Anadromous Arctic Char Microbiomes: Bioprospecting in the High Arctic

Northern populations of Arctic char (Salvelinus alpinus) can be anadromous, migrating annually from the ocean to freshwater lakes and rivers in order to escape sub-zero temperatures. Such seasonal behavior demands that these fish and their associated microbiomes adapt to changes in salinity, temperature, and other environmental challenges. We characterized the microbial community composition of anadromous S. alpinus, netted by Inuit fishermen at freshwater and seawater fishing sites in the high Arctic, both under ice and in open water. Bacterial profiles were generated by DNA extraction and high-throughput sequencing of PCR-amplified 16S ribosomal RNA genes. Results showed that microbial communities on the skin and intestine of Arctic char were statistically different when sampled from freshwater or saline water sites. This association was tested using hierarchical Ward's linkage clustering, showing eight distinct clusters in each of the skin and intestinal microbiomes, with the clusters reflecting sampling location between fresh and saline environments, confirming a salinity-linked turnover. This analysis also provided evidence for a core composition of skin and intestinal bacteria, with the phyla Proteobacteria, Firmicutes, and Cyanobacteria presenting as major phyla within the skin-associated microbiomes. The intestine-associated microbiome was characterized by unidentified genera from families Fusobacteriaceae, Comamonadaceae, Pseudomonadaceae, and Vibrionaceae. The salinity-linked turnover was further tested through ordinations that showed samples grouping based on environment for both skin- and intestine-associated microbiomes. This finding implies that core microbiomes between fresh and saline conditions could be used to assist in regulating optimal fish health in aquaculture practices. Furthermore, identified taxa from known psychrophiles and with nitrogen cycling properties suggest that there is additional potential for biotechnological applications for fish farm and waste management practices

New Insights on Polar Bear (Ursus maritimus) Diet from Faeces based on Next-Generation Sequencing Technologies

Practical tools to quantify range-wide dietary choices of the polar bear have not been well developed, thus impeding the monitoring of this species in a changing climate. Here we describe our steps toward non-invasive polar bear diet determination with the optimization of 454 pyrosequencing of a 136 base pair (bp) mitochondrial cytochrome b (cytb) fragment amplified from the extracts of captive and wild polar bear faeces. We first determine the efficacy, reliability, and accuracy of our method using five faecal samples from a captive polar bear fed a known diet at the Canadian Polar Bear Habitat in Cochrane, Ontario, Canada; 19 samples from three polar bears at the Metro Toronto Zoo, Toronto, Ontario, Canada; and seven samples from seven wild (unfed) polar bears from a holding facility in Churchill, Manitoba, Canada. We report 91% overall success in amplifying a 136 bp cytb amplicon from the faeces of polar bears. Our DNA analyses accurately recovered the vertebrate diet profiles of captive bears fed known diets. We then characterized multiyear vertebrate prey diet choices from free-ranging polar bears from the sea ice of the M’Clintock Channel polar bear management unit, Nunavut, Canada (n = 117 from an unknown number of bears). These data point to a diet unsurprisingly dominated by ringed seal (Pusa hispida) while including evidence of bearded seal (Erignathus barbatus), harbour seal (Phoca vitulina), muskox (Ovibos moschatus ssp.), Arctic fox (Vulpes lagopus), wolf (Canis lupus), Herring Gull (Larus argentatus), and Willow Ptarmigan (Lagopus lagopus). We found low levels of contamination (< 3% of sequences when present) and suggest specific process improvements to reduce contamination in range-wide studies. Together, these findings indicate that next-generation sequencing-based diet assessments show great promise in monitoring free-ranging polar bears in this time of climate change.   La réduction de la calotte glaciaire arctique suite au changement climatique risque d’avoir un effet direct sur la capacité des ours polaires à capturer les phoques, leurs principales sources de nourriture. Une surveillance précise des changements alimentaires des ours polaires s’avère ainsi essentielle pour mieux cerner l’impact des changements climatiques sur la survie de cette espèce. Nous détaillons dans cette étude, l’optimisation d’une méthode non invasive basée sur le séquençage de dernière génération (next generation sequencing - NGS) d’un fragment du gène mitochondrial cytochrome b (cytB) de 136 bp à partir de fèces d’ours polaires sauvages collectées en milieu naturel. Pour déterminer l’efficacité, la fiabilité et l’exactitude de notre méthode, nous avons analysé des fèces d›ours polaires en captivité dont le régime alimentaire était connu (Zoo Cochrane (n = 5), Toronto (Ontario, Canada) (n = 17) et des fèces d’ours polaires sauvages provenant de la ville de Churchill (Manitoba, Canada) (n= 7)) ainsi que de la région située au niveau du détroit de M’Clintock (Nunavut, Canada)(n= 117). Ces dernières fèces ont été analysées pour mieux cerner les choix alimentaires pluriannuels des ours polaires sauvages. Les profils alimentaires des ours captifs nourris avec des aliments connus ont été estimés avec précision et ont validé notre méthode. Notre étude sur les ours polaires sauvages du détroit de M’Clintock a révélé que même si le phoque annelé (Phoca hispidia) constituait la majorité de leur régime alimentaire, le phoque barbu (Erignathus barbatus), le phoque commun (Phoca vitulina), le boeuf musqué (Ovibos spp.), le renard arctique (Vulpes lagopus), le loup (Canis lupus), le goéland argenté (Larus argentatus) et le lagopède alpin (Lagopus lagopus) constituaient également des proies. Les risques de contaminations lors de l’utilisation de ces technologies NGS sont également discutés. De faibles degrés de contamination ont été observés (< 3 % des séquences lorsque la contamination était présente). Différentes stratégies sont proposées pour diminuer encore ces risques de contaminations. En conclusion notre étude démontre que les techniques de séquençage de dernière génération s’avèrent trés prometteuses pour l’étude de l’impact du changement climatique sur le régime alimentaire des ours polaires sauvages.&nbsp

