Optimisation and validation of a PCR to detect viable Tenacibaculum maritimum in salmon skin tissue samples

A PCR protocol was optimised and validated for the detection of viable Tenacibaculum maritimum cells in salmon skin tissue. Viability conventional (vPCR) and quantitative PCR (v-qPCR) assays both had a limit of detection of 103 CFU mL−1 viable cells. The v-qPCR assay showed a linear quantification over 4 log units. Conventional vPCR showed complete signal suppression when only dead cells were present at concentrations lower than 106 CFU mL−1. While the v-qPCR did not result in complete suppression when only dead cells were present, a method was developed to determine if viable cells were present based on the % Δ in cycle threshold (Ct) value. The procedure was validated for high-throughput processing and an enrichment protocol was validated to reliably detect low concentrations of viable cells both with and without a high background of dead cells. Performing this protocol on naturally infected tissues showed that vPCR and v-qPCR reduced the potential for false positives compared to using conventional PCR and qPCR. The optimised protocol developed for this study provides an efficient, reliable and robust alternative for the detection of viable T. maritimum in skin tissue

Early-onset progressive retinal atrophy associated with an IQCB1 variant in African black-footed cats (Felis nigripes)

African black-footed cats (Felis nigripes) are endangered wild felids. One male and full-sibling female African black-footed cat developed vision deficits and mydriasis as early as 3 months of age. The diagnosis of early-onset progressive retinal atrophy (PRA) was supported by reduced direct and consensual pupillary light reflexes, phenotypic presence of retinal degeneration, and a non-recordable electroretinogram with negligible amplitudes in both eyes. Whole genome sequencing, conducted on two unaffected parents and one affected offspring was compared to a variant database from 51 domestic cats and a Pallas cat, revealed 50 candidate variants that segregated concordantly with the PRA phenotype. Testing in additional affected cats confirmed that cats homozygous for a 2 base pair (bp) deletion within IQ calmodulin-binding motif-containing protein-1 (IQCB1), the gene that encodes for nephrocystin-5 (NPHP5), had vision loss. The variant segregated concordantly in other related individuals within the pedigree supporting the identification of a recessively inherited early-onset feline PRA. Analysis of the black-footed cat studbook suggests additional captive cats are at risk. Genetic testing for IQCB1 and avoidance of matings between carriers should be added to the species survival plan for captive management

Novel Viruses: Update on the significance of papillomavirus infections in cats

Practical relevance: Prior to 1990 papillomaviruses (PVs) were not recognised to infect or cause disease in domestic cats. Since this time, the use of histology, immunohistochemistry and, more recently, molecular techniques has revealed that PVs almost certainly cause feline viral plaques and Bowenoid in situ carcinomas, oral papillomas and feline sarcoids. In addition, there is increasing evidence that PVs play a significant role in the development of feline cutaneous squamous cell carcinomas, one of the most common skin cancers of cats. Recent studies have also revealed that most cats are asymptomatically infected with PVs. This raises a critical question that is currently unanswered: why do only a small proportion of infected cats develop disease? In the future it may be possible to prevent PV-induced diseases by using a vaccine to prevent PV infection. Alternatively, novel therapies may be developed that prevent PVs from causing clinical disease by stimulating the host immune response. Clinical challenges: A recognition of the skin diseases caused by PVs is important to more accurately predict disease progression. Unfortunately, there are currently no non-surgical treatments that have been proven to be beneficial in cats and clinical management of PV-induced skin disease in cats can be challenging. Global importance: PVs have a worldwide distribution and negatively impact feline health and welfare globally. Audience: This review is aimed at clinicians, especially those who regularly treat cats with skin disease. The review will also be useful to oncologists and researchers who have an interest in how cancer develops in cats. Evidence base: In producing this update the authors have drawn on recently published peer-reviewed literature

Effects of coral bleaching on the obligate coral-dwelling crab\ud Trapezia cymodoce

