Alphaherpesvirus infections in semidomesticated reindeer: A cross-sectional serological study

Alphaherpesviruses infect a wide range of animal species and cause diseases. Cervid herpesvirus 2 (CvHV-2) was originally isolated from reindeer in Finland but the impact of CvHV-2 infections on reindeer remains unclear. CvHV-2 infection could be partly responsible for calf losses as there are indications that it is associated with abortions and neonatal diseases. Previous serosurveys of reindeer (Rangifer tarandus tarandus) have shown that an alphaherpesvirus is circulating among reindeer in Norway. The aim of the present study was to determine the prevalence of CvHV-2 infection among reindeer in various herding districts in Finnmark, the largest reindeer area in Norway, and to identify factors associated with becoming infected with CvHV-2. A total of 3062 serum samples were tested using an ELISA and a sub-set of samples was further tested using a seroneutralization test. The ELISA revealed that 49% of samples were positive. Extrapolation of the results to the total population (111,350 animals; 66% of the Finnmark reindeer population) showed that the seroprevalence in the population was 48%. Seroprevalence varied from 7.6% to 90.7% between districts and was affected by age, weight and population density. ELISA-positive samples neutralized CvHV-2 at serum dilutions greater than those required for neutralization of bovine herpesvirus type 1. It is concluded that CvHV-2 is endemic throughout the reindeer herding districts of northern Norway.     

Antibody response to experimental Salmonella typhimurium infection in chickens measured by ELISA

An indirect ELISA has been developed to detect Salmonella typhimurium antibodies in chicken sera, using whole bacterial cell protein, flagellar protein or lipopolysaccharide as antigens. In experimental infections high concentrations of S typhimurium-specific IgG persisted after the faecal excretion of S typhimurium had ceased, whereas the specific IgM response was transitory. Some uninfected chickens placed in contact with experimentally infected birds developed high IgG titres in the absence of detectable faecal excretion. Other S typhimurium strains, which varied in their invasive abilities, also induced high titres of IgG. The ELISA allowed chickens infected experimentally with S typhimurium to be differentiated from chickens infected with 10 other serotypes, including S enteritidis. The use of whole blood in place of serum in the ELISA reduced the titres slightly. The storage of serum dried on to filter paper strips for four weeks produced little change in ELISA antibody titre, and the treatment of such strips with phenol or chloroform vapour had little or no effect on the antibody titre.     

Cervids as sentinel‐species for tick‐borne encephalitis virus in Norway ‐ A serological study

Tick‐borne encephalitis virus (TBEV) is the causative agent of tick‐borne encephalitis (TBE). TBEV is one of the most important neurological pathogens transmitted by tick bites in Europe. The objectives of this study were to investigate the seroprevalence of TBE antibodies in cervids in Norway and the possible emergence of new foci, and furthermore to evaluate if cervids can function as sentinel animals for the distribution of TBEV in the country. Serum samples from 286 moose, 148 roe deer, 140 red deer and 83 reindeer from all over Norway were collected and screened for TBE immunoglobulin G (IgG) antibodies with a modified commercial enzyme‐linked immunosorbent assay (ELISA) and confirmed by TBEV serum neutralisation test (SNT). The overall seroprevalence against the TBEV complex in the cervid specimens from Norway was 4.6%. The highest number of seropositive cervids was found in south‐eastern Norway, but seropositive cervids were also detected in southern‐ and central Norway. Antibodies against TBEV detected by SNT were present in 9.4% of the moose samples, 1.4% in red deer, 0.7% in roe deer, and nil in reindeer. The majority of the positive samples in our study originated from areas where human cases of TBE have been reported in Norway. The study is the first comprehensive screening of cervid species in Norway for antibodies to TBEV, and shows that cervids are useful sentinel animals to indicate TBEV occurrence, as supplement to studies in ticks. Furthermore, the results indicate that TBEV might be spreading northwards in Norway. This information may be of relevance for public health considerations and supports previous findings of TBEV in ticks in Norway.     

Evaluation of a novel enzyme-linked immunosorbent assay for detection of antibodies against Salmonella, employing a stable coating of lipopolysaccharide-derived antigens covalently attached to polystyrene microwells.

