Mercury Exposure Is Associated with Urbanization but not Salmonella Status in White Ibis (Eudocimus albus).

Mercury (Hg) is a ubiquitous contaminant in wetlands that can cause immunosuppression in birds, which may increase susceptibility to colonization with Salmonella spp. Previously, we found that White Ibis (Eudocimus albus), a recently urbanized wading bird, shed Salmonella spp. at a higher prevalence when captured at urban sites, compared with natural sites. In this study, we sought to determine if Hg burdens in ibis are related to Salmonella status or degree of urbanization or both. We analyzed feathers from 94 ibis in Palm Beach County, Florida, USA, along an urbanization gradient (0-68% urbanization) and from individuals with confirmed Salmonella spp. status (shedding or not shedding). We detected Hg in all ibis feathers (0.22-8.47 mg/kg; mean=1.96 mg/kg; SD=1.94). The Hg concentration was not significantly correlated to Salmonella spp. shedding status (Wilcoxon rank sum test, W=1170; P=0.596) but was negatively associated with capture site urbanization level (R2=0.327; P=0.026). Our findings may suggest that the immunosuppressive effects of Hg do not affect Salmonella shedding in the ibis or that Hg burdens were too low to affect Salmonella shedding status. Further, ibis that were captured in high urbanization sites appeared to have a lower risk of Hg exposure than ibis that were captured within low urbanization sites.

Human Salmonellosis Outbreak Linked to Salmonella Typhimurium Epidemic in Wild Songbirds, United States, 2020-2021.

Salmonella infection causes epidemic death in wild songbirds, with potential to spread to humans. In February 2021, public health officials in Oregon and Washington, USA, isolated a strain of Salmonella enterica serovar Typhimurium from humans and a wild songbird. Investigation by public health partners ultimately identified 30 illnesses in 12 states linked to an epidemic of Salmonella Typhimurium in songbirds. We report a multistate outbreak of human salmonellosis associated with songbirds, resulting from direct handling of sick and dead birds or indirect contact with contaminated birdfeeders. Companion animals might have contributed to the spread of Salmonella between songbirds and patients; the outbreak strain was detected in 1 ill dog, and a cat became ill after contact with a wild bird. This outbreak highlights a One Health issue where actions like regular cleaning of birdfeeders might reduce the health risk to wildlife, companion animals, and humans.

An Acute Mortality Event Associated with Novel Macrorhabdus ornithogaster Infection and Underlying Factors in a Newly-Established Captive Group of American White Ibis (Eudocimus albus) Nestlings.

Twenty-four American white ibis (Eudocimus albus) nestlings were collected in Florida (USA) on 17 April 2017 to establish a captive flock. On 7 May 2017, three birds died suddenly, following severe lethargy, hemorrhaging from the mouth and nares, anorexia, and production of bright-green colored feces. An additional ibis with delayed growth and pathological fractures was euthanized 18 May 2017. Severe ventriculitis associated with Macrorhabdus ornithogaster was noted in all four birds, bacterial sepsis was confirmed in one bird by culture and histologic examination, and bacterial endotoxemia was suspected in two birds based on gross and histologic examination, but no bacteria were isolated from these birds. Birds also had vitamin E liver levels consistent with coagulopathy previously described in pelicans. We sampled feces from 91 adult, free-living, healthy ibis in Florida in July 2017 and found 71% were shedding organisms with morphologic characteristics consistent with Macrorhabdus sp. Molecular characterization of the ibis-origin M. ornithogaster showed it was phylogenetically related to numerous M. ornithogaster sequences. It is unknown if M. ornithogaster infection resulted in clinical disease as a result of dietary or stress-related dysbiosis, or other factors. Macrorhabdus-associated disease has not previously been confirmed in wading birds. We discuss potential associations of gastric M. ornithogaster infection with morbidity and mortality in these cases and highlight the need for additional studies on this pathogen in free-living birds.

A single Haemoproteus plataleae haplotype is widespread in white ibis (Eudocimus albus) from urban and rural sites in southern Florida.