New insights on polar bear (Ursus maritimus) diet from faeces based on Next Generation Sequencing technologies

Practical tools to quantify range-wide dietary choices on the polar bear have not been well developed impeding the monitoring of this species in a changing climate. Here we describe our steps toward non-invasive polar bear diet determination with the optimization of 454 pyrosequencing of a 136 (base pair: bp) mitochondrial cytochrome b (cytB) fragment amplified from the extracts of captive and wild polar bear faeces. We first determine the efficacy, reliability and accuracy of our method using polar bear faeces from captive polar bears fed known diets at the Cochrane Polar Bear Habitat (Canada, n = 5 faeces from 1 bear) and Metro Toronto Zoo (Canada, n =19 from 3 polar bears); and from wild (unfed) polar bears from a holding facility in Churchill (Canada; n=7 from 7 polar bears). We report 91% overall success in amplifying a 136 bp cytB amplicon from the faeces of polar bears. Our DNA analyses accurately recovered the vertebrate diet profiles of captive bears fed known diets. We then characterized multiyear vertebrate prey diet choices from free-ranging polar bears from the sea ice of the M’Clintock Channel (MC) polar bear Management Unit (Canada) (n =117 from an unknown number of bears). These data point to a diet unsurprisingly dominated by ringed seal (Phoca hispidia) while including evidence of bearded seal (Erignathus barbatus), harbor seal (Phoca vitulina), muskox (Ovibos spp.), Arctic foxes (Alopex lagopus), wolves (Canis lupus), herring gull (Larus argentatus) and willow ptarmigan (Lagopus lagopus). We found low levels pf contamination (<3% of sequences when present), suggesting specific process improvements to reduce contamination in range-wide studies. Together, these findings indicate that next generation sequencing-based diet assessments show great promise in monitoring free ranging polar bears in this time of climate chang

ddRAD genotyping reveals hierarchical genetic population structure in anadromous Arctic char (Salvelinus alpinus) in the Lower Northwest Passage, Nunavut

Contemporary intraspecific patterns of genetic variation reflect the historical effects of population subdivision/expansion and forces like drift, gene flow and selection. We investigated the population structure of anadromous Arctic char in the Lower Northwest Passage (LNWP), Nunavut, using 3,074 genome-wide single nucleotide polymorphisms markers (SNPs). Overall, the genetic differentiation was weak to moderate among 18 sampling locales (global FST=0.037). Populations were structured hierarchically, with a deeper genetic division between King William Island (global FST=0.018) and mainland populations ~ 200 km away to the south (global FST=0.018), and some evidence of genetic subgroups within the former. These findings suggest a role for char migratory behaviour in shaping contemporary genetic population structure, with demographic modelling favouring an isolation with migration over a strict isolation scenario. Twenty-two SNPs were identified as potentially under divergent selection with putative functions including neurotransmission and bone development and growth during late embryogenesis. Our study is the first survey of Arctic char in the LNWP using genomics, and provides baseline data for the development of a sustainable fishery within this region.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

The optimization of microsatellite genotyping and genetic sexing of non-invasively collected polar bear tissue: Implications for monitoring and census.

