Scott Carver

Emerging infectious diseases of wildlife: a critical perspective

We review the literature to distinguish reports of vertebrate wildlife disease emergence with sufficient evidence, enabling a robust assessment of emergence drivers. For potentially emerging agents that cannot be confirmed, sufficient data on prior absence (or a prior difference in disease dynamics) are frequently lacking. Improved surveillance, particularly for neglected host taxa, geographical regions and infectious agents, would enable more effective management should emergence occur. Exposure to domestic sources of infection and human-assisted exposure to wild sources were identified as the two main drivers of emergence across host taxa; the domestic source was primary for fish while the wild source was primary for other taxa. There was generally insufficient evidence for major roles of other hypothesized drivers of emergence.

Three Pathogens in Sympatric Populations of Pumas, Bobcats, and Domestic Cats: Implications for Infectious Disease Transmission

Anthropogenic landscape change can lead to increased opportunities for pathogen transmission between domestic and non-domestic animals. Pumas, bobcats, and domestic cats are sympatric in many areas of North America and share many of the same pathogens, some of which are zoonotic. We analyzed bobcat, puma, and feral domestic cat samples collected from targeted geographic areas. We examined exposure to three pathogens that are taxonomically diverse (bacterial, protozoal, viral), that incorporate multiple transmission strategies (vector-borne, environmental exposure/ingestion, and direct contact), and that vary in species-specificity. Bartonella spp., Feline Immunodeficiency Virus (FIV), and Toxoplasma gondii IgG were detected in all three species with mean respective prevalence as follows: puma 16%, 41% and 75%; bobcat 31%, 22% and 43%; domestic cat 45%, 10% and 1%. Bartonella spp. were highly prevalent among domestic cats in Southern California compared to other cohort groups. Feline Immunodeficiency Virus exposure was primarily associated with species and age, and was not influenced by geographic location. Pumas were more likely to be infected with FIV than bobcats, with domestic cats having the lowest infection rate. Toxoplasma gondii seroprevalence was high in both pumas and bobcats across all sites; in contrast, few domestic cats were seropositive, despite the fact that feral, free ranging domestic cats were targeted in this study. Interestingly, a directly transmitted species-specific disease (FIV) was not associated with geographic location, while exposure to indirectly transmitted diseases – vector-borne for Bartonella spp. and ingestion of oocysts via infected prey or environmental exposure for T. gondii – varied significantly by site. Pathogens transmitted by direct contact may be more dependent upon individual behaviors and intra-specific encounters. Future studies will integrate host density, as well as landscape features, to better understand the mechanisms driving disease exposure and to predict zones of cross-species pathogen transmission among wild and domestic felids.

Detection of Chronic Wasting Disease Prions in Salivary, Urinary, and Intestinal Tissues of Deer: Potential Mechanisms of Prion Shedding and Transmission

ABSTRACT Efficient horizontal transmission is a signature trait of chronic wasting disease (CWD) in cervids. Infectious prions shed into excreta appear to play a key role in this facile transmission, as has been demonstrated by bioassays of cervid and transgenic species and serial protein misfolding cyclic amplification (sPMCA). However, the source(s) of infectious prions in these body fluids has yet to be identified. In the present study, we analyzed tissues proximate to saliva, urine, and fecal production by sPMCA in an attempt to elucidate this unique aspect of CWD pathogenesis. Oropharyngeal, urogenital, and gastrointestinal tissues along with blood and obex from CWD-exposed cervids (comprising 27 animals and >350 individual samples) were analyzed and scored based on the apparent relative CWD burden. PrPCWD-generating activity was detected in a range of tissues and was highest in the salivary gland, urinary bladder, and distal intestinal tract. In the same assays, blood from the same animals and unseeded normal brain homogenate controls (n = 116 of 117) remained negative. The PrP-converting activity in peripheral tissues varied from 10−11- to 100-fold of that found in brain of the same animal. Deer with highest levels of PrPCWD amplification in the brain had higher and more widely disseminated prion amplification in excretory tissues. Interestingly, PrPCWD was not demonstrable in these excretory tissues by conventional Western blotting, suggesting a low prion burden or the presence of protease-sensitive infectious prions destroyed by harsh proteolytic treatments. These findings offer unique insights into the transmission of CWD in particular and prion infection and trafficking overall.