The American white ibis (Eudocimus albus), a common bird species in Florida, has become increasingly urban, with many populations relying heavily on urban and suburban habitats, which may alter parasite transmission. Parasites of ibis, especially haemosporidians, are understudied. Avian haemosporidia can have a wide range of impacts on birds, including decreased reproductive success or increased mortality. Because southern Florida is subtropical and has a high diversity of potential vectors for haemosporidia, we hypothesized that there will be a high prevalence and genetic diversity of haemosporidia in white ibis. A total of 636 ibis from South Florida were sampled from 2010 to 2022, and blood samples were tested for haemosporidia by examination of Giemsa-stained thin blood smears and/or nested PCRs targeting the cytochrome b gene. A total of 400 (62.9%, 95% CI 59-66.7%) ibis were positive for parasites that were morphologically identified as Haemoproteus plataleae. Sequences of 302 positives revealed a single haplotype of Haemoproteus (EUDRUB01), which was previously reported from white ibis in South Florida and captive scarlet ibis (E. ruber) in Brazil. No Plasmodium or Leucocytozoon infections were detected. Parasitemias of the 400 positive birds were very low (average 0.084%, range 0.001%-2.16% [although only 2 birds had parasitemias >1%]). Prevalence and parasitemias were similar for males and females (68% vs. 61.6% and 0.081% vs. 0.071%, respectively). Prevalence in juveniles was lower compared with adults (52% vs. 67.4%) but parasitemias were higher in juveniles (0.117% vs. 0.065%). This data shows that H. plataleae is common in ibis in South Florida. Although parasitemias were generally low, additional research is needed to determine if this parasite has subclinical effects on ibis, if additional haplotypes or parasite species infect ibis in other regions of their range, or if H. plataleae is pathogenic for other sympatric avian species.

Surveillance for gastrointestinal, subcutaneous, and ectoparasites of invasive North American raccoons (Procyon lotor) in central Spain.

The American raccoon (Procyon lotor) is an invasive meso-carnivore which has been introduced and established in many European countries. Although the presence of the raccoon in the Iberian Peninsula was confirmed around 20 years ago, there are few data on pathogens of these animals in this region. For this work, 72 American raccoons from two subpopulations in the central region of the Iberian Peninsula were examined for selected parasites. Ectoparasite species richness (both fleas and ticks) increased during the sampling season and was highest in the Henares subpopulation and on males. Similarly, ectoparasite abundance increased during the sampling season and was highest in Henares and on adult raccoons. Four species of ticks were detected including Rhipicephalus pusillus (71%), followed by R. sanguineus sensu lato (24%), Ixodes ventalloi (3%), and Dermacentor marginatus (1.4%). Four species of fleas were detected including Pulex irritans (44%), Ctenocephalides felis (3%), C. canis (1.4%), and Paraceras melis (1.4%) infestations. A subset of raccoons (n = 56) was examined for intestinal parasites; low prevalence and diversity were found including Strongyloides procyonis (4%), Dilepis sp. (5%), Plagiorchis sp. (2%), and Moniliformis moniliformis (2%). Importantly, Baylisascaris procyonis was not found. Finally, no subcutaneous nematodes (i.e., Dracunculus and Dirofilaria spp.) were found in the 56 raccoons examined. The results of this work show that the invasive North American raccoons currently are infected with few endoparasites but are commonly infested with native ectoparasites, several of which can transmit pathogens relevant for public and veterinary health. However, the geographically distinct populations of raccoons in Spain have different introduction histories, thus additional surveillance for parasites is warranted.

West Nile Virus Infections in an Urban Colony of American White Ibises (Eudocimus albus) in South Florida, USA.

West Nile virus (WNV) is pathogenic in a wide range of avian hosts and is endemic in much of North America. This virus is responsible for population declines of some Passeriformes. We describe a WNV-associated mortality event in American White Ibis (Eudocimus albus) nestlings. This is a species, inherent to the Everglades ecosystem, which has recently begun nesting in urban areas. An urban colony in south Florida was monitored from March-July in 2020 as part of an ongoing study. Nestling carcasses were collected opportunistically and sent to the Southeastern Cooperative Wildlife Disease Study, University of Georgia within 24 h for diagnosis. Three ibis nestling deaths were confirmed to be caused by WNV infection based on histopathology, immunohistochemistry, and reverse transcription PCR. Serial plasma samples collected weekly from 36 healthy chicks of the same urban rookery were tested for WNV-neutralizing antibodies via plaque reduction neutralization test; four chicks were seropositive. Antibody titers in three seropositive chicks from which serial samples were collected waned over time, suggesting maternal antibody transfer. Ibis mortalities were consistent with a spike of WNV activity in this region of Florida. West Nile virus infection may be an important seasonal cause of mortality for wading bird nestlings.

