SARS-CoV-2 neutralising antibodies in dogs and cats in the United Kingdom.

Companion animals are susceptible to SARS-CoV-2 infection and sporadic cases of pet infections have occurred in the United Kingdom. Here we present the first large-scale serological survey of SARS-CoV-2 neutralising antibodies in dogs and cats in the UK. Results are reported for 688 sera (454 canine, 234 feline) collected by a large veterinary diagnostic laboratory for routine haematology during three time periods; pre-COVID-19 (January 2020), during the first wave of UK human infections (April-May 2020) and during the second wave of UK human infections (September 2020-February 2021). Both pre-COVID-19 sera and those from the first wave tested negative. However, in sera collected during the second wave, 1.4% (n â€‹= â€‹4) of dogs and 2.2% (n â€‹= â€‹2) of cats tested positive for neutralising antibodies. The low numbers of animals testing positive suggests pet animals are unlikely to be a major reservoir for human infection in the UK. However, continued surveillance of in-contact susceptible animals should be performed as part of ongoing population health surveillance initiatives.

SARS-CoV-2 neutralising antibodies in Dogs and Cats in the United Kingdom.

Companion animals are susceptible to SARS-CoV-2 infection and sporadic cases of pet infections have occurred in the United Kingdom. Here we present the first large-scale serological survey of SARS-CoV-2 neutralising antibodies in dogs and cats in the UK. Results are reported for 688 sera (454 canine, 234 feline) collected by a large veterinary diagnostic laboratory for routine haematology during three time periods; pre-COVID-19 (January 2020), during the first wave of UK human infections (April-May 2020) and during the second wave of UK human infections (September 2020-February 2021). Both pre-COVID-19 sera and those from the first wave tested negative. However, in sera collected during the second wave, 1.4% (n=4) of dogs and 2.2% (n=2) cats tested positive for neutralising antibodies. The low numbers of animals testing positive suggests pet animals are unlikely to be a major reservoir for human infection in the UK. However, continued surveillance of in-contact susceptible animals should be performed as part of ongoing population health surveillance initiatives.

A virtual biobank for companion animals: A parvovirus pilot study.

BACKGROUND: There is a lack of national population data concerning infectious disease in companion animals. Here, we piloted the feasibility of linking diagnostic laboratories, population surveillance and modern sequencing approaches to extract targeted diagnostic samples from laboratories before they were discarded, as a novel route to better understand national epidemiology of major small animal pathogens. METHODS: Samples tested for canine or feline parvovirus were requested from a national veterinary diagnostic laboratory and analysed by Sanger or next generation sequencing. Samples were linked to electronic health data held in the SAVSNET database. RESULTS: Sequences obtained from positive samples, together with associated metadata, provided new insights into the recent geographical distribution of parvovirus strains in circulation in the United Kingdom (UK). CONCLUSIONS: This collaboration with industry represents a 'National Virtual Biobank' that can rapidly be called on, to efficiently add new layers of epidemiological information of relevance to animal, and potentially human, population health.

SARS-CoV-2 Infections in Animals: Reservoirs for Reverse Zoonosis and Models for Study.

The recent SARS-CoV-2 pandemic has brought many questions over the origin of the virus, the threat it poses to animals both in the wild and captivity, and the risks of a permanent viral reservoir developing in animals. Animal experiments have shown that a variety of animals can become infected with the virus. While coronaviruses have been known to infect animals for decades, the true intermediate host of the virus has not been identified, with no cases of SARS-CoV-2 in wild animals. The screening of wild, farmed, and domesticated animals is necessary to help us understand the virus and its origins and prevent future outbreaks of both COVID-19 and other diseases. There is intriguing evidence that farmed mink infections (acquired from humans) have led to infection of other farm workers in turn, with a recent outbreak of a mink variant in humans in Denmark. A thorough examination of the current knowledge and evidence of the ability of SARS-CoV-2 to infect different animal species is therefore vital to evaluate the threat of animal to human transmission and reverse zoonosis.

Methods of Inactivation of SARS-CoV-2 for Downstream Biological Assays.

