Use of enzyme immunoassay in a serological survey of leptospirosis in sheep

A total of 731 serums, all from Merino rams from 20 farms, were tested for antibodies against Leptospira interrogans serovars hardjo, pomona and tarassovi using the microscopic agglutination test (MAT). The enzyme immunoassay (EIA) technique was used to test all serums for IgM and IgG antibodies to serovar hardjo. In the MAT, reactions to serovar hardjo were most common with 236 rams (32.3%) reacting at 1/100 or greater. Only 1.9% of serums reacted against serovar tarassovi and 1.1% against serovar pomona. The percentage of sheep with positive MAT reactions to serovar hardjo ranged from 0 0 to 94.9 between farms. When using EIA, 46 (6.2%) of the serums were positive for IgM antibody and 246 (33.6%) were positive for IgG antibody. Correlation of the EIA for detection of IgG antibody with the MAT was good. The EIA detection of IgG antibody was considered to be a good alternative test to the MAT for epidemiological studies in sheep.     

Evaluation of the enzyme-linked immunosorbent assay in the detection of cattle infected with Brucella abortus

One group of 51 cattle was vaccinated with B. abortus S19 (S19) and a further 51 cattle were vaccinated with B. abortus S45/20 (S45/20). Forty-eight cattle (24 from each group) and a control group of 12 cattle were subsequently challenged with B. abortus S544. The enzyme-linked immunosorbent assay (ELISA) was used to detect specific IgG and IgM antibodies in these groups. All cattle vaccinated with S19 had high levels of IgG and IgM, but the S45/20 vaccine produced detectable antibody in only a few cattle. In those cattle where the challenge induced infection, the mean levels of IgG and IgM were much higher than those of the uninfected cattle in the same groups. When the isolation of B. abortus was compared at slaughter with the serological results, the ELISA, when used to detect specific IgG, was more sensitive but less specific than the serum agglutination test, complement fixation test and indirect haemolysis test, and more sensitive and more specific than the Rose Bengal test.     

Specific antibodies to Besnoitia besnoiti precipitated from serum of cattle by live parasites and by soluble antigen

Specific precipitable antibodies of both IgG and IgM classes were detected in sera of cattle naturally infected with B. besnoiti. The amount of specific antibodies of the IgG class precipitated by soluble antigen was in the range of 17-50 micrograms/ml serum while that of the IgM class ranged between 5 and 24 micrograms/ml serum. Specific antibodies precipitated by live B. besnoiti parasites were in amounts of 10 to 22 micrograms/ml serum for IgG and 4 to 26 micrograms/ml serum for IgM. Different ratios of IgG/IgM were obtained by the two methods of precipitation. This might indicate that antibodies to B. besnoiti of the IgM class can be precipitated and detected in sera of naturally infected cattle in similar amounts either by live parasites or by solubilized antigen, whereas antibodies of the IgG class can be preferentially detected when solubilized antigen is used for precipitation.     

Immunoglobulins in porcine umbilical cord blood and maternal placenta.

Studies were made of the immunoglobulin (Ig) in serums from umbilical cord of newborn pigs and maternal placenta. The neutralization test for porcine parvovirus and Japanese encephalitis virus was carried out with the serum of the sow and that of the umbilical cord of the newborn pig. Comparative studies of the serums from the dam and the umbilical cord were also done with gel filtration. Of 20 umbilical cord serum samples, IgG was seen in 5 samples (25%), IgA in 1 sample (5%), and IgM in 9 samples (45%). The amount of any 1 of the 3 classes of Ig in the serums was between 13.5 and 28.0 mg/dl. Among the samples examined, 1 had both IgG and IgA and 1 had IgG and IgM, but none had both IgA and IgM and none had 3 classes of immunoglobulins (i.e., IgG, IgA, and IgM). Only 7 samples (35%) did not have any class of Ig. The IgG disappeared from the blood of hysterectomy-produced colostrum-deprived pigs at 3 days of age, and IgM disappeared when pigs were 7 days of age. Neutralization antibodies of porcine parvovirus and Japanese encephalitis virus in maternal serum were not transferred to the fetus through the placenta. Results of immunohistologic surveys indicated that the sow's Ig were not transferred to the fetus through the placenta. Therefore, it is believed that the Ig in the porcine fetus might be synthesized in certain cells in the placental tissue, and the degree of production of the Ig in the placental tissues may differ in each case. The component, which seems to be Ig, was observed as the obscure band of the beta- to gamma-globulin area in serum of the umbilical cord. Comparison was made, with gel filtration, of maternal serum and serum from the umbilical cord of the newborn pig originating from the same sow. Seemingly, the IgG in the umbilical cord serum is mainly in the lower molecular weight fraction, whereas IgG in the sow's serum was distributed in the high to low molecular weight fractions.     