Avian Influenza Virus Status and Maternal Antibodies in Nestling White Ibis (Eudocimus albus).

The White Ibis (Eudocimus albus), a nomadic wading bird, has increased its exploitation of urban habitats in South Florida, United States, and has recently established several urban breeding colonies. Certain characteristics of ibis ecology could position them in the natural cycle of the avian influenza virus (AIV). In fact, experimentally infected ibises were shown to be competent hosts for multiple AIV subtypes, and seroconversion to AIV has been documented in adult ibises in natural populations. However, the mechanisms of transmission and the timing of infection are unclear as we have yet to isolate AIV from a free-living ibis. To investigate the age-specific AIV dynamics of ibis, we captured nestlings (n = 115) weekly for 1-4 weeks from urban and natural settings in 2020 and 2021. We collected choanal/cloacal swabs for rRT-PCR and virus isolation, and plasma to screen for maternal AIV antibodies. AIV was not detected in any individual by virus isolation; however, maternal antibodies to AIV were detected in 95% of nestlings, with varying rates of catabolism. These results confirm that nestlings are afforded maternal antibodies from adults at rates reflective of higher adult seroprevalence than previously documented and that nestlings in breeding colonies may have some degree of protection and are unlikely to become infected with AIV.

Free-Living Aquatic Turtles as Sentinels of Salmonella spp. for Water Bodies.

Reptile-associated human salmonellosis cases have increased recently in the United States. It is not uncommon to find healthy chelonians shedding Salmonella enterica. The rate and frequency of bacterial shedding are not fully understood, and most studies have focused on captive vs. free-living chelonians and often in relation to an outbreak. Their ecology and significance as sentinels are important to understanding Salmonella transmission. In 2012-2013, Salmonella prevalence was determined for free-living aquatic turtles in man-made ponds in Clarke and Oconee Counties, in northern Georgia (USA) and the correlation between species, basking ecology, demographics (age/sex), season, or landcover with prevalence was assessed. The genetic relatedness between turtle and archived, human isolates, as well as, other archived animal and water isolates reported from this study area was examined. Salmonella was isolated from 45 of 194 turtles (23.2%, range 14-100%) across six species. Prevalence was higher in juveniles (36%) than adults (20%), higher in females (33%) than males (18%), and higher in bottom-dwelling species (31%; common and loggerhead musk turtles, common snapping turtles) than basking species (15%; sliders, painted turtles). Salmonella prevalence decreased as forest cover, canopy cover, and distance from roads increased. Prevalence was also higher in low-density, residential areas that have 20-49% impervious surface. A total of 9 different serovars of two subspecies were isolated including 3 S. enterica subsp. arizonae and 44 S. enterica subsp. enterica (two turtles had two serotypes isolated from each). Among the S. enterica serovars, Montevideo (n = 13) and Rubislaw (n = 11) were predominant. Salmonella serovars Muenchen, Newport, Mississippi, Inverness, Brazil, and Paratyphi B. var L(+) tartrate positive (Java) were also isolated. Importantly, 85% of the turtle isolates matched pulsed-field gel electrophoresis patterns of human isolates, including those reported from Georgia. Collectively, these results suggest that turtles accumulate Salmonella present in water bodies, and they may be effective sentinels of environmental contamination. Ultimately, the Salmonella prevalence rates in wild aquatic turtles, especially those strains shared with humans, highlight a significant public health concern.

Site Fidelity is Associated with Food Provisioning and Salmonella in an Urban Wading Bird.

Food provisioning can change wildlife pathogen dynamics by altering host susceptibility via nutrition and/or through shifts in foraging behavior and space use. We used the American white ibis (Eudocimus albus), a wading bird increasingly observed in urban parks, as a model to study synergistic relationships between food provisioning and infection risk across an urban gradient in South Florida. We tested whether Salmonella prevalence was associated with changes in ibis diet (stable isotope analysis), space use (site fidelity via GPS tracking), and local density (flock size). We compared the relative importance of these mechanisms by ranking candidate models using logistic regression. We detected Salmonella in 27% of white ibises (n = 233) sampled at 15 sites. Ibises with diets higher in anthropogenic food exhibited higher site fidelity. Salmonella prevalence was higher at sites where ibises exhibited greater site fidelity and Salmonella was more prevalent in soil and water. Overlap in Salmonella serotypes between ibises and soil or water also was more likely at sites where ibises exhibited higher site fidelity. Our results suggest that repeated use of foraging areas may increase Salmonella exposure for birds if foraging areas are contaminated from animal feces, human waste, or other bacterial sources. Limiting wildlife feeding in parks-perhaps best achieved through understanding the motivations for feeding, education, and enforcement-may reduce health risks for wildlife and the public.