The scientific community has responded to the coronavirus disease 2019 (COVID-19) pandemic by rapidly undertaking research to find effective strategies to reduce the burden of this disease. Encouragingly, researchers from a diverse array of fields are collectively working towards this goal. Research with infectious severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is undertaken in high-containment laboratories; however, it is often desirable to work with samples at lower-containment levels. To facilitate the transfer of infectious samples from high-containment laboratories, we have tested methods commonly used to inactivate virus and prepare the sample for additional experiments. Incubation at 80°C, a range of detergents, Trizol reagents, and UV energies were successful at inactivating a high titer of SARS-CoV-2. Methanol and paraformaldehyde incubation of infected cells also inactivated the virus. These protocols can provide a framework for in-house inactivation of SARS-CoV-2 in other laboratories, ensuring the safe use of samples in lower-containment levels.

Methods of inactivation of SARS-CoV-2 for downstream biological assays.

The scientific community has responded to the COVID-19 pandemic by rapidly undertaking research to find effective strategies to reduce the burden of this disease. Encouragingly, researchers from a diverse array of fields are collectively working towards this goal. Research with infectious SARS-CoV-2 is undertaken in high containment laboratories, however, it is often desirable to work with samples at lower containment levels. To facilitate the transfer of infectious samples from high containment laboratories, we have tested methods commonly used to inactivate virus and prepare the sample for additional experiments. Incubation at 80°C, and a range of detergents and UV energies were successful at inactivating a high titre of SARS-CoV-2. These protocols can provide a framework for in house inactivation of SARS-CoV-2 in other laboratories, ensuring the safe use of samples in lower containment levels.

Temporally separated feline calicivirus isolates do not cluster phylogenetically and are similarly neutralised by high-titre vaccine strain FCV-F9 antisera in vitro.

OBJECTIVES: Feline calicivirus (FCV) is a highly variable and globally important feline pathogen for which vaccination has been the mainstay of control. Here, we test whether the continued use of FCV-F9, one of the most frequently used vaccine strains globally, is driving the emergence of vaccine-resistant viruses in the field. METHODS: This study made use of two representative panels of field isolates previously collected from cats visiting randomly selected veterinary practices across the UK as part of separate cross-sectional studies from 2001 and 2013/2014. Phylogenetic analysis and in vitro virus neutralisation tests were used to compare the genetic and antigenic relationships between these populations and FCV-F9. RESULTS: Phylogenetic analysis showed a typically radial distribution dominated by 52 distinct strains, with strains from both 2001 and 2013/2014 intermingled. The sequence for FCV-F9 appeared to be integral to this phylogeny and there were no significant differences in the genetic distances within each studied population (intra-population distances), or between them (inter-population distances), or between each population and FCV-F9. A 1 in 8 dilution neutralised 97% and 100% of the 2001 and 2013/14 isolates, respectively, and a 1 in 16 dilution neutralised 87% and 75% of isolates, respectively. There was no significant difference either in variance between the FCV-F9 neutralising titres for the two populations, or in the distribution of neutralisation titres across the two populations. CONCLUSIONS AND RELEVANCE: Although FCV is a highly variable virus, we found no evidence for a progressive divergence of field virus from vaccine strain FCV-F9, either phylogenetically or antigenically, with FCV-F9 antisera remaining broadly and equally cross-reactive to two geographically representative and temporally separated FCV populations. We suggest this may be because the immunodominant region of the FCV capsid responsible for neutralisation may have structural constraints preventing its longer term progressive antigenic evolution.

A multi-national European cross-sectional study of feline calicivirus epidemiology, diversity and vaccine cross-reactivity.