Dryland Salinity and Ecosystem Distress Syndrome: Human Health Implications

Clearing of native vegetation for agriculture has left 1.047 million hectares of southwest Western Australia affected by dryland salinity, and this area may expand up to a further 1.7–3.4 million hectares if trends continue. Ecosystems in saline-affected regions display many of the classic characteristics of Ecosystem Distress Syndrome, one outcome of which has not yet been investigated in relation to dryland salinity: adverse human health implications. This article seeks to review existing information and identify potential adverse human health effects. Three key potential impacts on human health resulting from dryland salinity are identified: wind-borne dust and respiratory health; altered ecology of the mosquito-borne disease Ross River virus; and mental health consequences of salinity-induced environmental degradation. Given the predicted increase in extent and severity of dryland salinity over coming decades, adverse outcomes of salinity are likely to be further exacerbated, including those related to human health. There is a clear need to investigate the issues discussed in this review and also to identify other potential adverse health effects of dryland salinity. Investigations must be multidisciplinary to sufficiently examine the broad scope of these issues. The relationship between human health and salinity may also be relevant beyond Australia in other countries where secondary soil salinization is occurring.

Influence of hosts on the ecology of arboviral transmission: Potential mechanisms influencing dengue, Murray Valley encephalitis, and Ross River virus in Australia

Ecological interactions are fundamental to the transmission of infectious disease. Arboviruses are particularly elegant examples, where rich arrays of mechanisms influence transmission between vectors and hosts. Research on host contributions to the ecology of arboviral diseases has been undertaken within multiple subdisciplines, but significant gaps in knowledge remain and multidisciplinary approaches are needed. Through our multidisciplinary review of the literature we have identified five broad areas where hosts may influence the ecology of arboviral transmission: host immunity; cross-protective immunity and antibody-dependent enhancement; host abundance; host diversity; and pathogen spillover and dispersal. Herein we discuss the known and theoretical roles of hosts within these topics and then apply this knowledge to three epidemiologically important mosquito-borne arboviruses that occur in Australia: dengue virus (DENV), Murray Valley encephalitis virus (MVEV), and Ross River virus (RRV). We argue that the underlying mechanisms by which hosts influence arboviral activity are numerous and attempts to delineate these mechanisms further are needed. Investigations that focus on hosts of vector-borne diseases are likely to be rewarding, particularly where the ecology of vectors is relatively well understood. From an applied perspective, enhanced knowledge of host influences upon vector-borne disease transmission is likely to enable better management of disease burden. Finally, we suggest a framework that may be useful to identify and determine host contributions to the ecology of arboviruses.

Novel gammaherpesviruses in North American domestic cats, bobcats and pumas: identification, prevalence and risk factors