Demographic and Pathogens of Domestic, Free-Roaming Pets and the Implications for Wild Carnivores and Human Health in the San Luis Region of Costa Rica.

Habitat loss and degradation, restricted ranges, prey exploitation, and poaching are important factors for the decline of several wild carnivore populations and additional stress from infectious agents is an increasing concern. Given the rapid growth of human populations in some regions like Costa Rica, pathogens introduced, sustained, and transmitted by domestic carnivores may be particularly important. To better understand the significance of domestic carnivore pathogens for wildlife, we determine the prevalence of infection and possible mechanisms for contact between the two groups. The demographics, role in the household, and pathogens of pet dogs and cats was studied during three annual spay/neuter clinics in San Luis, Costa Rica. Most dogs were owned primarily as pets and guard animals, but ~10% were used for hunting. Cats were owned primarily as pets and for pest control. Both roamed freely outdoors. We detected high prevalences of some pathogens (e.g., carnivore protoparvovirus 1 and Toxoplasma gondii). Some pathogens are known to persist in the environment, which increases the probability of exposure to wild carnivores. This study demonstrated that domestic pets in San Luis, home to a number of protected and endangered wildlife species, are infected with pathogens to which these wild species are potentially susceptible. Additionally, results from our questionnaire support the potential for domestic and wild animal contact, which may result in disease spillover.

Surveillance and Genetic Characterization of Virulent Newcastle Disease Virus Subgenotype V.3 in Indigenous Chickens from Backyard Poultry Farms and Live Bird Markets in Kenya.

Kenyan poultry consists of ~80% free-range indigenous chickens kept in small flocks (~30 birds) on backyard poultry farms (BPFs) and they are traded via live bird markets (LBMs). Newcastle disease virus (NDV) was detected in samples collected from chickens, wild farm birds, and other domestic poultry species during a 2017-2018 survey conducted at 66 BPFs and 21 LBMs in nine Kenyan counties. NDV nucleic acids were detected by rRT-PCR L-test in 39.5% (641/1621) of 1621 analyzed samples, of which 9.67% (62/641) were NDV-positive by both the L-test and a fusion-test designed to identify the virulent virus, with a majority being at LBMs (64.5%; 40/62) compared to BPFs (25.5%; 22/62). Virus isolation and next-generation sequencing (NGS) on a subset of samples resulted in 32 complete NDV genome sequences with 95.8-100% nucleotide identities amongst themselves and 95.7-98.2% identity with other east African isolates from 2010-2016. These isolates were classified as a new sub-genotype, V.3, and shared 86.5-88.9% and 88.5-91.8% nucleotide identities with subgenotypes V.1 and V.2 viruses, respectively. The putative fusion protein cleavage site (113R-Q-K-R↓F 117) in all 32 isolates, and a 1.86 ICPI score of an isolate from a BPF chicken that had clinical signs consistent with Newcastle disease, confirmed the high virulence of the NDVs. Compared to genotypes V and VI viruses, the attachment (HN) protein of 18 of the 32 vNDVs had amino acid substitutions in the antigenic sites. A time-scaled phylogeographic analysis suggests a west-to-east dispersal of the NDVs via the live chicken trade, but the virus origins remain unconfirmed due to scarcity of continuous and systematic surveillance data. This study reveals the widespread prevalence of vNDVs in Kenyan backyard poultry, the central role of LBMs in the dispersal and possibly generation of new virus variants, and the need for robust molecular epidemiological surveillance in poultry and non-poultry avian species.

Experimental Susceptibility of North American Raccoons (Procyon lotor) and Striped Skunks (Mephitis mephitis) to SARS-CoV-2.

