SEASONAL OCCURRENCE OF NEOECHINORHYNCHUS EMYDIS (PHYLUM: ACANTHOCEPHALA) IN THE FRESHWATER SNAIL, PLANORBELLA CF. P. TRIVOLVIS, FROM OKLAHOMA.

The acanthocephalan Neoechinorhynchus emydis has a complex life cycle and infects turtle, ostracod, and snail hosts. However, little information is available on the seasonal distribution or the effects of N. emydis on freshwater snail hosts. To address this, we examined the seasonal distribution and melanization of acanthocephalans in Planorbella cf. Planorbella trivolvis snails from a single location in north-central Oklahoma. Seasonally, prevalence of N. emydis was 0% during the winter, increased to 50% during the summer, and declined to 17% in the fall. Mean abundance exhibited more variation but generally followed a similar pattern as prevalence. More important, all acanthocephalans located within the head/foot region of snail hosts contained melaninlike pigment surrounding each worm, suggesting that snails were mounting an immunological reaction to infections with N. emydis. Snail shell diameter was greatest during the fall and decreased during the winter, indicating that larger or older snails were dying during the winter. However, because field-collected snails were commonly infected with trematodes, and snail size varied significantly with season, it was unclear whether the observed seasonal dynamics of acanthocephalan infections were a result of snail mortality resulting from snail age, parasitic infections, or a combination of factors. To control for these factors, we exposed laboratory-reared Planorbella cf. P. trivolvis snails to naturally infected ostracods in field cages for 5-wk intervals during the winter, spring, and summer. Data from snail-cage infections were consistent with the seasonal field survey such that N. emydis infections were highest in the summer (20%) and lowest (0%) in the winter, suggesting that snails were not ingesting infected ostracods during the winter. However, fewer of our laboratory-reared snails survived in field cages during winter than during spring and summer, suggesting that snails may die more often during harsh winter conditions. Finally, we conducted a laboratory survival experiment by testing the life span and egg production of field-collected snails of various sizes that were naturally infected with acanthocephalans or trematodes or both. Our snail-survival experiment indicated that snail size but not infection status with acanthocephalans or trematodes affected snail survival, with larger snails surviving a shorter amount of time than smaller snails. In addition, snails infected with trematodes laid significantly fewer eggs compared with uninfected snails or snails infected with acanthocephalans. However, we found no significant difference in the number of eggs laid by acanthocephalan-infected and uninfected snails. Although other abiotic factors still need evaluation, we suggest that the occurrence of acanthocephalans in snails throughout the year may be partially influenced by the abundance of infected ostracods that snails may be ingesting and snail population fluctuations during the year.

Multi-population puma connectivity could restore genomic diversity to at-risk coastal populations in California.

Urbanization is decreasing wildlife habitat and connectivity worldwide, including for apex predators, such as the puma (Puma concolor). Puma populations along California's central and southern coastal habitats have experienced rapid fragmentation from development, leading to calls for demographic and genetic management. To address urgent conservation genomic concerns, we used double-digest restriction-site associated DNA (ddRAD) sequencing to analyze 16,285 genome-wide single-nucleotide polymorphisms (SNPs) from 401 pumas sampled broadly across the state. Our analyses indicated support for 4-10 geographically nested, broad- to fine-scale genetic clusters. At the broadest scale, the four genetic clusters had high genetic diversity and exhibited low linkage disequilibrium, indicating that pumas have retained genomic diversity statewide. However, multiple lines of evidence indicated substructure, including 10 finer-scale genetic clusters, some of which exhibited fixed alleles and linkage disequilibrium. Fragmented populations along the Southern Coast and Central Coast had particularly low genetic diversity and strong linkage disequilibrium, indicating genetic drift and close inbreeding. Our results demonstrate that genetically at risk populations are typically nested within a broader-scale group of interconnected populations that collectively retain high genetic diversity and heterogenous fixations. Thus, extant variation at the broader scale has potential to restore diversity to local populations if management actions can enhance vital gene flow and recombine locally sequestered genetic diversity. These state- and genome-wide results are critically important for science-based conservation and management practices. Our nested population genomic analysis highlights the information that can be gained from population genomic studies aiming to provide guidance for the conservation of fragmented populations.

