Screening of commercial milk samples using ELISA for immuno-epidemiological evidence of infection by the cattle grub (Diptera: Oestridae)

Pasteurised and ultra high temperature (UHT)-treated milk were tested over the period November 1997-February 1998 in order to evaluate the applicability of an hypodermosis ELISA test on commercial milk samples. Milk samples from six dairy companies were analysed and the development of anti-Hypoderma antibodies recorded for the period from September 1998 to June 1999. For one dairy (no. 3) bulk milk samples were also obtained from the tankers, transporting milk away from the farms, and analysed; the results were compared to the parasitological status of the cows whose milk had been collected on the farms. Out of 32 pasteurised samples tested, 23 were positive, exceeding the cut-off (22%), while 9 samples were negative. UHT milk was always negative. The antibody levels in milk samples from four of the six companies were highest during January and decreased gradually during February to May 1999. The bulk milk samples also had high antibody levels and 47.2% of milk producing cows had lesions of hypodermosis. It was concluded that testing commercial milk for antibodies is an efficient way of detecting the presence of hypodermosis in cattle, especially in those countries for which no data on this disease are available.     

奶牛新孢子虫病血清流行病学调查

本研究用酶联免疫吸附试验（ELISA）对采集自北京地区的94份奶牛血清（随机采集）和河北地区的55份奶牛血清（有流产史奶牛），进行Neospora caninum血清抗体检测。结果发现，北京地区随机采集的奶牛血清N．caninum抗体阳性率为18．1％（17／94），河北地区有流产史的奶牛N．caninum血清抗体阳性率为23．6％（13／55）。采用牛奶记录体系（DHI）对北京地区17头N．caninum血清抗体阳性牛进行了日产奶量、乳中蛋白率和乳脂率的测定，并与同群牛中134头阴性牛比较。结果表明，N．caninum血清抗体阳性牛日产奶量比阴性牛降低9．7％，乳中蛋白率和乳脂率分别降低20％和15．4％。初步证明N．caninum血清抗体阳性奶牛产奶量降低及奶品质的下降。对不同N．caninum抗体滴度阳性牛的泌乳期主要生产性能比较发现，其生产性能的变化与抗体滴度无明显相关性。     

Feasibility of serological bulk milk testing as a method for BVD surveillance

A commercial ELISA detecting antibodies against bovine viral diarrhoea Virus (BVDV) was analysed for its applicability for bulk-milk screening. Detection limits were analysed using native and concentrated milk samples (milk treated with rennet and ammonium sulfate precipitated) from 10 cows whose sera showed different reactivity levels in the ELISA and from two cows which gave birth to persistently infected calves during the last year. Further this and a second commercial ELISA were used to screen 591 randomly selected bulk-milk samples. To clarify discrepancies thirty-nine herds were included in a follow-up study. A second bulk-milk sample and serum samples from 10 young cattle of 6 to 28 month of age per herd were analysed for antibodies against BVDV. The results of this second testing and the detection of viremic animals in 4 herds confirmed the results from initial bulk-milk testing with both tests. The analysed test is suitable for bulk-milk testing although its application is limited by vaccination.     

Informative Value of an Indirect Enzyme‐Linked Immunosorbent Assay (ELISA) for the Detection of Bovine Viral Diarrhoea Virus (BVDV) Antibodies in Milk

Bulk and individual milk samples from 117 herds located in Brittany (west France) were used to assess: (i) the performance characteristics of an indirect enzyme‐linked immunosorbent assay (ELISA) applied to individual milk for the detection of antibodies to bovine viral diarrhoea virus (BVDV); and (ii) the relationship between the bulk milk result obtained from this test and the within‐herd prevalence of antibody‐positive lactating cows. This ELISA test was based on a monoclonal antibody directed against non‐structural protein NS2‐3 of pestiviruses. At the individual level, based on 1113 matched milk/serum samples, the sensitivity and specificity of this test applied to milk, compared with the virus neutralization test on serum, were 95.0 and 97.7%, respectively. At the herd level, the relationship between the optical density percentage (OD%) of bulk milk and the within‐herd prevalence of antibody‐positive lactating cows was assessed using the receiver operating characteristics (ROC) analysis. Classes of OD% of bulk milk were determined so that they were associated with minimum intraclass and maximum between‐class variances of within‐herd prevalence of antibody‐positive cows. The ROC analysis resulted in two classes of bulk milk results corresponding to different expected levels of within‐herd prevalence. Herds with an OD% of bulk milk <75% and ≥75% had a mean observed prevalence of antibody‐positive cows of 8.9 and 60.6%, respectively. Herds with a bulk milk result <75% were expected to be BVDV free, whereas large variations in prevalence of antibody‐positive cows existed in the herds with OD% ≥75%. The test described in this study is suitable to identify herds likely to have a low prevalence of BVDV antibody‐positive cows.     