Corals are an essential and threatened habitat for a diverse range of reef-associated animals. Episodes of coral bleaching are predicted to increase in frequency and intensity over coming decades, yet the effects of coral-host bleaching on the associated animal communities remain poorly understood. The present study investigated the effects of host-colony bleaching on the obligate coral-dwelling crab, Trapezia cymodoce, during a natural bleaching event in the lagoon of Lizard Island, Australia. Branching corals, which harbour the highest diversity of coral associates, comprised 13% of live coral cover at the study site, with 83% affected by bleaching. Crabs on healthy and bleached colonies of Pocillopora damicornis were monitored over a 5-week period to determine whether coral bleaching affected crab density and movement patterns. All coral colonies initially contained one breeding pair of crabs. There was a significant decline in crab density on bleached corals after 5 weeks, with many corals losing one or both crabs, yet all healthy colonies retained a mating pair. Fecundity of crabs collected from bleached and healthy colonies of P. damicornis was also compared. The size of egg clutches of crabs collected from bleached hosts was 40% smaller than those from healthy hosts, indicating a significant reduction in fecundity. A laboratory experiment on movement patterns found that host-colony bleaching also prompted crabs to emigrate in search of more suitable colonies. Emigrant crabs engaged in aggressive interactions with occupants of healthy hosts, with larger crabs always usurping occupants of a smaller size. Decreased densities and clutch sizes, along with increased competitive interactions, could potentially result in a population decline of these important coral associates with cascading effects on coral health

Novel feline viruses: Emerging significance of gammaherpesvirus and morbillivirus infections

Practical relevance: New technologies capable of sequencing the genetic material in any given biological sample, combined with computer-based algorithms for sequence assembly and analysis, have revolutionised infectious disease research. The rate at which novel viruses are being discovered now exceeds our understanding of their clinical relevance. Novel viruses may contribute to diseases that are major causes of feline morbidity and mortality, including cancer and chronic kidney disease. The identification of new viral pathogens raises the prospect of not only improved patient outcomes through specific treatment but even disease prevention through viral control measures. Clinical challenges: It can be difficult to determine the role of a novel virus in disease development. Disease may be an occasional outcome, often years after infection. A high prevalence of infection in the general population can make disease associations harder to identify and almost impossible to rule out. Host cofactors such as immune dysfunction, genetic background or coinfections may be required for manifestation of disease, and one virus species may be linked to a range of pathological sequelae. Establishing causality relies on evaluating accumulating evidence from multiple investigations, which is often hard to access by practitioners. Global importance: The worldwide distribution of gammaherpesvirus and morbillivirus infections in domestic cats underlines the potential of these viruses to negatively impact feline health and welfare globally. Evidence base: This review relies on grade la—III evidence

Use of fluorescence in situ hybridization to detect Felis catus papillomavirus type 2 in feline Bowenoid in situ carcinomas

Objectives: Papillomaviruses (PVs) are ubiquitous host- and site-specific viruses. PV infections in cats are associated with oral papillomas, viral plaques, Bowenoid in situ carcinomas (BISCs), squamous cell carcinomas and sarcoids; this association is primarily based on PCR detection of PV DNA within said lesions. PV DNA is frequently detectable on normal feline skin; thus, it is possible that some of the implicated DNA is commensal rather than associated with lesion formation. Therefore, the aim of the present study was to use fluorescence in situ hybridization (FISH) to localize PV DNA within feline BISCs, to provide additional evidence that PV infection may influence the development of these neoplasms. Methods: FISH probes targeting Felis catus papillomavirus type 2 (FcaPV2) DNA were used to localize FcaPV2 DNA within 42 BISCs from which FcaPV2 DNA had previously been amplified via PCR. Results: Fifteen of 42 BISC lesions (35.7%) demonstrated intralesional FcaPV2 using FISH. Probe annealing was predominantly located within the nuclei of koilocytes found in the upper strata of the epidermis. Probes were typically scattered multifocally within the lesions; most commonly this was near the periphery of the BISCs. Conclusions and relevance: These results confirm that a proportion of BISCs contain FcaPV2 DNA. These results further support a causative association between FcaPV2 and BISCs in cats

Detection of papillomaviral DNA sequences in a feline oral squamous cell carcinoma

Oral squamous cell carcinomas (OSCCs) are common and often fatal feline neoplasms. Factors that predispose to neoplasm development in cats are poorly defined. Around 25% of human OSCCs are caused by papillomaviruses (PVs). To determine if PVs are associated with OSCCs in cats, three sets of consensus primers were used to evaluate 20 feline OSCCs and 20 non-neoplastic feline oral lesions for the presence of PV DNA. Papillomaviral sequences were detected within one OSCC, but no non-neoplastic lesion. Sequencing of the amplified DNA revealed a previously unreported PV that was most similar to human PV type 76. This is the first time PV DNA has been amplified from the oral cavity of a cat. However, while these results suggest that feline gingival epithelial cells can be infected by PVs, they do not support a causal association between viral infection and the development of feline OSCCs