to this was first
instar Prosimulium which were rarely collected on
tiles, probably because they do not filter feed at Table 6. Comparison of the number of preimaginal black flies collected on tiles and rocks using a random sampling design. •H Eh i—l X od 0 co o o CO * * * * * * * ❖ * * * •H 00 P Pd O -p Pd o £ £ 0 0 o cd W) cd £ cd £ £ § hD 0 0 £ £ £ cd -3- 00 00 00 M3 ft >A 'A ^A t—1 t—1 CA ft t—1 na na ^A CA ca CA ft ca ft ^—1 ON ca 1 1 • • -3* -3- NO VA ft ft NA O ON ft >A T-1 o O CA ft ft 1 1 • • -3* -3- -3- NO -3- ft NA CA o ft O CA XA T—1 NA \—1 ft O CA o 1 1 • • -=t NO VA ft o ft tH O- NA ft \—1 o O ON ft 1 1 • Pd P to p £ W) cd 0 O £ cd £ cd cd • * ft \ S p *1—3 H P Td to p p tH p rO o o 0 £ O cd CD cd P 0 CD o Ph £ cd cd £ cd CD 0 £ o £ £ 0 £ o £ * * \ V-/ /—. •H S •H P i—1 P & P P 1—1 I—I £ £ 0 cd o £ £ 0 £ > O 0 O >5 o £ CD o £ 0 O 0 o cd o £ £ o co ft 0 I * 00 * o * ^ •h Pd •H ,£ •H *i—1 g cd ^ d) P o £3 tH P On p • P Q p 0 •H fccO P 0 TO 0 •H p cd i—1 r—1 >5 P *H •H £j ro p •H £ r£ >)P p ft cd i— 0 H CD NO -p -P cd CD o p £ £ -P cd 0 CD O O CD £ £ cd 0 £ £ O i—| o & £ *H 6 fcio Cd ,£ to £ £ ft £ 0 p i £ CO cd m o 39 On P •H i—l X' o o O CD o £ cd 4o
cq P o ft o -P CQ £ cd 0 EH £ ft o £ cd • p o ft CD CM ft £ u ft cd £ o •H O -p £ ft ft o fcuOft cd o •H ft o CQ a CO CQ 0 0 *H 0 0 1—1 £ CQ •H ft ft 0 I—I i—I £ ft ft ft O CQ cd £ P P o o P £ 0 cd O 0 £ i—l cd i—l o ft 0 P P £ ft £ 0 H P cd ft £ o or •H >3 £ £ 0 P 0 tiO ft cd 0 i—i p ft P ft £ ft 0 0 •H ;> ft 0 p O u o ft >3 ft O cd ft P ft 0 o ft ft p o P £ 0 CQ <: £ 0 O & ft ft £ O 0 •H 0 i—1 P CQ CQ 0 £ P i—1 P •f—3 CQ £ o ft £ ft cd £ S P o CQ 0 0 cd —^ £ P •H • ■ 0 ft ft ft •H cd cd >5 CQ -p p ft ft ft p CQ g ft ft o o cd cd ft 0 £ £ p £ o o ft £ • £ i cd £ ft o o CQ ft I—I P 0 ft o CQ cq cd 0 i—i cd •H cd £ CQ > 0 >3 ft P £ 0 I—I 0 P cd t*o £ i—I ft •H £ o ft £ ft ft cd ft £ •H £ £ ft 0 P £ ft p o ra 0 O 2 0 CQ £ o £ CQ £ i > o CQ cd ft o o 0 P O •H 0 • 0 cd ft £ ft £ 0 cd cd o ft rH P 0 ft W) 0 ft * ft £ 0 * * P p cd § * * * 0 CQ cd £ ft * * * * P cd eh cd * * * * ft £ 41
Table 8. Comparison of the percentage of the total of each black fly species captured on tiles vs. rocks at station 1.
Random_ Stratified. Species Rocks Tiles Rocks Tiles
S. decorum 4 4 2 4 S. tuberosum 21 23 71 67 S. verecundum 37 35 11 12 S. vittatum 38 38 16 17
^Sampled June 26, 1978*
•ft# Sampled July 10, 1978.
Table 9. Comparison of the proportion of the total of each S. tuberosum instar collected on 25 natural versus 25 tile samples at station 4.
■* Instar Substrate Total 1 2 3 4 5 6 Pupae
rocks 181 9.4 72.4 7.2 3-3 3-3 3-3 1.1
tiles 1104 11.9 78.4 4.7 2.6 0.8 0.5 1.1
*Chi-square test for difference among proportions - 6.47 (not significant). 42
that stage.
Importance of current velocity and tile angle. The
importance of current velocity over a tile was
demonstrated by the fact that the average velocity
was about twice as fast on stratified samples as
random samples (Table 5). Mean depth of the tiles
was the same for both treatments. However, current
velocity is not necessarily a good predictor of the
number of black flies which will accumulate on tiles.
Least squares linear regression of larvae and pupae
captured (N + 1 log^Q transformed, Elliot 1977) versus current velocity of the random and stratified
random samples combined indicated that only station 1,
a high density site, had a significant correlation (Table
10) .
Table 10. Least squares linear regression of the number of preimaginal black flies (N + 1 log1Q transformed) versus current velocity of the data from random and stratified random samples combined.
r2 Station N Line Equation Significance
1 20 Y = 0.94 + 0.02X 0.57 P £0.01 2 20 Y = 4.78 - 0.02X 0.003 N.S. 3 20 Y = -0.03 + 0.01X 0.01 N.S. 4 20 Y = 9.46 + 0.12X 0.10 N.S. 43
More larvae were captured on tiles placed at
20° than at 0, 40, 60 and 90° though the difference was not significant (Table 11). Numbers captured were low because tests were run in mid-September when S. tuberosum populations were small. The most uniform distribution of larvae on the tiles was on the 0 and 20° tiles. On tiles at 40, 60 and 90° larvae were only near the edges (Fig. 4). These
results are most likely explained by the way tile
angle affects current flow. Tiles placed flat on the board (0°) did not interrupt proximal current
(Table 12) but because they did not proturude into the water column there was no substrate for the silk
strands of larvae to catch onto. At 40, 60 and 90° the tiles protruded in the current which caused a reduction in velocity against their upstream surface and increased velocity around their edges (Table 12).
