Clinical syndromes associated with the circulation of multiple serotypes of bluetongue virus in dairy cattle in Israel.

From 2008 to 2011, seven distinct bluetongue virus (BTV) serotypes (BTV-2, BTV-4, BTV-5, BTV-8, BTV-15, BTV-16 and BTV-24) have been identified to be circulating in diseased sheep and cattle in Israel. This paper describes the array of clinical manifestations caused by BTV in cattle in Israel. Each set of clinical manifestations has been categorised as a syndrome and six distinct clinical syndromes have been observed in dairy cattle: 'footrot-like syndrome', 'sore nose syndrome', 'subcutaneous emphysema syndrome', 'red/rough udder syndrome', 'bluetongue/epizootic haemorrhagic disease systemic syndrome' and 'maladjustment syndrome'.

Epizootic heamorragic disease.

Epizootic haemorrhagic disease (EHD) is an infectious non-contagious viral disease transmitted by insects of the genus Culicoides which affects wild and domestic ruminants. The causative agent, the epizootic haemorrhagic disease virus (EHDV), belongs to the family Reoviridae, genus Orbivirus and shares many morphological and structural characteristics with the other members of the genus such as bluetongue, African horse sickness and equine encephalosis viruses. In recent years EHD outbreaks have been reported in countries bordering the European Union. They caused disease in cattle and severe repercussion on the livestock industry of the affected countries. In the light of recent European bluetongue epizootic these events pose an increasing threat to the European Union. This review includes the most recent information regarding the virus and the disease as well as tools for its diagnosis and control. It is our conviction that more attention should be drawn to both EHDV and the disease itself in order to fulfil all these gaps and not to be unprepared in case future possible incursions.

No evidence for involvement of sheep in the epidemiology of cattle virulent epizootic hemorrhagic disease virus.

Epizootic hemorrhagic disease virus (EHDV) is an Orbivirus. While not previously considered as an important disease in cattle, several EHDV serotypes (EHDV-6 and 7) have recently been implicated in disease outbreaks. The involvement of sheep in the epidemiology of EHDV is still not understood. In this study we compared the prevalence of antibodies to EHDV and bluetongue virus (BTV) in sheep to their prevalence in cattle after an outbreak of EHDV that occurred in Israel during 2006. Sixty-six sheep and lambs scattered in seven herds were compared to 114 cows and calves scattered in 13 dairy cattle herds, matched to the sheep herds by location. While antibody prevalence to EHDV was high in cattle (35.2% within the outbreak zone) no evidence of exposure to EHDV was found in sheep (p<0.0001). Antibodies to BTV were apparent in both cattle and sheep though in the former it was significantly higher (63.2%, 16.7% respectively, p<0.0001), suggesting higher exposure of cattle to biting Culicoides midges. Taken together, these results imply that sheep have a negligible role in the epidemiology of EHDV.

Description of the pathology of a gazelle that died during a major outbreak of foot-and-mouth disease in Israel.

Naturally occurring foot-and-mouth disease (FMD) in wildlife is a relatively mild condition but occasionally it can be devastating as has been documented in impala in South Africa and in mountain gazelles in Israel. This report describes pathological changes in an adult male gazelle with FMD from an outbreak in the Nature Reserve of Ramot-Issachar region and the lower Galilee in Israel. The outbreak was characterised by the malignant form of the disease, which is uncommon among domestic animals. Lesions observed included, ulceration in the oral cavity, oesophagus and ruminal pillars, coronitis, multifocal cardiac necrosis and pancreatic necrosis and inflammation. Pneumonia, caused by Muellerius capillaries was an incidental finding.

Options for decentralized testing of suspected secondary outbreaks of foot-and-mouth disease.

This article reviews the options for use of virus detection techniques for decentralized testing of samples from suspected secondary outbreaks of foot-and-mouth disease (FMD). These options have been expanded by the advent of new tests including disposable lateral flow devices (LFDs) that detect viral proteins and portable RT-PCR equipment that detects viral RNA. LFDs have been developed with similar sensitivity to antigen detection ELISA but with the ability to provide a result 1-30 min after the addition of epithelium or vesicular fluid. Portable RT-PCR platforms are being developed that can detect FMD viral RNA in blood, epithelium or other materials with minimal sample processing and with high sensitivity, in as little as 60 min in some cases. These devices may be used on infected farms as pen-side tests, in regional, local or mobile laboratories, or in National Reference Laboratories (NRL). Advantages and disadvantages of different testing options are considered to inform decisions on the optimal strategies for different national circumstances. Issues include validation and quality control, containment needs, availability of test devices and reagents, the decision tree for declaring an outbreak, training issues and provision of samples for subsequent viral characterization. Tests to confirm the diagnosis of the index case of an outbreak of FMD should continue to be carried out in the NRL.

