Aujeszky's Disease Virus Research Articles

Virome analysis of ticks in a forest region of Liaoning, China: characterization of a novel hepe-like virus sequence

BackgroundTicks (class Arachnida, subclass Acari) are vectors of transmitting a broad range of pathogenic microorganisms, protozoa, and viruses affecting humans and animals. Liaoning Province is rich in forests where different animals and, abundant Haemaphysalis longicornis ticks exist.MethodsUsing viral metagenomics, we analyzed the virome in 300 Haemaphysalis longicornis ticks collected from June to August 2015 in the forested region of Liaoning Province, China.ResultsFrom the 300 ticks, 1,218,388 high-quality reads were generated, of which 5643 (0.463%) reads showed significant sequence identity to known viruses. Sequence and phylogenetic analysis revealed that viral sequences showing a close relationship with Dabieshan tick virus, Aleutian mink disease virus, adeno-associated virus, Gokushovirus, avian gyrovirus 2 were present in the virome of these ticks. However, the significance of these viruses to human and animal health requires further investigation. Notably, an hepe-like virus, named tick-borne hepe-like virus sequence, was obtained and was highly prevalent in these ticks with a rate of 50%. Nevertheless, one constraint of our study was the limited geographical distribution of the sampled ticks.ConclusionOur study offers an overview of the virome in ticks from a forest region of Liaoning Province and provides further awareness of the viral diversity of ticks.

Genetic and phenotypic parameters for Aleutian disease tests and their correlations with pelt quality, reproductive performance, packed-cell volume, and harvest length in mink.

Aleutian disease (AD), caused by the Aleutian mink disease virus (AMDV), is a major health concern that results in global economic losses to the mink industry. The unsatisfactory outcome of the culling strategy, immunoprophylaxis, and medical treatment in controlling AD have urged mink farmers to select AD resilient mink based on several detection tests, including enzyme-linked immunosorbent assay (ELISA), counterimmunoelectrophoresis (CIEP), and iodine agglutination test (IAT). However, the genetic analysis of these AD tests and their correlations with pelt quality, reproductive performance, packed-cell volume (PCV), and harvest length (HL) have not been investigated. In this study, data on 5,824 mink were used to estimate the genetic and phenotypic parameters of four AD tests, including two systems of ELISA, CIEP, and IAT, and their genetic and phenotypic correlations with two pelt quality, five female reproductive performance, PCV, and HL traits. Significances (P < 0.05) of fixed effects (sex, year, dam age, and color type), covariates (age at harvest and blood sampling), and random effects (additive genetic, permanent environmental, and maternal effects) were determined under univariate models using ASReml 4.1 software. The genetic and phenotypic parameters for all traits were estimated under bivariate models using ASReml 4.1 software. Estimated heritabilities (±SE) were 0.39 ± 0.06, 0.61 ± 0.07, 0.11 ± 0.07, and 0.26 ± 0.05 for AMDV antigen-based ELISA (ELISA-G), AMDV capsid protein-based ELISA, CIEP, and IAT, respectively. The ELISA-G also showed a moderate repeatability (0.58 ± 0.04) and had significant negative genetic correlations (±SE) with reproductive performance traits (from -0.41 ± 0.16 to -0.49 ± 0.12), PCV (-0.53 ± 0.09), and HL (-0.45 ± 0.16). These results indicated that ELISA-G had the potential to be applied as an indicator trait for genetic selection of AD resilient mink in AD endemic ranches and therefore help mink farmers to reduce the adverse effects caused by AD.

Effects of health related farm-level factors on skin size and quality in commercial mink (Neovison vison) production

