Cara N. Love

32 species validation of a new Illumina paired-end approach for the development of microsatellites

Development and optimization of novel species-specific microsatellites, or simple sequence repeats (SSRs) remains an important step for studies in ecology, evolution, and behavior. Numerous approaches exist for identifying new SSRs that vary widely in terms of both time and cost investments. A recent approach of using paired-end Illumina sequence data in conjunction with the bioinformatics pipeline, PAL_FINDER, has the potential to substantially reduce the cost and labor investment while also improving efficiency. However, it does not appear that the approach has been widely adopted, perhaps due to concerns over its broad applicability across taxa. Therefore, to validate the utility of the approach we developed SSRs for 32 species representing 30 families, 25 orders, 11 classes, and six phyla and optimized SSRs for 13 of the species. Overall the IPE method worked extremely well and we identified 1000s of SSRs for all species (mean = 128,485), with 17% of loci being potentially amplifiable loci, and 25% of these met our most stringent criteria designed to that avoid SSRs associated with repetitive elements. Approximately 61% of screened primers yielded strong amplification of a single locus.

Development of 31 polymorphic microsatellite markers for the mole salamander (Ambystoma talpoideum) using Illumina paired-end sequencing

We isolated and characterized a total of 31 microsatellite loci from the mole salamander, Ambystoma talpoideum. Loci were screened in 20 individuals from a single location in Aiken, South Carolina. The number of alleles per locus ranged from 3 to 11, observed heterozygosity ranged from 0.000 to 0.700, and the probability of identity values ranged from 0.031 to 0.400. These new loci will provide tools for examining the genetic diversity, structure, mating system, and adult morph determination of A. talpoideum.

Development of polymorphic microsatellite markers for blue king crab (Paralithodes platypus)

We isolated and characterized a total of 23 microsatellite loci from the blue king crab, Paralithodesplatypus. Loci were screened in 24 individuals from St. Matthew Island. The number of alleles per locus ranged from 4 to 17, observed heterozygosity ranged from 0.050 to 1.000, and the probability of identity values ranged from 0.015 to 0.339. These new loci will provide tools for examining the genetic population structure of the species throughout its range.

Development of polymorphic microsatellite markers for the North American porcupine, Erethizon dorsatum, using paired-end Illumina sequencing

We isolated and characterized a total of 19 microsatellite loci from the North American porcupine, Erethizon dorsatum. Loci were screened in 22 individuals from St. Lawrence and Franklin Counties in northern New York State. The number of alleles per locus ranged from 4 to 14, observed heterozygosity ranged from 0.316 to 1.000, and the probability of identity values ranged from 0.028 to 0.243. These new loci will provide tools for examination of the effects of habitat fragmentation, for instance by roads and other infrastructure, on gene flow and population subdivision in this species.

Development and characterization of thirty novel microsatellite markers for the critically endangered Myanmar Roofed Turtle, Batagur trivittata, and cross-amplification in the Painted River Terrapin, B. borneoensis, and the Southern River Terrapin, B. affinis, using paired-end Illumina shotgun sequencing

We isolated and characterized 30 microsatellite loci from the critically endangered Myanmar Roofed Turtle, Batagur trivittata. Loci were screened in 9 B. trivittata samples and in the congeners the Painted River Terrapin, Batagur borneoensis, with 22 of 30 amplifying, and the Southern River Terrapin, B. affinis, with 15 of 30 amplifying. In the B. trivittata samples, the number of alleles per locus ranged from 3 to 10 and the probability of identity values ranged from 0.031 to 0.354. These new loci will provide tools for captive management and reintroduction programs for B. trivittata and the other five species of Batagur, particularly B. affinis in Cambodia.

Isolation and characterization of 18 novel polymorphic microsatellite markers from the Mayan cichlid (Cichlasoma urophthalmus)

We isolated and characterized 18 microsatellite loci from the Mayan cichlid, Cichlasoma urophthalmus. Loci were screened for 24 specimens from a total of seven sites in south Florida, Mexico, Belize and Honduras. The number of alleles per locus ranged from 3 to 21, observed heterozygosity ranged from 0.208 to 0.875, and the probability of identity values ranged from 0.012 to 0.203. These new loci will provide tools for identifying the source population(s) for the introduction of Mayan cichlids in south Florida and for comparing population genetic structure of Mayan cichlids within and among subpopulations in its native (Central America) and introduced ranges (south Florida). Mayan cichlids are an invasive species in south Florida so identifying source populations may reveal pathways that can be managed to prevent further introductions. Mayan cichlids may also be useful as a model system in which to examine the relationship between introduction history, population genetic diversity, and invasibility.

Patterns of amphibian infection prevalence across wetlands on the Savannah River Site, South Carolina, USA.