The Immune Response of Calves given Mycoplasma bovis Antigens

Seven calves seven to 30 days of age were given Mycoplasma bovis antigen by different routes. Immunization was in two phases. The first consisted of single or multiple SC, IV or oral doses of antigen for two to four weeks. The second phase consisted of multiple SC or ID injections given from the eighth to the 19th week. The experiment was terminated at 26 weeks. Antibody titers were followed by indirect hemagglutination, growth inhibition and tetrazolium reduction inhibition. Total serum protein, protein fractions and IgG and IgM concentrations were determined in serums of one calf and the distribution of indirect hemagglutination antibodies in IgG and IgM classes were determined in serums of two of the calves.     

Antibody response of cattle to rinderpest vaccine

Antibody production was studied in cattle infected with rinderpest vaccine virus. Vaccinated cattle produced both IgM and IgG serum antibodies. The IgG antibodies were mainly those of IgG2 subclass. No IgA antibody response was detected in vaccinated animals.     

An abortion storm in cattle associated with neosporosis in Taiwan

An abortion storm associated with acute neosporosis involving 18 cattle was observed in a dairy farm in Taiwan. Aborted fetus age ranged from 3 to 8 months. Of the 38 cattle in that farm examined during the abortion storm, 52.6% (20/38), 13.2% (5/38) and 10.5% (4/38) contained both IgG and IgM, only IgG and only IgM antibodies to Neospora caninum, respectively. No antibody to N. caninum was detected prior to the abortion storm. Follow-up study conducted a year later showed that 23 out of 28 cattle had sero-converted. Since some cattle were positive to either only IgG or IgM, we suggest that both IgG and IgM should be tested for diagnosing neosporosis. Neosporosis surveillance of naive cattle herd is recommended.     

Trypanosome non-specific IgM antibodies detected in serum of Trypanosoma congolense-infected cattle are polyreactive.

Serum Ig from Trypanosoma congolense-infected cattle were affinity-purified using immobilised trypanosome or non-trypanosome antigens (beta-galactosidase, cytochrome C and ferritin). The bound and unbound IgG and IgM fractions were collected and tested in ELISA for reactivity to each antigen. The results indicated that the presence of reactivity to non-parasite antigens in serum of infected cattle is due to polyreactive IgM antibodies. However, the IgG fraction only bound to trypanosome antigens and was only present in post-infection sera, indicating that it was induced by the infecting trypanosomes. Since the polyreactive IgM antibodies were also present in pre-infection sera, it is probable that they were natural antibodies that were not induced but only amplified by the trypanosome infection.     

Preparation of monospecific antiserums against porcine immunoglobulins, using agarose-linked immunosorbents.

Immunoglobulins IgG, IgA, and IgM were isolated from porcine serum and milk, and antiserums against the 3 immunoglobulin classes were prepared. Monospecificity of the antiserums for the gamma-, alpha-, and mu-chains was obtained by absorbing them in agarose-linked immunosorbent columns. These immunosorbents were prepared by linking IgG or IgA-IgM to CNBr-activated agarose. Contaminating anti-alpha2-macroglobulin antibodies in the anti-IgA and anti-IgM serums were removed with agarose-linked fetal globulins.     