This study investigated the potential effects of management and health related factors on the productivity in the commercial mink production, during 2015−2018. Data were available from the database at Kopenhagen Fur, the national veterinary prescription database, VetStat, and the laboratory database at the Center for Diagnostics, Technological University of Denmark. A cross-sectional study, including 1.464 min. farms grouped into 1.187 epidemiological units, was applied. Data were analyzed in two models with different outcomes representing productivity on the mink farms, namely skin size and economical value (value sum) of the produced skin. The studied risk factors included use of vaccines and antibacterials, herd size, associated feed producer, purchases and sales of live animals, breeding results (litter size after weaning), Aleutian mink disease virus antibody (AMDV status) and stamping out, and laboratory test results.Vaccination against mink enteritis parvovirus and high breeding results were found to have a positive association with both outcomes, skin size and value sum. Both outcomes also varied significantly between farm clusters associated with different feed producers. Significant effects of antibacterial treatment were found, but the results were complex with both positive and negative associations with the outcome variables, depending on season and interactions with feed producer. Positive effects on antibacterial prescription on skin size were observed, except for farms associated with two small feed producers, known to have a variable microbiological feed quality. In farms receiving feed of very high quality, the positive effect of antibacterial prescription was marginal. ConclusionsThe use of mink data has allowed us to assess the impact of feed quality as well as antibacterial prescription on productivity. The results showed a positive quantitative effect of vaccination against mink enteritis parvovirus on skin size and value, with an optimal effect by vaccination of the whole litter. Antibacterial prescription in the growth period, particularly around weaning, was found to have a positive quantitative effect on productivity in some farms, and the results suggest that the effect was associated with the feed quality. Use of antibacterials to counteract negative effects of low feed quality is not in accordance with principles for prudent use of antibacterials.

Разработка и получение иммунодиагностикума к вирусу алеутской болезни норок

Разработка и получение иммунодиагностикума к вирусу алеутской болезни норок

Aptamer-targeting of Aleutian mink disease virus (AMDV) can be an effective strategy to inhibit virus replication

Aleutian mink disease (AMD), which is caused by Aleutian mink disease virus (AMDV), is an important contagious disease for which no effective vaccine is yet available. AMD causes major economic losses for mink farmers globally and threatens some carnivores such as skunks, genets, foxes and raccoons. Aptamers have exciting potential for the diagnosis and/or treatment of infectious viral diseases, including AMD. Using a magnetic beads-based systemic evolution of ligands by exponential enrichment (SELEX) approach, we have developed aptamers with activity against AMDV after 10 rounds of selection. After incubation with the ADVa012 aptamer (4 μM) for 48 h, the concentration of AMDV in the supernatant of infected cells was 47% lower than in the supernatant of untreated cells, whereas a random library of aptamers has no effect. The half-life of ADVa012 was ~ 32 h, which is significantly longer than that of other aptamers. Sequences and three dimensions structural modeling of selected aptamers indicated that they fold into similar stem-loop structures, which may be a preferred structure for binding to the target protein. The ADVa012 aptamer was shown to have an effective and long-lasting inhibitory effect on viral production in vitro.

Pseudorabies (Aujeszky's disease) virus DNA detection in swine nasal swab and oral fluid specimens using a gB-based real-time quantitative PCR

In this study, the detection of PRV DNA in nasal swab (n = 440) and oral fluid (n = 1,545) samples collected over time from experimentally PRV vaccinated and/or PRV inoculated pigs (n = 40) was comparatively evaluated by real-time PCR. Serum samples (n = 440) were tested by PRV gB/gE blocking ELISAs (Pseudorabies Virus gB Antibody Test Kit and Pseudorabies Virus gpI Antibody Test Kit, IDEXX Laboratories, Inc., Westbrook, ME) to monitor PRV status over time. Following exposure to a gE-deleted modified live vaccine (Ingelvac® Aujeszky MLV, Boehringer Ingelheim, Ridgefield, CT) and/or a wild-type virus (3 CR Ossabaw), PRV gB DNA was detected in oral fluid specimens in a pattern similar to that of nasal swabs. For quantitative analyses, PRV PCR quantification cycle (Cq) results were re-expressed as “efficiency standardized Cqs (ECqs)” as a function of PCR efficiency using plate-specific positive amplification controls. ROC analyses of the PRV gB PCR ECqs results showed a similar performance of the PRV gB PCR for nasal swab and oral fluid specimens (area under the ROC curve = 85 % vs 83 %) and, based on an ECq cutoff of 0.01 a diagnostic specificity of 100 % and diagnostic sensitivities for oral fluid and nasal swab specimens of 53 % (95 % CI: 43 %, 62 %) and 70 % (95 % CI: 55 %, 83 %), respectively. Thus, the results described herein demonstrated the detection of PRV gB DNA in swine oral fluid and supported the use of this specimen in PRV diagnosis and surveillance.

