Development and application of a SYBR Green RT-PCR for first line screening and quantification of porcine sapovirus infection

BACKGROUND: Sapoviruses are single stranded positive sense RNA viruses belonging to the family Caliciviridae. The virus is detected in different species including the human and the porcine species as an enteric pathogen causing asymptomatic to symptomatic enteritis. In this study, we report the development of a rapid real time qRT-PCR based on SYBR Green chemistry for the diagnosis of porcine sapovirus infection in swine. RESULTS: The method allows the detection of porcine sapoviruses and the quantification of the genomic copies present in stool samples. During its development, the diagnostic tool showed good correlation compared with the gold standard conventional RT-PCR and was ten-fold more sensitive. When the method was applied to field samples, porcine noroviruses from genogroup 2 genotype 11b were also detected. The method was also applied to swine samples from the Netherlands that were positive for PoSaV infection. Phylogenetic results obtained from the samples showed that PoSaV sequences were genetically related to the currently described genogroup III, to the proposed genogroup VII and also to the MI-QW19 sequence (close to the human SaV sequences). CONCLUSIONS: A rapid, sensitive, and reliable diagnosis method was developed for porcine sapovirus diagnosis. It correlated with the gold standard conventional RT-PCR. Specificity was good apart for genogroup 2 genotype 11b porcine noroviruses. As a first line screening diagnosis method, it allows a quicker and easier decision on doubtful samples

Repeated examination of natural sapovirus infections in pig litters raised under experimental conditions

Porcine sapovirus, belonging to the family Caliciviridae, is an enteric virus that is widespread in the swine industry worldwide. A total of 14 sapovirus genogroups have been suggested and the most commonly found genogroup in swine is genogroup III (GIII). The goal of the present experiment was to examine the presence of sapovirus in 51 naturally infected pigs at two different time points. The pigs were kept under experimental conditions after weaning. Previous studies on sapovirus have primarily been of a cross sectional nature, typically prevalence studies performed on farms and abattoirs. In the present study, faecal samples, collected from each pig at 5½ weeks and 15-18 weeks of age, were analysed for sapovirus by reverse transciptase polymerase chain reaction and positive findings were genotyped by sequencing. At 5½ weeks of age, sapovirus was detected in the majority of the pigs. Sequencing revealed four different strains in the 5½ week olds-belonging to genogroups GIII and GVII. Ten to 13 weeks later, the virus was no longer detectable from stools of infected pigs. However, at this time point 13 pigs were infected with another GIII sapovirus strain not previously detected in the pigs studied. This GIII strain was only found in pigs that, in the initial samples, were virus-negative or positive for GVII. At 5 weeks of age 74 % of the pigs were infected with sapovirus. At 15-18 weeks of age all pigs had cleared their initial infection, but a new sapovirus GIII strain was detected in 25 % of the pigs. None of the pigs initially infected with the first GIII strain were reinfected with this new GIII strain, which may indicate the presence of a genogroup-specific immunity

Hepatitis E virus: a zoonosis adapting to humans

Hepatitis E virus (HEV) infection is gaining global attention, not only because of the increasing burden of the disease in low endemicity countries, in terms of morbidity and mortality rates, but also due to recent advances in the molecular virology and epidemiology of this emerging pathogen. HEV infection spread can be described as the evolution of a zoonosis towards an established human infection. As known from other viruses, such as the human immunodeficiency virus or the influenza viruses, crossing the species barriers from animals to humans is a recurrent phenomenon. Albeit slow at the beginning, once the virus has adapted to humans, the person-to-person spread can proceed very quickly. Although an optimal cell culture system for HEV is not yet available, outstanding progress has been made with the in vitro expression of HEV-like particles. These new tools have fostered new research to understand the molecular, structural and immunological aspects of human HEV infection. Although some promising data from Phase II vaccine trials are available, recent discoveries will certainly open new avenues for HEV-specific prophylaxis and therap

Protocol for a Scoping/Systematic Review: Scoping Review of Vaccination for the Prevention of Calf Scours in Cow-Calf Operations