Development and evaluation of a highly sensitive and specific blocking enzyme-linked immunosorbent assay and polymerase chain reaction assay for diagnosis of bovine leukemia virus infection in cattle

OBJECTIVE: To develop a blocking ELISA for detection of bovine leukemia virus (BLV) antibodies that is comparable to a radioimmunoprecipitation (RIP) assay, to evaluate use of this ELISA for identification of BLV-infected herds, and to develop a polymerase chain reaction (PCR) assay for direct diagnosis of infection with BLV. SAMPLE POPULATION: Serum samples and pooled bulk-tank milk samples from cattle. PROCEDURE: The blocking ELISA was developed, using BLV gp51 as antigen, captured by a selected bovine polyclonal serum. A nested PCR was conducted with primers specific for a segment of the pol region of the BLV genome. RESULTS: Sensitivity and specificity of the ELISA were comparable to those of the RIP assay. Use of the ELISA on pooled milk samples allowed identification of herds in which prevalence of BLV infection among lactating cows was as low as 2.5%. Pooled milk samples from BLV-free herds did not react in the ELISA. All cattle that had positive results for the nested PCR had BLV antibodies, but cattle with consistantly low antibody titers required examination of sequential DNA samples to detect viral sequences. None of the 63 antibody-negative cattle had positive results for the PCR. CONCLUSIONS AND CLINICAL RELEVANCE: This ELISA is a highly specific and sensitive assay for the detection of BLV antibodies in serum and milk samples of cattle. Examination of pooled milk samples with the ELISA provides a reliable, practical, and economic procedure for identification of BLV-infected herds. The nested PCR also constitutes a specific procedure for direct diagnosis of infection with BLV.     

Comparison of milk and serum enzyme-linked immunosorbent assays for diagnosis of Mycobacterium avium subspecies paratuberculosis infection in dairy cattle.

Milk and serum samples from 35 dairy herds in 17 states were evaluated for cow- and herd-level Mycobacterium avium subspecies paratuberculosis (MAP) antibody test agreement. Evaluation of 6,349 samples suggested moderate agreement between milk and serum enzyme-linked immunosorbent assay (ELISA) results, with a kappa value of 0.50. Cow-level sensitivity (Se) for 18 dairy operations with 1,921 animals was evaluated relative to fecal culture results. At the cow level, the milk ELISA relative Se was not significantly different from that of the serum ELISA (21.2 and 23.5%, respectively). Logistic regression models revealed a positive association between lactation number and milk ELISA status. Non-Holstein cows were more likely to test milk ELISA positive than Holstein cows. Cows in the first 2 weeks of lactation and after week 45 of lactation were more likely to test milk ELISA positive than cows between 3 and 12 weeks of lactation. Milk production > 80% of herd average was negatively associated with testing milk ELISA positive. Animals in the West and Midwest regions were less likely than animals in the Southeast region to test ELISA positive by either test. Estimates for herd-level sensitivity for the milk and serum ELISA, relative to fecal culture results, ranged from 56 to 83%. At the cow and herd levels, milk ELISA performed equivalent to serum ELISA using fecal culture as a reference for MAP infection and has the advantage of decreased labor costs on farms that use Dairy Herd Improvement Association testing.     