Occurrence of Aujeszky’s disease in wild boar (Sus scrofa) in Slovakia

Along with the rising population of wild boars (Sus scrofa scrofa Linnaeus, 1758) in Slovakia, a higher rate of exposition to Aujeszky’s disease virus (ADV) was noted. The high density of animals is one of the main influencing factors of ADV persistence. Wild boar tissue samples were obtained by local hunters from different locations of Central and Eastern Slovakia. In total, muscle tissue samples from 139 individual wild boars were collected. Muscle fluid was obtained from these samples and subjected to serological analysis; a commercial ADV Ab-ELISA kit was used. In total, 53 samples were proved to be seropositive; out of these 19 samples of piglets, 13 samples of juveniles and 21 samples of adults. Out of 139 samples tested by ELISA, 53 were seropositive, which represents 38.12 % of positive samples. No significant difference was found between the two sexes; however, piglets have shown to have a significantly higher occurrence compared to adults. Our study confirmed the presence of ADV antibodies in the wild boar population in central and eastern Slovakia.

Detection of selection signatures for response to Aleutian mink disease virus infection in American mink

Aleutian disease (AD) is the most significant health issue for farmed American mink. The objective of this study was to identify the genomic regions subjected to selection for response to infection with Aleutian mink disease virus (AMDV) in American mink using genotyping by sequencing (GBS) data. A total of 225 black mink were inoculated with AMDV and genotyped using a GBS assay based on the sequencing of ApeKI-digested libraries. Five AD-characterized phenotypes were used to assign animals to pairwise groups. Signatures of selection were detected using integrated measurement of fixation index (FST) and nucleotide diversity (θπ), that were validated by haplotype-based (hap-FLK) test. The total of 99 putatively selected regions harbouring 63 genes were detected in different groups. The gene ontology revealed numerous genes related to immune response (e.g. TRAF3IP2, WDR7, SWAP70, CBFB, and GPR65), liver development (e.g. SULF2, SRSF5) and reproduction process (e.g. FBXO5, CatSperβ, CATSPER4, and IGF2R). The hapFLK test supported two strongly selected regions that contained five candidate genes related to immune response, virus–host interaction, reproduction and liver regeneration. This study provided the first map of putative selection signals of response to AMDV infection in American mink, bringing new insights into genomic regions controlling the AD phenotypes.

Specific humoral immunity in mink infected with the Aleutian disease virus (AMDV) and vaccinated against distemper

The Aleutian mink disease (AD) is one of the most serious threats to modern mink breeding. In farms where AD is endemic, a significant effect on breeding and reproductive effects is caused by permanent AMDV infection. The aim of this study was investigating the influence of subclinical infection of AMDV on the functional immune status and on the state of postvaccinal immunity against mink distemper. The analyses involved 30 brown mink females aged 12 weeks and allocated into an experimental group (group I) and control group (group II), with 15 animals in each. The minks from group I originated from the farm with an endemic form of Aleutian disease. The control minks from group II originated from the AMDV-uninfected farm. The dynamics of the growth of specific antibodies was examined after vaccination of the minks from both groups against distemper. Serum levels were determined in both groups with a seroneutralisation assay (SN). Obtaining positive values in the SN test in both groups of animals confirmed the induction of a specific immune response. There were significant differences in the magnitude of the positive SN assay values between the minks from group I and group II at all days of determination. The significantly lower SN values noted in the group minks with subclinical course of AMDV reveal that Aleutian disease can cause a suppressive effect on humoral response after vaccination.

Aleutian mink disease: Spatio-temporal variation of prevalence and influence on the feral American mink.

Pathogens are one of the factors driving wildlife population dynamics. The spread of pathogens in wildlife is currently highly related to the transmission of pathogens from farmed animals, which has increased with the constant development of farming. Here, we analysed the spatio‐temporal variation in the prevalence of Aleutian mink disease virus (AMDV) antibodies in feral American mink (Neovison vison) populations in Poland (1,153 individuals from nine sites) in relation to mink farming intensity. AMDV was detected in feral mink at all study sites and the prevalence ranged from 0.461 in the northern region to 0.826 in the western region. Mink males and adults were infected more often than females and subadults; the infection was also more frequent during the mink breeding season than during non‐breeding. The prevalence of AMDV changed non‐linearly in consecutive years and the peak of prevalence was every 3–4 years. The predicted AMDV prevalence was low at sites where the number of farmed mink was also low and increased linearly with the increase in the number of mink kept on farms. The predicted AMDV prevalence at sites with low mink farming intensity strongly varied between years, whereas at sites with high mink farming intensity, the predicted prevalence did not change significantly. AMDV infection affected the mink's body condition and caused an increase in the size of the spleen, liver and kidneys. This study shows that Aleutian mink disease strongly affects feral mink but the spatio‐temporal variation of its prevalence is complex and partly related to the transmission of the virus from farmed mink to feral populations. The study highlights the complexity of AMDV circulation in feral mink populations and implicates a potential spillover of the virus to native species.