The distribution patterns of larvae on the tiles
indicate that they respond to this and attach near the edges where current is fastest (Fig. 4). It is possible that the reason fewer larvae were collected on the tiles at sharper angles is that water flowing around them created a physical barrier that prevented the silk strands of passively drifting larvae from making contact. Tiles at 20° protruded into the Table 11. Comparison of the number of S. tuberosum larvae captured on tiles placed at different angles on redwood boards. •H < i—i i—i £ 0 tin CD No o C\] o CT\ O o o -3- VO m n 00 U~\ CO CNJ +1 co co o vo £N- vo vo vo VO + I o- o o o ON co C\2 ON co o o o co +1 + I +1 + I o £ cd • • • • Ch 00 -p X—I CO CM o tH co o o cd
Four repetitions for each angle. Mann-Whitney U-test of 0 versus each of the other angles tested indicated no significant differences. 45
Figure 4. Distribution of Simulium tuberosum larvae on the upstream surface of tiles placed at 0, 20, 40, 60, and 90 degree angles relative to the stream bottom on a redwood board. Data is combined from 4 repetitions. Tops of the figures represent the tile edges nearest the water surface. Direction of stream flow is indicated by the arrow. 46
0°
STREAM FLOW Table 12. Comparison of current velocity measured at 4 areas near a ceramic tile on a redwood board at 0, 20, 40, 60, and 90 degrees. •H •H ft o -p PC ft -p ft £1 £ tuo 0 £ 0 0 PI 0 Cti o £ O £ £ Pi O O 0 -t •H *H ft £ (1) OQ ft 0 ft -p 0) pc in -p Q ft r-H £ ft o O CQ CD *H >>ft tuO ft 0 ft £ o £ o S ft P 0 S £ cd ft -3" 00 O-0s-NOCO vo ^-3-^o vo -3-no^o on covocm VO ft-COCMft ON COo0-00 o CO On CMMD CM -3-NOON O 47 48
water column without interrupting flow so silk strands would have been more likely to make contact allowing larvae to attach.
Tile colonization. Colonization experiments were run for only 6 days but the data suggest that tiles were approaching equilibrium on day 6. Sampling ended on day 6 because the riffle had become overcrowded with tiles. There was a significant increase in cap¬ tures between 24-48 h and 48-98 h but not between
96_l44 h (Table 13). The total black flies and other
insects collecting on the tiles increased at a lower
rate with time (Table 13)*
Captures on old versus new tiles. During winter (Feb.
and early March) there was no significant differences
in larvae captured on old versus new tiles (Table 14).
The species and percentage of the total captured were:
STATION 1 - P. mixtum/fuscum (78), P. magnum (2),
St. mutata (13), S. venustum (l), and S. vittatum (6),
and STATION 2 - P. mixtum/fuscum (85), P. magnum (8),
St. mutata (6), and S. venustum (1). In summer
samples (Aug.) about half as many larvae and pupae
were captured on old versus new tiles though only
station 1 samples were significantly different (Table 14).
Species and their percentages in the river at the time
of summer sampling were: STATION 1 - S. decorum (1), Table 13. Colonization rate of black flies and other insects on ceramic tiles at station 4. * * * •H ,G O -P -P 1—1 o I—I I—I -P 001 G CD 0 G cq O CQ o G 0 CQ o 0 0 o ^ G o 00 G CQ 0 G -P H 00 Q •H § s + l G G 0 0 G o 0 CQ cd 0 cd o G CQ Q) cd G 0 cd • --4 ^4 M0 \A \—1 *A CA CM f>- O- O + l CM O CM • • • ! -4 CAr-i -4" CArH ^A ^ 4-w CM O O ca vH ON m -4 -4* ^4 M0 'A O- -4- On^4 £N- On CA 0 £N- 00 MO^4 + 1 O CM • • • * * CM O- O ON O • M0 MO *A tH ^A 'A vH VA O £N- O CM On 0 + 1 CM • • • s 00 -4* • • VA CM CA 0 • -4" •4" -4- MO 00 ''A *A tH VA CM CM O CM O T—1 + • m • * o Ph O y/ m * * X/ 'A O O Ph O • * * >!< Families and percentage of each collected were: Diptera: Chironomidae (46),
Trichoptera: Hydropsychidae (42), Ephemeroptera: Baetidae (6), Ephemeroptera: 49 Heptageniidae (3)1 and others (3) • 50
£ o • • GO GO GO m * a • • a * a a t—1 o\ -P CM 00 (ft O 'ft • • 0 -a I—I T—1 tH t—1 Oft I—I o a o l—I o o 4- c°i oo ft 0 ft ft ft cd ft -S • • • ft ft CM CM ft cd 0 ^—I £ ft a o ft o TJ £ cd 62 'ft O NO • o 'ft O- ft • • • CD 00 Cd 0 GO o -3* Cft NO > T—1 a CM Cft £ •H + i a cd EH + 1 + i + 1 + 1 i—I ■ £ £ m a cd o o CM NO o 0 o o 00 00 £ >>a i—I • • H -P o • • ft ft £ CM Oft NO o- ON CM V—1 a ftl 0 o ft cd £ cd VO CN- ft ^± i—i CD oo co o eft £ I o ft £ m • ft o- o i—I o m 0 o ft 'ft 'ft cd Q o £ a •H GO CD £ EH + 1 + 1 + 1 + 1 o rQ cd + I •H NO 00 Cft ft ft £ £ 0 £ o 00 CN- NO m £ 0 a cd • • • • •H ft 0 'ft ft NO JN- ft o £ o CM Cft Cft cd -£ *H \—1 ft ft £ m i ft tuD o o o o ft O £ a CM CM CM CM •H a £ £ 0 o £ £ ft -p CQ a O w £ •H •H 0 a £ tQ ft CM ft CM ft •H cd 0 cd I £ ft H ft a ft £ ft GO w o ft •H o 0 a a £ ft >5 ft >s • o l—I •H r—I -4- £ a i—I ft id cd cd £ 0 ft 0 cd ft 0 CQ ft ft £ O ■s * a £ a cd a cd 0 0 cd £ * ft £ GO ft S < * 51
S. tuberosum (78), S. verecundum (20), and S. vittatum
(1), and STATION 2 - S. tuberosum (99) » and S. verecundum (l). The difference between summer and winter captures may either be due to species diff¬ erences or to the fact that in summer old tiles become coated with more periphyton and insects during the 7 day sampling period than new tiles. This was not observed for winter samples.
Discussion
A number of factors involving both tiles and the ecology and behavior of larvae were found to affect black fly sampling in a rocky bottom stream.
The most important factor was the negative binomial distribution of larvae. Many black fly species are reported to have a contagious distribution (Grenier
19^9» Wolfe and Peterson 1958, Maitland and Penney
1967, Elliot 1971» Disney 1972, Ross and Merritt 1978,
Colbo 1979) but Ross and Merritt (1978) were the only previous investigators to describe this distribution statistically as a negative binomial.
The reason preimaginal black flies are so clumped is due to the importance of current. Larvae are predominately filter feeders (Kurtack 1978) and require flowing water. Most species that have been 52
studied have been found to prefer certain current speeds (Harrod 1965» Phillipson 195^» 1957» Lewis and Bennett 1975, Colbo and Moorehouse 1979)- Sec¬
ondly, current provides the major means of dispersal
for larvae via drift (Waters 1972). Black fly larvae
cannot swim and rely on silk strands to stick to
substrates and halt downstream movement (Rubstov
1964, Ruhm 1970, Tarshis and Neil 1970). Therefore
water converging between two rocks and increasing
in velocity acts like a funnel and draws in drifting
larvae. These habitats tend to be the areas of
highest density. Most tiles placed in stratified
random positions were in this type of habitat. At
times of low discharge these habitats become more
abundant and owing to a decrease in other suitable
habitats will affect density estimates (Disney 1972).
Ross and Merritt (1978) observed a negative correlation
between discharge and the number of larvae captured
on tiles. Gersabeck and Merritt (1978) showed stream
velocity will affect the number and instar distribution
of larvae attaching to artificial substrates.