Amphibian diseases, such as chytridiomycosis caused by Batrachochytrium dendrobatidis (Bd) and ranaviral disease caused by ranaviruses, are often linked to global amphibian population declines, yet the ecological dynamics of both pathogens are poorly understood. The goal of our study was to determine the baseline prevalence, pathogen loads, and co-infection rate of Bd and ranavirus across the Savannah River Site (SRS) in South Carolina, USA, a region with rich amphibian diversity and a history of amphibian-based research. We tested over 1000 individuals, encompassing 21 amphibian species from 11 wetlands for both Bd and ranavirus. The prevalence of Bd across individuals was 9.7%. Using wetland means, the mean (±SE) Bd prevalence was 7.9 ± 2.9%. Among toad species, Anaxyrus terrestris had 95 and 380% greater odds of being infected with Bd than Scaphiopus holbrookii and Gastrophryne carolinensis, respectively. Odds of Bd infection in adult A. terrestris and Lithobates sphenocephalus were 75 to 77% greater in metal-contaminated sites. The prevalence of ranavirus infections across all individuals was 37.4%. Mean wetland ranavirus prevalence was 29.8 ± 8.8% and was higher in post-metamorphic individuals than in aquatic larvae. Ambystoma tigrinum had 83 to 85% higher odds of ranavirus infection than A. opacum and A. talpoideum. We detected a 4.8% co-infection rate, with individuals positive for ranavirus having a 5% higher occurrence of Bd. In adult Anaxyrus terrestris, odds of Bd infection were 13% higher in ranavirus-positive animals and odds of co-infection were 23% higher in contaminated wetlands. Overall, we found the pathogen prevalence varied by wetland, species, and life stage.

Testing for Associations between Hematozoa Infection and Mercury in Wading Bird Nestlings

Abstract Several wading bird species in the southeastern US have a history of infection by hematozoa/avian malaria as well as mercury accumulation through their diet, and thus may be exposed to two, generally sublethal, yet chronic, stressors. We analyzed nestling wading birds (n = 171) of varying size and trophic position from the southeastern US, and a smaller sample (n = 23) of older, free-ranging birds, to look for potential interrelationships between infection by hematozoa and mercury (Hg) uptake. Only one nestling was PCR positive for hematozoa (Plasmodium/Haemoproteus) whereas nine (39%) of the older wading birds were positive. Sequencing indicated that both nestling and adult positives were infected with Plasmodium. Given the low infection rate of the nestlings, there was no association between Hg and malaria. The older birds exhibited a possible malaria/Hg association, but it may be confounded by their greater potential exposure period and large-scale movements.

Development and characterization of thirty-three microsatellite markers for the Patagonian sprat, Sprattus fuegensis (Jenyns, 1842), using paired-end Illumina shotgun sequencing

We isolated and characterized a total of 33 microsatellite loci from the Patagonian sprat Sprattusfuegensis, a recent exploited marine resource with a conservation status unknowing. Loci were screened in 24 individuals from the inshore waters of the Aysén Fjord, Chile. The number of alleles per locus ranged from 7 to 24, observed heterozygosity ranged from 0.217 to 0.875, and the probability of identity values ranged from 0.006 to 0.133. These new loci will provide tools for examining population genetic structure, estimating effective population size and provide information to fisheries management and conservation.

Development of twenty-one polymorphic microsatellite markers for the fungus-growing ant, Mycocepurus goeldii (Formicidae: Attini), using Illumina paired-end genomic sequencing

Obligate social parasites, or inquilines, exploit the colonies of free-living social species and evolved at least 80 times in ants alone. Most species of the highly specialized inquiline social parasites are rare, only known from one or very few, geographically isolated populations, and the sexual offspring of most inquiline species mates inside the maternal colony. Therefore, inquiline populations are believed to be small and genetically homogeneous due to inbreeding. To comparatively study the genetic diversity of the socially parasitic fungus-growing ant, Mycocepurus castrator, and its only known host species, Mycocepurus goeldii, and to infer the parasite’s conservation status, we developed 21 microsatellite markers for the host species, M. goeldii, and evaluated whether these markers cross-amplify in the social parasite, M. castrator. We isolated and characterized a total of 21 microsatellite loci for M. goeldii. The loci were screened for 24 individuals from geographically distant and genetically divergent populations in Brazil. The number of alleles per locus ranged from 18 to 4, the observed heterozygosity ranged from 0.25 to 0.636, and the probability of identity values ranged from 0.011 to 0.146. Preliminary analyses show that these markers cross amplify in the closely related social parasite species M. castrator. These newly developed loci provide tools for studying the genetic diversity and the evolution of social parasitism in the Mycocepurus host–parasite system.