Dietary supplementation of Ascophylum nodosum improved kidney function of mink challenged with Aleutian mink disease virus

BackgroundFeed additives which can ease the negative effects of infection by the Aleutian mink disease virus (AMDV) are of interest to mink farmers. The effects of kelp meal (Ascophylum nodosum) supplementation on immune response, virus replication and blood parameters of mink inoculated with AMDV were assessed. AMDV-free black mink (n = 75) were intranasally inoculated with a local strain of AMDV and fed a commercial pellet supplemented with kelp meal at the rates of 1.5% or 0.75% of the feed or were kept as controls (no kelp) for 451 days. Blood was collected on days 0 (pre-inoculation), 31, 56, 99, 155, 366 and 451 post-inoculation (dpi).ResultsNo significant difference was observed among the treatments for the proportion of animals positive for antibodies against the virus measured by the counter-immunoelectrophoresis (CIEP), viremia measured by PCR, antibody titer measured by quantitative ELISA, total serum protein measured by a refractometer or elevated levels of gamma globulin measured by iodine agglutination test at the sampling occasions. At the termination of the experiment on 451 dpi, there were no differences among treatments for antibody titer measured by CIEP, total serum protein, albumin, globulins, albumin:globulin ratio, alkaline phosphatase, gamma-glutamyl transferase, and proportions of PCR positive spleen, lymph node or bone marrow samples, but blood urea nitrogen and creatine levels were significantly lower in the 1.5% kelp supplemented group than in the controls.ConclusionKelp supplementation improved kidney function of mink infected with AMDV with no effect on liver function, immune response to infection by AMDV or virus replication.

PSXII-24 Identification of selection signatures for response of American mink to Aleutian mink disease virus infection

Abstract Aleutian disease (AD) is one of the most important health problems in the mink industry worldwide, leading to economic losses. We used a set of single nucleotide polymorphisms (SNPs) to detect the genomic regions potentially under selection for response to Aleutian mink disease virus (AMDV) infection in black American mink. A total of 191 mink which were inoculated with a local strain of AMDV and survived until pelting were genotyped using genotyping-by-sequencing technique. The presence of viral DNA in the spleen samples were tested by polymerase chain reaction. After filtering, 47,800 SNPs at 171 individuals were used for further analyses. Signatures of selection for response to AMDV infection were detected using fixation index (FST) and nucleotide diversity (θπ) statistics measured between negative and positive groups. The overlap of top 1% SNPs obtained from both FST and θπ scores were considered as potential selection signs. This measurement identified a total of 21 candidate regions containing 11 genes which were likely subjected to selection for viral clearance. Several identified genes were those that modulate immune system (TCF4), reproductive process (CATSPERB, MAS1 and IGF2R), response to stimulus (WNT11 and MAS1), and functions of heart (TENM4 and WNT11) and liver (IGF2R). In addition, gene ontology showed that 63.6% of detected genes (seven) were involved in binding activities (GO:0005488). These genes can be used in molecular assessment of viral clearance in American mink. The results indicated that selection for viral clearance and thus AD tolerant animals can be a feasible strategy to deal with the global AMDV infection on mink farms.