BACKGROUND: Feline calicivirus (FCV) is an important pathogen of cats for which vaccination is regularly practised. Long-term use of established vaccine antigens raises the theoretical possibility that field viruses could become resistant. This study aimed to assess the current ability of the FCV-F9 vaccine strain to neutralise a randomly collected contemporary panel of FCV field strains collected prospectively in six European countries. METHODS: Veterinary practices (64) were randomly selected from six countries (UK, Sweden, Netherlands, Germany, France and Italy). Oropharyngeal swabs were requested from 30 (UK) and 40 (other countries) cats attending each practice. Presence of FCV was determined by virus isolation, and risk factors for FCV shedding assessed by multivariable logistic regression. Phylogenetic analyses were used to describe the FCV population structure. In vitro virus neutralisation assays were performed to evaluate FCV-F9 cross-reactivity using plasma from four vaccinated cats. RESULTS: The overall prevalence of FCV was 9.2%. Risk factors positively associated with FCV shedding included multi-cat households, chronic gingivostomatitis, younger age, not being neutered, as well as residing in certain countries. Phylogenetic analysis showed extensive variability and no countrywide clusters. Despite being first isolated in the 1950s, FCV-F9 clustered with contemporary field isolates. Plasma raised to FCV-F9 neutralized 97% of tested isolates (titres 1:4 to 1:5792), with 26.5%, 35.7% and 50% of isolates being neutralized by 5, 10 and 20 antibody units respectively. CONCLUSIONS: This study represents the largest prospective analysis of FCV diversity and antigenic cross-reactivity at a European level. The scale and random nature of sampling used gives confidence that the FCV isolates used are broadly representative of FCVs that cats are exposed to in these countries. The in vitro neutralisation results suggest that antibodies raised to FCV-F9 remain broadly cross-reactive to contemporary FCV isolates across the European countries sampled.

Improving access to care for women veterans suffering from chronic pain and depression associated with trauma.

OBJECTIVE: Access to care has become a priority for the Veterans Administration (VA) health care system as a significant number of veterans enrolled in the VA health care system reside in rural areas. The feasibility and effects of a novel clinical intervention that combined group therapy and biofeedback training was evaluated on women veterans living in rural areas. METHODS: The study was conducted at selected community-based outpatient clinics (CBOCs) in Texas. Thirty four women veterans with chronic pain and comorbid depression and/or posttraumatic stress disorder (PTSD) were recruited. Five sessions of education/therapy were delivered via telemedicine in combination with daily home practice of a portable biofeedback device (Stress Eraser®, Helicor, New York, NY, USA). Participants responded to self-report questionnaires at baseline, at posttreatment, and at 6-week follow-up. Daily practice logs were also maintained by participants. RESULTS: The clinical protocol was acceptable, easy to administer, and associated with statistically significant decreases in self-reported pain unpleasantness, pain interference, depressive symptoms, PTSD symptoms, and sleep disturbance at posttreatment. Improvements were maintained at 6-week follow-up. Qualitative analyses indicated that many participants 1) wished to continue to meet as a support group in their respective CBOCs and 2) felt less isolated and more empowered to cope with their problems of daily living as a result of the treatment. CONCLUSIONS: It is feasible to provide treatment to women veterans living in rural areas by utilizing video-teleconferencing technology between larger VA medical centers and facilities at CBOCs in more rural settings. A controlled trial of the intervention is warranted.

A subset of mucosa-associated Escherichia coli isolates from patients with colon cancer, but not Crohn's disease, share pathogenicity islands with urinary pathogenic E. coli.

Adherent and invasive mucosa-associated Escherichia coli have been implicated in the pathogenesis of colon cancer and inflammatory bowel diseases. It has been reported that such isolates share features of extraintestinal E. coli (ExPEC) and particularly uropathogenic E. coli (UPEC). We used suppression subtractive hybridization (SSH) to subtract the genome of E. coli K-12 from that of a colon cancer mucosal E. coli isolate. Of the subtracted sequences, 53 % were present in the genomes of one or more of three sequenced UPEC strains but absent from the genome of an enterohaemorrhagic E. coli (EHEC) strain. Of the subtracted sequences, 80 % matched at least one UPEC genome, whereas only 4 % were absent from the UPEC genomes but present in the genome of the EHEC strain. A further genomic subtraction against the UPEC strain 536 enriched for sequences matching mobile genetic elements, other ExPEC strains, and other UPEC strains or commensals, rather than strains associated with gastrointestinal disease. We analysed the distribution of selected subtracted sequences and UPEC-associated pathogenicity islands (PAIs) amongst a panel of mucosa-associated E. coli isolated from colonoscopic biopsies of patients with colon cancer, patients with Crohn's disease and controls. This enabled us to identify a group of isolates from colon cancer (30-40 %) carrying multiple genes previously categorized as UPEC-specific and implicated in virulence.