ABSTRACT Gammaherpesviruses (GHVs) are a diverse and rapidly expanding group of viruses associated with a variety of disease conditions in humans and animals. To identify felid GHVs, we screened domestic cat (Felis catus), bobcat (Lynx rufus), and puma (Puma concolor) blood cell DNA samples from California, Colorado, and Florida using a degenerate pan-GHV PCR. Additional pan-GHV and long-distance PCRs were used to sequence a contiguous 3.4-kb region of each putative virus species, including partial glycoprotein B and DNA polymerase genes. We identified three novel GHVs, each present predominantly in one felid species: Felis catus GHV 1 (FcaGHV1) in domestic cats, Lynx rufus GHV 1 (LruGHV1) in bobcats, and Puma concolor GHV 1 (PcoGHV1) in pumas. To estimate infection prevalence, we developed real-time quantitative PCR assays for each virus and screened additional DNA samples from all three species (n = 282). FcaGHV1 was detected in 16% of domestic cats across all study sites. LruGHV1 was detected in 47% of bobcats and 13% of pumas across all study sites, suggesting relatively common interspecific transmission. PcoGHV1 was detected in 6% of pumas, all from a specific region of Southern California. The risk of infection for each host varied with geographic location. Age was a positive risk factor for bobcat LruGHV1 infection, and age and being male were risk factors for domestic cat FcaGHV1 infection. Further characterization of these viruses may have significant health implications for domestic cats and may aid studies of free-ranging felid ecology. IMPORTANCE Gammaherpesviruses (GHVs) establish lifelong infection in many animal species and can cause cancer and other diseases in humans and animals. In this study, we identified the DNA sequences of three GHVs present in the blood of domestic cats (Felis catus), bobcats (Lynx rufus), and pumas (Puma concolor; also known as mountain lions, cougars, and panthers). We found that these viruses were closely related to, but distinct from, other known GHVs of animals and represent the first GHVs identified to be native to these feline species. We developed techniques to rapidly and specifically detect the DNA of these viruses in feline blood and found that the domestic cat and bobcat viruses were widespread across the United States. In contrast, puma virus was found only in a specific region of Southern California. Surprisingly, the bobcat virus was also detected in some pumas, suggesting relatively common virus transmission between these species. Adult domestic cats and bobcats were at greater risk for infection than juveniles. Male domestic cats were at greater risk for infection than females. This study identifies three new viruses that are widespread in three feline species, indicates risk factors for infection that may relate to the route of infection, and demonstrates cross-species transmission between bobcats and pumas. These newly identified viruses may have important effects on feline health and ecology.

Does Chytridiomycosis Disrupt Amphibian Skin Function

Abstract Chytridiomycosis, a disease caused by the fungus Batrachochytrium dendrobatidis (Bd), potentially disrupts osmoregulation or respiration across the skin of amphibians it infects, releases toxins into the host, or both. We investigated whether infection with Bd alters water balance or metabolic rate of the hylid frog Litoria raniformis. Frogs were held in laboratory conditions simulating those in which Bd epizootics had been observed in the field. We inoculated six frogs with infective Bd zoospores, held the subjects in individual containers, and compared their course of infection and associated physiological measures with those of six controls. Experimental subjects exhibited clinical signs of chytridiomycosis during the early period of infection, one week after they were inoculated, possibly due to invasion of Bd into the skin. These clinical signs were accompanied by significant inhibition of rehydration through the skin. However, we detected no changes in metabolic rate attributable to chytridiomycosis after one week. Five months after inoculation, all but one of the infected subjects had survived. Molecular testing confirmed that surviving frogs, although aclinical, still were infected. Control and infected subjects showed no difference in water balance or metabolism. These results provide evidence of inhibited rehydration in individuals exhibiting clinical signs of chytridiomycosis. However, aclinical chytridiomycosis does not severely affect amphibian skin function. Frogs that survive infection by Bd, even if they remain infected, may suffer no significant impairment in their physiological responses. The disease progression, with initial clinical signs of chytridiomycosis followed by apparent full recovery, is consistent with an adaptive immune response to Bd infection. Further research is needed to determine how Bd causes clinical chytridiomycosis and the immunological mechanisms by which hosts respond to Bd.

Prion-seeding activity in cerebrospinal fluid of deer with chronic wasting disease.