12 Genetic and phenotypic parameters for Aleutian disease tests and their correlations with growth and pelt quality traits in American mink

Abstract Aleutian disease brings tremendous financial losses to the mink industry. The ineffective immunoprophylaxis, medication, and culling strategies have urged the mink industry to select mink with low quantitative enzyme-linked immunosorbent assay (qELISA) score or negative counterimmunoelectrophoresis (CIEP) test result. However, little is known about the heritabilities of qELISA and CEIP as well as their relationships with growth and pelt quality traits. The traits, including qELISA, CIEP, body length at harvest (HLEN), the size of dried pelt (SIZE), the overall quality of dried pelt (QUA), and the nap length of dried pelt (NAP), were measured on 1,683 American mink from the Canadian Center for Fur Animal Research (Nova Scotia, Canada) and Millbank Fur Farm (Ontario, Canada). Significance (P &amp;lt; 0.05) of fixed effects (sex, farm, age, and color) and random effects (common litter, permanent environment, and dam) were determined by univariate analyses, while genetic and phenotypic parameters for all traits were estimated under bivariate analyses using ASREML 4.1. Estimated heritabilities (±SE) were 0.41±0.07 for qELISA, 0.06±0.06 for CIEP, 0.39±0.06 for HLEN, 0.46±0.07 for SIZE, 0.25±0.06 for QUA, and 0.46±0.08 for NAP. The qELISA showed non-significant (P &amp;gt; 0.05) genetic correlations with HLEN (0.05±0.13) and dried pelt traits (0.02±0.18 with SIZE, -0.21±0.20 with QUA, and -0.13±0.16 with NAP). The CIEP only showed a significant (P &amp;lt; 0.05) negative genetic correlation with SIZE (-0.85±0.33). The moderate-to-high heritabilities of qELISA, HLEN, SIZE, QUA, and NAP indicated that these traits can be genetically improved through a genetic/genomic selection. The low and non-significant heritability of CIEP indicated the ineffectiveness of direct selection for this trait. The estimated genetic parameters for qELISA suggested that selection for lower qELISA scores may not interfere with the selection of pelt size and quality in the genetic improvement programs of American mink.

PSVII-39 Late-Breaking Abstract: Enhancing production and Aleutian disease resilience in mink through advanced genomics

Abstract The fur industry is one of the oldest and the most historically significant industries in Canada. The industry has used American mink (Neovison vison) as the major source of fur for decades because of their high-quality fur and wide range of colours. This project will seek to (1) create the first accurate whole-genome sequence assembly of mink using next-generation sequencing technology to help understanding the biology and evolution of the order Carnivora, (2) design a robust and informative SNP assay for genomics discovery in mink, (3) discover genome structure and signature of selection as well as identify new genetic variants explaining variation in economically important traits, and (4) identify the genetic relationships among these traits including feed efficiency, Aleutian disease resilience, fur quality, reproductive performance, growth rate and pelt size. One hundred mink DNA samples from the Canadian Centre for Fur Animal Research at Dalhousie Agriculture Campus (Truro, Nova Scotia), and one breeding population (Millbank Fur Farm Limited, Rockwood, Ontario) were sequenced using next-generation whole-genome sequencing with more than 30x coverage to create the first SNP assay for American mink. A DNA panel composed of these sequenced mink from five color-types were assembled to identify the most homozygous individual as the reference animal for whole-genome sequence assembly development. The phenotypic data and DNA samples from 3,323 animals were collected and will be genotyped using the customized assay. The ultimate objective is to develop new tools for implementation of marker assisted selection or genomic selection in mink breeding programs for development of superior, highly efficient, and healthy animals. This approach will help improve the overall performance of the North American mink industry, which is now in difficulty due to several economic factors such as the high price of feed, declining price of fur and prevalence of diseases.

Development of an antigen-capture enzyme-linked immunosorbent assay for diagnosis of Aleutian mink disease virus.

Aleutian mink disease (AMD), caused by Aleutian mink disease virus (AMDV), is a very important infectious disease of mink. Currently, elimination of antibody- or antigen-positive animals is the most successful strategy for eradicating AMD, but the claw-cutting method of blood sampling is difficult to perform and painful for the animal. In this study, we aimed to establish an antigen capture enzyme-linked immunosorbent assay (AC-ELISA) method for the efficient detection of AMDV antigens using fecal samples. A purified mouse monoclonal antibody (mAb) was used as the capture antibody, and a rabbit polyclonal antibody (pAb) was used as the detection antibody. The assay was optimized by adjusting a series of parameters. Using a cutoff value of 0.205, the limit of detection of the AC-ELISA for strain AMDV-G antigen was 2 μg/mL, and there was no cross-reaction with other mink viruses. The intra- and inter-assay standard deviations were below 0.046, and the correlation of variance (CV) values were 1.24-7.12% when testing fecal samples. Compared with conventional PCR results, the specificity and sensitivity were 91.5% and 90.6%, respectively, and the concordance rate between the two methods was 91.1%.