Assessment of cattle grub (Hypoderma spp.) prevalence in northeastern Italy: an immunoepidemiological survey on bulk milk samples using ELISA

Hypodermosis is a myiasis of cattle mainly found in the Northern Hemisphere. In Italy it has been reported primarily in southern regions with a farm prevalence reaching a peak of 85%, while few data are available in the north of the country.This paper reports the results of a serological survey carried out on hypodermosis in Vicenza province (northeastern Italy). During January and February 1999, bulk milk samples were collected from 224 dairy farms and tested, using an hypodermosis ELISA kit. Epidemiological data, such as farm location, herd size, number of cows, grazing management, treatments against Hypoderma larvae, presence of imported animals, were gathered and statistically analysed in relation to the serological results. In 15 farms the presence of warbles was monitored by manual palpation and larvae collected for identification.Ninety-seven (43.3%) samples were detected as positive, whereas 12 (5.4%) and 115 (51.3%) were inconclusive and negative, respectively.Thirteen farms were positive upon parasitological examination and the larvae collected were identified as Hypoderma bovis.Seroprevalence was highly correlated with the practice of free grazing and slightly, but significantly, correlated with the number of cows on the farm. The mean antibody level of positive herds was significantly higher on farms without imported animals. On farms with warbled animals there was a high inverse correlation between antibody level and quantity of milk in the tanks and the number of lactating cows.The ELISA test on bulk milk samples has been shown, under field conditions, to be a useful, cost-effective and easy test for the diagnosis of bovine hypodermosis.     

Improving the sensitivity of the IBR-gE ELISA for testing IBR marker vaccinated cows from bulk milk

The low sensitivity of the IBR-gE ELISA compared to other diagnostic ELISA tests for IBR is a major disadvantage of IBR control programmes based on IBR marker vaccination. Therefore the IBR-gE ELISA is not generally recommended for testing pooled or bulk milk samples.The aim of this study was to determine the performance of a commercially available kit for concentrating and purifying antibodies in milk in order to improve the sensitivity of detecting IBR-gE antibody positive cows from pooled and bulk milk samples. A single IBR-gE positive cow is likely to remain undetected in a pool of 49 negative milk samples without concentration. By contrast, the bulk milk concentration procedure improved sensitivity from 5.4% to 75.7% in a positive herd. Milk samples with a high or moderate positive signal are more likely to be detected after pool milk concentration compared to weak positive samples. Whereas a follow up study involving a monthly testing of bulk milk samples from three marker vaccinated IBR-gE negative herds over a period of seven months yielded negative results each month, bulk milk from a herd containing <5% IBR-gE positive cows always detected positive after concentration. Although the milk concentration procedure had no impact on specificity, it significantly enhanced the sensitivity of the detection of IBR-gE positive milk in pooled and bulk milk samples. After further evaluation this procedure could allow a cost efficient and reliable method of monitoring IBR marker-vaccinated herds for IBR-gE antibodies.     

Evaluation of enzyme-linked immunosorbent assay using milk samples as a potential screening test of bovine tuberculosis of dairy cows in Korea

An enzyme-linked immunosorbent assay (ELISA) using bulk tank milk samples was evaluated as a screening test for bovine tuberculosis (TB), a contagious chronic disease of cattle. An ELISA with MPB70, a major antigen of Mycobacterium bovis was performed using paired sets of milk and sera samples from 33 tuberculin-positive and 43 tuberculin-negative cattle. Anti-MPB70 antibodies were detected in milk samples and there was a significant correlation between seroreactivities of milk and sera samples (R² = 0.83). Using the tuberculin skin test as the reference test, the sensitivities of ELISA using milk and sera samples were 87.8% and 81.8%, respectively, and the specificities were 97.7% and 100%, respectively.In the screening test using bulk tank milk samples from 931 dairy herds in Whasung, Gyeonggi-do, Korea, the positive rate for anti-MPB70 antibody was 4.5% (42/931) and the tuberculin-positive rate was 2.8% (26/931). Individual milk samples (n = 253) were collected from randomly selected 8 problematic and 3 negative herds (positive and negative in the screening test by MPB70 ELISA using bulk tank milk samples, respectively) and tested by MPB70 milk ELISA. In the problematic herds, positive rates were 10.5% (20/190) for anti-MPB70 antibodies in milk ELISA and 2.1% (4/190) in the tuberculin skin test. More than one dairy cows were positive by milk ELISA among the problematic herds, and all tuberculin-positive dairy cows were positive in the milk ELISA. Further, no positive cows were detected in negative herds both by milk ELISA and tuberculin skin test. These results suggest that an ELISA, using bulk tank milk samples, might be a potential efficient screening test for bovine TB of dairy cows.     