Polysaccharides derived from Salmonella typhimurium lipopolysaccharide (LPS) representing the O-antigen factors 1, 4, 5, and 12 and the O-antigen factors 6 and 7 from Salmonella choleraesuis LPS were derivatized with the photoreactive compound anthraquinone and subsequently covalently coupled to microtiter polystyrene plates by ultraviolet irradiation. Both polysaccharide antigens could be coupled simultaneously to the same microtiter plate. The coated surface was used in indirect ELISA for the determination of serum antibodies from pigs infected with bacteria of the two Salmonella groups and from uninfected pigs. This ELISA proved itself by having a good long-term durability and a high degree of reproducibility, including low day-to-day variations and low interplate variations. Furthermore, the ELISA showed good specificity and sensitivity when data were compared with the optical density levels of a panel of pig sera as determined by a conventional ELISA on the basis of passive coating of the two Salmonella LPS antigens (the mix-ELISA). The covalent anthraquinone mix-ELISA shows promise as a stable and durable alternative to the existing conventional ELISA for serological surveillance of Salmonella infections in pigs.     

Differences in the seroprevalence of Salmonella spp. in free-ranging and corralled semi-domesticated reindeer (Rangifer tarandus tarandus) in Finland

Serum samples from 1032 semi-domesticated reindeer (Rangifer tarandus tarandus) from Finland were examined for the occurrence of Salmonella-antibodies by use of an indirect ELISA. The majority of samples originated from clinically healthy slaughter reindeer, kept extensively (n = 802; year of sampling: 1996). The remaining samples (n = 230) came from a research herd, permanently kept intensively, with repeated outbreaks of diarrhoea. In this study, 29 of the examined serum samples showed an OD above the determined cut-off. The prevalence in the clinically healthy slaughter reindeer was 0.9%, in the research herd 4.2% in 1996, 10.5% in 1997 and 12.9% in 1998. It must be assumed that the intensive husbandry in the corralled research herd may favour the spreading of infectious agents and eventually outbreaks of crowding diseases in the herd. This investigation is complemented by a review on the occurrence of Salmonella in wild and semi-domesticated cervids.     

Occurrence of Salmonella serotype Typhimurium DT104 on a commercial swine farm before, during, and after depopulation and repopulation

OBJECTIVE: To determine whether depopulation-repopulation could be used to eradicate Salmonella serotype Typhimurium DT104 from a commercial swine farm in the midwestern United States. DESIGN: Observational study SAMPLE POPULATION: A commercial swine farm undergoing depopulation-repopulation to eliminate porcine reproductive and respiratory syndrome virus and Mycoplasma hyopneumoniae. PROCEDURE: Pooled fecal samples, tissue samples, and serum samples were collected from pigs on the farm before and after depopulation-repopulation. When there were no pigs on the farm, environmental swab specimens were collected for bacterial culture. Serum was analyzed for anti-Salmonella antibodies with an indirect ELISA. Salmonella isolates obtained by bacterial culture of fecal, tissue, and environmental samples were characterized by means of serotyping, phage typing, pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility testing. RESULTS: 167 Salmonella isolates representing 9 serotypes were recovered from the farm. Results of PFGE and antimicrobial susceptibility testing suggested that S. Typhimurium DT104 strain was not eradicated from the farm. However, seroprevalence of anti-Salmonella antibodies and the percentage of pooled fecal samples positive for Salmonella spp were significantly decreased following repopulation. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that depopulation-repopulation in conjunction with stringent cleaning and disinfection, attention to biosecurity procedures, control of other diseases, and changes in feed management may reduce the occurrence of, but likely will not eliminate, Salmonella spp in commercial swine herds.     

Serological response of chickens naturally infected with Salmonella typhimurium detected by ELISA.