In addition to location in the stream, the way
tiles are placed is important because water can
create a physical barrier around a substrate which 53
prevents attachment of the silk strands of drifting larvae. This was illustrated by the capture rate and distribution of larvae on tiles at various angles (Table 11, Fig. 4). Wolfe and Peterson
(1958) reported that more larvae and the evenest
distribution was on tiles at 30° (0, 30> 40, 60, and
90° tested). Colbo (1979) demonstrated that micro¬
currents determine distribution of larvae on a sub¬
strate .
The rate at which artificial substrates are
colonized by black flies has been examined by several
investigators with results generally similiar to
the present study (Table 13)• Fredeen and Spurr
(1978) reported mesh plates attained maximum density
after 7 days while ropes took 2 weeks and floats
and polyethylene-covered plates continued to increase
until the end of the tests (3 weeks). Boobar and
Granett (1978) found that polyethylene tapes reached
a peak after 6 days and remained stable for 21 days.
Gersabeck and Merritt (1978) observed a peak on tiles
and tapes of P. mixtum/fuscum and Cn. dacotensis on
tiles after 5-7 days.
Intraspecific (Disney 1972* Gersabeck and Merritt
1978, Chapter IV) and interspecific interactions 54
(Disney 1972, Chapter IV) are known to influence
"black fly microdistribution and drift (Chapter IV) and are often listed as reasons for equilibrium densities on artificial substrates (Disney 1972,
Fredeen and Spurr 1978, Colbo and Moorehouse 1979)*
It is noteworthy that in the present study the percentage increase of black flies and other insects on the tiles decreased with time (Table 13)* It is also important to consider that tiles at sharp angles have less suitable surface area available to larvae due to alteration of current flow (Table 12). This might be important at times of high population density when greater competition for space is more likely to occur.
Only a few quantitative studies of black fly captures on artificial subtrates compared with natural population density have been made. Lewis and Bennett (1974) reported nearly identicle rock versus tile density in Newfoundland but they did no consider surface area differences. Boobar and
Granett (1978) collected S. penobscotensis Snoddy and Bauer and S. nyssa Stone and Snoddy on polyethylene tapes in similar numbers to those collected on vegetation. In the present study there was a close correlation between 100 cm of tile and rock when 55
sampling was done in a stratified random design.
Significantly more preimaginal black flies were collected on tiles than rocks when the data were not corrected for surface area (Tables 5 and 6).
Zahar (1951) and Carlsson (1962) reported that artificial substrates became fouled with algal slime after a period of time in the stream. This problem was not observed in the present study, probably because of weekly sampling. However, it was observed that during the summer, tiles became stained after about a month.
They then seemed to attract more periphyton and insects and resulted in fewer black fly larvae being captured compared to new tiles (Table 14). To prevent this problem tiles may be alternated and cleaned in bleach and detergent to remove stains.
Conclusions
1. Rust-colored tiles provided accurate estimates
of the species and density of preimaginal black
flies in a rocky bottom stream. Highest capture
rates and best correlation with natural population
density were attained when tiles were placed
in the preferred habitats of larvae using a
stratified random sampling design. 56
2. Although current speed was twice as fast on
stratified random samples, least squares linear
regression of random and stratified random sample
data indicated it is not always a good predictor
of capture rates, especially at low population
densities.
3. Tile angle affects both the number of larvae
captured and distribution on the tiles. Tiles
at 40, 60, and 90° angles divert proximal current
toward their edges which might create a physical,
barrier and prevent larval silk strands from
attaching. Larvae congregated on the edges
of the tiles that were at 40, 60 and 90° but were
evenly distributed on the tiles at 0 and 20°
which did not interrupt current flow. Owing to
intraspecific interactions this could affect
density estimates if populations were large since
larvae only congregate in areas of suitable
microcurrents and tiles at sharp angles have
effectively less surface area available to larvae
and pupae.
4. Colonization data from this study and others suggests
maximum density is attained in about 1 week.
5. Long term use of tiles for bionomic studies should
include periodic replacement or cleaning of tiles 57
in a bleach and detergent solution to remove
stains which were found to reduce captures of
black fly larvae and pupae by 50% during summer.
6. Due to the large variation associated with
sampling highly clumped preimaginal black fly
populations it is unlikely that sampling results
will yield anything more than qualitative
density estimates. A prohibitively large number
of sampling units would be required to obtain
statistical accuracy (Elliot 1977)* CHAPTER IV
DRIFTING BEHAVIOR OF SIMULIUM TUBEROSUM COMPLEX
AND OTHER BLACK FLIES AND
ITS PROBABLE CAUSE
Introduction
Next to mayflies, black fly larvae are generally the most abundant insect group collected in the invertebrate drift in streams (Waters 1972). Exp¬ erimental examination of the causes of black fly drift are lacking but it frequently has been hypothesized that it is associated with activity of other inver¬
tebrates (Minshall and Winger 1968, Elliot 1969»
Kureck 1969, Disney 1972, Colbo and Moorehouse 19791
Wotton et al. 1979» Colbo and Wotton 1981). Disney
(1972) provided the only experimental evidence for this by showing an increase in Simulium spp. relocation in bamboo troughs when nocturnal mayflies were added.
Since black flies have many natural predators
(Twinn 1939, Peterson i960, Peterson and Davies i960,
Burton and McRae 1972) it is logical to question
58 59
whether drift might he a predator escape response.
Predator:prey interactions were not considered an important cause of drift until recently, when it was demonstrated that mayflies drift to escape stonefly predators (Peckarsky 1980, Walton 1981).
Black fly larvae congregate in dense patches in photopositive zones of the substratum and since
they feed continuously (Kureck 1969) they do not move to different areas of the substrate during day or night, as do most other aquatic insects (Hynes 1970).
However, many black flies have nocturnal drift peaks
(Anderson 1966, Muller 1966, Chaston 1969, Bishop and
Hynes 1969, Elliot 1969» Hynes 1970, Reisen 1977), which coincide with foraging of nocturnal insects
(Hynes 1970). Black fly larvae are legless and move in a slow, looping geometrid-like fashion, suggesting substrate detachment (=drift) would be their only means to quickly escape predation.
The objectives of this study were to: 1) dem¬ onstrate the diel drift pattern of Simulium tuberosum
Lundstrom complex in nature, 2) determine the foraging periodicity of perlid stonefly predators in an artificial
stream, 3) determine the prey consumed by stonefly predators versus that which is available to them 6o
in their natural habitats in nature, and 4) quantity the role of drift in response to stonefly predators in an artificial stream. Simulium tuberosum was used because it inhabits rocky riffles where stone- flies are abundant (Simmons unpubl. data) and was present in low to medium densities at the study site during summer. Observations on the defensive behavior of Prosimulium mixtum/fuscum Syme and
Davies, Stegonterna mutata (Malloch), and Simulium damnosum Theobald complex are also included.
Materials and Methods
Field study area. Field investigations on S. tuberosum drift were conducted on the Saw Mill River, a cool- water, 4th order stream in Franklin Co., Massachusetts.