Bovine IgM and IgM response to Leptospira interrogans serovar hardjo as measured by enzyme immunoassay

The enzyme-linked immunosorbent assay (ELISA) was used to detect specific IgG and IgG antibodies in the sera of cattle infected or immunized with Leptospira interrogans serovar hardjo. IgM appeared first but was quickly followed by IgG which persisted longer than IgM. The levels of antibody detectable by ELISA and by the microscopic agglutination test (MAT) did not correlate, suggesting that the two techniques measured different antigen-antibody systems. The transient nature of the IgM response as measured by ELISA indicates potential usefulness as a serodiagnostic test for detecting current leptospiral infections.     

Brucella abortus-specific immunoglobulin in isotypes in serum and vaginal mucus from cattle vaccinated with strain 19 and challenge exposed with virulent strain 2308

The immunoglobulins (IgG1, IgG2, IgM, and IgA) of the Brucella-specific antibody response of 69 crossbred beef heifers were studied after Brucella abortus strain 19 vaccination and strain 2308 challenge exposure. The immunoglobulin isotype responses in serum and vaginal mucus were measured by use of fluorescent immunoassay. Serum antibody responses were detected also by 3 standard serologic tests (complement fixation [CF], Rivanol precipitation, and the CARD test] and 2 primary bindings assays that detect IgG antibodies. One month after vaccination, mean antibody titers for all immunoglobulin isotypes were higher for vaccinated cattle (n = 46) than for nonvaccinated controls (n = 23). After vaccination, IgA antibody responses in vaccinated cattle were only 2-fold higher than those for controls, whereas IgG1, IgG2, and IgM antibody responses were 3- to 90-fold greater than those for controls. Measurement of IgA antibody responses classified 21 of 39 vaccinates as seropositive after vaccination, whereas the other isotypes classified 28 or 34 cattle as seropositive. Three months after challenge exposure, the mean antibody responses for each isotype were higher in cattle that aborted or were culture positive than in cattle that did not abort and were culture negative. Although IgG1, IgG2, and IgM antibody titers were each of benefit in identifying B abortus-infected cattle, it did not appear that the magnitude of the antibody responses provided sufficient discrimination between S19-vaccinated cattle and S2308 challenge-exposed cattle. Serum IgA antibody responses were 10-fold higher after challenge exposure than after vaccination and may be a response to mucosal infection with the virulent organism.(ABSTRACT TRUNCATED AT 250 WORDS)     

IMMUNOGLOBULIN CLASSES IN SERUM ANTIBODY REACTIONS IN CATTLE FOLLOWING VACCINATION WITH BRUCELLA ABORTUS STRAIN 19 AND KILLED 45/20 VACCINES

SUMMARY A group of 4 cows was vaccinated with Brucella abortus strain 19, followed 8 weeks later by a single dose of B. abortus 45/20 vaccine. A similar group received 2 doses of B. abortus 45/20 vaccine 8 weeks apart. The antibody responses of the groups were compared by testing whole serums and separated IgM and IgG fractions by the Rose Bengal Plate (RBP) agglutination and the complement fixation tests (CFT) using rough and smooth B. abortus antigens. Animals that had received B. abortus strain 19 responded to the 45/20 vaccine with increased titres to the smooth antigen. These relevant antibodies were predominantly of the IgG class. Standard CFT and RBP test antibodies could be detected in IgM and IgG fractions after the primary inoculation with B. abortus strain 19 vaccine.     

A microplate enzyme-linked immunorsorbent assay for measuring antibody to Anaplasma marginale in cattle serum

An enzyme-linked immunosorbent assay (ELISA) method is described for measuring antibody against Anaplasma marginale in cattle serum. This method was more sensitive and objective than a previously described ELISA method for A. marginale and possible reasons for this are discussed. All 83 cattle experimentally infected with A. marginale (81) or A. centrale (2) developed demonstrable specific antibody but the serums of 98.8% of 839 cattle from cattle tick-free areas did not react by ELISA; 378 serums containing antibody to Babesia bovis were tested for cross reactions in the A. marginale ELISA. There were no significant cross-reactions except when cattle had been inoculated at least twice with B. bovis-infected erythrocytes, presumably due to antibodies reacting with erythrocyte material in the ELISA antigen. The ELISA detected antibodies for more than 3 years after infection, at least 2 years longer than did a complement fixation test. When A. marginale infections in cattle were eliminated by long acting oxytetracycline, their serums ceased to react by ELISA. An ELISA score for serum antibody level was shown to have a statistically significant correlation with ELISA titre.     