Four laying flocks of chickens in Britain, each with a history of Salmonella typhimurium infection, were investigated serologically and bacteriologically. Blood samples were taken from identified birds from a single house on each site and sent to the Central Veterinary Laboratory, Weybridge for serological examination using enzyme-linked immunosorbent assays (ELISA) and rapid slide agglutination test (RST) using stained S. pullorum. The identified birds were taken to the local Veterinary Investigation Centre for bacteriological examination. On site A no salmonellae were recovered from birds in the house chosen for serological examination. Of these birds approximately 20% had antibodies to S. typhimurium in ELISA which used either a lipopolysaccharide (LPS) or heat-extract (HE) antigen from S. typhimurium. S. typhimurium was recovered from birds in one other of the four houses on the same site; these birds were not tested serologically. On site B, S. typhimurium was isolated from 8% of the birds examined. Of the total tested serologically, a third to half were seropositive by S. typhimurium ELISA using the LPS and HE antigen respectively. A small proportion of birds was seropositive by S. enteritidis ELISA and RST. No salmonellae were isolated from the other two sites although about 10% of birds tested on site C were seropositive in S. typhimurium ELISA. Cross-reactions were seen between S. typhimurium antigens in the ELISA and experimentally prepared antiserum to S. enteritidis. The S. enteritidis ELISA was generally more specific although cross-reacting antibodies were detected in sera from birds on sites A and B.     

A disc ELISA for the detection of salmonella group D antibodies in poultry

An ELISA using lipopolysaccharide antigens prepared from Salmonella gallinarum and S enteritidis was developed for the serological diagnosis of fowl typhoid and S enteritidis infection in poultry. There was good agreement between the results of the ELISA and conventional serological tests when samples from naturally infected birds and S enteritidis immunised birds were tested. Some cross reactions were observed when serum samples from S typhimurium infected birds were tested by ELISA. Subsequently a disc ELISA, using filter paper discs, was developed to facilitate sampling and testing of poultry. There was good correlation between the results of the disc and serum ELISAs and the test is recommended for the field testing of birds.     

Aspects of bovine herpesvirus-1 infection in dairy and beef herds in the Republic of Ireland

Background

Infection with bovine herpesvirus-1 (BHV-1) causes a wide range of disease manifestations, including respiratory disease and abortion, with world-wide distribution. The primary objective of the present study was to describe aspects of BHV-1 infection and control on Irish farms, including herd-level seroprevalence (based on pooled sera) and vaccine usage.

Methods

The characteristics of a diagnostic indirect BHV-1 antibody ELISA test when used on serum pools were evaluated using laboratory replicates for use in the seroprevalence study. The output from this indirect ELISA was expressed as a percentage positivity (PP) value. A proposed cut off (PCO) PP was applied in a cross-sectional study of a stratified random sample of 1,175 Irish dairy and beef cattle herds in 2009, using serum pools, to estimate herd seroprevalence. The study was observational, based primarily on the analysis of existing samples, and only aggregated results were reported. For these reasons, ethical approval was not required. Bulk milk samples from a subset of 111 dairy herds were analysed using the same ELISA. Information regarding vaccine usage was determined in a telephone survey.

Results

A PCO PP of 7.88% was determined to give 97.1% sensitivity and 100% specificity relative to the use of the ELISA on individual sera giving maximization of the prevalence independent Youden''s index, on receiver operating characteristics analysis of replicate results. The herd-level BHV-1 seroprevalence was 74.9% (95% CI - 69.9%-79.8%), with no significant difference between dairy and beef herds. 95.5% agreement in herd classification was found between bulk milk and serum pools. Only 1.8 percent of farmers used BHV-1 marker vaccine, 80% of which was live while 75% of vaccinated herds were dairy.A significant association was found between herd size (quartiles) and seroprevalence (quartiles).

Conclusions

The results from this study indicate BHV-1 infection is endemic, although BHV-1 vaccines are rarely used, in the cattle population in Ireland.     

Apparent Seroprevalence of Salmonella spp. in Harp Seals in the Greenland Sea as Determined by Enzyme-linked Immunosorbent Assay

An indirect ELISA was developed as a possible tool for surveillance of the seroprevalence of Salmonella spp. in harp seals. This species is hunted for human consumption and thus transmission of disease to humans cannot be excluded. To cover a broad spectrum of serogroups, a mixture of the lipopolysaccharides (LPS) of S. typhimurium and S. choleraesuis was used as the antigen in this pilot study. Chicken anti-harp-seal immunoglobulin horseradish peroxidase conjugate served as the immunoconjugate. Sera from four captive harp seals, which were Salmonella culture-negative and had no clinical or historical evidence of salmonellosis, were used as negative controls. After immunization with an inactivated S. typhimurium vaccine, further sera from these seals were used as positive controls, as no serum from naturally infected animals was available. Serum samples from 93 harp seals caught in the Greenland sea in 1999 were examined, and anti-Salmonella antibodies were found in the samples from two individuals (seroprevalence 2.2%). Although Salmonella has been isolated from other pinniped species, this is the first documentation of Salmonella-seropositive harp seals. This study contributes to the evaluation of the importance of salmonellosis in arctic marine mammals and thus to the prevention of potential outbreaks of this important zoonosis.     