The sampling station was 9 km below the outlet of
Lake Wyola in a zone 8-10 m wide, 10-30 cm deep with
a heterogeneous rock and cobble substratum containing no trailing vegetation. Simulium tuberosum generally
was the only species present at this station during
summer months.
Sampling of S. tuberosum drift. When black fly larvae
drift they extrude a silk strand from their salivary
glands that attaches onto objects and halts their
downstream movement (Rubztov 1964, Ruhm 1970, Tarshis 61
and Neil 1970). To sample drifting S. tuberosum larvae ceramic tiles were placed 8-10 cm below the water surface on a redwood board held in position in the water column with steel posts anchored in cement blocks that were buried flush with the stream bed
(Fig. 3). Two sample boards with 5 tiles each were placed 5 m apart in a stream channel ca. 33 ® wide and 30 cm deep, and below a riffle with an S. tub¬ erosum population. Obstructions in the channel were removed so water flowed evenly over each tile. The
samplers were designed so only larvae drifting in the water column were sampled.
Tiles were the same as those described in Chapter
III. The contoured side was placed face-up and grooves were perpendicular to stream flow. Tile depth was
controlled by adjusting board depth. New tiles were
used for each experiment or repetition.
Artificial stream. The artificial stream used to
study interactions between stonefly predators and
S. tuberosum larvae was constructed and operated as
follows: water was pumped from a lower reservoir
(50 liter capacity) to an upper reservoir (13 liters)
from which it flowed over a plexiglas sluice (12 cm
wide X 5 cm long) and down a plexiglas trough
(12 cm wide X 45 cm long X 5 cm deep) with a slope 62
of 10°; two nylon screens (0.25 mm mesh opening), one fitting tightly within the other, were placed at a 3°° angle beneath the lip of the lower end of the trough to capture detached larvae; the inner screen was readily removed to count larvae.
A riffle was simulated with a removable substrate that was 18 cm long X 1.5 cm deep with a frame made of a 1.3 mm square acrylic rod. To create a dark region for stoneflies to rest in during the day the top of the riffle had four strips of black polyethylene with 0.5 cm spaces between each strip and the interior was loosely packed with stones ca. 2-3 cm long and 1 cm thick. Stoneflies readily sought refuge among the stones and black fly larvae attached to the surface of the polyethylene, an arrangement similar to their daytime niches in nature. The animals could be clearly observed through the side of the trough.
Water velocity over the substrate was ca. 35 cm/sec
(determined by timing the flow of a cork, Schwoerbel
1970), discharge was 0.4? L/s, and the depth was 3-4 cm.
Determination of S. tuberosum1s drift pattern. Twenty- four hour samples were taken August 2-3, 6-7, 22-23,
1979, and June 27-28, and July 5-6, 1980. Tiles were set at 1600 h and counted every 3 h beginning at 1900 h. Tiles on the downstream sampler were counted 63
first by individually removing each tile from the board. Night counts were made with the aid of a
flashlight. All insects on the tiles were counted
and placed in 9S7° ethanol.
After all tiles were examined, water temperature,
velocity (General Oceanics Model 2030 current meter)
and depth at 5 equidistant points of the stream transect
were measured. Boards were then cleaned with a nylon
brush to remove any insects which had settled on them.
Simulium tuberosum larvae were sorted to instar
(Chapter II). An occasional Simulium jenningsi
complex Malloch larva was collected in 1979 and
these were subtracted from the total field counts.
Plecoptera were indentified to species (Hitchcock 1976),
Ephemeroptera to genus (Hilsenhoff 1975)» and Trichoptera
to genus or species (Flint 1962, Hilsenhoff 1975»
Schuster and Etnier 1978)* Discharge was determined
by the formula of Davis (in Hynes 1970, p. 4l).
To determine the change in S. tuberosum drift
during the crespuscular-to-dark period, samples
were taken as in the diel experiments but at 20 min
intervals beginning at 1920 h (tiles set at 1900 h)
during mid-July 1980, Light readings (Panlux Elec¬
tronic Luxmeter) were made prior to each sample.
Counting could be completed in ca. 2-3 min. 64
The foraging time of perlid stoneflies was determined by taking at least 25 rock samples during afternoon and night in a section of stream below the drift samplers. Rocks were selected from preferred microhabitats of the perlids, which are also the habitats with greatest concentrations of S. tuberosum larvae (Simmons unpubl. data). Rocks were approached and removed carefully from the water so as not to disturb the stoneflies before their position was noted.
A subjective decision of stonefly position based on the rock surface being darker and rock position relative to the stream bed was made. Stoneflies were put in 95% ethanol. Perlids were later iden¬ tified, measured, and the alimentary canal removed
(Siegfried and Knight 1976). The location and digested status of prey in the foregut, midgut and hindgut were determined. Prey that were not macerated and were readily identified to at least family were considered undigested.
Prey selection by Paragentina immarginata
(Say), the most abundant stonelfy predator at the
study site, was also determined. Twenty-five rocks were sampled from a riffle with a large S. tuberosum
and P. immarginata population during early morning
by brushing all insects on them into a bucket containing 65
95$ ethanol. Paragentina immarginata nymphs were also collected from rocks. The nymphs were later dissected and all identifiable prey in the foregut and midgut was counted and measured. All insects collected on the rocks that were in the size range consumed hy P. immarginata were then counted. Prey selection was determined by calculating coefficients of electivity (Ivlev 1961) for prey items. Electivity was calculated after Ivlev (1961):
E = (ri - pi)/(ri + pi) where r^ is the fraction of the diet made up by species i, and p^ is the fraction of the total prey available composed by species i. Electivity therefore ranges from +1 to -lj zero indicates no selection or avoidance, that is, the prey item appears in the diet in the same proportion as it appears in the food complex; positive values indicate selection and hegative values indicate avoidance or inaccessibility of the prey.
Observation of predator:prey interaction in the artificial stream. The artificial stream was set outdoors under natural light. Observations made in the dark were with a red filtered battery powered light focused indirectly on the stream so the minimum light to view the insects 66
was used. A control test showed that S. tuberosum larvae did not drift or relocate in response to the red light.
Simulium tuberosum larvae and stoneflies
(when used) were collected from the field site the morning of each test. Stream water also was put in the artificial stream and kept at field temperatures
(19-22° C) with a water cooler (Aqua-chiller Model
365). One hundred and fifty 3rd-5th instar S. tuberosum were placed in the stream on the surface of the polyethylene. Stoneflies were placed in the substrate beneath the polyethylene. Stonefly species and numbers used per test were: P. immarginata
(l), Phasganophora canitata (Pictet) (1), and Acron- euria abnormis (Newman) (2). These species are
common at the study site and are known predators
(Hitchcock 1976, Peckarsky 1979)*
Simulium tuberosum drift was measured in the artificial stream with and without stoneflies present in early August 198O. The distribution pattern of S. tuberosum on the substrate was plotted
at the beginning and end of each test. Observations
began at 1600 h and the animals were observed con¬
tinuously for the duration of each test. Drift was
quantified by counting the number of larvae in the screen every 10 min, which could he done in ca.
30 s. Light readings were taken before each count.
Two repetitions were conducted for each treatment.
Stoneflies were recorded as: 1) resting beneath the polyethylene, 2) actively seeking prey
and 3) on the surface but not seeking prey. Prey
searching and attack behavior of the stoneflies
and the response of S. tuberosum was noted.