Minimum habitat thresholds required for conserving mountain lion genetic diversity.

Jointly considering the ecology (e.g., habitat use) and genetics (e.g., population genetic structure and diversity) of a species can increase understanding of current conservation status and inform future management practices. Previous analyses indicate that mountain lion (Puma concolor) populations in California are genetically structured and exhibit extreme variation in population genetic diversity. Although human development may have fragmented gene flow, we hypothesized the quantity and quality of remaining habitat available would affect the genetic viability of each population. Our results indicate that area of suitable habitat, determined via a resource selection function derived using 843,500 location fixes from 263 radio-collared mountain lions, is strongly and positively associated with population genetic diversity and viability metrics, particularly with effective population size. Our results suggested that contiguous habitat of ≥10,000 km2 may be sufficient to alleviate the negative effects of genetic drift and inbreeding, allowing mountain lion populations to maintain suitable effective population sizes. Areas occupied by five of the nine geographic-genetic mountain lion populations in California fell below this habitat threshold, and two (Santa Monica Area and Santa Ana) of those five populations lack connectivity to nearby populations. Enhancing ecological conditions by protection of greater areas of suitable habitat and facilitating positive evolutionary processes by increasing connectivity (e.g., road-crossing structures) might promote persistence of small or isolated populations. The conservation status of suitable habitat also appeared to influence genetic diversity of populations. Thus, our results demonstrate that both the area and status (i.e., protected or unprotected) of suitable habitat influence the genetic viability of mountain lion populations.

Pronghorn population genomics show connectivity in the core of their range.

Preserving connectivity in the core of a species' range is crucial for long-term persistence. However, a combination of ecological characteristics, social behavior, and landscape features can reduce connectivity among wildlife populations and lead to genetic structure. Pronghorn (Antilocapra americana), for example, exhibit fluctuating herd dynamics and variable seasonal migration strategies, but GPS tracking studies show that landscape features such as highways impede their movements, leading to conflicting hypotheses about expected levels of genetic structure. Given that pronghorn populations declined significantly in the early 1900s, have only partially recovered, and are experiencing modern threats from landscape modification, conserving connectivity among populations is important for their long-term persistence in North America. To assess the genetic structure and diversity of pronghorn in the core of their range, we genotyped 4,949 genome-wide single-nucleotide polymorphisms and 11 microsatellites from 398 individuals throughout the state of Wyoming. We found no evidence of genetic subdivision and minimal evidence of isolation by distance despite a range that spans hundreds of kilometers, multiple mountain ranges, and three interstate highways. In addition, a rare variant analysis using putatively recent mutations found no genetic division between pronghorn on either side of a major highway corridor. Although we found no evidence that barriers to daily and seasonal movements of pronghorn impede gene flow, we suggest periodic monitoring of genetic structure and diversity as a part of management strategies to identify changes in connectivity.

GENETIC DIVERSITY OF YELLOW-BILLED MAGPIES ( PICA NUTALLI) BEFORE AND AFTER A WEST NILE VIRUS EPIDEMIC.

