Surveillance of West Nile virus in equids in eastern Austria

Dissertation - Veterinärmedizinische Universität Wien - 2021 Aus rechtlichen Gründen sind nicht alle Teile dieser Arbeit frei zugänglich. Der Zugriff auf den elektronischen Volltext ist auf Angehörige der Veterinärmedizinischen Universität Wien beschränkt. Bitte einloggen!West Nil Virus (WNV) ist ein durch Stechmucken ubertragenes Flavivirus mit nahezu weltweiter Verbreitung. Es zirkuliert naturlicherweise zwischen Stechmucken und Vogeln. Menschen und Pferde sind Endwirte, die nach dem Stich infizierter Mucken erkranken, das Virus aber nicht mehr weitergeben konnen. Erste auf WNV zuruckzufuhrende neurologische Krankheitsfalle beim Pferd traten 2016 auf. In dieser Dissertation werden die ersten sieben Falle von WNV Encephalomyelitis in Osterreich beschrieben. In zusatzlich durchgefuhrten entomologischen Untersuchungen wurden WNV-postive Steckmucken im Stall des Index-Falls gefunden und 2 von 14 Kontaktpferden seropositiv getestet. Das Ziel dieser Dissertation war die Flavivirus-Seropravalenz und WNV RNA-Pravalenz in der osterreichischen – vor allem Ost-osterreichischen – Pferdepopulation festzustellen und Risikofaktoren fur Seropositivitat zu identifizieren. Eine Querschnitt Pravalenz-Studie mit Riskofaktor-Analyse umfasste eine willkurliche Stichprobe und Proben von hospitalisierten Pferden aus Ost-Osterreich aus dem Jahre 2017. PferdebesitzerInnen und -versorgerInnen ubermittelten den Equidenpass und fullten einen Fragenbogen aus. Pferdesera wurden mittels RT-qPCR auf Vorliegen von WNV-RNA und mittels kommerzieller IgG und IgM ELISA Kits (ID Vet, Grabels) auf Flavivirus-spezifische Antikorper untersucht. ELISA-reaktive Proben wurden weiter mittels Plaque Reduction Neutralisationstest analysiert um zwischen Infektionen mit WNV, Fruhsommer Meningoenzephalitisvirus (FSME) und Usutuvirus (USUV) zu differenzieren. Logistische Regression wurde als statistisches Modell genutzt. 348 Equiden (334 Pferde, 14 Esel) wurde beprobt. Die Seropravalanz betrug nach Ausschluss geimpfter Tiere 5,3% fur WNV (autochthone Infektionen 1,2%), 15,5% fur TBEV und 0% fur USUV. Die FSME-Seroprevalenz in Eseln war 28.6%, Keine IgM-Antikorper oder WNV RNA wurden detektiert. Auch keine Risikofaktoren wurden identifiziert. Die WNV Seropravalanz in Osterreich liegt niedriger als jene im Nachbarland Slowakei (8,3%). Die FSME Seropravalenz ist hoch im Vergleich zu einer Studie aus Deutschland (2,9%) und der Slowakei (3,4%). Equiden stellen somit sentinels fur Flavivirus-Aktivitat dar. Die beispiellos hohe WNV-Aktivitat in Europa im Jahre 2018 beweist wie wichtig ein gesamt-osterreichisches FlavivirusUberwachungsprogramm ist.Dissertation - University of Veterinary Medicine Vienna - 2020 The full text is only available to university members. Please log in!West Nile virus (WNV) is an arthropod-borne flavivirus with near global distribution, circulating between mosquitoes and birds. Humans and horses are dead-end hosts, susceptible to infection by mosquitoes. The emergence of WNV in Austria manifested in the diagnosis of the first equine case of West Nile neurologic disease (WNND) in 2016 and seasonal outbreaks similar to central and southern Europe. The first seven confirmed cases of equine WNND in Austria are described in this thesis. In addition, flavivirus seroprevalence and WNV RNA prevalence in Austrian equines and mosquitos were identified. Furthermore, serologic evidence of WNV infection was found in 2/14 in-contact horses at the stable which was the location of the Austrian index case. We aimed to investigate flavivirus seroprevalence and WNV RNA prevalence in Austrian equines and to identify risk factors for seropositivity. This cross-sectional prevalence study with risk factor analysis included convenience samples and samples from hospitalized equines in eastern Austria collected during 2017. Owners and caretakers provided the equine passports and completed questionnaires. Sera were analysed for WNV RNA with RT-qPCR and for flavivirus-specific antibodies with commercial IgG and IgM ELISAs (ID Vet, Grabels). Samples which reacted to ELISA were further analysed with 80 % plaque-reduction neutralisation tests to differentiate WNV, tick-borne encephalitis virus (TBEV) and Usutu virus (USUV) infections. Logistic regression was used for statistical analysis. A total of 348 equines (334 horses, 14 donkeys) were sampled. The seroprevalence values were 5.3 % for WNV (autochthonous infections = 1.2 %), 15.5 % for TBEV, and 0 % for USUV, excluding vaccinated equids. Donkey TBEV seroprevalence was 28.6 %. No IgM and WNV RNA were detected and no risk factors were identified. The WNV seroprevalence in Austria is lower than in neighbouring Slovakia (8.3 %). The TBEV seroprevalence is high compared to non-outbreak situations reported in Germany (2.9 %) and Slovakia (3.4 %). Equids can serve as sentinels for flavivirus activity. An increase in flavivirus-related disease and the importance of continuous surveillance are highlighted by the unprecedented magnitude of the 2018 European WNV outbreak