Characterization of epithelial IL-8 response to inflammatory bowel disease mucosal E. coli and its inhibition by mesalamine.

BACKGROUND: Mucosally adherent E. coli are found in inflammatory bowel disease (IBD) and colon cancer. They promote release of the proinflammatory cytokine interleukin-8 (IL-8). We explored mechanisms for this release and its inhibition by drugs. METHODS: IL-8 release from colon epithelial cells in response to mucosal E. coli isolates from IBD, colon cancer, and controls was characterized at the cellular and molecular level. RESULTS: IL-8 response of HT29 cells was greater with Crohn's disease (689 +/- 298 [mean +/- SD] pg IL-8/mL at 4 hours, n = 7) and colon cancer isolates (532 +/- 415 pg/mL, n = 14) than with ulcerative colitis (236 +/- 58 pg/mL, n = 6) or control isolates (236 +/- 100 pg/mL, n = 6, P < 0.0001). Bacterial supernatants contained shed flagellin that triggered IL-8 release. For whole bacteria the IL-8 response to E. coli that agglutinate red blood cells (548 +/- 428 pg IL-8/mL, n = 16), a function that correlates with epithelial invasion, was greater than for nonhemagglutinators (281 +/- 253 pg/mL, n = 17; P < 0.0001). This was particularly marked among E. coli that, although flagellate, could not release IL-8 from TLR5-transfected HEK293 cells. IL-8 release was mediated by extracellular-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) and inhibited by mesalamine, but not hydrocortisone, at therapeutic concentrations. CONCLUSIONS: Mucosa-associated E. coli shed flagellin that elicits epithelial IL-8 release but this may only become relevant when the mucosal barrier is weakened to expose basolateral TLR5. Adherent and invasive IBD and colon cancer E. coli isolates also elicit a flagellin-independent IL-8 response that may be relevant when the mucosal barrier is intact. The IL-8 release is MAPK-dependent and inhibited by mesalamine.

Maspin - the most commonly-expressed gene of the 18q21.3 serpin cluster in lung cancer - is strongly expressed in preneoplastic bronchial lesions.

Maspin, SCCA1/2 and hurpin were identified by cDNA microarray analyses as dramatically differentially expressed transcripts in primary non-small cell lung cancer (NSCLC). These sequences are located within a 10-gene serpin cluster on 18q21.3. Using comparative RT-PCR, we have investigated the expression of each of these serpins, including their flanking loci, in an independent NSCLC series. Whereas six of the genes (maspin, SCCA1, SCCA2, hurpin, megsin and pAI-2) were commonly differentially expressed in primary lesions, each significantly more often in squamous cell tumours, maspin was identified as the most frequently over-represented sequence in both squamous cell carcinoma and adenocarcinoma. Using a well-characterized monoclonal antibody, we have shown strong maspin expression in tumour protein extracts, detected multiple isoforms of the 42 kDa protein and shown that maspin is localized specifically to the tumour cells within neoplastic lesions. In contrast, most cells in non-neoplastic lung tissue appear not to express the gene, with the exception of the multipotent basal epithelial cells that line the bronchial airway. These reserve cells generally show strong predominantly nuclear localization of maspin. Strong nuclear expression of maspin within primary tumour cells is correlated with increased survival (P=0.05) and a longer remission duration (P=0.02) in resectable-staged patients. However, within the airways of cancer patients and somewhat in contrast to this observation, such expression was more frequently detected in the superficial cells of preneoplastic over non-neoplastic epithelia (P<0.0001), consistent with a role for the protein in early neoplasia.