Recent spillback events of SARS-CoV-2 from humans to animals has raised concerns about it becoming endemic in wildlife. A sylvatic cycle of SARS-CoV-2 could present multiple opportunities for repeated spillback into human populations and other susceptible wildlife. Based on their taxonomy and natural history, two native North American wildlife species -the striped skunk (Mephitis mephitis) and the raccoon (Procyon lotor) -represent a high likelihood of susceptibility and ecological opportunity of becoming infected with SARS-CoV-2. Eight skunks and raccoons were each intranasally inoculated with one of two doses of the virus (103 PFU and 105 PFU) and housed in pairs. To evaluate direct transmission, a naïve animal was added to each inoculated pair 48 h post-inoculation. Four control animals of each species were handled like the experimental groups. At predetermined intervals, we collected nasal and rectal swabs to quantify virus shed via virus isolation and detect viral RNA via rRT-PCR and blood for serum neutralization. Lastly, animals were euthanized at staggered intervals to describe disease progression through histopathology and immunohistochemistry. No animals developed clinical disease. All intranasally inoculated animals seroconverted, suggesting both species are susceptible to SARS-CoV-2 infection. The highest titers in skunks and raccoons were 1:128 and 1:64, respectively. Low quantities of virus were isolated from 2/8 inoculated skunks for up to day 5 post-inoculation, however no virus was isolated from inoculated raccoons or direct contacts of either species. Neither species had gross lesions, but recovering mild chronic pneumonia consistent with viral insult was recorded histologically in 5/8 inoculated skunks. Unlike another SARS-CoV-2 infection trial in these species, we detected neutralizing antibodies in inoculated raccoons; thus, future wildlife serologic surveillance results must be interpreted with caution. Due to the inability to isolate virus from raccoons, the lack of evidence of direct transmission between both species, and low amount of virus shed by skunks, it seems unlikely for SARS-CoV-2 to become established in raccoon and skunk populations and for virus to spillback into humans. Continued outbreaks in non-domestic species, wild and captive, highlight that additional research on the susceptibility of SARS-CoV-2 in wildlife, especially musteloidea, and of conservation concern, is needed.

Urban specialization reduces habitat connectivity by a highly mobile wading bird.

BACKGROUND: Mobile animals transport nutrients and propagules across habitats, and are crucial for the functioning of food webs and for ecosystem services. Human activities such as urbanization can alter animal movement behavior, including site fidelity and resource use. Because many urban areas are adjacent to natural sites, mobile animals might connect natural and urban habitats. More generally, understanding animal movement patterns in urban areas can help predict how urban expansion will affect the roles of highly mobile animals in ecological processes. METHODS: Here, we examined movements by a seasonally nomadic wading bird, the American white ibis (Eudocimus albus), in South Florida, USA. White ibis are colonial wading birds that forage on aquatic prey; in recent years, some ibis have shifted their behavior to forage in urban parks, where they are fed by people. We used a spatial network approach to investigate how individual movement patterns influence connectivity between urban and non-urban sites. We built a network of habitat connectivity using GPS tracking data from ibis during their non-breeding season and compared this network to simulated networks that assumed individuals moved indiscriminately with respect to habitat type. RESULTS: We found that the observed network was less connected than the simulated networks, that urban-urban and natural-natural connections were strong, and that individuals using urban sites had the least-variable habitat use. Importantly, the few ibis that used both urban and natural habitats contributed the most to connectivity. CONCLUSIONS: Habitat specialization in urban-acclimated wildlife could reduce the exchange of propagules and nutrients between urban and natural areas, which has consequences both for beneficial effects of connectivity such as gene flow and for detrimental effects such as the spread of contaminants or pathogens.

A Review of Pathogen Transmission at the Backyard Chicken-Wild Bird Interface.

Habitat conversion and the expansion of domesticated, invasive species into native habitats are increasingly recognized as drivers of pathogen emergence at the agricultural-wildlife interface. Poultry agriculture is one of the largest subsets of this interface, and pathogen spillover events between backyard chickens and wild birds are becoming more commonly reported. Native wild bird species are under numerous anthropogenic pressures, but the risks of pathogen spillover from domestic chickens have been historically underappreciated as a threat to wild birds. Now that the backyard chicken industry is one of the fastest growing industries in the world, it is imperative that the principles of biosecurity, specifically bioexclusion and biocontainment, are legislated and implemented. We reviewed the literature on spillover events of pathogens historically associated with poultry into wild birds. We also reviewed the reasons for biosecurity failures in backyard flocks that lead to those spillover events and provide recommendations for current and future backyard flock owners.

Effects of an anthropogenic diet on indicators of physiological challenge and immunity of white ibis nestlings raised in captivity.