A pressure algometer is a useful tool to objectively monitor the effect of diagnostic palpation by a physiotherapist in Warmblood horses

There is a lack of scientific evidence for objective evaluation of neck and back musculoskeletal sensitivity in horses, although pressure algometry has been described as an objective tool to quantify musculoskeletal responses by mechanical nociceptive threshold (MNT) values. This study aimed to evaluate the use of pressure algometry for objectively quantifying the effect of diagnostic palpation applied by physiotherapists on the musculoskeletal function of the equine neck and back. The inter-examiner repeatability of animal physiotherapists was tested, and their subjective clinical scores for the vertebral column area were objectively compared with MNT values measured at the same locations to investigate the potential clinical implementation of the pressure algometer in daily equine rehabilitation practice. Six adult Dutch Warmblood riding school mares were randomly assigned to an experimental or a control group. The MNT of all horses was measured on 35 pre-defined sites on the vertebral column in the morning and in the evening of the same day. In the experimental group (n = 3), neck and back surface "temperature", "pain", "muscle tone", and passive "mobility" were scored through palpation by three certified physiotherapists and related to MNT measurements at the same vertebral column locations. Agreement between the physiotherapists was determined from Spearman's rank correlation coefficients (P<.05). These correlation coefficients showed a significant agreement between the scores of individual physiotherapists and with objective MNT measurements. The three physiotherapists agreed best in their subjective gradings of "pain", but less for "temperature" and "muscle tone", and least for "mobility". There was also a significant difference in MNT between individual horses. The physiotherapeutic diagnostic intervention did not significantly alter the MNT of the experimental group compared with the control group. There was a significant difference, however, between morning (7.4 kg/cm(2)) and evening (6.9 kg/cm(2)) MNT-measurements within the combined group (n = 6, P<.05). In conclusion, a pressure algometer proved to be a useful tool to objectively monitor the palpation of individual Warmbloods by individual physiotherapists. The correlation of their scores to the objective MNT measurements elucidated that there were differences on which scale ("pain", "temperature", "muscle tone", "mobility") they merely relied upon in their palpation. Significant effects of physiotherapeutic diagnostic palpation on MNT, however, were not found. The lower MNT of the horses at the second trial in the evening could be a sensitization of the measurement location because of bruising, a learning effect of the horses, or a diurnal fluctuation. The use of pressure algometry has both a potential to quantify clinical neck and back musculoskeletal sensitivity in horses possibly leading to dysfunction, as veil as to objectively evaluate treatment results. Repeated measurements on the same day and on the same location along the vertebral column may influence absolute MNT values. The algometer can be used with success provided that the operator has proper and frequent training

West Nile Virus and Tick-Borne Encephalitis Virus Are Endemic in Equids in Eastern Austria

The emergence of West Nile virus (WNV) and Usutu virus (USUV) in addition to the autochthonous tick-borne encephalitis virus (TBEV) in Europe causes rising concern for public and animal health. The first equine case of West Nile neuroinvasive disease in Austria was diagnosed in 2016. As a consequence, a cross-sectional seroprevalence study was conducted in 2017, including 348 equids from eastern Austria. Serum samples reactive by ELISA for either flavivirus immunoglobulin G or M were further analyzed with the plaque reduction neutralization test (PRNT-80) to identify the specific etiologic agent. Neutralizing antibody prevalences excluding vaccinated equids were found to be 5.3% for WNV, 15.5% for TBEV, 0% for USUV, and 1.2% for WNV from autochthonous origin. Additionally, reverse transcription quantitative polymerase chain reaction (RT-qPCR) was performed to detect WNV nucleic acid in horse sera and was found to be negative in all cases. Risk factor analysis did not identify any factors significantly associated with seropositivity