The appearance of West Nile virus (WNV) coincided with declines in California, US bird populations beginning in 2004, and particularly affected corvid populations, including Yellow-billed Magpies ( Pica nutalli), an endemic species to California. Our objective was to determine if the timing of the WNV epidemic correlated with changes in the genetic diversity or population structure of magpies. We hypothesized the declines in magpie abundance from WNV would lead to genetic bottlenecks and reduced genetic diversity, but not to changes in population genetic structure. To test these hypotheses, we genetically typed magpie samples collected during the Dead Bird Survey before WNV arrived (2002-04), immediately after WNV arrived in late 2004 (2006-08), and several generations after the onset of the epidemic (2009-11). For each of these three time periods, we tested for genetic bottlenecks, estimated genetic heterozygosity, allelic richness, relatedness, effective population sizes, and genetic structure, with the use of 10 nuclear microsatellite loci. Although there was no evidence for spatial or temporal genetic structure, genetic-diversity estimates were similar or below estimates for endangered corvid species. Measures of genetic diversity were consistent across time periods. In contrast to our expectation, we detected a genetic bottleneck prior to the WNV epidemic, which may have coincided with severe drought conditions in California, increasing human population size in magpie range, and an estimated 33% decrease in population size. We found weak evidence to support a bottleneck after the introduction of WNV in California. Our results suggest the WNV epidemic did not have additional catastrophic effects on the neutral genetic diversity of P. nutalli in the sampled areas. However, because we detected lower heterozygosity in Yellow-billed Magpies than has been reported in closely related endangered species, this species is of conservation concern and should be monitored to detect further population declines or loss of genetic diversity.

Measures of effective population size in sea otters reveal special considerations for wide-ranging species.

Conservation genetic techniques and considerations of the evolutionary potential of a species are increasingly being applied to species conservation. For example, effective population size (N e) estimates are useful for determining the conservation status of species, yet accurate estimates of current N e remain difficult to obtain. The effective population size can contribute to setting federal delisting criteria, as was done for the southern sea otter (Enhydra lutris nereis). After being hunted to near extinction during the North Pacific fur trade, the southern sea otter has recovered over part of its former range, but remains at relatively low numbers, making it desirable to obtain accurate and consistent estimates of N e. Although theoretical papers have compared the validity of several methods, comparisons of estimators using empirical data in applied conservation settings are limited. We combined thirteen years of demographic and genetic data from 1,006 sea otters to assess multiple N e estimators, as well as temporal trends in genetic diversity and population genetic structure. Genetic diversity was low and did not increase over time. There was no evidence for distinct genetic units, but some evidence for genetic isolation by distance. In particular, estimates of N e based on demographic data were much larger than genetic estimates when computed for the entire range of the population, but were similar at smaller spatial scales. The discrepancy between estimates at large spatial scales could be driven by cryptic population structure and/or individual differences in reproductive success. We recommend the development of new delisting criteria for the southern sea otter. We advise the use of multiple estimates of N e for other wide-ranging species, species with overlapping generations, or with sex-biased dispersal, as well as the development of improved metrics of genetic assessments of populations.

Founder events, isolation, and inbreeding: Intercontinental genetic structure of the domestic ferret.

Domestication and breeding for human-desired morphological traits can reduce population genetic diversity via founder events and artificial selection, resulting in inbreeding depression and genetic disorders. The ferret (Mustela putorius furo) was domesticated from European polecats (M. putorius), transported to multiple continents, and has been artificially selected for several traits. The ferret is now a common pet, a laboratory model organism, and feral ferrets can impact native biodiversity. We hypothesized global ferret trade resulted in distinct international genetic clusters and that ferrets transported to other continents would have lower genetic diversity than ferrets from Europe because of extreme founder events and no hybridization with wild polecats or genetically diverse ferrets. To assess these hypotheses, we genotyped 765 ferrets at 31 microsatellites from 11 countries among the continents of North America, Europe, and Australia and estimated population structure and genetic diversity. Fifteen M. putorius were genotyped for comparison. Our study indicated ferrets exhibit geographically distinct clusters and highlights the low genetic variation in certain countries. Australian and North American clusters have the lowest genetic diversities and highest inbreeding metrics whereas the United Kingdom (UK) cluster exhibited intermediate genetic diversity. Non-UK European ferrets had high genetic diversity, possibly a result of introgression with wild polecats. Notably, Hungarian ferrets had the highest genetic diversity and Hungary is the only country sampled with two wild polecat species. Our research has broad social, economic, and biomedical importance. Ferret owners and veterinarians should be made aware of potential inbreeding depression. Breeders in North America and Australia would benefit by incorporating genetically diverse ferrets from mainland Europe. Laboratories using ferrets as biomedical organisms should consider diversifying their genetic stock and incorporating genetic information into bioassays. These results also have forensic applications for conserving the genetics of wild polecat species and for identifying and managing sources of feral ferrets causing ecosystem damage.