Appearance of skin lesions in cattle populations vaccinated against lumpy skin disease: statutory challenge.

The ultimate goal of a vaccine is to protect vaccinated animals against re-exposure to the same pathogen and provide sterile immunity. However, a cutaneous clinical manifestation appeared, following re-exposure of cattle that had been vaccinated with the RM65 strain, to LSDV infection during an epidemic in 2006-2007. Four thousand six hundred and seven vaccinated cows entered the study after being re-exposed to LSDV infection. Of them, 513 (11%) presented lumps, and there was a marked difference between the proportions of dairy and feedlot animals that were affected: 146 out of 3517 and 367 out of 1090 (6.6 and 33.7%, respectively). This data suggests that the potency of the vaccine need to be re-assessed for beef cattle.

Temporospatial clustering of foot-and-mouth disease outbreaks in Israel and Palestine, 2006-2007.

Foot-and-mouth disease (FMD) is endemic to the Middle East and there is a perception that political instability and limited resources have led to the uncontrolled circulation of FMD virus throughout the region. Certain aspects of FMD epidemiology in the Middle East remain unknown. The goal of this study was to identify the geographical location, temporal extent and direction of spread of clusters of 70 FMD outbreaks reported in Israel and Palestine from February 4, 2006, through July 15, 2007. The space-time permutation model of the scan statistic test detected nine significant (P < 0.1) clusters. Significant (P < 0.05) direction of spread was identified in four of the nine clusters. The Gaza Strip, where no outbreaks were reported, or a nearby location, seemed to be the origin of a cluster of outbreaks located in Hadarom (April 2007); a cluster of outbreaks centered in West Bank (February 2006) may be linked with spread from Northern Israel; a cluster in Hazafon (January 2007) seemed to have originated from nearby the Jordan borders; and a cluster located in Northern Hazafon was likely related to areas next to the Lebanon and Syrian borders. The association between the clusters in West Bank and earlier Israeli samples and between the cluster in Hazafon and Jordan was also supported (P < 0.05) by phylogenetic analysis of samples collected from the outbreaks. These results suggest that the FMD outbreaks reported in Israel and Palestine in 2006 and 2007 were likely a consequence of different epidemics associated with the circulation and spread of FMD virus strains from different regions of the Middle East.

The NSP immune response of vaccinated animals after in-field exposure to FMDV.

The aim was to examine the immune response (IR) to non-structural proteins (NSPs), in order to assess the validity of the detection of antibodies to NSPs as a means of diagnosing foot and mouth disease (FMD infection) infection when vaccinated populations are in close contact with clinically sick animals. The study was performed during FMD outbreaks in Israel in January 2004; the IR was examined in vaccinated dairy and feedlot cattle herds under natural field exposure to FMDV, and in vaccinated and unvaccinated sheep flocks. During the 2004 outbreaks, clinical signs were age-related and were noted only among imported calves, although they had been vaccinated; such signs were not found among the local dairy cattle populations. The NSP IR among the feedlot cattle that had been vaccinated more than 4 months prior to the in-field exposure was 86%, compared with only 30% among those feedlot cattle that had received one dose of vaccine less than 4 months before the field exposure. The prevalence of NSP IR indicates that animals vaccinated once, less than 4 months prior to exposure, were clinically resistant to FMDV infection, although possibly still susceptible to subclinical infections, whereas those vaccinated more than 4 months prior to the in-field exposure presented clinical manifestations. This situation is unlikely to occur among repeatedly vaccinated livestock; these remained refractory to FMD exposure, as reflected in the absence of clinical manifestations and a relatively low prevalence of NSP IR compared with that in imported calves.

Foot-and-mouth disease non-structural protein serology in cattle: use of a Bayesian framework to estimate diagnostic sensitivity and specificity of six ELISA tests and true prevalence in the field.

The diagnostic performance of six foot-and-mouth disease (FMD) assays for detection of antibodies to the non-structural proteins (NSP) of the FMD virus (FMDV) was estimated using a Bayesian analysis on field sera from cattle of unknown infection status originating from post-FMDV outbreak situations in Israel and Zimbabwe. Estimations of the disease prevalence in both populations were also obtained. The diagnostic sensitivity estimates did not differ between both field studies, although overall Bayesian estimates were markedly higher than those previously reported based on sera from comparable experimentally infected (vaccinated) cattle populations. All NSP-based assays demonstrated a lower diagnostic specificity when applied to the Zimbabwean sera compared to both published specificities and similar Bayesian specificity estimates derived for the Israeli dataset. In Israel, the disease prevalence was estimated at 23.9% (95% credibility interval: 19.5-28.8%), whereas 65.4% (59.0-72.5%) was found in Zimbabwe. The need for reliable diagnostic test performance estimates and the benefits of Bayesian analysis in obtaining them are also addressed.