The monitoring of Polar Bears in Canada has been completed largely through management unit (MU) wide capture-mark-recapture (CMR) surveys. While this data is very useful at the time of collection, these surveys are expensive and take time to plan and execute; cannot be feasibly executed across the polar bear range at intervals that reflect the expected rapid environmental changes in the Arctic; and are disdained by the Inuit as being invasive. As part of recent efforts to explore less expensive and non-invasive methods to monitor polar bears (see Wong et al & Van Coevderden de Groot et al this conference) we are evaluating genetic information obtained from non-invasively collected polar bear tissue. In this work we report on the genetic data obtained from non-invasively collected harisnags recovered from sampling stations erected between May-June 2006-2009 in M’Clintock Channel, Nunavut. Across the 4 years 344 hair snags were collected; following Paetkau (2004) we optimized 6 microsatellite loci to reliably amplify polar bear DNA from this tissue and we modified the procedure of Pages et al (2009) to reliably genetically sex these tissues. Our estimates for two common errors with this type of tissue across all loci – allelic dropout (0.026) and false allele (0.03) - were both less than p =.05. This suggests these errors are not going to significantly affect the accuracy of the consensus genotypes collected from these data. Using consensus genotypes from relevant hairsnags, we posit a minimum of 59 (max 82) unique bears entered our sampling stations. Of these, 24% were female, 64% were male, and 12% could not be sexed. We resampled 2 bears in 2006, 1 in 2007, 0 bears in 2008 and 14 bears in 2009 – the 2009 value reflects significantly increased sampling effort in 2009. Five bears were re-sampled between the non-invasive surveys in 2006-2009. When comparing our data to a subset of cubs and subadults captured during the Taylor et al. (2006) CMR survey of M’Clintock Channel (MU), we found 6 genotype matches. Our sampling stations may have a male bias as the sex ratio from the 1998-2000 CMR study was 42.1% ♂ (Taylor et al 2006) vs. 64% ♂ (this study). We cannot accurately determine the age bias (but see Wong et al this conference). Genetic data from Polar bear faecal samples may provide an unbiased sex and age sample of polar bears in any MU. Any data from these samples will help refine hairsnag derived MKNA estimate of polar bears from any MU. Here we report on our efforts to genotype and genetically sex 95 faecals we have collected from M’Clintock Channel from 2006-2009. Finally, we discuss the implications of these findings, results from other noninvasive work (Wong et al & Van Coevderden de Groot et al this conference) and ongoing/proposed work in the context of i) a non-invasive Inuit-based polar bear activity and health survey, and ii) a more rigorous census method which may allow more precise adjustments of harvest levels than currently possible using infrequently collected CMR data only

Unexpected and undesired conservation outcomes of wildlife trade bans—An emerging problem for stakeholders?

CITES regulates international trade with the goal of preventing over-exploitation, thus the survival of species are not jeopardized from trade practices; however it has been used recently in nontrade conservation measures. As an example, the US proposed to up-list polar bears under CITES Appendix I, despite that the species did not conform to the biological criteria. Polar bears were listed as ‘threatened’ under US ESA in 2008, in response to loss of sea-ice and warming temperatures. In Nunavut, where most of Canada’s polar bears are harvested, the resulting trade ban did not decrease total harvest after the ESA listing but reduced US hunter participation and the proportion of quotas taken by sport hunters from specific populations. Consequently, the import ban impacted livelihoods of Arctic indigenous communities with negative conservation — reduced tolerance for dangerous fauna and affected local participation in shared management initiatives. The polar bear may be the exemplar of an emerging problem: the use of trade bans in place of action for non-trade threats, e.g., climate change. Conservation prospects for this species and other climate-sensitive wildlife will likely diminish if the increasing use of trade bans to combat non-trade issues cause stakeholders to lose faith in participatory management

Free-living, psychrotrophic bacteria of the genus Psychrobacter are descendents of pathobionts

Host-adapted microorganisms are generally assumed to have evolved from free-living, environmental microorganisms, as examples of the reverse process are rare. In the phylum Gammaproteobacteria, family Moraxellaceae, the genus Psychrobacter includes strains from a broad ecological distribution including animal bodies as well as sea ice and other nonhost environments. To elucidate the relationship between these ecological niches and Psychrobacter’s evolutionary history, we performed tandem genomic analyses with phenotyping of 85 Psychrobacter accessions. Phylogenomic analysis of the family Moraxellaceae reveals that basal members of the Psychrobacter clade are Moraxella spp., a group of often-pathogenic organisms. Psychrobacter exhibited two broad growth patterns in our phenotypic screen: one group that we called the “flexible ecotype” (FE) had the ability to grow between 4 and 37°C, and the other, which we called the “restricted ecotype” (RE), could grow between 4 and 25°C. The FE group includes phylogenetically basal strains, and FE strains exhibit increased transposon copy numbers, smaller genomes, and a higher likelihood to be bile salt resistant. The RE group contains only phylogenetically derived strains and has increased proportions of lipid metabolism and biofilm formation genes, functions that are adaptive to cold stress. In a 16S rRNA gene survey of polar bear fecal samples, we detect both FE and RE strains, but in in vivo colonizations of gnotobiotic mice, only FE strains persist. Our results indicate the ability to grow at 37°C, seemingly necessary for mammalian gut colonization, is an ancestral trait for Psychrobacter, which likely evolved from a pathobiont