Active equine parvovirus‐hepatitis infection is most frequently detected in Austrian horses of advanced age

Background: Equine parvovirus-hepatitis (EqPV-H) research is in its infancy. Information regarding prevalence, geographical distribution, genetic diversity, pathogenesis and risk factors enhances understanding of this potentially fatal infection. Objectives: Determining the prevalence of EqPV-H in Austrian equids. Investigating factors increasing probability of infection, liver-associated biochemistry parameters, concurrent equine hepacivirus (EqHV) infection and phylogenetic analysis of Austrian EqPV-H variants. Study design: Cross-sectional study. Methods: Sera from 259 horses and 13 donkeys in Austria were analysed for anti-EqPV-H VP1-specific antibodies by luciferase immunoprecipitation system (LIPS) and EqPV-H DNA by nested polymerase chain reaction (PCR). Associations between infection status, sex and age were described. Glutamate dehydrogenase (GLDH), gamma-glutamyl transferase (GGT), bile acids and albumin concentrations were compared between horses with active infection and PCR-negative horses. PCR targeting partial EqPV-H NS1 was performed and phylogenetic analysis of Austrian EqPV-H variants was conducted. Complete coding sequences (CDS) of four Austrian variants were determined by next-generation sequencing (NGS) and compared with published sequences. Results: Horses' EqPV-H seroprevalence was 30.1% and DNA prevalence was 8.9%. One horse was co-infected with EqHV. Significantly, higher probability of active EqPV-H infection was identified in 16- to 31-year-old horses, compared with 1- to 8-year-old horses (P = 0.002; OR = 8.19; 95% CI = 1.79 to 37.50) and 9- to 15-year-old horses (P = 0.03; OR = 2.96; 95% CI = 1.08 to 8.17). Liver-associated plasma parameters were not significantly different between horses with active infection and controls. Austrian EqPV-H variants revealed high similarity to sequences worldwide. No evidence of EqPV-H was detected in donkeys. Main limitations: Equids' inclusion depended upon owner consent. There was only one sampling point per animal and the sample of donkeys was small. Conclusions: EqPV-H antibodies and DNA are frequently detected in Austrian horses, without associated hepatitis in horses with active infection. The risk of active EqPV-H infection increases with increasing age. Phylogenetic evidence supports close relation of EqPV-H variants globally, including Austrian variants

No Evidence of Mosquito Involvement in the Transmission of Equine Hepacivirus (Flaviviridae) in an Epidemiological Survey of Austrian Horses

Prevalence studies have demonstrated a global distribution of equine hepacivirus (EqHV), a member of the family Flaviviridae. However, apart from a single case of vertical transmission, natural routes of EqHV transmission remain elusive. Many known flaviviruses are horizontally transmitted between hematophagous arthropods and vertebrate hosts. This study represents the first investigation of potential EqHV transmission by mosquitoes. More than 5000 mosquitoes were collected across Austria and analyzed for EqHV ribonucleic acid (RNA) by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Concurrently, 386 serum samples from horses in eastern Austria were analyzed for EqHV-specific antibodies by luciferase immunoprecipitation system (LIPS) and for EqHV RNA by RT-qPCR. Additionally, liver-specific biochemistry parameters were compared between EqHV RNA-positive horses and EqHV RNA-negative horses. Phylogenetic analysis was conducted in comparison to previously published sequences from various origins. No EqHV RNA was detected in mosquito pools. Serum samples yielded an EqHV antibody prevalence of 45.9% (177/386) and RNA prevalence of 4.15% (16/386). EqHV RNA-positive horses had significantly higher glutamate dehydrogenase (GLDH) levels (p = 0.013) than control horses. Phylogenetic analysis showed high similarity between nucleotide sequences of EqHV in Austrian horses and EqHV circulating in other regions. Despite frequently detected evidence of EqHV infection in Austrian horses, no viral RNA was found in mosquitoes. It is therefore unlikely that mosquitoes are vectors of this flavivirus

Active equine parvovirus-hepatitis infection is most frequently detected in Austrian horses of advanced age.

