Quantitative analysis of waterfowl parvoviruses in geese and Muscovy ducks by real-time polymerase chain reaction: correlation between age, clinical symptoms and DNA copy number of waterfowl parvoviruses

<p>Abstract</p> <p>Background</p> <p>Waterfowl parvoviruses cause serious loss in geese and ducks production. Goose parvovirus (GPV) is infectious for geese and ducks while Muscovy duck parvovirus (MDPV) infects Muscovy ducks only. So far, for these viruses' sensitive detection polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) were applied. However, there was no molecular biology method for both waterfowl parvoviruses detection and quantification which could unify the laboratory procedures. The level of GPV and MDPV replication and distribution plays a significant role in the parvoviral infection progress and is strictly correlated to clinical symptoms. Meanwhile, experiments conducted previously on GPV distribution in geese, performed as animal trial, did not involve epidemiological data from the disease field cases. The study on the correlation between age, clinical symptoms and viral DNA copy number may be benefitable in understanding the GPV and MDPV infection. Such data may also aid in determination of the stage and severity of the infection with parvoviruses. Therefore the aim of this study was to develop quantitative real-time PCR for parallel detection of GPV and MDPV in geese and Muscovy ducks and to determine the correlation between the age of the infected birds, clinical symptoms and DNA copy number for the estimation of the disease stage or severity.</p> <p>Results</p> <p>In order to develop quantitative real-time PCR the viral material was collected from 13 farms of geese and 3 farms of Muscovy ducks. The designed primers and <it>Taqman </it>probe for real-time PCR were complementary to GPV and MDPV inverted terminal repeats region. The pITR plasmid was constructed, purified and used to prepare dilutions for standard curve preparation and DNA quantification. The applied method detected both GPV and MDPV in all the examined samples extracted from the heart and liver of the infected birds. The conducted correlation tests have shown relationship between age, clinical symptoms during parvoviral infection and the DNA copy number of these pathogens. The method allowed for a sensitive detection of GPV and MDPV even in 1-week old infected goslings or 2-week old ducklings before observation of any disease symptoms.</p> <p>Conclusions</p> <p>The developed method was found to be a valuable tool for the unification of laboratory procedures and both parvoviruses parallel detection and quantification. The conducted analysis revealed significant correlation between the age of the infected birds, the observed clinical symptoms and DNA copy number of GPV and MDPV in the examined organs. The obtained data may aid in better understanding of the pathogenesis and epidemiology of Derzsy's disease and 3-w disease as well as estimation of the infection's severity and stage of the disease.</p

How do genetic relatedness and spatial proximity shape African swine fever infections in wild boar?

The importance of social and spatial structuring of wildlife populations for disease spread, though widely recognized, is still poorly understood in many host-pathogen systems. In particular, system-specific kin relationships among hosts can create contact heterogeneities and differential disease transmission rates. Here, we investigate how distance-dependent infection risk is influenced by genetic relatedness in a novel host-pathogen system: wild boar (Sus scrofa) and African swine fever (ASF).We hypothesized that infection risk would correlate positively with proximity and relatedness to ASF-infected individuals but expected those relationships to weaken with the distance between individuals due to decay in contact rates and genetic similarity.We genotyped 323 wild boar samples (243 ASF-negative and 80 ASF-positive) collected in north-eastern Poland in 2014–2016 and modelled the effects of geographic distance, genetic relatedness and ASF virus transmission mode (direct or carcass-based) on the probability of ASF infection. Infection risk was positively associated with spatial proximity and genetic relatedness to infected individuals with generally stronger effect of distance. In the high-contact zone (0–2 km), infection risk was shaped by the presence of infected individuals rather than by relatedness to them. In the medium-contact zone (2–5 km), infection risk decreased but was still associated with relatedness and paired infections were more frequent among relatives. At farther distances, infection risk further declined with relatedness and proximity to positive individuals, and was 60% lower among un-related individuals in the no-contact zone (33% in10–20 km) compared among relatives in the high-contact zone (93% in 0–2 km). Transmission mode influenced the relationship between proximity or relatedness and infection risk. Our results indicate that the presence of nearby infected individuals is most important for shaping ASF infection rates through carcass-based transmission, while relatedness plays an important role in shaping transmission rates between live animals

Molecular characteristics of Polish field strains of Marek's disease herpesvirus isolated from vaccinated chickens