A single migrant enhances the genetic diversity of an inbred puma population.

Migration is essential for maintaining genetic diversity among populations, and pumas (Puma concolor) provide an excellent model for studying the genetic impacts of migrants on populations isolated by increasing human development. In densely populated southern California, USA, puma populations on the east and west side of interstate highway 15 (I-15) have become fragmented into a small inbred population on the west side (Santa Ana Mountains) and a relatively larger, more diverse population on the east side (Eastern Peninsular Range). From 146 sampled pumas, genetic analyses indicate seven pumas crossed I-15 over the last 15 years, including four males from west to east, and three males from east to west. However, only a single migrant (named M86) was detected to have produced offspring and contribute to gene flow across the I-15 barrier. Prior to the M86 migration, the Santa Ana population exhibited inbreeding and had significantly lower genetic diversity than the Eastern Peninsular Range population. After M86 emigrated, he sired 11 offspring with Santa Ana females, decreasing inbreeding measures and raising heterozygosity to levels similar to pumas in the Eastern Peninsular Range. The emigration of M86 also introduced new alleles into the Santa Ana population, although allelic richness still remained significantly lower than the Eastern Peninsular population. Our results clearly show the benefit of a single migrant to the genetics of a small, isolated population. However, ongoing development and habitat loss on both sides of I-15 will increasingly strengthen the barrier to successful migration. Further monitoring, and potential human intervention, including minimizing development effects on connectivity, adding or improving freeway crossing structures, or animal translocation, may be needed to ensure adequate gene flow and long-term persistence of the Santa Ana puma population.

Atrazine reduces the transmission of an amphibian trematode by altering snail and ostracod host-parasite interactions.

Trematodes are ubiquitous members of aquatic environments, have many functional roles in ecosystems, and can cause diseases in humans, livestock, and wild animals. Despite their importance and reports of parasite population declines, few studies have concurrently assessed the effects of aquatic contaminants on multiple hosts, multiple parasite life cycle stages, and on transmission-related host-parasite interactions. Here, we test the effects of environmentally relevant concentrations of the herbicide atrazine (0, 3, 30 µg/L) on the establishment and development of an amphibian trematode (Halipegus eccentricus) in a first-intermediate snail host (Physa acuta) and in a second-intermediate ostracod host (Cypridopsis sp.). Additionally, we test the interactive effects of atrazine and parasitism on snail and ostracod survival. Our results indicate that atrazine negatively affects trematode transmission by altering snail and ostracod host-parasite interactions. Although atrazine did not affect the survival of uninfected snails alone, atrazine acted synergistically with parasitism to reduce the longevity of infected snails. As a result, the number of cercariae (i.e., larval trematodes) produced by snails was 50.7 % (3 µg/L) and 14.9 % (30 µg/L) relative to controls. Atrazine exhibited direct negative effects on ostracod survival at 30 µg/L. However, when ostracods were also exposed to trematodes, the negative effects of atrazine on survival were diminished. Although infected ostracod survival remained high, trematode development was significantly reduced, resulting in reduced infectivity of metacercariae (i.e., nongravid adult cysts infective to definite host) to 32.2 % (3 µg/L) and 28.6 % (30 µg/L) relative to the controls. The combination of reduced cercaria production and reduced metacercarial infectivity in the 3 and 30 µg/L atrazine treatment groups reduced the net number of infective worms produced to 16.4 and 4.3 % (respectively) relative to the control. These results demonstrate the complex nature of pesticide effects on trematode infections and indicate that trematodes can affect their first- and second-intermediate hosts differently under different pesticide concentrations. Our work has broad implications for parasite transmission and conservation and provides a testable mechanism for understanding trematode population declines in contaminated wetlands.

Effects of trematode parasitism on the shell morphology of snails from flow and nonflow environments.