Observations using a needle to simulate predators were
also made on S. tuberosum in the artificial
stream and field.
Other observations of black fly defensive behavior.
Observations on the defensive behavior of P. mixtum/
fuscum were made on larvae on rocks and tiles in ca.
15 cm of smooth water in the Saw Mill River in mid-March. Stegopterna mutata was observed in Tyler
Brook, Franklin Co., in about 5 cm of water in late
March. Simulium damnosum s.l. were from field
eggs and F^-F^ larvae an a laboratory stream (Simmons
and Edman 1982). Observations were made using a needle to simulate predators. 68
Results
Diel drift pattern of S. tuberosum. Simulium tuberosum had a drift pattern characterized by a 2.5 fold increase at night (Fig. 5, Table 15). The night versus day difference in drift rate was significant for all 6 diel samples (Table 16).
The S. tuberosum poulation studied has 6 larval
instars (Chapter II). No obvious difference in the diel drift pattern between instars was noted (Fig.
6) except 6th (last) instars which made up a sig¬ nificantly greater proportion of night compared to
day drift (Table 17)* Also, there was no significant
difference in the percentage of each instar captured
in 24 h drift samples versus random collections
from natural substrates of the standing population
(Table 18). Few first instar S. tuberosum were
collected on the tiles and in random samples (Table 18).
Data indicate that the drift pattern is constant,
despite fluctuations in stream temperature and dis¬
charge (Table 15» Fig. 7)* There was a significant
increase in S. tuberosum drift during the day-dark
periods (1600-1900 h versus 1900-2200 h samples) on
each sample date except August 22 (Table 19)-
Conversely, a significant decrease in drift was observed 69
Figure 5. Average drift pattern of S. tuberosum larvae based on the mean percent of the total larvae and pupae captured on tiles for each sample period of each of the six diel test dates. Lines around each mean are standard deviations. 70 hour
“□iq/aaunidvo nvioi jo % x Table 15* Comparison of the percent of the total larvae and pupae captured and temperature measurements during each sample period of each of the six diel test dates. ^ -d ft ft -P ft Td ft O o cd cd P > — cd o 0 o P cd d ft 0 cd cd co m * H 0 Eh cd -p i—1 d o -p Td Eh -P •H ft ft £ cd 0 Td cd Q ft-P O ft cd d P ft P P 0 cd cd 0 P P 0 6 ft 0 cd P ft o P 0 0 PJ d o Ufr No o Td o On o CM CM o o o o o O O O P- o o o o o cp r—i o O O o 0 ft ft- 00 00
f
Figure 6. Diel drift pattern of S. tuberosum instars based on the percentage of each instar cap¬ tured for each of the 8 sample periods during a diel (1 = 1900 h, 2 = 2200 h, 3 = 0100 h, 4 = 0400 h, 5 = 0?00 h, 6 = 1000 h, 7 = 1300 h, 8 = l600_h; the letters A-E correspond to the sample dates listed in Fig. 7-). 60 H n—r © O o ^ (M aoia3d 37dNvs/63anidvo (D o o 3VAdV~l IVlOl30°/o c\j O \r
**Day and night sample periods (hour) correspond to those in Table 75 ***Significant difference between day and night samples (P'$0.05). Table 18. Comparison of the population age structure of S. tuberosum larva 0 as determined from random substrate collections and 24 h drift tile samples (1900 - 1900 h). o P CO EH £ -P H 3 (P o p to d 0 O O Pj p > m CD O CD P Ph p cd P ft P P P cd cd cd H rP EH p 1—1 P 0 CD cd VO ■3- * *A 0 d P p ca EH Q p -p CM P 0 0 0 p p 0 P £ Ph cd cd cd p- CO CM O NO CM CA CACM O cm ca
Figure 7. Drift pattern of S. tuberosum larvae for each of the six diel periods. The solid line is the mean number of larvae per tile and the hatched line is discharge. (A = Aug. 2-3» B = Aug. 6-7, C = Aug. 7-8, D = Aug. 22-23, E = June 26-27, F = July 5-6). 78
600
500
400
400
300 200
700
600
500
700
600
500 DISCHARGE (LITRES/SEC.) 700
600
500
700
600
500
HOUR 79
* * * * * * * * * * * * * * * * * o o o O o O * A ON A 'A o O • • • • • • pi VO A vO A On 00 0 P- O- £N- O- {N- o- 0 £ -p A VO fP- o -P 00 0 -P o T—1 VO A £N- ON CM p P, o wi {N- CA CA O o CA CO •H O p «—I I +1 +l +1 +1 +1 +1 0 o I-1 & o O O O o o o PH Q p- P- VO C\i VO o ON i o p !N- p~ p- ^—1 T—1 ON CO p cd -p Q O -3* On T—1 CM D- q-H On A ON A p- A •H o p o P-’ CA CA ^—1 CM On p- o +1 +1 +1 +1 +1 +1 P I o o O O o O o O o A A CM VO T—1 ON P 0 o CM {>- O CM A CM p rH -rH A p -p Ph s{« sje # cd * * * * O * * * * * n'a. * * * * 0 • * * ❖ * * * cd ra O o o o o o > Td O A o A A o P O Cd *H * rH £N- ■p- A rH VO H P ON 00 A A ON 0 T—1 s Ph p to •p H—1 CM A VO ON o o p VO CM VO rH A ON p W) o 0 •H p o CA P- A CM A P p CM rH P cd CM • -P p I +1 +1 +1 +1 +1 -H -P I o m • i -p o o a O O 0 C/2|P p„ ON oo rH ON O O A EH P tud • • • o A • Ph cd •H ON CM A • • A 0 o Td rH \—1 rH A A A i—i Ph P Td $ o P Q A VO o A CM co cd CA A VO rH O A 00 •rH o • m • • • • 1 P P o CM x—1 ^—1 rH rH ON O cd P ON 3? Ph cd +1 +1 +1 +1 +1 +1 &H S p I O I o O o o o O o >5 rH o -p o CM o rH -3- co CM 0 o p VO • • • • • • P tH o UjO A CM rH o A -P AO • rH •H O • o 0\ H P rH • o CM VO :§ • o CM VO 00 CM CM A S? i o V/ 7 Ph 0 Ph * • • • N/- 0 • 0 >5 c Ph * * P 0 -p W) w tlD W) P r—1 p p * * * cd p cd P p P p P P s * * * * Eh -P Q between the night-dark samples (0100-0400 h versus 0400-0700 h samples) on each sample date except June 29 (Table 19). Data for thes dates indicated a trend towards significance (P Q.l) but population densities were low (Table 16, Fig. 7 D,E). The percent of the total S. tuberosum larvae captured every 20 min during the day-night phase in mid-July 1980 is illustrated in Fig. 8. Drift increased after 2020 h when light intensity dropped below ca. 100 lux and peaked when light readings dropped below 0.005 lux at 2120 h. There is a significant difference between larvae captured below ca. 0.05 lux and above it (Mann-Whitney Two-Sample test, Sokal and Rohlf 19&9, U = 180.5» 0.001). Test criteria were established a posteriori based on reported light activity thresholds of perlid stoneflies (Chaston 1968, Bishop 1969)* Furthur evidence which suggests light or factors associated with it are important in S. tuberosum drift comes from analysis of drift during moonlight. On Aug. 2-3 (quarter moon) it was noticably brighter at 2200 and 0100 h when the moon was out than 0400 h when the moon was in. Fewer larvae were captured at 2200 and 0100 h than at 0400 h (Fig. ?A) but these differences were not significant (Mann-Whitney U-test, U = 72 and 75» respectively, P^O.01). On Aug. 6-7 81 Figure 8. Drift of S. tuberosum larvae during the daylight-to-dark period in mid-July 1980. Lines about the means are standard deviations. 