Antibody responses of naive cattle to two inactivated bovine viral diarrhoea virus vaccines,measured by indirect and blocking ELISAS and virus neutralisation

Two groups of naive heifers were given primary courses of two inactivated bovine viral diarrhoea (BVD) virus vaccines licensed for use in the UK. Their humoral responses in serum and milk were assayed by means of an indirect ELISA detecting antibodies to structural viral glycoproteins, a blocking ELISA specific for antibodies to the non-structural protein NS2-3 and the virus neutralisation test (VNT). For each assay, the numbers of serum or milk samples testing positive at each sample point and the mean values were determined. In both vaccine groups, serum antibody responses were detected by the indirect ELISA and the VNT, with both the numbers of seropositive animals and mean values peaking five weeks after the second vaccination. In the 23 heifers vaccinated with Bovilis BVD, the mean NS2-3-specific ELISA values remained low throughout the trial, with no serum or milk samples testing positive. In the 24 heifers vaccinated with Bovidec, the mean NS2-3 responses peaked below the level of positivity five weeks after the second vaccination, before declining again; NS2-3-specific antibodies were detected in one serum sample and one milk sample from two heifers in this group. A pooled milk sample from each vaccine group tested negative by both ELISAS 12 weeks after the second vaccination.     

Immune response of pregnant cows to bovine rotavirus immunization

Fifteen pregnant Holstein cows were freely assigned to 3 experimental groups (5 cows in each group). Cows in group I were inoculated IM and intramammarily (IMm) with Ohio Agricultural Research and Development Center (OARDC) tissue culture-propagated modified-live Nebraska calf diarrhea bovine rotavirus with added adjuvant (OARDC vaccine-immunized cows). Group II cows were given IM injections of a commercial modified-live rotavirus-coronavirus vaccine (commercial vaccine-immunized cows), and the remaining 5 cows were noninoculated controls (group III). Rotavirus antibody in colostrum and milk was mainly associated with immunoglobulin (Ig)G1, and less so with IgG2, IgA, and IgM, as analyzed by the enzyme-linked immunosorbent assay (ELISA), using monospecific anti-bovine IgG1, IgG2, IgM, and IgA sera. In serum, the rotavirus antibody was distributed almost equally between IgG1 and IgG2. The same relationships appeared in both immunized and nonvaccinated cows. All OARDC vaccine-injected cows had virus-neutralization (VN) and ELISA IgG1 rotavirus antibody titers in serum and mammary secretions at significantly increased levels (at least 100-fold; P less than 0.05) compared with the titers in groups II (commercial vaccine-immunized cows) and III (controls). Serum, colostrum, and milk antibody titers from these latter 2 groups did not differ statistically. The ELISA IgG2, IgA, and IgM rotavirus antibody titers also were significantly greater in mammary secretions from OARDC vaccine-immunized cows than in groups II and III cows. There was a high correlation between ELISA IgG1 and VN rotavirus antibody titers for all samples tested (r = 0.97, P less than 0.001), but ELISA IgG1 antibody titers were consistently higher than VN titers. The ELISA IgG1 and VN antibody titers of milk samples collected from cows 30 days after parturition were higher from the OARDC vaccine-immunized cows (ELISA IgG1, geometric mean titer (GMT) = 3,511; VN GMT = 1,689) than were titers from the group II cows (ELISA IgG1 GMT = 39; VN GMT = 33) or group III cows (ELISA IgG1 GMT = 21; VN GMT = 19). These results indicate that IM plus IMm immunization of pregnant cows, using modified-live bovine rotavirus with added adjuvant, may significantly enhance serum, colostrum, and milk rotavirus antibody titers, whereas IM vaccinal inoculation of pregnant cows with a commercial modified-live rotavirus-coronavirus vaccine may not.     