Background: The use of antimicrobials in the livestock industry has been a topic of increasing concern in the last  few years. Calf scours is one of the main causes of mortality among calves younger than 1 month and affects the development of the animal, representing severe economic losses to the producers. The causes of the disease include a variety of viral, bacterial and protozoal pathogens, and other non-infectious causes. The treatment efficacy, in particular of antimicrobial therapy, depends on the type of infection. Blanket antimicrobial treatment for calf scours without identification of the pathogen may contribute to antimicrobial resistance. In this review we explore the available literature for evidence of effective methods to prevent calf scours in cow-calf operations in California through the use of vaccination.Objectives:  The objective of this scoping review is to examine and describe the existing literature on vaccination for the prevention of calf scours that might reduce the incidence of calf scours by different causes, and therefore reduce the use of antimicrobials due to calf scours.Design: Primary research on vaccination for pathogens that cause calf scours will be considered for inclusion, such as studies conducted in pre-weaned calves that report the efficacy of vaccines for the prevention of calf scours. The process for selection and inclusion of the studies will be reported in a flow chart according to the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA). The results will be summarized in tables and charts describing study types, interventions and outcomes

Waterborne outbreak of Norwalk-like virus gastroenteritis at a tourist resort, Italy.

In July 2000, an outbreak of gastroenteritis occurred at a tourist resort in the Gulf of Taranto in southern Italy. Illness in 344 people, 69 of whom were staff members, met the case definition. Norwalk-like virus (NLV) was found in 22 of 28 stool specimens tested. The source of illness was likely contaminated drinking water, as environmental inspection identified a breakdown in the resort water system and tap water samples were contaminated with fecal bacteria. Attack rates were increased (51.4%) in staff members involved in water sports. Relative risks were significant only for exposure to beach showers and consuming drinks with ice. Although Italy has no surveillance system for nonbacterial gastroenteritis, no outbreak caused by NLV has been described previously in the country

Skin diseases in cetaceans. Scientific Committee document SC/60/DW8, International Whaling Commission, June 2008, Santiago, Chile

Micro-organisms that are known or suspected to cause skin diseases in cetaceans are briefly reviewed. Viruses belonging to four families i.e. Caliciviridae, Herpesviridae, Papillomaviridae and Poxviridae were detected by electron microscopy, histology and molecular techniques in vesicular skin lesions, black dots perceptible by the touch, warts and tattoos in several species of odontocetes and mysticetes. Herpesviruses, poxviruses and likely a cutaneous papillomavirus are cetacean specific. Among bacteria, Dermatophilus spp., Erysipelothrix rhusiopathiae, Mycobacterium marinum, Pseudomonas spp., Streptococcus iniae and Vibrio spp. were isolated from ulcerative dermatitis, pyogranulomatous dermatitis and panniculitis, diamond skin disease and slow-healing ulcers and abscesses. Aeremonas spp., Mycobacterium marinum, Pseudomonas spp. and Vibrio spp. are normally present in the marine environment while Erysipelothrix rhusiopathiae and Streptococcus iniae are fish pathogens that may also infect captive dolphins. Most seem to be opportunistic pathogens, exploiting some break-down in the host’s defenses to initiate an infection. Selection of antibiotic-resistant bacteria through the prophylactic use of antibiotics in aquaculture is suggested to be a growing problem in South America and may account for the emergence of unusual cutaneous conditions. At least four groups of fungi i.e. Candida albicans, Fusarium spp., Trichophyton spp. and Lacazia loboi cause skin diseases. Candidiasis occurs predominantly in captive odontocetes. The lesions are often localized around the body orifices and may become extensive, granulating and ulcerated. Fusariosis is characterized by firm, erythematous, cutaneous nodules. Trichophyton spp. was isolated from widespread superficial nodules in an Atlantic T. truncates kept in captivity in Japan. Lobomycosis or lacaziosis is distinguished by grayish, whitish to slightly pink, verrucuous lesions, often in pronounced relief that may ulcerate. While initially described only in Tursiops truncates and Sotalia guianensis from the Americas, lobomycosis seems to be expanding to other continents. The role of ballast water in transporting fungi worldwide should be investigated. Finally, ciliated protozoans, likely Kyaroikeus cetarius, caused invasive dermatitis in small cetaceans from the USA and Korea. The aquatic environment of cetaceans is naturally home to bacteria and fungi but cetacean skin has several mechanisms to impede invasion. Chemical contaminants may affect natural skin barriers and depress the immune system. Wounds and specific viral infection (poxvirus, herpesvirus) may provide routes of entry