Transmissible spongiform encephalopathies (TSEs), or prion diseases, are a uniformly fatal family of neurodegenerative diseases in mammals that includes chronic wasting disease (CWD) of cervids. The early and ante-mortem identification of TSE-infected individuals using conventional western blotting or immunohistochemistry (IHC) has proven difficult, as the levels of infectious prions in readily obtainable samples, including blood and bodily fluids, are typically beyond the limits of detection. The development of amplification-based seeding assays has been instrumental in the detection of low levels of infectious prions in clinical samples. In the present study, we evaluated the cerebrospinal fluid (CSF) of CWD-exposed (n=44) and naïve (n=4) deer (n=48 total) for CWD prions (PrPd) using two amplification assays: serial protein misfolding cyclic amplification with polytetrafluoroethylene beads (sPMCAb) and real-time quaking induced conversion (RT-QuIC) employing a truncated Syrian hamster recombinant protein substrate. Samples were evaluated blindly in parallel with appropriate positive and negative controls. Results from amplification assays were compared to one another and to obex immunohistochemistry, and were correlated to available clinical histories including CWD inoculum source (e.g. saliva, blood), genotype, survival period, and duration of clinical signs. We found that both sPMCAb and RT-QuIC were capable of amplifying CWD prions from cervid CSF, and results correlated well with one another. Prion seeding activity in either assay was observed in approximately 50% of deer with PrPd detected by IHC in the obex region of the brain. Important predictors of amplification included duration of clinical signs and time of first tonsil biopsy positive results, and ultimately the levels of PrPd identified in the obex by IHC. Based on our findings, we expect that both sPMCAb and RT-QuIC may prove to be useful detection assays for the detection of prions in CSF.

Felis catus gammaherpesvirus 1; a widely endemic potential pathogen of domestic cats.

Felis catus gammaherpesvirus 1 (FcaGHV1), recently discovered in the USA, was detected in domestic cats in Australia (11.4%, 95% confidence interval 5.9-19.1, n=110) and Singapore (9.6%, 95% confidence interval 5.9-14.6, n=176) using qPCR. FcaGHV1 qPCR positive cats were 2.8 times more likely to be sick than healthy. Risk factors for FcaGHV1 detection included being male, increasing age and coinfection with pathogenic retroviruses, feline immunodeficiency virus (FIV) or feline leukaemia virus. FcaGHV1 DNA was detected in multiple tissues from infected cats with consistently high virus loads in the small intestine. FcaGHV1 viral load was significantly higher in FIV-infected cats compared with matched controls, mimicking increased Epstein-Barr virus loads in human immunodeficiency virus-infected humans. FcaGHV1 is endemic in distant geographic regions and is associated with being sick and with coinfections. Horizontal transmission of FcaGHV1 is supported, with biting being a plausible route. A pathogenic role for FcaGHV1 in domestic cats is supported.

Detection of Chronic Wasting Disease in the Lymph Nodes of Free-Ranging Cervids by Real-Time Quaking-Induced Conversion

ABSTRACT Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of deer, elk, and moose, is the only prion disease affecting free-ranging animals. Since the disease was first identified in northern Colorado and southern Wyoming in 1967, new epidemic foci of the disease have been identified in 20 additional states, as well as two Canadian provinces and the Republic of South Korea. Identification of CWD-affected animals currently requires postmortem analysis of brain or lymphoid tissues using immunohistochemistry (IHC) or an enzyme-linked immunosorbent assay (ELISA), with no practical way to evaluate potential strain types or to investigate the epidemiology of existing or novel foci of disease. Using a standardized real-time (RT)-quaking-induced conversion (QuIC) assay, a seeded amplification assay employing recombinant prion protein as a conversion substrate and thioflavin T (ThT) as an amyloid-binding fluorophore, we analyzed, in a blinded manner, 1,243 retropharyngeal lymph node samples from white-tailed deer, mule deer, and moose, collected in the field from areas with current or historic CWD endemicity. RT-QuIC results were then compared with those obtained by conventional IHC and ELISA, and amplification metrics using ThT and thioflavin S were examined in relation to the clinical history of the sampled deer. The results indicate that RT-QuIC is useful for both identifying CWD-infected animals and facilitating epidemiological studies in areas in which CWD is endemic or not endemic.