<p>Abstract</p> <p>Background</p> <p>Twenty-nine Marek's disease virus (MDV) strains were isolated during a 3 year period (2007-2010) from vaccinated and infected chicken flocks in Poland. These strains had caused severe clinical symptoms and lesions. In spite of proper vaccination with mono- or bivalent vaccines against Marek's disease (MD), the chickens developed symptoms of MD with paralysis.</p> <p>Because of this we decided to investigate possible changes and mutations in the field strains that could potentially increase their virulence. We supposed that such mutations may have been caused by recombination with retroviruses of poultry - especially reticuloendotheliosis virus (REV).</p> <p>Methods</p> <p>In order to detect the possible reasons of recent changes in virulence of MDV strains, polymerase chain reaction (PCR) analyses for <it>meq </it>oncogene and for long-terminal repeat (LTR) region of REV were conducted. The obtained PCR products were sequenced and compared with other MDV and REV strains isolated worldwide and accessible in the GeneBank database.</p> <p>Results</p> <p>Sequencing of the <it>meq </it>oncogene showed a 68 basepair insertion and frame shift within 12 of 24 field strains. Interestingly, the analyses also showed 0.78, 0.8, 0.82, 1.6 kb and other random LTR-REV insertions into the MDV genome in 28 of 29 of strains. These genetic inserts were present after passage in chicken embryo kidney cells suggesting LTR integration into a non-functional region of the MDV genome.</p> <p>Conclusion</p> <p>The results indicate the presence of a recombination between MDV and REV under field conditions in Polish chicken farms. The genetic changes within the MDV genome may influence the virus replication and its features <it>in vivo</it>. However, there is no evidence that <it>meq </it>alteration and REV insertions are related to the strains' virulence.</p

Dynamics of African swine fever virus (ASFV) infection in domestic pigs infected with virulent, moderate virulent and attenuated genotype II ASFV European isolates

This study aimed to compare the infection dynamics of three genotype II African swine fever viruses (ASFV) circulating in Europe. Eighteen domestic pigs divided into three groups were infected intramuscularly or by direct contact with two haemadsorbent ASFVs (HAD) from Poland (Pol16/DP/ OUT21) and Estonia (Est16/WB/Viru8), and with the Latvian non-HAD ASFV (Lv17/WB/Rie1). Parameters, such as symptoms, pathogenicity, and distribution of the virus in tissues, humoral immune response, and dissemination of the virus by blood, oropharyngeal and rectal routes, were investigated. The Polish ASFV caused a case of rapidly developing fatal acute disease, while the Estonian ASFV caused acute to sub-acute infections and two animals survived. In contrast, animals infected with the ASFV from Latvia developed a more subtle, mild, or even subclinical disease. Oral excretion was sporadic or even absent in the attenuated group, whereas in animals that developed an acute or sub-acute form of ASF, oral excretion began at the same time the ASFV was detected in the blood, or even 3 days earlier, and persisted up to 22 days. Regardless of virulence, blood was the main route of transmission of ASFV and infectious virus was isolated from persistently infected animals for at least 19 days in the attenuated group and up to 44 days in the group of moderate virulence. Rectal excretion was limited to the acute phase of infection. In terms of diagnostics, the ASFV genome was detected in contact pigs from oropharyngeal samples earlier than in blood, independently of virulence. Together with blood, both samples could allow to detect ASFV infection for a longer period. The results presented here provide quantitative data on the spread and excretion of ASFV strains of different virulence among domestic pigs that can help to better focus surveillance activities and, thus, increase the ability to detect ASF introductions earlier.This study was supported by the INIA (Projects RTA2015-00033-C02-01, AT2015-002) and the European Union Reference laboratory for ASF (Grant no.: UE- LR PPA/03). We would like to thank all the staff at the INIA-CISA who worked in the animal facilities department.Peer reviewe

Current status of porcine epidemic diarrhoea (PED) in European pigs

Porcine epidemic diarrhoea (PED) is a highly contagious and devastating enteric disease of pigs caused by porcine epidemic diarrhoea virus (PEDV), an enveloped, single-stranded RNA virus belonging to the Alphacoronavirus genus of the Coronaviridae family. The disease is clinically similar to other forms of porcine gastroenteritis. Pigs are the only known host of the disease, and the occurrence of PED in wild boars is unknown. The virus causes acute diarrhoea, vomiting, dehydration, and high mortality in suckling piglets reaching 100%. Heavy economic losses in the pig-farming industry were sustained in the USA between 2013 and 2015 when PEDV spread very quickly and resulted in epidemics. The loss in the US pig industry has been estimated at almost seven million pigs. The purpose of this review is a description of the current status of porcine epidemic diarrhoea in European pigs and the risk presented by the introduction of PEDV to Poland in comparison to the epidemics in the USA