Discovery and genomic characterization of a novel bat sapovirus with unusual genomic features and phylogenetic position

Sapovirus is a genus of caliciviruses that are known to cause enteric disease in humans and animals. There is considerable genetic diversity among the sapoviruses, which are classified into different genogroups based on phylogenetic analysis of the full-length capsid protein sequence. While several mammalian species, including humans, pigs, minks, and dogs, have been identified as animal hosts for sapoviruses, there were no reports of sapoviruses in bats in spite of their biological diversity. In this report, we present the results of a targeted surveillance study in different bat species in Hong Kong. Five of the 321 specimens from the bat species, Hipposideros pomona, were found to be positive for sapoviruses by RT-PCR. Complete or nearly full-length genome sequences of approximately 7.7 kb in length were obtained for three strains, which showed similar organization of the genome compared to other sapoviruses. Interestingly, they possess many genomic features atypical of most sapoviruses, like high G+C content and minimal CpG suppression. Phylogenetic analysis of the viral proteins suggested that the bat sapovirus descended from an ancestral sapovirus lineage and is most closely related to the porcine sapoviruses. Codon usage analysis showed that the bat sapovirus genome has greater codon usage bias relative to other sapovirus genomes. In summary, we report the discovery and genomic characterization of the first bat calicivirus, which appears to have evolved under different conditions after early divergence from other sapovirus lineages.published_or_final_versio

Detection of RHDV strains in the Iberian hare (Lepus granatensis): earliest evidence of rabbit lagovirus cross-species infection

This is an Open Access article distributed under the terms of the Creative Commons Attribution License.-- et al.Rabbit hemorrhagic disease virus (RHDV) is a highly lethal Lagovirus, family Caliciviridae, that threatens European rabbits (Oryctolagus cuniculus). Although a related virus severely affects hares, cross-species infection was only recently described for new variant RHDV in Cape hares (Lepus capensis mediterraneus). We sequenced two strains from dead Iberian hares (Lepus granatensis) collected in the 1990s in Portugal. Clinical signs were compatible with a Lagovirus infection. Phylogenetic analysis of the complete capsid gene positioned them in the RHDV genogroup that circulated on the Iberian Peninsula at that time. This is the earliest evidence of RHDV affecting a species other than European rabbits.This work was supported by FCT (Fundação para a Ciência e a Tecnologia; research project ref.: FCT-ANR/BIABIC/0043/2012). FCT also supported the doctoral grants of AML and AP (refs.: SFRH/BD/78738/2011 and SFRH/BD/71252/2010) and the FCT Investigator grant of JA (ref.: IF/01396/2013). “Genomics Applied To Genetic Resources” co-financed by North Portugal Regional Operational Programme 2007/2013 (ON.2 – O Novo Norte), under the National Strategic Reference Framework (NSRF), through the European Regional Development Fund (ERDF), also supported this work.Peer Reviewe

Molecular screening of blue mussels indicated high mid-summer prevalence of human genogroup II Noroviruses, including the pandemic “GII.4 2012” variants in UK coastal waters during 2013

Pandemic norovirus in coastal blue mussels during summer in UK This molecular study is the first report, to the best of our knowledge, on identification of norovirus, NoV GII.4 Sydney 2012 variants, from blue mussels collected from UK coastal waters. Blue mussels (three pooled samples from twelve mussels) collected during the 2013 summer months from UK coastal sites were screened by RT-PCR assays. PCR products of RdRP gene for noroviruses were purified, sequenced and subjected to phylogenetic analysis. All the samples tested positive for NoVs. Sequencing revealed that the NoV partial RdRP gene sequences from two pooled samples clustered with the pandemic “GII.4 Sydney variants” whilst the other pooled sample clustered with the NoV GII.2 variants. This molecular study indicated mussel contamination with pathogenic NoVs even during mid-summer in UK coastal waters which posed potential risk of NoV outbreaks irrespective of season. As the detection of Sydney 2012 NoV from our preliminary study of natural coastal mussels interestingly corroborated with NoV outbreaks in nearby areas during the same period, it emphasizes the importance of environmental surveillance work for forecast of high risk zones of NoV outbreaks