ckground: Equine parvovirus-hepatitis (EqPV-H) research is in its infancy. Information regarding prevalence, geographical distribution, genetic diversity, pathogenesis and risk factors enhances understanding of this potentially fatal infection. Objectives: Determining the prevalence of EqPV-H in Austrian equids. Investigating factors increasing probability of infection, liver-associated biochemistry parameters, concurrent equine hepacivirus (EqHV) infection and phylogenetic analysis of Austrian EqPV-H variants. Study design: Cross-sectional study. Methods: Sera from 259 horses and 13 donkeys in Austria were analysed for anti-EqPV-H VP1-specific antibodies by luciferase immunoprecipitation system (LIPS) and EqPV-H DNA by nested polymerase chain reaction (PCR). Associations between infection status, sex and age were described. Glutamate dehydrogenase (GLDH), gamma-glutamyl transferase (GGT), bile acids and albumin concentrations were compared between horses with active infection and PCR-negative horses. PCR targeting partial EqPV-H NS1 was performed and phylogenetic analysis of Austrian EqPV-H variants was conducted. Complete coding sequences (CDS) of four Austrian variants were determined by next-generation sequencing (NGS) and compared with published sequences. Results: Horses' EqPV-H seroprevalence was 30.1% and DNA prevalence was 8.9%. One horse was co-infected with EqHV. Significantly, higher probability of active EqPV-H infection was identified in 16- to 31-year-old horses, compared with 1- to 8-year-old horses (P = 0.002; OR = 8.19; 95% CI = 1.79 to 37.50) and 9- to 15-year-old horses (P = 0.03; OR = 2.96; 95% CI = 1.08 to 8.17). Liver-associated plasma parameters were not significantly different between horses with active infection and controls. Austrian EqPV-H variants revealed high similarity to sequences worldwide. No evidence of EqPV-H was detected in donkeys. Main limitations: Equids' inclusion depended upon owner consent. There was only one sampling point per animal and the sample of donkeys was small. Conclusions: EqPV-H antibodies and DNA are frequently detected in Austrian horses, without associated hepatitis in horses with active infection. The risk of active EqPV-H infection increases with increasing age. Phylogenetic evidence supports close relation of EqPV-H variants globally, including Austrian variants

Changes in the population of methicillin-resistant Staphylococcus pseudintermedius and dissemination of antimicrobial-resistant phenotypes in the Netherlands

<p>Methicillin-resistant Staphylococcus pseudintermedius (MRSP), which is often multidrug resistant (MDR), has recently emerged as a threat to canine health worldwide. Knowledge of the temporal distribution of specific MRSP lineages, their antimicrobial resistance phenotypes, and their association with clinical conditions may help us to understand the emergence and spread of MRSP in dogs. The aim of this study was to determine the yearly proportions of MRSP lineages and their antimicrobial-resistant phenotypes in the Netherlands and to examine possible associations with clinical conditions. MRSP was first isolated from a canine specimen submitted for diagnostics to the Faculty of Veterinary Medicine of Utrecht University in 2004. The annual cumulative incidence of MRSP among S. pseudintermedius increased from 0.9% in 2004 to 7% in 2013. MRSP was significantly associated with pyoderma and, to a lesser extent, with wound infections and otitis externa. Multilocus sequence typing (MLST) of 478 MRSP isolates yielded 39 sequence types (ST) belonging to 4 clonal complexes (CC) and 15 singletons. CC71 was the dominant lineage that emerged since 2004, and CC258, CC45, and several unlinked isolates became more frequent during the following years. All but two strains conferred an MDR phenotype, but strains belonging to CC258 or singletons were less resistant. In conclusion, our study showed that MDR CC71 emerged as the dominant lineage from 2004 and onward and that less-resistant lineages were partly replacing CC71.</p

Severe Neurologic Disease in a Horse Caused by Tick-Borne Encephalitis Virus, Austria, 2021

As evidenced by sero-epidemiological studies, infections of horses with the tick-borne encephalitis virus (TBEV) occur frequently in TBEV-endemic areas. However, there are only very few reports of clinical cases. A possible underreporting may be due to a variety of diagnostic challenges. In this study, ELISA and neutralization tests were applied to serum samples. Brain tissue samples were investigated for the presence of nucleic acids of TBEV, Equid alphaherpesvirus 1, Borna disease virus 1, West Nile and Usutu viruses, rustrela virus, as well as Eastern, Western, and Venezuelan equine encephalitis viruses with RT-qPCR, RT-PCR, and qPCR, respectively. TBEV-specific amplification products were subjected to Sanger sequencing. In addition, a direct fluorescent antibody test for rabies was performed. Clinical and patho-histological findings are reported. Using specific RT-qPCR and RT-PCR assays, TBEV nucleic acids were demonstrated in brain tissue samples. Sequencing revealed the Western (formerly Central) European subtype of TBEV as the etiological agent. A high titer of TBEV-specific neutralizing antibodies was found in the serum. RNAscope in situ hybridization revealed TBEV RNA confined to neuronal cell bodies and processes. No other pathogens or nucleic acids thereof could be detected. Diagnostic procedures need to be carried out early after the onset of neurological signs to allow for a final etiological diagnosis of acute TBEV infections in horses