Effects of dietary kelp (Ascophylum nodosum) supplementation on survival rate and reproductive performance of mink challenged with Aleutian mink disease virus

Infection with Aleutian mink disease virus (AMDV) has negative effects on reproductive performance and survival rate of American mink (Neovison vison). The objectives of this study were to assess the effects of kelp (Ascophylum nodosum) supplementation on survival, growth rate, and reproductive performance of mink challenged with AMDV. AMDV-free female black mink (n = 75) were intranasally inoculated with a local AMDV strain. Mink were fed a commercial pellet supplemented with 1.5% or 0.75% kelp or were kept as controls (received no kelp) for 451 d. Body weight and rectal temperature were recorded on days 0, 31, 56, 99, 155, 366, and 451 post inoculation (PI). Annual mortality rates were 13.6%, 20.0%, and 31.8% for mink fed 1.5%, 0.75%, or 0.0% kelp, respectively (P = 0.29). Mink which were fed 1.5% kelp had a significantly (P &lt; 0.01) greater daily weight loss during breeding and post-breeding periods (days 155–366 PI), and outperformed (P &lt; 0.01) the other groups in regard to litter sizes at birth and weaning. Differences among treatments were not significant for the number of females mated, or whelped of those exposed to males, kit survival from birth to weaning, or rectal temperature. It was concluded that 1.5% kelp supplementation had beneficial effects on survival rate of adult mink and litter size.

Molecular epidemiology of Aleutian mink disease virus from fecal swab of mink in northeast China

BackgroundAleutian mink disease parvovirus (AMDV) causes Aleutian mink disease (AMD), which is a serious infectious disease of mink. The aim of this study was to get a better understanding of the molecular epidemiology of AMDV in northeast China to control and prevent AMD from further spreading. This study for the first time isolated AMDV from fecal swab samples of mink in China.ResultsA total of 157/291 (54.0%) of the fecal swab samples were positive for AMDV. Of these, 23 AMDV positive samples were randomly selected for sequence alignment and phylogenetic analysis based on the acquired partial fragments of VP2 gene with the hypervariable region. Comparative DNA sequence analysis of 23 AMDV isolates with a reference nonpathogenic (AMDV-G) strain revealed 8.3% difference in partial VP2 nucleotide sequences. Amino acid alignment indicated the presence of several genetic variants, as well as one single amino acid residue deletion. The most concentrated area of variation was located in the hypervariable region of VP2 protein. According to phylogenetic analysis, the Chinese AMDV strains and the other reference AMDV strains from different countries clustered into three groups (clades A, B and C). Most of the newly sequenced strains were found to form a Chinese-specific group, which solely consisted of Chinese AMDV strains.ConclusionThese findings indicated that a high genetic diversity was found in Chinese AMDV strains and the virus distribution were not dependent on geographical origin. Both local and imported AMDV positive species were prevalent in the Chinese mink farming population. The genetic evidence of AMDV variety and epidemic isolates have importance in mink farming practice.

Multi-host dispersal of known and novel carnivore amdoparvoviruses.