Evaluation of an O antigen enzyme-linked immunosorbent assay for screening of milk samples for Salmonella dublin infection in dairy herds.

Levels of antibodies to the O antigens (O:1,9,12) of Salmonella dublin were tested in 1355 serum, 1143 cow milk and 160 bulk milk samples from dairy herds using an enzyme-linked immunosorbent assay (ELISA). In order to define the background reaction, milk samples from all lactating cows and serum samples from 9 animals were collected in each of 20 salmonellosis-free herds located on the island of Bornholm, where cattle salmonellosis has not been reported. Similar samples were collected from all stalled animals in 10 herds with recent (< 6 months) outbreaks of salmonellosis located in Jutland, where salmonella infection is enzootic. Using herd history of salmonellosis, herd location and clinical status of the herds as criteria, the optimal cutoff in the milk ELISA was determined as being at least 5% of the samples having optical density > 0.5, resulting in herd sensitivity of 1.0 and herd specificity of 0.95. While none of the sera in the herds from Bornholm was ELISA positive, 2 herds had a few reactors in the milk ELISA. Using the same cutoff, all but 1 bulk milk sample from 150 herds on Bornholm was ELISA-negative, and all 10 salmonellosis-positive herds from Jutland were ELISA-positive. A significant correlation was found between ELISA reactions in milk and in serum of cows (34% and 32% respectively, rs = 0.69, P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Enzyme-linked immunosorbent assay for detecting antibodies to Brucella abortus in bovine milk and serum

An enzyme-linked immunosorbent assay (ELISA) method was evaluated for the detection of antibodies to Brucella abortus in cows milk. Milk samples from seropositive or -negative cows were sed to determine the distribution of absorbance values to classify milk as ELISA positive or ELISA negative. Brucella abortus was isolated from milk samples from 10 (45%) of the 22 cows whose milk and serum were ELISA positive. The ELISA was evaluated and determined to be an appropriate method for detecting antibodies to B abortus in bovine milk.     

Use of bulk milk for detection of Neospora caninum infection in dairy herds in Thailand

The relationship between the level of Neospora caninum antibodies in bulk milk and the seroprevalence in lactating cows was investigated. Bulk milk was also used to estimate the prevalence of N. caninum infection in dairy herds in the northeast and north Thailand. Bulk milk and individual serum from all lactating cows in 11 herds as well as 220 bulk milk samples from nine milk collection centres were analysed for presence of N. caninum antibodies using an iscom ELISA. In the 11 herds the bulk milk absorbances ranged between 0.04 and 0.89 and the seroprevalences varied between 0 and 46%. Five herds had milk absorbances below 0.20, among those were the two herds housing only seronegative lactating cows. In the remaining three herds with such low bulk milk absorbances one or two cows (5-14%) were seropositive. Six of the investigated herds had bulk milk absorbances above 0.20. In the two herds with the highest bulk milk absorbances more than 30% of the cows were seropositive. Using an absorbance of 0.20 to discriminate between negative and positive herds, 102 (46%) of 220 bulk milk samples were judged positive. There was no significant difference in mean bulk milk absorbance between the milk collection centres within each region. However, the proportion of herds with bulk milk absorbances > or =0.50 in the north was statistically (P < 0.01) higher than that in the northeast. It was concluded that bulk milk antibody testing can be used to identify N. caninum-infected herds and that N. caninum is a common infection in dairy herds in Thailand.     

