Dynamics of an experimental microbial invasion Francisco Acostaa,b, Richard M. Zamora,b,1, Fares Z. Najarc,d, Bruce A. Roec,d, and K. David Hambrighta,b,2 aProgram in Ecology and Evolutionary Biology, Department of Biology, University of Oklahoma, Norman, OK 73019; bPlankton Ecology and Limnology Laboratory, Department of Biology, University of Oklahoma, Norman, OK 73019; cAdvanced Center for Genome Technology, University of Oklahoma, Norman, OK 73019; and dDepartment of Chemistry and Biochemistry, University of Oklahoma, Norman, OK 73019 Edited by Daniel S. Simberloff, The University of Tennessee, Knoxville, TN, and approved August 3, 2015 (received for review March 14, 2015) The ecological dynamics underlying species invasions have been a Traditional macroorganism-based ecological theory suggests major focus of research in macroorganisms for the last five decades. that invasion success tends to be highest in exotic species char- However, we still know little about the processes behind invasion by acterized by high dispersal abilities, growth rates, and resource unicellular organisms. To expand our knowledge of microbial in- efficiencies (12), and in native communities characterized by low vasions, we studied the roles of propagule pressure, nutrient supply, species diversity and high disturbance levels, including fluctua- and biotic resistance in the invasion success of a freshwater invasive tion of resources (13). Once an invasion occurs, it can induce alga, Prymnesium parvum, using microcosms containing natural significant change in the invaded community, including modifi- freshwater microbial assemblages. Microcosms were subjected to a cation of community structure and loss of species and ecosystem factorial design with two levels of nutrient-induced diversity and function (13). three levels of propagule pressure, and incubated for 7 d, during Although the applicability of macrobial principles to microbial P. parvum which densities and microbial community composition systems is debatable (1, 4), they constitute a general framework of were tracked. Successful invasion occurred in microcosms receiving study and readily testable hypotheses, such as whether microbial high propagule pressure whereas nutrients or community diversity invasions are limited by dispersal or whether high diversity in played no role in invasion success. Invaded communities experienced native communities provides resistance to invasion by exotics. distinctive changes in composition compared with communities Indeed, few available experimental studies of microbial invasions where the invasion was unsuccessful. Successfully invaded microbial communities had an increased abundance of fungi and ciliates, and have been designed to directly test such hypotheses, but, in- decreased abundances of diatoms and cercozoans. Many of these terestingly, they tend to suggest that interspecific interactions, ECOLOGY changes mirrored the microbial community changes detected during rather than diversity per se, play critical roles in invasion resistance – anaturalP. parvum bloom in the source system. This role of propa- (14 16), although nutrient supply (17) and propagule pressure gule pressure is particularly relevant for P. parvum in the reservoir- (18) have also been shown to facilitate microbial invasions. dominated southern United States because this species can form Unfortunately, there are limitations in these previous studies, large, sustained blooms that can generate intense propagule pres- such as the use of artificial communities with very low species sures for downstream sites. Human impact and global climate numbers. By not taking into account the enormous diversity, vari- change are currently causing widespread environmental changes in ability, and stochasticity inherent to natural communities, which most southern US freshwater systems that may facilitate P. parvum establishment and, when coupled with strong propagule pressure, Significance could put many more systems at risk for invasion. Current models for biological invasions are predominantly microbial ecology | diversity | invasion resistance | propagule pressure | based on macroorganisms. Few invasion model assumptions Prymnesium have been validated for microbial systems. Further research on microbial invasion dynamics is required to determine whether icrobial species invasions, thought to occur worldwide in macrobial models are appropriate for microbes, as well as to Mterrestrial and aquatic systems and involve both patho- understand present and future distributions of invasive mi- genic and free-living taxa, represent an emerging challenge to croorganisms, particularly in the face of contemporary envi- our understanding of the interplay between biodiversity and ronmental