Amdoparvoviruses (family Parvoviridae) are ssDNA viruses that cause an immune complex-mediated wasting syndrome in carnivores. They are multi-host pathogens and cross-species infection is facilitated by the fact that viral entry is mediated by cellular Fc receptors recognizing antibody-coated viruses. We developed a pan-amdoparvovirus PCR and screened tissue samples from 666 wild carnivores (families Felidae, Canidae, and Mustelidae) from Newfoundland or Labrador (Canada) and molecularly characterized the identified strains. Fifty-four out of 666 (8.1%) animals were amdoparvovirus-positive. Infection rate was the highest in American mink (34/47, 72.3%), followed by foxes (Arctic and red foxes, 13/311, 4.2%), lynx (2/58, 3.5%), and American martens (5/156, 3.4%). No virus was detected in samples from 87 coyotes and 17 ermines. Viruses from Newfoundland were classified as Aleutian mink disease virus (AMDV). Mink harvested near AMDV-affected fur farms had higher prevalence (24/24, 100%) than other mink (10/23, 43.5%; P < 0.001) and their viruses were phylogenetically closely related to those from farms, while most viruses from other mink were in other clades. Strains from three foxes and two lynx were highly related to mink strains. This proves that farms disperse AMDV that subsequently spreads among wild mink (maintenance host) and transmits to other spillover carnivore hosts. In Labrador two novel viruses were identified, Labrador amdoparvovirus 1 (LaAV-1) found in foxes (9/261, 3.5%) and martens (5/156, 3.4%), and LaAV-2 found in one fox (0.4%). LaAV-1 fulfills all requirements to be classified as a novel species. LaAV-1 was most similar to viruses of mink and skunks (AMDV and skunk amdoparvovirus (SKAV)) while LaAV-2 was more closely related to other viruses infecting canids. LaAV-1 capsid proteins were almost indistinguishable from those of AMDV in some regions, suggesting that LaAV-1 could be a virus of mustelids that can infect foxes. While intensive farming practices provide occasions for inter-species transmission in farms, niche overlap or predation could explain cross-species transmission in the wild, but competition among sympatric species reduces the chances of direct contacts, making this an infrequent event. Pan-amdoparvovirus detection methods in wide epidemiological investigations can play a crucial role in defining amdoparvoviral ecology and evolution and discovering novel viruses.

Propagation of the Aujeszky’s disease virus isolate in continuous cell lines

The objective of this study was to investigate the cultural properties of Aujeszky's disease virus isolate and the optimization of the conditions of obtaining virus-containing material in the most prospective cell cultures. As a result, it was established that the study virus isolate is highly pathogenic for laboratory animals: rabbits, guinea pigs, and rats. The following continuous cell lines were sensitive to the virus isolate: MDBK, Taurus-1 and ВНК-21. CPE (cytopathogenic effect of viruses) nature was similar in different cell cultures. The ability for propagation in cultures of continuous cell lines of the test virus isolates reached the maximum value to 4-6 passage, equaling to 6.85-7.65 lgTCD50/cm3. The obtained results show that the dynamics of the virus isolate dynamics had no significant differences, and following the adaptation, the virus isolate maintained the stable inherent propagation activity level throughout the study. As the study result, the most sensitive cell culture Taurus-1 was selected at passaging, with the highest virus accumulation observed, the infection activity was 7.55±0.12 lg TCD50/cm3. The presence of Aujeszky's disease virus was confirmed by the bioassay and the other laboratory methods, including PCR (polymerase chain reaction). The PCR method allowed defining the virus genome in the virus-containing cultural and organ and tissue material. PCR allowed amplifying the target gene fragment with the size of 194 base pairs (b.p.) in samples containing Aujeszky's disease virus DNA. No non-specific reactions were observed; PCR products did not exhibit the expected size

Dose response of black American mink to Aleutian mink disease virus.

IntroductionAleutian mink disease virus (AMDV) causes a serious health problem for mink globally. The disease has no cure nor an effective vaccine and selection for tolerance using antibody titer is adopted by many mink farmers. The objective of this study was to investigate the effects of various doses of a local AMDV isolate on the response of black American mink to infection with AMDV.MethodsEight black American mink were each inoculated intranasally with 0.5 mL of eight serial 10‐fold dilutions (100 to 10−7) of a 10% spleen homogenate containing a local AMDV isolate. Blood samples were collected on days 0, 20, 35, 56, 84, 140, and 196 postinoculation (dpi). Anti‐AMDV antibodies and viral DNA were tested by counter‐immunoelectrophoresis (CIEP) and PCR, respectively. Animals that were PCR or CIEP positive at 196 dpi (n = 41) were killed at 218 dpi, and samples of blood and seven organs were tested by CIEP and PCR.ResultsAntibody production persisted in all seroconverted mink until the termination of the experiment, whereas 71.1% of the mink showed short‐lived viremia. Significant associations were observed between inoculum dose and the incidence of viremia until 84 dpi which disappeared thereafter, whereas associations between inoculum dose and the incidence of seropositive mink were significant on all sampling occasions. Antibody titer at 218 dpi significantly decreased with decreasing inoculum dose. AMDV DNA was detected in the bone marrow, lymph nodes, and spleen samples of almost all mink inoculated at every dose but was not detected in other organs of some mink.ConclusionsCIEP is more accurate than PCR for detecting AMDV infection in mink. Using antibody titer in naturally infected mink may not be accurate for the identification of tolerant mink.