Detection of antibody to Salmonella enteritidis and S typhimurium in the yolk of hens' eggs

Enzyme-linked immunosorbent assays (ELISAs) have been developed to detect IgG antibodies to Salmonella enteritidis and S typhimurium in the yolk of hens' eggs. Better discrimination and more consistent results were obtained between eggs from experimentally infected and uninfected hens by using saline-dilution of yolk rather than chloroform extraction. Threshold absorbance values were determined in three salmonella-free flocks, and on the basis of these results ELISA optical density values greater than 0.25 were considered to be positive for antibodies to salmonella. Four flocks with a history of salmonella infection were examined; three contained birds which were seropositive for S enteritidis by ELISA and from which S enteritidis was isolated, and a large proportion of eggs from these birds contained antibody to S enteritidis. Eggs from the fourth flock had no detectable antibody, although serum antibody was detected in some birds. No salmonellae were isolated from the yolks of the eggs from any of the four flocks.     

Incidence of Salmonella infection in healthy dogs in Gifu Prefecture,Japan

A total of 1,013 feces samples and 8 mesenteric lymphonodus samples obtained from apparently healthy dogs were examined for the incidence of salmonella infection. One strain of S. typhimurium (ST) was isolated from feces of one dog, and S. enteritidis (SE) was isolated from the mesenteric lymphonodus of one dog. Sera obtained from 330 apparently healthy dogs were examined for Salmonella antibodies using an ELISA with heated whole cells of SE and ST. Fifty-one of the 330 serum samples were considered to be positive for salmonella antibodies, including 12 which were SE-positive and 39 which were ST-positive. These results indicate that dogs cause possible environmental problems as Salmonella carriers.     

Seroprevalence and antibiotic sensitivity of serotypes of Salmonella enterica in Greek pig herds

Blood samples were taken from 50 pigs in each of 59 farrow-to-finish production herds and from 40 pigs in each of four of five registered multiplying herds. Samples of feed and faeces were also collected from 17 of the production herds and from the four multiplying herds. The sera were tested for antibodies to Salmonella enterica by the Danish mix-ELISA, and the organisms were isolated, serotyped and sensitivity tested by standard techniques. The average within-herd seroprevalence was 3.4 per cent and at least one pig tested seropositive in 21 of the 59 herds. In the multiplying herds, only a single seroreactor was detected. Salmonellae were isolated from only five of 95 feed samples, from two of the 17 herds sampled, Salmonella tennessee in four of five samples from one herd and an untypable strain in one of five samples from another. Four infected faecal samples were detected in four herds; they harboured Salmonella typhimurium, Salmonella bredeney or Salmonella london. No salmonellae were isolated from the samples of feed and faeces taken from the multiplying herds. The S london and S typhimurium had a low sensitivity to streptomycin, kanamycin and neomycin, and the S typhimurium also had low sensitivity to amoxycillin, ticarcillin, piperacillin, amoxycillin + clavulanic acid, cefalotin and cefoperazone. The other isolates were sensitive to all the antimicrobial agents tested.     

Serological response of chickens to infection with Salmonella gallinarum-S. pullorum detected by enzyme-linked immunosorbent assay.