changes. We studied the establishment of an ecosystem function, particularly under pressures of global envi- invasive protist in natural microbial assemblages in replicate ronmental change (1). Despite assertions of limitless dispersal experimental microcosms and found that, under adequate en- capability, sensu Baas-Becking (2), many microorganisms seem vironmental conditions, invasion success was determined by not to be cosmopolitan, and biogeographic studies suggest a the number of invading propagules rather than resource significant effect of ecological drift and dispersal limitation in availability and the diversity of the invaded communities. This study is among the first to test invasibility hypotheses using an their distributions (3, 4). actual invasive microbial species in natural communities. Invasions of microbial species are hard to track because small, inconspicuous species are routinely overlooked in most assess- Author contributions: R.M.Z. and K.D.H. designed research; R.M.Z. performed research; ments of invasive species (5) and are detected only when they F.Z.N. and B.A.R. contributed new reagents/analytic tools; F.A., R.M.Z., and K.D.H. ana- have conspicuous impacts, such as the formation of blooms. In- lyzed data; F.A. and K.D.H. wrote the paper; and R.M.Z., F.Z.N., and B.A.R. conducted vasions from pathogenic microbes have been relatively well- community sequencing. studied because they are comparatively easy to track, in part, due The authors declare no conflict of interest. to strong, observable impacts (1). A particularly well-studied This article is a PNAS Direct Submission. example is the invasion of the human gut microbiota by patho- Freely available online through the PNAS open access option. genic bacteria (6, 7). We know, however, much less about in- Data deposition: The sequences reported in this paper have been deposited in the National Center for Biotechnology Information (NCBI) Sequence Read Archive under BioProject vasion by nonpathogenic microbes, even though we have evidence PRJNA271537 and Biosamples SAMN03274828–SAMN03274857. that they do occur in nature. For example, in the last two decades, 1Present address: Ecosystems and Lake Management, Grand River Dam Authority, Langley, a number of invasions by aquatic microbial species have been OK 74350. – documented (8 11). Although there are some studies on the eco- 2To whom correspondence should be addressed. Email: [email protected]. logical impacts of these invasions (8), we still know little about their This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. potential consequences. 1073/pnas.1505204112/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1505204112 PNAS Early Edition | 1of6 Downloaded by guest on September 29, 2021 Table 1. Chlorophyll and diversity conditions for microcosms Results before inoculation of propagule treatments Community Diversity Manipulation. Microbial assemblages were Low nutrient High nutrient created from composite mixtures of lake water collected from Indicator treatment treatment P three different sites in Lake Texoma (Oklahoma and Texas, United States) during the summer non-P. parvum bloom season. Chlorophyll a 32.2 ± 2.84 98.3 ± 7.02 <0.001 Salinity and temperature were adjusted to 2.3 parts per thousand Eukaryotic richness 703 ± 12.4 593 ± 37.2 0.008 (ppt) and 15 °C to simulate the ambient winter lake conditions Eukaryotic alpha diversity 60.1 ± 4.49 28.4 ± 4.34 <0.001 conducive to P. parvum blooms (29). Three days later, commu- Bacterial richness 4162 ± 197 3892 ± 489 0.425 nity resistance to invasion was manipulated in half of the mi- Bacterial alpha diversity 479 ± 35.6 408 ± 85.6 0.259 crocosms by supplementing nitrogen and phosphorus levels to Values are mean ± SD; n = 9 per treatment for chlorophyll (μg/L), n = 3 per reduce community diversity and increase resource availability. treatment for richness (Chao1) diversity (inverse Simpson) estimates. Bold Nutrient manipulation resulted in clear differences in experi- font indicates a significant difference between low and high nutrient treat- mental microcosms after 5 d (day 8) of incubation (Table 1). Total ments; for more detail on statistical analyses, consult Table S1. chlorophyll was higher [generalized linear model (GLM), F-test, P < 0.001] whereas richness and alpha diversity of eukaryotic communities were lower (GLM, F-test, P = 0.008 and P < 0.001, can have thousands of interacting bacterial and protistan species, as respectively) (Table S1) in microcosms that received nutrient ad- well as viruses, it is unclear how applicable these earlier experi- ditions. Communities contained 6,543 bacterial and 1,079 eukaryote mental studies are with respect to actual invasion dynamics. Ad- operational taxonomic units