Control measures in officially acknowledged brucellosis-free and leukosis unsuspected dairy herds on the basis of bulk milk samples in combination with ELISA tests

1. EC- and National Regulations. Since 1988 the EC-regulations accept in addition to the on Agar Gel Immunodiffusion test (AGIDT) based blood serum testing of cattle herds that are filed as "free from Enzootic Bovine Leucosis" the use of ELISA for this purpose. The regular testings in dairy cattle herds can be done alternatively with single or pooled milk samples, in other herds with pooled blood sera using ELISA. General condition is only a minimal sensitivity of the test to detect the European EBL Antibody Standard ("E4") in a dilution of 1:10 in negative serum or 1:250 in negative milk. Adequate national regulations are in preparation. The present limitation of pool sizes, blood maximum 50 animals without preparation steps 20, and milk after concentration treatment 50 cows is neutralized by proceedings in development of higher sensitive ELISA tests. This limitation should be canceled. Herd bulk milk samples without size limitations are accepted to be tested with "Milk Ring Test" by EC for the regular testings in filed "Brucellosis Free Dairy Cattle Herds". The alternative use of more sensitive (and more specific) ELISA tests for this purpose including the technical conditions is in a final discussion. 2. Scientific-Technical Base for Using the Chances of the Proceeding in the EC-Regulations. The realisation of the EC accepted or final discussed ELISA based bulk milk testing to control filed "EBL- and/or Brucellosis Free Herds" depends on some basic conditions like sensitivity, specificity, and variability of the ELISA systems. Field trials of more than 20,000 bulk milk samples in case of Brucellosis and more than 2,000 in case of EBL show the feasibilities and the limits of the ELISA systems in defining the status of the herds. The Brucellosis respectively the EBL situations of the dairy cattle herds tested in this trail were well known by history and by investigation of single animal blood samples using conventional tests. Special test run variations of pretested assays demonstrated the possibilities to define the EBL status of dairy cattle herds up to 50 lactating cows without preparation of the bulk milk sample and up 100 after concentration of the antibodies by the rennet-ammonium sulfate method. The concentration limit for detection of Brucellosis antibodies is 100 lactating cows. The bulk milk of smaller herds can be tested without concentration. On principle the evaluation of the test values bases on defined relations to a "weak positive" reference.(ABSTRACT TRUNCATED AT 400 WORDS)     

Challenges for bovine viral diarrhoea virus antibody detection in bulk milk by antibody enzyme-linked immunosorbent assays due to changes in milk production levels

Background

Bovine viral diarrhoea (BVD) is considered eradicated from Denmark. Currently, very few (if any) Danish cattle herds could be infected with BVD virus (BVDV). The Danish antibody blocking enzyme-linked immunosorbent assay (ELISA) has been successfully used during the Danish BVD eradication program, initiated in 1994. During the last decade, the cattle herd size has increased while the prevalence of BVDV has decreased. In this study, we investigated how these changes could affect the performance of the Danish blocking ELISA and of the SVANOVIR^®BVDV-Ab indirect ELISA. The latter has successfully been used to eradicate BVD in Sweden.Data (2003–2010) on changes in median herd size and milk production levels, occurrence of viremic animals and bulk milk surveillance were analysed. Additionally, the Danish blocking ELISA and the SVANOVIR ELISA were compared analyzing milk and serum samples. The prevalence of antibody positive milking cows that could be detected by each test was estimated, by diluting positive individual milk samples and making artificial milk pools.

Results

During the study period, the median herd size increased from 74 (2003) to 127 cows (2010), while the prevalence of BVDV infected herds decreased from 0.51 to 0.02 %. The daily milk yield contribution of a single seropositive cow to the entire daily bulk milk was reduced from 1.61 % in 2003 to 0.95 % in 2010 due to the increased herd size. It was observed that antibody levels in bulk milk decreased at national level. Moreover, we found that when testing bulk milk, the SVANOVIR^®BVDV-Ab can detect a lower prevalence of seropositive lactating cows, compared to the Danish blocking ELISA (0.78 % vs. 50 %). Values in the SVANOVIR^®BVDV-Ab better relate to low concentrations of antibody positive milk (R² = 94-98 %), than values in the blocking ELISA (R² = 23–75 %). For sera, the two ELISAs performed equally well.

Conclusions

The SVANOVIR ELISA is recommended for analysis of bulk milk samples in the current Danish situation, since infected dairy herds e.g. due to import of infected cattle can be detected shortly after BVDV introduction, when only few lactating cows have seroconverted. In sera, the two ELISAs can be used interchangeably.     