Detection of pseudorabies virus antibodies: time of appearance by two serotests

This report describes a time-course comparison of detection of pseudorabies virus antibodies in experimentally infected swine by the virus-neutralization (VN) and indirect hemagglutination tests. Specific antibody titers were observed by the IHA test at 5 days after swine were inoculated, but not until 12 days by the VN test. The predominant immunoglobulin (Ig) class present in the serums of the swine at 5 and 7 days after inoculation was IgM, as determined by sulfhydryl reductions. The VN test lacked sensitivity to early Ig levels (IgM) in these experimentally infected swine, while the indirect hemagglutination test was highly sensitive to these same levels. On the basis of these results, it is possible that the VN test may read early infections as pseudorabies virus negative, due to the low presence of IgG in these samples.     

Differential antigen recognition by serum antibodies from three bovid hosts of Mycobacterium bovis infection

Cattle, bison and buffaloes are susceptible to Mycobacterium bovis, the causative agent for bovine tuberculosis. Accurate and timely identification of infected animals is critical for improved management and control of disease in these species. Bovids develop humoral immune responses to M. bovis infection making serological tests attractive for tuberculosis screening. However, optimization and validation of antibody assays designed for various animal species require understanding of antigen recognition patterns in each target host. The objective of this study was to characterize serological reactivity profiles generated by cattle, American bison, and African buffaloes in tuberculosis. Serum samples from M. bovis-infected animals were tested for the presence of IgM and IgG antibodies to MPB70/MPB83 and CFP10/ESAT6 chimeric proteins using Dual-Path Platform technology. All three host species showed IgG responses of higher magnitude and frequency than IgM responses; further, IgM seroreactivity was limited to MPB70/MPB83, whereas IgG antibodies recognized both test antigens. In cattle, the IgM and IgG responses were elicited mainly by MPB70/MPB83, whereas bison and buffaloes showed similar IgG seroreactivity rates for MPB70/MPB83 and CFP10/ESAT6 antigens. The findings demonstrate distinct patterns of predominant antigen recognition by different bovid species in M. bovis infection.     

A SURVEY OF ANTIBODY TO AINO VIRUS IN CATTLE AND OTHER SPECIES IN AUSTRALIA

SUMMARY A serological survey of healthy cattle in Australia showed that antibodies to Aino virus were present in serums from cattle in northern Australia and down the east coast as far as central New South Wales in 1975, 1976 and 1977, but occurred with a lower frequency than antibodies to Akabane virus. in contrast to the findings with Akabane virus, no neutralising antibodies to Aino virus were detected in serums from camels, dogs or horses. Antibodies to both viruses were detected in buffaloes and sheep, but not in humans or any of the Australian indigenous species so far tested. All positive serums originated from within the known range of Culicoides brevitarsis.     

Antibody response in bovine pharyngeal fluid following foot-and-mouth disease vaccination and, or, exposure to live virus

Samples of pharyngeal fluid and serum were collected from cattle after exposure to live foot-and-mouth disease (FMD) virus (with or without prior vaccination) or after subcutaneous vaccination with inactivated virus. The pharyngeal fluid samples were examined for FMD neutralising activity and specific anti-FMD IgG, IgM and IgA antibodies. The neutralising activity of the serum was also monitored. A peak of neutralising activity which occurred in the pharyngeal fluid of unvaccinated cattle seven days after virus exposure corresponded to a rise in specific IgM and IgA antibodies. This peak appeared to be due to serum and tissue fluid escaping from the damaged mucosa during the acute inflammatory phase of infection. At later stages (20 to 60 days after virus exposure) the pharyngeal fluid neutralising activity corresponded to a rise in specific IgA antibodies, suggesting that active local antibody production was taking place. The pharyngeal fluid neutralising activity detected after revaccination with oil emulsion or aqueous vaccines, without exposure to live virus, corresponded to a rise in specific IgG and IgM antibody levels and this may have been due to serum transudation.     