When wildlife forage and/or live in urban habitats, they often experience a shift in resource availability and dietary quality. Some species even use human handouts, such as bread, as well as human refuse, as a large part of their new diets; yet the influences of this nutritional shift on health and survival remain unclear. American white ibises are increasingly being seen in urban areas in Florida; they collect handouts, such as bread and other food items, from humans in parks, and are also found foraging on anthropogenic sources in trash heaps. We hypothesized that the consumption of these new anthropogenic food sources may trigger increases in indicators of physiological challenge and dampen immune responses. We tested this experimentally by raising 20 white ibis nestlings in captivity, and exposing 10 to a simulated anthropogenic diet (including the addition of white bread and a reduction in seafood content) while maintaining 10 on a diet similar to what ibises consume in more natural environments. We then tested two indicators of physiological challenge (corticosterone and heat shock protein 70), assessed innate immunity in these birds via bactericidal assays and an in vitro carbon clearance assay, and adaptive immunity using a phytohemagglutinin skin test. The anthropogenic diet depressed the development of the ability to kill Salmonella paratyphi in culture. Our results suggest that consuming an anthropogenic diet may be detrimental in terms of the ability to battle a pathogenic bacterial species, but there was little effect on indicators of physiological challenge and other immunological measures.

Gut microbiome shifts with urbanization and potentially facilitates a zoonotic pathogen in a wading bird.

Microbial communities in the gastrointestinal tract influence many aspects of host health, including metabolism and susceptibility to pathogen colonization. These relationships and the environmental and individual factors that drive them are relatively unexplored for free-living wildlife. We quantified the relationships between urban habitat use, diet, and age with microbiome composition and diversity for 82 American white ibises (Eudocimus albus) captured along an urban gradient in south Florida and tested whether gut microbial diversity was associated with Salmonella enterica prevalence. Shifts in community composition were significantly associated with urban land cover and, to a lesser extent, diets higher in provisioned food. The diversity of genera was negatively associated with community composition associated with urban land cover, positively associated with age class, and negatively associated with Salmonella shedding. Our results suggest that shifts in both habitat use and diet for urban birds significantly alter gut microbial composition and diversity in ways that may influence health and pathogen susceptibility as species adapt to urban habitats.

The movements of a recently urbanized wading bird reveal changes in season timing and length related to resource use.

The American White Ibis (Eudocimus albus) is a nomadic wading bird that is increasing the amount of time spent foraging in urban areas, relying on artificial wetlands and other anthropogenic resources year-round. In this study, we explore whether and how American White Ibis association with urban environments is predictive of variation in the timing and length of behavioral seasons. Other urbanized species exhibit altered annual cycles such as loss of migratory behavior and year-round breeding related to consistent resource abundance, often related to intentional and unintentional provisioning. To determine if these same patterns of behavior were also present in White Ibis, we used behavioral change point analysis to segment the tracks of 41 ibis equipped with GPS backpacks to identify the initiation and duration of four behavioral seasons (non-breeding, pre-breeding, breeding, post-breeding) the degree of urban association. We found that intraspecific variation in urban habitat use had strong carryover effects on the timing and duration of behavioral seasons. This study revealed ibis with higher use of urban habitats in non-breeding seasons had longer non-breeding seasons and shorter breeding seasons that began earlier in the year compared to ibis that primarily use wetland habitats. The timing and duration of seasons also varied with ibis age, such that ibis spent more time engaged in breeding-related seasons as they aged. Juvenile and subadult ibis, though considered to be reproductively immature, also exhibit behavioral shifts in relation to breeding seasons. The behavioral patterns found in this study provide evidence that ibis are adapting their annual cycles and seasonal behaviors to exploit urban resources. Future research is needed to identify the effect of interactions between ibis urban association and age on behavioral season expression.

Epidemiological Investigation of a Mortality Event in a Translocated Gopher Tortoise (Gopherus polyphemus) Population in Northwest Florida.

