Investigation to determine staff exposure and describe animal bite surveillance after detection of a rabid zebra in a safari lodge in Kenya, 2011

Introduction: Rabies is a fatal viral infection, resulting in >55,000 deaths globally each year. In August 2011, a young orphaned zebra at a Kenyan safari lodge acquired rabies and potentially exposed >150  tourists and local staff. An investigation was initiated to determine exposures among the local staff, and to describe animal bite surveillance in the affected district.Methods: We interviewed lodge staff on circumstances surrounding the zebra's illness and assessed  their exposure status. We reviewed animal bite report forms from the outpatient department at the district hospital. Results: The zebra was reported bitten by a dog on 31st July 2011, became ill on  23rdAugust, and died three days later. There were 22 employees working at the lodge during that time. Six (27%) had high  exposure due to contact with saliva (bottle feeding, veterinary care) and received four doses of rabies  vaccine and one of immune-globulin, and 16 (73%) had low exposure due to casual contact and received only four doses of rabies vaccine. From January 2010 to September 2011, 118 cases of animal bites were reported in the district; 67 (57%) occurred among males, 65 (57%) in children <15 years old, and 61  (52%) were inflicted in a lower extremity. Domestic and stray dogs accounted for 98% of reported bites.Conclusion: Dog bites remains the main source of rabies exposure in the district, but exposure can  result from wildlife. This highlights the importance of a one health approach with strong communication between wildlife, veterinary, and human health sectors to improve rabies prevention and control.Key words: Rabies, outbreak, epidemiology, East Africa 

Considerations for studying transmission of antimicrobial resistant enteric bacteria between wild birds and the environment on intensive dairy and beef cattle operations

Background Wild birds using livestock facilities for food and shelter may contribute to dissemination of enteric pathogens or antimicrobial resistant bacteria. However, drivers of microbial exchange among wildlife and livestock are not well characterized. Predisposition for acquiring and retaining environmental bacteria may vary among species because of physiologic or behavioral differences, complicating selection of a bacterial model that can accurately characterize microbial connections among hosts of interest. This study compares the prevalence and antibiotic resistance phenotypes of two potential model bacterial organisms isolated from wild birds and their environments. Methods We compared prevalence and resistance profiles of Escherichia coli and Enterococcus species isolated from environmental swabs and bird feces on a residential control site, a confinement dairy, a pasture-based beef farm, and a confinement beef farm. Results Bird feces at all sites had low-to-moderate prevalence of Escherichia coli (range: 17–47%), despite potential for exposure on farms (range: 63–97%). Few Escherichia coli were isolated from the control environment. Enterococcus faecalis was dominant in birds at both beef farms (62% and 81% of Enterococcus isolates) and low-to-moderately prevalent at the dairy and control sites (29% and 23% of isolates, respectively). Antimicrobial resistance prevalence was higher in farm samples compared to those from the residential control, but distribution of resistant isolates varied between the bacterial genera. Birds on all farms carried resistant Enterococcus at similar rates to that of the environment, but resistance was less common in bird-associated Escherichia coli despite presence of resistant isolates in the farm environment. Discussion Bacterial species studied may affect how readily bacterial exchange among populations is detected. Selection of microbial models must carefully consider both the questions being posed and how findings might influence resulting management decisions

Salmonella Strains Isolated from Galápagos Iguanas Show Spatial Structuring of Serovar and Genomic Diversity

It is thought that dispersal limitation primarily structures host-associated bacterial populations because host distributions inherently limit transmission opportunities. However, enteric bacteria may disperse great distances during food-borne outbreaks. It is unclear if such rapid long-distance dispersal events happen regularly in natural systems or if these events represent an anthropogenic exception. We characterized Salmonella enterica isolates from the feces of free-living Galápagos land and marine iguanas from five sites on four islands using serotyping and genomic fingerprinting. Each site hosted unique and nearly exclusive serovar assemblages. Genomic fingerprint analysis offered a more complex model of S. enterica biogeography, with evidence of both unique strain pools and of spatial population structuring along a geographic gradient. These findings suggest that even relatively generalist enteric bacteria may be strongly dispersal limited in a natural system with strong barriers, such as oceanic divides. Yet, these differing results seen on two typing methods also suggests that genomic variation is less dispersal limited, allowing for different ecological processes to shape biogeographical patterns of the core and flexible portions of this bacterial species' genome

Evolution of Salmonella enterica Virulence via Point Mutations in the Fimbrial Adhesin

Whereas the majority of pathogenic Salmonella serovars are capable of infecting many different animal species, typically producing a self-limited gastroenteritis, serovars with narrow host-specificity exhibit increased virulence and their infections frequently result in fatal systemic diseases. In our study, a genetic and functional analysis of the mannose-specific type 1 fimbrial adhesin FimH from a variety of serovars of Salmonella enterica revealed that specific mutant variants of FimH are common in host-adapted (systemically invasive) serovars. We have found that while the low-binding shear-dependent phenotype of the adhesin is preserved in broad host-range (usually systemically non-invasive) Salmonella, the majority of host-adapted serovars express FimH variants with one of two alternative phenotypes: a significantly increased binding to mannose (as in S. Typhi, S. Paratyphi C, S. Dublin and some isolates of S. Choleraesuis), or complete loss of the mannose-binding activity (as in S. Paratyphi B, S. Choleraesuis and S. Gallinarum). The functional diversification of FimH in host-adapted Salmonella results from recently acquired structural mutations. Many of the mutations are of a convergent nature indicative of strong positive selection. The high-binding phenotype of FimH that leads to increased bacterial adhesiveness to and invasiveness of epithelial cells and macrophages usually precedes acquisition of the non-binding phenotype. Collectively these observations suggest that activation or inactivation of mannose-specific adhesive properties in different systemically invasive serovars of Salmonella reflects their dynamic trajectories of adaptation to a life style in specific hosts. In conclusion, our study demonstrates that point mutations are the target of positive selection and, in addition to horizontal gene transfer and genome degradation events, can contribute to the differential pathoadaptive evolution of Salmonella