82 (Xmn3A31 1H9I1 o o o O o o o o o o o O o O O O in o m o in O O r> m OJ CM in o 83 (night before a full moon) the moon had not risen above the trees at 2200 h but was shining brightly at 0100 h. There was a significant drop in drift between the 2 samples (U = 78.5» P^0.25» Fig* 78). About 0300 h the moon set below a hill, darkening the study site considerably. Drift increased at 0400 h but the difference between this time and 0100 h is not significant (U = 68.5» P^ 0.1, Fig. 7B). On the next night, (full moon) it was clouded until ca. 0200 h after which the moon showed inter¬ mittently through broken clouds and began to set at ca. 0300 h. There was no significant difference between the 2200 and 0100 h samples when it was overcast (U = 51)* Though there was a noticeable drop in drift between 0100 h and 0400 h the difference between these samples is not significant (U = 62.5» Table 16, Fig. 70). On nights of other samples, moon light was absent (Table 15) and no obvious fluctuations in drift were observed (Table 15, Fig. 7D, E, and F). Three genera of Ephemeroptera, Baetis sp., Pseudocleon sp., and Epeorus sp., collected during the 4 diel samples in 1979 also had peak drift at 2200 h and little day drift (Fig. 9)* Occasional small (1-3 mm) Sympitopsyche sp. (Trichoptera: Hydropsychidae), Chironomidae (Diptera), and Phasganophora sp. (Plecoptera: 84 Figure 9* Drift pattern of 3 genera of Ephemeroptera based on the mean percent per time period of the total number captured for each diel period sampled in 1979* 85 Pi Q - LjJ LjJ LlI < s O< Q CL- t CO LlI LlI < QQ O Q_* w LlJ CO cl: co (J CO O 3 CO Q QJ CO I— 3 o LlJ LU u LlJ < CO CL Q QQ Q_ U n HOUR 1900 2200 0100 0400 0700 1000 1300 1600 iBiq^aaynidVD.ivioi jo% x 86 Perlidae) were also collected at night. Perlids probably were the most abundant invertebrate predator of black flies at the study site. Ryacophila fus- cula Walker (Trichoptera: Ryacophilidae), and Ryacophila sp. were also common. This impression is based on extensive but unquantified rock, kick net and surber sampling. The fact that predators were not collected on the tiles is interesting because the cumulative number of larvae captured in the eight 3-hour sample periods were within 20% of the number captured in subsequent 24 h samples taken at 1900 h (Table 20). This suggests larvae may drift to the tiles and, in the absence of predators, remain there during subsequent light:dark changes. Still, the possibility of larvae arriving and leaving tiles at equal rates cannot be discounted. Foraging behavior of perlid stoneflies. All perlid stoneflies found during the day were on the underside (~ photonegative zone) of rock substrates whereas 58% of those found at night were on the top side (Table 21). Additionally„ all 100 P. immarginata collected during day for electivity determination were on the underside of the rocks. Of the 14 P. immarginata captured at night, 93% had undigested prey in the foregut and midgut but only 14% had fresh prey Table 20. Total number of black fly larvae and pupae collected in eight contiguous 3 h samples over a 24 h period compared to those captured in single 24-h samples on the 2 subsequent days. / ft CN- VP* VP* o\ ON ft ft < £ ttO •H •H •H ft ft (T\ ■p ft rH X I—I x' -P -p ft o ft H 0 ft £ ft ft > i—l * CO ft S cd £ £ £ o £ m 0 tuo Q W) 0 * o 0 o £ ON ft £ I 0 ON o 0 - ft C\j T-I O £ ft > o 0 v—I o cd £ Cd ft o 0 £ CQ cd £ S fi ft -3- o o ft 0 87 Table 21. Location of perlid stoneflies on rocks during day and night. -p EH P -p o I—I I—1 s o d 0 0 o 0 o o P EH •H P P "d H •H 0 o ft m &H 0 0 m £ Q -P 0 s 0 d vo OtH^-0--M- O tH o- o o- o CM tHM000H eft 3 d 1 - VO 00 O o o ^ tH -p O CM Q P i—i ■p >? M Cu *H m o d 00 O P -P 89 in the hindgut. In contrast, only 11% of 19 P. immarginata in late afternoon had undigested prey in their foregut, and the midgut and hindgut contained only digested prey, mostly insect cuticle. Phasganophora capitata and Ac. abnormis captured in the day also had few prey but none were collected at night for comparison with (Table 22) . In the artificial stream, the 3 stonefly species remained in the photonegative zone of the substrate during the daylight and crepuscular periods. Paragnetina immarginata became active at dark (2100 h) and foraged among the S. tuberosum (Figs. 10 and 11). Phasganophora capitata and A. abnormis became active after dark but did not forage. They either drifted or retreated when approached by larger P. immarginata. On 2 occasions, P. immarginata was observed to tap A. abnormis with its antennae which caused it to drift. All 3 species drifted or retreated beneath the substrate when touched with a needle or when a non-filtered light was shone on the stream. Paragentina immarginata appeared to locate prey with their antennae. When searching they swept their antennae from side-to-side while alternately tapping the substrate. They also moved their heads Table 22. Digested status of prey items in the guts of perlid stoneflies collected during day and night. •H •H * ft T) -p "d ,£ ft -P •H i—1 !>: CD d 0 £ £ ti£ cq ft O ft *H 0 O -P S H •H •H K -P ft P> •H £ fcuo 3 OQ fcuO ft o 0 CQ £ 0 O ft o o o o •H •H rH \—I*—I On VO -p ftl ft £ ixL d g £ d g d • •H -p ft -P Q ft d o d d & •H £i <1 ftl £ O g CQ £ d • CXI •H •H o •H -P ft -p S £ g g £ d d d W) • ^Digested status of prey in the gut was assessed as follows: 1) undigested II 0 ft •p prey not macerated and still identifiable, and 2) digested = tissue macera and reduced mostly to fluid. Most speciemens collected during the day had the midgut and hindgut full of sclerotized integument only. 91 pigur*6 10. Number of black fly larvae drifting into the screen of the artificial stream correlated with stonefly activity. 92 HOUR 1930 2000 2030 2100 2130 2200 2230 2300 2330 2400 0030 93 Figure 11. Number of black fly larvae drifting into the screen of the artificial stream correlated with stonefly activity. 9 4 CD 2: CD ACTIVE o >- Li_ >-, > INACTIVE LJ- (J LU ^ z O h- uo SUNSET 0.05 LUX 10 - LlI LlJ €C CD LO kJ LlI 5 - cr < _J O 0 T 11 1930 2000 2030 2100 2130 2200 2230 HOUR ~ 95 and bodies from side-to-side, probably to scan as much area a^s possible. When a prey was found it was lined up between the antennae. A lunge was then made for it. Prey escaped by detaching from the substrate i # and drifting. When P. immarginata moved through a patch of S. tuberosum they often lowered their mouths to the substrate and tried to grab larvae with their mandibles. If prey were not located in a short time searching increased rapidly and if no prey were encountered they moved off the substrate. Simulium tuberosum drift and evasive behavior in the artificial stream. In 2 trials with no predators only 2 larvae drifted into the screen, both during dark (Table 23). No other larval movement e.g. looping, was observed. When predators were present drift was 0.07/larvae/l0 min during crepuscular periods and increased to 3*19 larvae/lO min after dark when predators foraged (Table 23)* However, drift during dark occurred primarily when predators were active (Figs. 10 and 11) indicating it is a direct response to predator activity. Thirty-seven percent of the larvae drifted into the screen in trials with predators compared to less than 1% when they were absent (Table 23). Release by larvae from their substrate in response to predators was actually much 23 • Drift of* S. tuberosum larvae in an outdoor artificial stream with and without stonefly predators. r *H ft O rH £h*H 0 > EH ft 1—1 0CO ft Mit ft *H O ft < ^ cd a ftm £ 0 ft 0 0 ft £0 0 O O ft0 U > cd cd 0 ft 0 Eh O ft 0 > 1-1 W)^ 0 OGO £-1 cd0 o -p higher than indicated by counts of larvae in the screen since most larvae did not drift into the screen but descended a few centimeters on a silk-strand and re-attached. At the end of predator tests the position of the remaining larvae was almost com¬ pletely re-arranged whereas in the non-predator tests it was