Nokuse Plantation, a 22,055 ha private conservation preserve in northwest Florida, is a recipient site for gopher tortoises translocated from development sites in Florida. Since 2006, Nokuse has received over 5,000 tortoises from multiple development sites. During 2013-2015, 52 tortoises were found sick (n = 14) or dead (n = 38) in multiple soft-release enclosures in which tortoises consistently exhibited clinical signs, with additional sick (n = 5) and dead (n = 5) tortoises presenting similarly during 2016-2017. When found alive, tortoises behaved abnormally (e.g., frequently out of burrows during cold weather, pacing along enclosure fencing), appeared emaciated, were lethargic, and had developed redness under plastron scutes. Similar numbers of male (n = 28) and female (n = 32) tortoises were recovered along with two of unidentified sex, including mainly adults (n = 59) and three subadults. Physical examination, blood analysis, and other diagnostics were indicative of starvation and dehydration. Most sick tortoises provided with supportive care recovered. Necropsy findings generally confirmed starvation, with no evidence of infectious pathogens or contaminants. There were no apparent differences in quality of habitat, plant community, or soil or water among affected and unaffected enclosures. Botanical surveys indicated adequate forage quality and quantity, with no poisonous exotic or native plants detected. No land management practices changed prior to this event. Analysis of epidemiological data and demographic factors from before and during this mortality event identified initial density of tortoises in the enclosures as exerting the strongest influence on detection of tortoise morbidity and mortality. We believe that the stress associated with mixing tortoises from different populations and at higher densities during translocation impacted an individual tortoise's ability to obtain or absorb adequate nutrients from foraging, ultimately leading to a wasting condition consistent with starvation. Based on our findings, we recommend a maximum of 3 gopher tortoises per ha in soft-release enclosures for translocation, but further research is warranted to investigate the complexity of stress and social pressures associated with translocation.

Pathogen Surveillance and Detection of Ranavirus (Frog virus 3) in Translocated Gopher Tortoises (Gopherus polyphemus).

Emerging pathogens may pose additional threats to already vulnerable populations of chelonians, such as gopher tortoises (Gopherus polyphemus). In response to a mortality event on a translocation site in northwest Florida, US during 2013-15, 13 gopher tortoises were necropsied and their tissues were screened for 12 pathogens, including Mycoplasma agassizii, Mycoplasma testudineum, and Frog virus 3-like ranavirus (FV3). The DNA of FV3 was detected via quantitative PCR in the gastrointestinal tract of three tortoises. Subsequently, pathogen surveillance was performed on whole blood and oral-cloacal swab samples of live translocated tortoises from two different enclosures within the site (n=68), rehabilitated tortoises from the site (n=18), and tortoises prior to release on site (n=35) during 2015-17. Mycoplasma spp. were present in all groups and years of live tortoises tested. The DNA of FV3 was detected in 15 individuals both with and without clinical signs of disease in 2016. We recaptured 20 tortoises and captured an additional 20 tortoises in 2017 for surveillance, yet FV3 DNA was no longer detected, even in those that had previously tested positive (n=7). The results of this study contribute to the epidemiology of ranavirus in chelonians and suggests that gopher tortoises could be reservoirs for FV3. We recommend that the status of Ranavirus infection should be included for health screens for gopher tortoises in translocation programs.

The effect of diet on the gastrointestinal microbiome of juvenile rehabilitating green turtles (Chelonia mydas).

Threatened and endangered green turtles (Chelonia mydas) are unique because as juveniles they recruit from pelagic to near-shore waters and shift from an omnivorous to primarily herbivorous diet (i.e. seagrass and algae). Nevertheless, when injured and ill animals are admitted to rehabilitation, animal protein (e.g. seafood) is often offered to combat poor appetite and emaciation. We examined how the fecal microbiome of juvenile green turtles changed in response to a dietary shift during rehabilitation. We collected fecal samples from January 2014 -January 2016 from turtles (N = 17) in rehabilitation at the Georgia Sea Turtle Center and used next generation sequencing to analyze bacterial community composition. Samples were collected at admission, mid-rehabilitation, and recovery, which entailed a shift from a mixed seafood-vegetable diet at admission to a primarily herbivorous diet at recovery. The dominant phyla changed over time, from primarily Firmicutes (55.0%) with less Bacteroidetes (11.4%) at admission, to primarily Bacteroidetes (38.4%) and less Firmicutes (31.8%) at recovery. While the microbiome likely shifts with the changing health status of individuals, this consistent inversion of Bacteroidetes and Firmicutes among individuals likely reflects the increased need for protein digestion, for which Bacteroidetes are important. Firmicutes are significant in metabolizing plant polysaccharides; thus, fewer Firmicutes may result in underutilization of wild diet items in released individuals. This study demonstrates the importance of transitioning rehabilitating green turtles to an herbivorous diet as soon as possible to afford them the best probability of survival.