Site effects on wildlife enteric bacterial diversity: you are where you eat?

The study of microbial biogeography has made vast strides in recent years due, in part, to advances in the technological capacity to document microscopic biodiversity at the community, population and genomic levels. In this dissertation, we apply a combination of molecular and cultivation approaches to the study of enteric microbial diversity in a unique system of sister iguanid species, the Galápagos land and marine iguanas (Conolophus spp. and Amblyrhynchus subcristatus, respectively). We explored the spatial diversity of enteric bacteria in these two species at multiple levels, from communities to genetic traits. The unique host population history and geography of this island chain informed hypotheses about expected biogeographical patterns and the underlying processes that shape microbial diversity in this system. In Chapter 1, we reviewed current understanding of microbial biogeography and explored how incorporation of such ecological theory might benefit understanding of enteric microbial community structure and function across space and time. Chapter 2 explored the spatial community diversity of marine and land iguana enteric communities using a molecular approach based on 454 pyrosequencing. Firstly, we demonstrated that while host species was the strongest force shaping these communities, within a host species, geographical proximity also determined overlap in community composition at the genus level. In addition, we found that the degree of contact among host species can produce distinct local effects in community richness and composition, especially when host species have amplified opportunities for microbial exchange such as higher population densities or limited habitat area. Next in Chapter 3, we explored taxonomically finer-scale diversity patterns among marine iguana populations for the rich enteric genus Clostridium. In contrast to the expectation that each host population should have relatively distinct Clostridium communities, we found a surprising amount of phylogenetic conservation across all sites, despite also demonstrating evidence suggesting on-going taxonomic turnover – forces which might otherwise lead to rapid divergence of enteric communities in allopatric host populations. We then applied a more traditional cultivation and molecular genetic approach in Chapter 4 in order to document Salmonella enterica strain diversity among sites. We serotyped and genotyped Salmonella enterica isolates carried by land and marine iguanas across a geographical gradient and found nearly complete isolation among strain pools. However, we also found suggestions of geographically-dependent genomic similarity among sites, possibly due to long-distance transport of genetic elements by oceanic currents. Finally, in Chapter 5 we explored genetic trait biogeography in this system by documenting phenotypic and genetic patterns of antibiotic resistance. We found that sites farther from high densities of humans (i.e. major port towns) harbored fewer resistant bacteria. We also noted that these antibiotic resistance traits may not be retained within the broader endemic bacterial community of Galápagos wildlife for any appreciable length of time, as Salmonella enteric isolates did not share resistance traits found in Escherichia coli within the same site or even within the same individual fecal sample. Through this work as a whole we demonstrated that, within the context of a single study system, biogeographical patterns and mechanisms can vary widely across bacterial taxonomic levels, with both ecological and evolutionary forces acting in concert on enteric biodiversity. Increased understanding of the interplay among these forces for shaping microbial community form and function across taxonomic scales has potential to improve not only theoretical understanding of microbial ecology but also to advance management of pressing issues such as novel disease emergence or antibiotic resistance dissemination

Appendix C. Supplemental analyses: fungal communities.

Supplemental analyses: fungal communities

Appendix A. tRFLP analysis laboratory methodology.

tRFLP analysis laboratory methodology

Data from: Ecological drift and local exposures drive gastrointestinal bacterial community differences among Galápagos iguana populations

Diet strongly influences the intestinal microbial communities through species sorting. Alternatively, these communicates may differ because of chance variation in local microbial exposures or species losses among allopatric host populations (i.e. ecological drift). We investigated how these forces shape enteric communities of Galápagos marine and land iguanas. Geographically proximate populations shared more similar communities within a host ecotype, suggesting a role for ecological drift during host colonization of the islands. Additionally, evidence of taxa sharing between proximate heterospecific host populations suggests that contemporary local exposures also influence the gut community assembly. While selective forces such as host-bacterial interactions or dietary differences are dominant drivers of intestinal community differences among hosts, historical and contemporary processes of ecological drift may lead to differences in bacterial composition within a host species. Whether such differences in community structure translate into geographic variation in benefits derived from these intimate microbial communities remains to be explored

Marine Iguana Sequences

Contains bacterial community 454 pyrosequencing reads obtained from fecal samples of marine iguanas (MI) living on Isla Fernandina (F), Isla Plaza Sur (P), Isla Santa Fe (S), and two sites on Isla San Cristobal - La Lobería (L) and Punta Carola (C) in the Galápagos Islands, Ecuador