The serological response to Salmonella pullorum and S. gallinarum infection in chickens was studied with an indirect enzyme-linked immunosorbent assay (ELISA). In broiler chickens, a more virulent strain of S. pullorum produced a significantly lower serum IgG titer than did a less virulent strain. In laying hens, the serum and egg-yolk IgG titers were very similar. In chickens infected with S. gallinarum, high IgG titers persisted for 30 weeks. In chickens reinfected with this strain, each reinfection was followed by transitory increases in IgG lasting no longer than 2 weeks. Serum samples from Brazil taken from a laying flock with evidence of fowl typhoid showed much higher antibody levels than did those from three uninfected flocks. Using lipopolysaccharide as the detecting antigen, infections caused by these salmonellae could be differentiated from those caused by other groups. Incorporation of the appropriate flagella antigen in the ELISA allowed differentiation between infections caused by S. pullorum and S. enteritidis.     

Dynamics of Salmonella contamination in a commercial quail operation.

Control of carcass contamination requires knowledge of the source and dynamics of spread of Salmonella in commercial poultry production. We examined Salmonella contamination at a U.S. commercial quail operation. Pulsed-field gel electrophoresis (PFGE) was used to type isolates in order to trace Salmonella throughout this production environment. During a 6-mo survey, Salmonella serotypes hadar, typhimurium, typhimurium variant Copenhagen, and paratyphi were encountered within this poultry operation. Ninety-four percent of the Salmonella isolated from breeder and production houses and from carcass rinses belonged to Salmonella serotypes typhimurium variant Copenhagen and hadar. There were six distinct S. typhimurium variant Copenhagen genetic types, as identified by PFGE, present within this particular poultry operation. Seventy-nine percent of S. typhimurium variant Copenhagen identified from the environment of the breeder and production houses produced the same PFGE pattern. Thirty-eight percent of S. typhimurium Copenhagen isolated from carcass rinses and the breeder house shared the same PFGE DNA pattern. This study demonstrates the transmission of salmonellae throughout this production environment, from the breeders to their progeny and to the birds ultimately processed for human consumption.     

Some aspects of the epidemiology and control of Salmonella typhimurium infection in outwintered suckler cows.

Two outbreaks of Salmonella typhimurium infections affected outwintered, spring-calving suckler cows in late pregnancy. The infections spread rapidly both within and between groups of stock on the affected farms, with morbidity in the infected groups varying from 14.5 per cent to over 60 per cent, and mortality in adult cattle varying from 0 to 14.3 per cent. Prophylactic measures included the use of antibiotics and killed vaccines against Escherichia coli, Salmonella dublin, S typhimurium, and Pasteurella multocida. In one outbreak, use was also made of a polyvalent serovaccine and hyperimmune serum against E coli, S typhimurium, and S dublin. In both outbreaks no new cases were reported in the affected groups after the administration of the second dose of vaccine, and there was no resurgence of disease on the affected farms within 18 months of the primary outbreaks.     

Prevalence of Bovine Viral Diarrhoea Virus antibodies among the industrial dairy cattle herds in suburb of Mashhad-Iran

Mashhad is a major dairy production in Iran. The subject of this study was to survey the seroprevalence of Bovine Viral Diarrhea Virus (BVDV) infection using an indirect Enzyme-linked immunosorbent assay (ELISA) test in industrial dairy cattle herds in suburb of Mashhad-Iran. Totally, 141 serum samples were tested. None of the herds had been vaccinated against BVDV. Commercial indirect ELISA kit was used. The herds divided to 3 sizes as cow population. They were included: small, medium and large herds. Data were analyzed using Chi-square test. Ninety-seven (68.79%) cows were ELISA seropositive. However, the true BVDV seroprevalence was 72.25%. All of the herds were antibody positive against BVDV. The prevalence ranged from 66 to 100% within the herds. There were no significant differences between the presence of antibodies to BVDV and the herd size (P > 0.05). The prevalence in animals lower than 2 years old differed significantly with cows higher than 2 years old (P < 0.05). According to the results, it is concluded that it is likely the presence of persistently infection (PI) animal(s) within the herds in suburb of Mashhad-Iran, which is responsible for the presence antibody.     