unchanged. Observations on the release and drift behavior of 4 black fly species, when needles were used to mimic predators, strongly suggest drift is mainly a predator escape response. When disturbed by vibration or being touched in the head, thorax or upper abdomen, larvae curled into a C shape flat against the substrate and appeared to touch their mouth to it (= "warned")* This may be to attach an anchor point for a silk strand ("life-line", Hynes 1970), though observations did not confirm this. Soon after the disturbance stopped, larvae returned to normal feeding posture. Continued touching of the head, thorax, or upper abdomen caused larvae to jerk from side-to-side or to crawl away. In no case did touching in these body regions cause larvae to drift. However, immediate release from the substrate occurred when the lower abdomen near the last abdominal segment was touched (= "attacked"). If larvae were first "warned” and then 98 "attacked" they released hut only drifted several centimeters because of a silk-strand anchored to the substrate. Larvae that were "attacked" but had not been "warned" or were "warned" but had returned to normal posture, drifted into the screen since they were without a "life-line". The reaction of S. tuberosum to P. immarginata foraging and attack was identical to the needle observations. When touched by predator antennae, S. tuberosum curled into a C shape but did not drift. Drift occurred when larvae were touched in the lower abdomen as P. immarginata lunged or when their legs touched this region during prey-searching. None of the stoneflies in the artificial stream captured an S. tuberosum larva, despite numberous attempts. It is noteworthy that of 167 perlids collected at the field site with identifiable prey only 6.6^ contained black fly larvae (Table 24). The electivity coefficient for black flies (all were S. tuberosum) was -0.29 which indicates they were consumed by P. immarginata at a rate lower than they were available in the food complex (Table 25) and suggests avoidance by the predator or that black flies escape to avoid being consumed. The electivity coefficient for Baetidae was -O.605 indicating even Table 24. Prey found in the alimentary canal of most common perl ids found at the study site. •H •H •H •H »H ft ft CO ft £ M ft -P stronger avoidance (Table 25) • Chironomids were the most abundant food available to P. immarginata. The electivity coefficient was O.0383 indicating they were consumed in equal proportion to their abundance in the food complex (Table 25)• The electivity coefficient for Hydropsychidae was +0.66 indicating they are selected by P. immarginata in greater proportion than their availability (Table 25)* Simulium tuberosum and S. damnosum induced to release onto a silk-strand in artificial streams oriented with their heads upstream and their bodies parallel to the substratum. To re-attach to the substrate, larvae angled their heads down such that the current forced them back down onto the substrate and they could attach a new anchor point with silk from their mouths. In the field S. tuberosum, P. mixtum/fuscum, and St. mutata were observed to re-attach to the substrate the same way. If no substrate was directly below they crawled back up the silk strand using their mouth and prolegs to re-attach to their original substrate. Also, as in the artificial streams, if induced to release without being "warned" all three species drifted downstream because no silk strand was attached to the substrate. 102 Discussion Behavioral and ecological data collected in this study indicate S. tuberosum drift is related to predator avoidance behavior. The release response of larvae touched on the lower abdomen where predators attack results in instantaneous escape from predation 1 in fast flowing water. Geometrid looping is too slow for escape. A consequence of this behavior is that contact with other invertebrates, such dorso- ventrally flattened Heptageniids (Ephemeroptera), would also stimulate release. C-curling to attach a silk strand anchor permits escape without complete loss of substrate contact. Reisen (1977) observed S. virgattum and S. trivittatum curl into a C-shape and release on a silk-strand when attempts were made to collect them. Reisen and Prins (1972) and Reisen (1977) observed no periodicity in S. tuberosum drift in an upper Piedmont stream. Though this does not agree with my results, they reported no predaceous stoneflies among 9 other taxa collected in drift nets. My. observations with S. damnosum s.l. support Disney's (1972) observation of increased relocation of S. damnosum complex and other Simulium sp. when nocturnal mayflies 103 were added to troughs. It is noteworthy that all observed S. tuberosum larvae attacked by P. immarginata in the artificial stream escaped and only 8.3^ of the perlids collected at the study site contained black fly larvae (Table 24). The electivity coefficient of -0.29 f°r S. tuberosum strongly suggests they are capable of avoiding predation in nature. Siegfried and Knight (1976a) reported negative electivity values of black flies for 5 monthly collections with nothing in the gut for 2 months over a year for Acroneuria californica Banks. It also is interesting that the electivity coefficient of Baetidae was -O.605 (Table 23) since they were collected on the tiles in a pattern similiar to S. tuberosum (Fig. 9)* Siegfried and Knight (1976a) reported negative electivity values for Baetidae as well. Recently, it was shown that mayfly drift increased when perlids were added to a laboratory stream (Corkum and Pointing 1979)* Mayflies respond defensively to stonefly attack in several ways, including drifting (Peckarsky 1980, Walton 1981, Simmons unpubl. data). Chironomids comprise the major portion of perlid diet (Table 25, Richardson and Gaufin 1971» Siegfried and Knight 1976 a,b). Chironomids are abundant reophilic 104 inhabitants but drift very little (Waters 1972). Hydropsychids also are seldom collected in the drift (Waters 1972) and in this study they had an electivity coefficient of +0.66, which would suggest they are ineffective at avoiding predation or that stoneflies selectivity feed on them (Table 25). The field and laboratory data show that the 3 major perlid species inhabiting the field study site forage and feed at night (Tables 21 and 22), which generally is true for most perlids (Chaston 1968, Bishop 1969, Hynes 1970). Prey location with antennae, which also has been reported previously (Hynes 1948, Brinck 1949, Richardson and Gaufin 1971> Gath and Richardson 1975> Hitchcock 1976, Allan 1978, Peckarsky 1979), supports this. Paragentina immarginata first became active in the artificial stream about 30 min after the natural light intensity fell below 0.05 lux. This coincides with the significant increase in S. tuberosum drift in the field during the crepuscular-to-dark period (Fig. 9)* In the artificial stream S. tuberosum larvae only drifted at this time if P. immarginata was foraging (Table 23» Figs. 10 and 11). There also was a peak in drift of the 3 mayfly genera collected on the tiles at 2200 h (Fig. 9)• Chaston (1968) found the mean light threshold at which 105 drift occurred with Simulium and 4 other