The primary function of the gastropod shell is protection. However, shells that function well in one environment may be maladaptive in another. Upon infection, the snail shell protects internal parasites and it is to the parasite's advantage to optimize, or not interfere with, shell functionality. However, parasites, particularly trematodes, are often pathogenic and it is not clear if parasitism will induce environment-dependent or -independent changes to gastropod shells. We conducted a field study and a complementary laboratory experiment to examine the effects of trematode parasitism on shell characteristics (shape, size, and crush resistance) of Physa acuta snails in flow and nonflow environments using geometric morphometrics and crush assays. Field results indicate wetland (nonflow) snails had large, crush resistant shells with narrow apertures and tall spires. In contrast, stream (flow) snails had small, weak shells with wide apertures and short spires. Parasitism had no apparent effect on the crush resistance of wetland snails but significantly reduced the crush resistance of stream snails. Parasitism had no significant effect on overall shell shape in stream or wetland snails. Similar to the results of our field study, nonflow tank snails had significantly more crush resistant shells than flow tank snails. Additionally, the shapes of flow and nonflow tank snails significantly differed where nonflow tank snails resembled wetland snails and flow tank snails resembled stream snails. For laboratory snails, parasitism reduced crush resistance regardless of flow/nonflow treatment. Our results demonstrate that habitat and/or flow treatment was the primary factor affecting P. acuta shell morphology and that trematode parasitism played a secondary role.

The effects of the herbicide atrazine on freshwater snails.

Atrazine has been shown to affect freshwater snails from the subcellular to community level. However, most studies have used different snail species, methods, endpoints, and atrazine exposure concentrations, resulting in some conflicting results and limiting our understanding. The goal of this study was to address these concerns by (1) investigating the acute and chronic effects of atrazine on four species of freshwater snails (Biomphalaria glabrata, Helisoma trivolvis, Physa acuta, and Stagnicola elodes) using the same methods, endpoints, and concentrations, and (2) summarizing the current literature pertaining to the effects of atrazine on freshwater snails. We conducted a 48 h acute toxicity test with an atrazine concentration higher than what typically occurs in aquatic environments (1000 µg/L). Additionally, we exposed snails to environmentally relevant atrazine concentrations (0, 0.3, 3, and 30 µg/L) for 28 days and assessed snail survival, growth, and reproduction. We also summarized all known literature pertaining to atrazine effects on freshwater snails. The literature summary suggests snails are often affected by environmentally relevant atrazine concentrations at the subcellular and cellular levels. These effects are typically not transitive to effects on survival, growth, or reproduction at the same concentrations. Our acute exposures corroborate the general trend of no direct effect on snail populations as atrazine did not directly affect the survival of any of the four snail species. Similarly, environmentally relevant concentrations did not significantly affect the survival, growth, or reproduction of any snail species. These results indicate that, in the absence of other possible stressors, the direct effects of environmentally relevant atrazine concentrations may not be realized at the snail population level.

Effects of host species and life stage on the helminth communities of sympatric northern leopard frogs (Lithobates pipiens) and wood frogs (Lithobates sylvaticus) in the Sheyenne National Grasslands, North Dakota.

We studied helminth communities in sympatric populations of leopard frogs (Lithobates pipiens) and wood frogs (Lithobates sylvaticus) and assessed the effects of host species and life stage on helminth community composition and helminth species richness. We examined 328 amphibians including 218 northern leopard frogs and 110 wood frogs collected between April and August of 2009 and 2010 in the Sheyenne National Grasslands of southeastern North Dakota. Echinostomatid metacercariae were the most common helminths found, with the highest prevalence in metamorphic wood frogs. Host species significantly influenced helminth community composition, and host life stage significantly influenced the component community composition of leopard frogs. In these sympatric populations, leopard frogs were common hosts for adult trematodes whereas wood frogs exhibited a higher prevalence of nematodes with direct life cycles. Metamorphic frogs were commonly infected with echinostomatid metacercariae and other larval trematodes whereas juvenile and adult frogs were most-frequently infected with directly transmitted nematodes and trophically transmitted trematodes. Accordingly, helminth species richness increased with the developmental life stage of the host.