Comparative evaluation of six ELISAs for the detection of antibodies to the non-structural proteins of foot-and-mouth disease virus.

To validate the use of serology in substantiating freedom from infection after foot-and-mouth disease (FMD) outbreaks have been controlled by measures that include vaccination, 3551 sera were tested with six assays that detect antibodies to the non-structural proteins of FMD virus. The sera came from naïve, vaccinated, infected and vaccinated-and-infected animals; two-thirds from cattle, the remainder from sheep and pigs. The assays were covariant for sensitivity, but not necessarily for specificity. A commercial kit from Cedi-diagnostics and an in-house assay from IZS-Brescia were comparable to the NCPanaftosa-screening index method described in the Diagnostic Manual of the World Animal Health Organisation. Using these three tests the specificity and sensitivity for the detection of carriers in vaccinated cattle approaches or exceeds 99% and 90%, respectively.

VP2-segment sequence analysis of some isolates of bluetongue virus recovered in the Mediterranean basin during the 1998-2003 outbreak.

The complete nucleotide sequences of the VP2 segments of bluetongue virus (BTV) isolates recovered from Italy, Greece and Israel, from 1998 to 2003, were determined. Phylogenetic analysis of these sequences, those from related viruses and the South African vaccine strains, were used to determine the probable geographic origin of BTV incursions into Italy. Results indicated that viruses from each of the four serotypes isolated in Italy (2, 4, 9 and 16) possibly had a different origin. Analysis of the bluetongue virus serotype 2 (BTV-2) isolates gave evidence that this serotype probably moved from Tunisia. BTV-4 results showed probable incursion from the southwest and not from Greece or Israel. BTV-9 isolates clearly have an eastern origin (most probably Greece), whereas BTV-16 isolates are indistinguishable from the BTV-16 live attenuated vaccine strain. The phylogenetic findings were supported by polyacrylamide gel electrophoresis (PAGE) analysis of the complete amplified genome of each isolate except for BTV-16 Italian field isolate, which showed a slightly different PAGE profile. A combination of the complete VP2 sequencing and PAGE analysis of complete genomes, allowed not only phylogenetic analysis, but also vaccine detection and assessment of reassortment events.

The detection of an unidentified type of adenovirus in the stools of calves with weak calf syndrome by use of a commercial kit designed for the detection of human adenoviruses.

An outbreak of polyarthritis in newborn calves in a large collective dairy herd was characterized by intra-articular blood-tinged synoviae, blood tainted faeces and massive subcorneal haemorrhages. Faecal samples from eight clinical newborn cases, 10 from unrelated dairy farms and 10 faecal samples from healthy calves were examined by the Rida Quick rotavirus/adenovirus-combi test . A specific adenovirus antigen precipitin-line was seen in the reaction in all the faecal samples from the diseased calves (n = 8), while all the others (n = 20) were negative. In addition, the same positive reaction was noted when one aqueous humor and two synovial samples were tested with this kit. Several other enteropathogens were found sporadically, but no conclusive significance could be attributed to their presence. Bovine viral diarrhoea and infectious bovine rhinothracheitis viruses as well as Chlamydia spp. and Mycoplasma spp. were not involved in this episode.

Serological and clinical evidence of a teratogenic Simbu serogroup virus infection of cattle in Israel, 2001-2003.