Serologic detection of experimental Salmonella enteritidis infections in laying hens by fluorescence polarization and enzyme immunoassay

Detection of infected poultry flocks is essential for controlling eggborne transmission of Salmonella enteritidis to humans. The present study evaluated the detection of antibodies in the sera of experimentally infected chickens by a fluorescence polarization assay with a tracer prepared from the O-polysaccharide of S. enteritidis and an enzyme-linked immunosorbent assay (ELISA) with an S. enteritidis flagellin antigen. In two trials, groups of specific-pathogen-free laying hens were infected orally with either 10(6) or 10(8) colony-forming units (CFU) of S. enteritidis (phage type 13a) or with 10(8) CFU of Salmonella typhimurium. Serum samples were collected before inoculation and at five subsequent weekly intervals. Both assays successfully detected the majority of hens infected with S. enteritidis at either dose level, but they also identified a substantial number of hens infected with S. typhimurium as seropositive. The fluorescence polarization test detected S. enteritidis infection significantly more often and cross-reacted with sera from hens infected with S. typhimurium significantly less often than the ELISA. The fluorescence polarization assay also offered advantages in terms of speed and methodologic simplicity.     

Validation of a commercial ELISA for the detection of bluetongue virus (BTV)-specific antibodies in individual milk samples of Dutch dairy cows

A recently developed indirect ELISA for the detection of bluetongue virus (BTV)-specific antibodies in bovine milk samples was compared to that of the routinely used competitive ELISA on serum samples. During the bluetongue outbreak in the Netherlands in 2006, caused by BTV serotype 8, coupled serum and milk samples were obtained from 470 individual cows from 10 BTV-infected farms with an average seroprevalence of 57%. In addition, bulk milk samples of the same farms, and historically BT-negative samples were tested. Compared to the ELISA for sera, the relative specificity and sensitivity of the ELISA for milk samples is 96.5% and 98.9%, respectively when using a S/P% cut-off value of 50% as advised by the manufacturer. The optimal cut-off value was found at S/P% of 90% revealing an optimal specificity (99.0%) combined with an optimal sensitivity (98.1%). Titres in positive individual milk samples ranged from 1 to 2048 with a peak titre of 128. Bulk milk samples contained antibodies with titres ranging from 64 to 512. The ELISA for milk samples was found to be a reliable and robust test. This diagnostic tool is very useful, and may replace the ELISA for serum samples as first choice in order to get insight into the status of lactating individual animals and therewith of the entire herd with respect to BTV infection.     

Culling decisions of dairy farmers during a 3-year Salmonella control study

Salmonella enterica subsp. enterica-serotypes lead to periodically increased morbidity and mortality in cattle herds. The bacteria can also lead to serious infections in humans. Consequently, Denmark has started a surveillance and control programme in 2002. The programme focuses on Salmonella Dublin which is the most prevalent and most persistent serotype in the Danish cattle population. A field study in 10 dairy herds with persistent Salmonella infections was carried out over three years to gain experience with control procedures including risk assessment, targeted control actions and test-and-cull procedures. From autumn 2003 until end of 2006 quarterly milk quality control samples from all lactating cows and biannual blood samples from all young stock above the age of three months were tested using an indirect antibody ELISA. The most recent and previous test results were used to categorise all animals into risk groups. These risk groups and all individual ELISA-results were communicated to the farmers as colour-coded lists four to six times per year. Farmers were advised to manage the risk of Salmonella transmission from cattle with repeatedly high ELISA results (flagged as "red") or cows with at least one recent moderately high ELISA result (flagged as "yellow") on the lists. Risk management included, e.g. culling or separation of the cows at calving. We analysed culling decisions using two models. For heifers a hierarchical multivariable logistic model with herd as random effect evaluated if animals with red and yellow flags had higher probability of being slaughtered or sold before first calving than animals without any risk flags. For adult cows a semi-parametric proportional hazard survival model was used to test the effect of number of red and yellow flags on hazards of culling at different time points and interactions with prevalence in the herd while accounting for parity, stage of lactation, milk yield, somatic cell count and the hierarchical structure of the data with animals clustered at herd level. This study illustrates how investigation of culling decisions made by herd managers when they have access to test-status of individual animals and overall apparent prevalence during control of an infection can lead to useful new knowledge. Overall herd managers were more likely to cull cattle with increasing number of yellow and red flags than animals with no flags. However, cattle were more likely to be culled with yellow and red flags during times with low or medium high within-herd seroprevalence than at times with high seroprevalence. These results are valuable knowledge for modelling and planning of control strategies and for making recommendations to farmers about control options.