insect genera to "be 1.584 lux. Bishop (1969) found the light threshold for drift in an artificial stream was between 0.01 and 0.001 lux for P. capitata and 3 genera of Ephemeroptera. The decrease in S. tuberosum drift when there was bright moonlight (Table 16, Fig. 7A-C) also suggests black fly drift is a function of invertebrate activity. Russell (1916) measured full moonlight at 65° to be 0.03 lux. This is above the light level recorded for peak S. tuberosum drift in the field (Fig. 5) and activity of P. immarginata in the artificial stream. Bishop and Hynes (1969) observed drift suppression with moonlight of 0.01-0.02 lux. Anderson (1966) observed a significant decrease in Simulium drift during a full moon. Since nocturnal stoneflies become active at light thresholds below the value reported by Russell (1916) for a full moon (Chaston 1968, Bishop 1969, P. immarginata this study) this is a plausible explanation for the decrease in drift during bright moonlight. Insects collected in drift frequently are larger than conspecifics in the substratum (Hynes 1970, Waters 1972, Muller 1974). Black flies have various patterns of instar drift which may be due to migration from eclosion sites to larval habitat or pupation 106 sites (Colbo and Wotton 1981)- No significant difference in the proportion of instars collected in the drift samples versus those in the standing population was observed in the present study (Table 18) but there was a significant difference in the proportion of mature larvae drifting at night versus day (Table 17)• This may be due to movement to pupation sites. In a laboratory colony of S. decorum (Simmons and Edman 1981) mature larvae move about the rearing chamber and frequently pupate away from their larval feeding sites in areas of reduced current. An increase in drift of larger larvae may also be influenced by the fact that larger body size makes them more susceptible to predation. Low drift among first instars may be due to their small size and/or lack of encounter with predators. Also, they may not need to migrate to feeding sites since females oviposit in preferred larval habitat (Simmons unpubl. data). Drift to feeding sites by first instars has been observed with some species which lay their eggs in masses on substrates (Rubztov 1965» Kureck 1969, Ruhm 1970, Colbo and Moorehouse 1979, Wotton et al. 1979, Colbo and Wotton 1981). Lack of a pattern of instar drift also supports the idea of predator induced drift Predators foraging indiscriminantly among patches of 107 larvae should cause random release by all instars present. Black fly drift has been correlated with temperature (Pearson and Franklin 19^8, Reisen and Prins 1972, Muller 197*0. water discharge (Carlsson 1967, Minshall and Winger 1968) and population density (Pearson and Franklin 1968, Reisen and Prins 1972, Hildebrand 197*0 • One must be cautious of multiple regression analysis of drift data since correlation does not necessarily indicate cause. The possibility that S. tuberosum drift is triggered by diel changes in stream conditions cannot be ruled out. However, constancy of drift pattern in spite of variable stream conditions (Table 5> Fig- 8) suggests other factors control most behavioral drift (Waters 1972). Gersabeck and Merritt (1979) showed increased relocation of P. mixtum/fuscum and Cnephia dacotensis with changing current velocities in an artificial stream. Daytime drift of S. tuberosum may possibly be due to day active predators such Rhyacophila spp(Trichoptera: Rhyacophilidae). Rhyacophila fuscula Walker frequently was taken in daytime collections from patches of S. tuberosum larvae and 108 was found to have ingested larvae. Other Trichoptera known to feed on hlack flies at the study site are Symphitopsyche bronta (Ross), S. morosa (Hagen), and S. sparna (Ross). A study of Trichopteran predators would be interesting since they have small peg-like antennae and probably rely heavily on vision for prey location. Thus, black fly larvae would not be "warned” of attack. A review of trichopteran predation on black flies is included in Burton and McCrae (1972). Black fly larvae are often found at extremely high densities (Tarshis and Neil 1970) and intraspecific competition for substrates can occur (Fredeen and Schemanchuck i960, Kureck 1969)• Gersabeck and Merritt (1979) observed fighting and subsequent relocation of P. mixtum/fuscum larvae. Disney (1972) also observed that large Simulium spp., including S. damnosum, clear away smaller.adjacent larvae and often "spar" during this process. On 3 occasions S. damnosum were observed to "fight" in a laboratory stream (Simmons, unpubl. data). The aggressor pushed at the lower abdomen of the defender, pushed, and induced it to release. It is interesting that this behavior is directed at the abdominal region which induced other test larvae 109 to drift when touched with a needle or when attacked hy a predator (in the case of S. tuberosum). Perhaps S. damnosum regulate their spatial dispersion within a substrate by "preying" on conspecifics. Conclusions 1. The diel drift pattern of S. tuberosum indicated 2.5 times as many larvae drifted at night versus day. Only last instars drifted in significantly greater proportion during night versus day. 2. Despite varying flucuations in stream temperature and discharge, the drift pattern was similiar on each of the 6 diel periods sampled. 3. Field data indicates drift is associated with light intensity. A significant increase and decrease during the day-night and night-day period respectively, was observed on 5 of 6 diel sample periods. During bright moonlight a significant decrease in drift also was noted. Drift samples taken every 20 minutes during the crepuscular- to-dark period revealed a significant increase in drift when light intensity fell below ca. 100 lux and a peak in drift 20 minutes after it fell to 0.05 lux. 110 Stonefly predators became active at night when light intensity fell below 0.05 lux. Drift of S. tuberosum in the artificial stream was in direct response to stonefly foraging and not light changes. No S. tuberosum larvae were caught by stoneflies in the artificial stream, despite numerous attempts. Only 6.6% of the stoneflies captured in the field had black fly larvae in their guts. The electivity coefficient for stoneflies feeding on black flies was -0.29, further suggesting that black flies effectively avoid stonefly predation. Observations on S. tuberosum and S. damnosum in artificial streams and St. mutata and P. mixturn/ fuscum in the field revealed that larvae which were "warned" (= touching by predator or needle) before attack would curl and attach an anchor point for a "life-line", which allows them to instantaneously escape danger without being carried away into the current because no life-line was attached to the substrate. It is concluded that nocturnal increase in S. tuberosum drift is a result of stimulation by contact with foraging insects, particularly stoneliy predators. Black fly drift response has probably evolved as an escape mechanism to avoid predation. LITERATURE CITED All, S. H., P. 0. Burbutis, W. F. Ritter and R. W. Lake. 1974. 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