During the last 35 years, two major outbreaks of Akabane virus (AKAV) infection were recorded in cattle in Israel in 1969/1970 and 2002/2003. Congenital malformations of calves characterised by the appearance of an arthrogryposis and hydranencephaly syndrome first appeared in Israel in 1969. Based on epidemiological, clinical, pathological, histopathological and serological data, this syndrome was strongly correlated with seroreactivity to AKAV, a member of the Bunyaviridae, Simbu serogroup. In February 2002, the first cases of 'blind newborn calves' (BNC) were observed on farms located in the northern valleys of Israel. Microtitre serum neutralisation (SN) tests of serum from malformed calves and their dams were conducted using Akabane and Aino viruses (AINOV). The first SN test was performed at the reference laboratory of the Clinical Virology Section, Kyushu Research Station, National Institute of Animal Health, Kagoshima, Japan. The clear-cut findings of seroreactivity to AKAV by cattle located in the affected zone, in contrast to negative findings in cattle from unaffected farms (87% and 3.7%, respectively) was indicative of AKAV infection. In contrast, seroreactivity to Aino virus was relatively low in both affected and non-affected areas during the 2002 outbreak. In order to establish Israeli laboratory standards for Simbu serogroup diagnosis, 57 serum samples tested by the Japanese laboratory were retested by SN in Israel. An almost complete homology (96.5%) was found between the two SN panels of sera (kappa = 0.92). SN and ELISA kits enabled the surveillance of this arbovirus epidemic in the second consecutive year (2003). Moreover, AKAV was identified in trapped midges by hemi-nested PCR and real-time PCR. With these techniques, the geographical limits of the BNC epidemic that appeared in some areas of Israel was identified for the first time and was recorded in the Arava Rift Valley, 400 km south of the epicentre of the 2002 outbreak. The reintroduction of AKAV into this region, together with some evidence of AINOV activity and epidemics of bluetongue (BT) in the southern parts of Europe and the eastern Mediterranean, and renewed outbreaks of West Nile virus infection in Israel, Italy and southern France, are all evidence of the potential spread of arbovirus activity into southern Europe from the Mediterranean Basin.

VP2 gene sequence analysis of some isolates of bluetongue virus recovered in the Mediterranean Basin during the 1998-2002 outbreak.

Since 1998, five serotypes of bluetongue virus (BTV), BTV-1, BTV-2, BTV-4, BTV-9 and BTV-16, have been reported in countries surrounding the Mediterranean Basin. Preliminary data on the sequencing analysis of the VP2-genes of BTV isolates recovered during the 1998-2002 epizootic of BT in Italy, Greece and Israel were studied. The VP2-genes of the Italian BTV-2 and BTV-9, Greek BTV-4 and BTV-9, Israeli BTV-4 and BTV-16 and South African BTV-2, BTV-4, BTV-9 and BTV-16, together with those of their corresponding South African serotype reference and vaccine strains, were cloned and the sequences of their terminal ends determined. These sequences, as well as those of all BTV VP2-gene sequences currently available on GenBank, were used to compile a phylogenetic tree to determine the probable geographic origins of the BTV incursions into Europe. The Italian isolates included in this study were from different regions, animal hosts and years (2000-2002). The results demonstrated that sequencing of the terminal end of the VP2-gene of BTV can be used for topotyping. According to the phylogenetic analysis, the Italian BTV-2 and BTV-9 isolates were stable across all species, irrespective of geographic origin and year of isolation. The sequencing data of the Italian isolates were identical to those of a BTV-2 isolate from Corsica. There was 97% homology between the Italian and Corsican BTV-2 isolates and the BTV-2 vaccine and reference isolates from South Africa. Italian BTV-9 isolates were also identical to the Greek BTV-9 isolates (99% homology). Surprisingly these BTV-9 isolates had only 67% homology with the reference BTV-9 isolate from South Africa. Conversely, BTV-9 field isolates from Australia and elsewhere in Europe had 89% homology with the Italian isolate at the nucleic acid level. Greek and Israeli BTV-4 isolates were almost identical (98% homology) and shared a 90% homology with the BTV-4 South African reference and vaccine strains. Israeli BTV-16 and South African BTV-16 reference strains were also similar. From these results, it may be concluded that Italian and Corsican BTV-2, Israeli and Greek BTV-4, and South African and Israeli BTV-16 had a common origin. The Greek BTV-9 isolate had more than 99% homology with the isolates from Italy, indicating these isolates to have had a common origin. The European BTV-9 isolates, grouped as 'eastern isolates', were more similar to the Australian isolates than to the South African reference strains.

Application of diagnostic procedures to epidemiological situations with special reference to arboviral infections.

The rationale behind the methodology employed to investigate a new disease that appears in an area hitherto unaffected is fundamentally different from that applied in an endemic disease situation. Special consideration must also be given to disease agents that appear and reappear at cyclical intervals. The authors present three separate approaches applicable in three different epidemiological situations. Each one may be used initially to determine the identity of the causal agent and the nature of the disease. These approaches, although described in a stepwise (flowchart) manner, are not meant to be applied rigidly, but rather should serve as a guideline for investigators. Special consideration is given to situations in which disease appears intermittently, using a sentinel model. Although this latter approach is expensive and time-consuming, it can yield excellent and reliable results when applied correctly.