Antibody response to toxic shock syndrome toxin-1 of Staphylococcus aureus in dairy cows

Antibody response to toxic shock syndrome toxin-1 (TSST-1) of Staphylococcus aureus in dairy cows was examined by enzyme linked immunosorbent assay (ELISA). Serum antibody to TSST-1 was not detected in 39 (76.5%) of 51 calves, which were 1-6 months of age. In contrast, TSST-1 antibody was demonstrated in 1728 (72.6%) of 2380 lactating cows housed on 36 dairy farms. The ELISA values of antibody ranged from 0.2 to 3.0 OD and presented a distribution with the peak at 1.6 OD. The mean ELISA value differed between farms, and it increased slightly along with parturient history. Somatic cell counts of milk from 174 lactating cows was compared with TSST-1 antibody and tst1,000,000 cells per ml. The mean ELISA values in milk were lower than those of sera, but they rose as somatic cells increased. The tst gene of S. aureus detected in 76.0-86.2% of the milk samples containing somatic cells > 500,000 cells per ml, a level which indicates mastitis. The data suggests that many lactating cows may be infected by TSST-1- producing S. aureus.     

Enzyme-linked immunosorbent assay for detection of milk immunoglobulins to leukocidin toxin of Staphylococcus aureus

Leukocidin toxin from a bovine strain of Staphylococcus aureus was partially purified by ion exchange chromatography. An enzyme-linked immunosorbent assay was developed to quantitate antibodies specific for leukocidin in bovine milk. This was used to assay quarter samples from 88 cows in a S aureus-infected herd for antibody levels to the toxin. Milk samples from 65 cows with S aureus infections in at least one quarter produced a mean optical density of 1.054, whereas milk samples from 23 cows that were free of bacteria on cultural examination had a mean optical density of 0.584. There was a significant difference (P less than 0.001) in milk anti-leukocidin levels between these 2 groups. Evaluation of serum samples from 40 of these cows indicated that the milk anti-leukocidin concentrations were reflective of systemic anti-leukocidin values. The capability of 57 milk samples to neutralize the cytolytic effect of minimal amounts of leukocidin on bovine peripheral blood neutrophils was examined. Good correlation existed between the enzyme-linked immunosorbent assay antibody concentration and toxin-neutralizing capability of individual milk samples.     

Detection of antibodies to mycoplasmas using an immunoenzyme method

Detection of the antibodies to the species Mycoplasma bovis in the serum and milk of dairy cows coming from a mastitis-infected herd is a good example of utilization of the ELISA immunoenzymologic method in the mycoplasmology. Examining the samples from 75 dairy cows and applying the indirect hemagglutination test, good correlation of the results of the two tests was determined. The antibodies to the species Ureaplasma diversum were demonstrated by the ELISA method both in the bovine serum and in the milk of dairy cows infected slightly with mastitis. We chosen that strain which detected the maximum titres in the selected samples of the sera out of four antigens prepared from various strains of U. diversum. Rabbit sera hyperimmune to 26 strains of the mycoplasmas of various species were used to identify two antigens (after removing the antibodies to the components of the media). Specific reaction was obtained with the antisera to M. hyorhinis and M. arginini.     

Evaluation of an enzyme-linked immunosorbent assay for detection of antibodies to Fasciola hepatica in milk

Antibodies against Fasciola hepatica were detected in serum and individual milk samples of dairy cattle using an ELISA. Percentage positivity (PP) values in milk samples were related to serum PP values and were not influenced by days into lactation. The correlation coefficient between serum and individual milk samples was highly significant (r=0.84, P<0.005). The correlation coefficient between herd seroprevalence and herd milk antibody prevalence was 0.96. The correlation coefficient between prevalence measured by faecal egg count and both seroprevalence and milk antibody prevalence within the herd was 0.87. The diagnostic sensitivity and specificity for milk were 92% (95% CI=89-96) and 88% (95% CI=85-91), respectively, when the serum test was considered as a gold standard. In conclusion, the level of antibody to F. hepatica in milk is significantly correlated with the antibody level in serum and this ELISA is suitable as a means of routine veterinary diagnosis of exposure to F. hepatica in cattle and an alternative to testing sera.