A SEROLOGICAL SURVEY FOR BLUETONGUE VIRUS ANTIBODY IN WESTERN AUSTRALIA

A serological survey was carried out to detect specific (serotype 20) and a group bluetongue virus antibody in cattle and sheep serums collected in Western Australia during the period January 1 1978 to June 30 1979. Of 18,849 cattle serums examined by the gel diffusion precipitin test (GDPT), 9.7% were positive and 6.1% gave doubtful results. All 1949 sheep serums tested were negative. Precipitin antibody was demonstrated in 22.5% of serums from Kimberley cattle and 3.6% of cattle serums from the Northwest. Serums collected from cattle in the South were consistently negative in GDPT. When 915 serums that reacted in the GDPT were further tested by the complement fixation test (CFT), 164 were positive. The percentage of CFT positive serums increased as the GDPT reaction became stronger. 2467 serums collected from cattle in Kimberley and Northwest areas and tested by the CFT, 175 (7.1%) were positive. These 175 positive serums were also examined by GDPT and 164 doubtful or positive reactions were obtained. The virus neutralisation (VNT) using serotype 20 virus was carried out on 3804 serums, including all serums that reacted in the GDPT, and 57 were positive. When the VNT positive serums were examined in the other 2 tests, 47 serums were either positive or doubtful in the GDPT and 8 were positive in the CFT. The presence of bluetongue virus group antibody in cattle serums closely followed the suggested distribution pattern of Culicoides brevitarsis but specific serotype 20 neutralising antibody was limited to cattle serums from stations situated north of latitude 17 degrees S in an area of mean annual rainfall higher than 700 mm.     

Ultracentrifugal Studies of Sera from Cattle Vaccinated or Naturally Infected with Brucella Abortus

In conformity with the findings of previous investigators, it was shown by density gradient ultracentrifugation that the antibodies in sera collected from calves shortly after vaccination with Brucella abortus, strain 19, were entirely or mainly rapidly-sedimenting. These macroglobulin (19S or IgM) antibodies showed complement-fixing as well as agglutinative activity with Br. abortus antigen. In later bleedings from the same vaccinated calves, antibodies with an intermediate sedimentation rate, (IgG), were present, as well as IgM. Sera from 15 of 22 non-vaccinated, relatively recent field cases of brucellosis appeared to contain only the IgG class of antibodies. In one herd, however, two cows with IgM only and five with both IgM and IgA were found; all seven of these cattle had been serologically negative before their introduction into this known infected herd a few months earlier. The agglutinative activity of sera from four cases of brucellosis of long standing and from eight cows, 4 to 13 years of age, that had been vaccinated as calves, was confined to the IgG fraction.     

Antibodies to prion and Acinetobacter peptide sequences in bovine spongiform encephalopathy

An amino acid sequence homology has been identified between the bovine prion sequence (RPVDQ) and the Acinetobacter calcoaceticus enzyme, uridine-diphosphate-N-acetyl glucosamine-1-carboxy-vinyl-transferase which also contains (RPVDQ). Class-specific IgA, IgG and IgM antibodies against synthetic peptides containing the structurally related sequences present in bovine prion and A. calcoaceticus were measured in 189 bovine spongiform encephalopathy (BSE) positive cattle, 127 BSE negative cattle and 87 healthy control animals using an ELISA technique. Class-specific IgA, IgG and IgM antibodies against the structurally related synthetic peptides were significantly elevated in BSE positive cattle when compared to BSE negative cattle (P < 0.001) and healthy control animals (P < 0.001). These autoantibodies may have a role in the pathogenesis of BSE.