Studies on haemophilus infection in swine I. Application of the latex agglutination test to the diagnosis of Haemophilus pleuropneumoniae (H. Parahaemolyticus) infection

A latex agglutination test was evaluated as a method for the detection and titration of antibodies against swine Haemophilus infection and it was found that the test is applicable to the etiologic diagnosis of Haemophilus infection in swine. In swine infected experimentally with H. pleuropneumoniae (H. parahaemolyticus), the micromethod of agglutination using latex particles coated with antigens of H. pleuropneumoniae was found to be comparable agar-gel immunodiffusion and complement-fixation tests, which have previously been used for the etiologic diagnosis of the disease. Antibody titers determined by the latex agglutination test corrlated well with those determined by the other serological tests. The latex agglutination test tended to detect antibodies earlier than any of the other tests. By the latex agglutination test, weak cross-reactions were observed among different serotypes of H. pleuropneumoniae, whereas no cross-reaction was demonstrated between H, pleurophneumoniae and H. parasuis. The latex agglutination test was found to be simple and useful for the serological survey of swine Haemophilus infection, especially when dealing with a large number of samples.     

A rapid agglutination assay for canine brucellosis using antigen coated beads

Brucella canis is the causative agent of canine brucellosis and facultative intracellular pathogen. The diagnosis of canine brucellosis is based on bacteriological examination and serological methods including agglutination and gel diffusion tests. In this study, crude antigens were extracted from B. canis using hot saline, coated on to latex beads and their usefulness in the serological diagnosis of canine brucellosis was examined. Mixing the antigen coated latex beads with the sera of dogs infected with B. canis produced clear agglutination, but this was not so for B. canis free dog sera. N-terminal amino acid sequence analysis of the crude hot saline extracts, showed that they contained copper-zinc superoxide dismutase, ribose ABC transporter and hypothetical protein of Brucella as antigens. A serological survey of canine serum samples conducted by means of an agglutination test using the antigen coated latex beads, showed that this method was more specific than the tube agglutination test using whole bacterial cell antigens. Although these results suggest that our method in which crude hot saline extracted antigens are coated on to latex beads would be useful in the serological diagnosis of canine brucellosis, we need further investigation using more serum samples to confirm the usefulness of our method.     

Rapid latex agglutination (RLA) test for the diagnosis of Babesia Argentina

A rapid test, utilizing latex particles (0.81-μm diameter), sensitized with Babesia argentina antigens, proved to be effective in the diagnosis of B. argentina in natural and experimental infections. Two drops of plasma or serum and one drop of B. argentina antigen placed on a glass plate were used in the test. Reaction was observed after 3—10 min rotation. The positive agglutination reaction was characterized by the formation of fine latex particle clumss. In experimental infections with B. argentina, the first detectable positive agglutination reactions coincided with the appearance of parasitemia in thin blood films. Plasma from animals with natural infections of B. argentina, proven by blood smears and indirect fluorescent antibody and complement fixation tests, also showed a reaction to the latex agglutination test.     

A latex agglutination test for field diagnosis of contagious caprine pleuropneumonia

Latex beads were sensitised with a polysaccharide isolated from a F38 culture supernatant and used in a slide agglutination test to detect serum antibodies in goats with contagious caprine pleuropneumonia. The latex agglutination test detected antibodies in the sera of goats by 22 +/- 2 (mean +/- 1 sd) days after contact exposure to contagious caprine pleuropneumonia, whereas the complement-fixation test detected antibodies by 24 +/- 4 days after contact exposure. Both tests were negative with 181 sera from a farm which was free of the disease. When the same tests were done on 763 sera from two different farms with outbreaks of classical contagious caprine pleuropneumonia, 63 per cent were positive by the latex agglutination test and 23 per cent were positive by the complement-fixation test. Besides being more sensitive than complement fixation, the latex agglutination test can be performed in the field using undiluted serum or whole blood and a result obtained within two minutes.     

A latex agglutination test for the detection of Echinococcus multilocularis coproantigen in the definitive hosts

A latex agglutination test for detecting Echinococcus multilocularis coproantigen in definitive hosts was developed using latex beads sensitized with EmA9 monoclonal antibody raised against somatic antigens of adult E. multilocularis. A primary test (LA 1) was performed on 82 fecal samples of necropsied foxes, of which 46 were infected, and resulted in 61% sensitivity and 86% specificity. To increase the sensitivity, 4 ng/mL of excretory/secretory antigens of adult worms was added to the samples in a secondary test (LA 2), resulting in 91% sensitivity and 61% specificity. The positive predictive value of the LA 1 test and the negative predictive value of the LA 2 test were both 85%. The combination of the LA 1 and LA 2 tests is applicable and practical for use in situations that require quick diagnosis or screening based on the following interpretation: the samples that are positive in the LA 1 test are positive; the samples that are negative in the LA 2 test are negative; and the samples that are negative in the LA 1 test and positive in the LA 2 test are classified as suspicious.     

Relationships of total protein, specific gravity, viscosity, refractive index and latex agglutination to immunoglobulin G concentration in mare colostrum

A colostrum sample was collected within 24 h after foaling from 27 mares and from 10 other mares a milk sample was collected several weeks post partum. Immunoglobulin G concentrations were determined quantitatively by radial immunodiffusion and semi-quantitatively using a commercial latex agglutination test. Total protein, specific gravity, viscosity and refractive index were determined and their relationships to the immunoglobulin G concentration analysed. All parameters correlated with the immunoglobulin G concentration. The latex agglutination test divided the colostrum samples into three groups with different means for immunoglobulin G and total protein concentrations. Specific gravity and the latex agglutination test were found to be the methods best suited for on-farm evaluation of colostrum quality.     

Comparison of Latex Agglutination, Indirect Hemagglutination, and ELISA Techniques for the Detection of Toxoplasma gondii-specific Antibodies in the Serum of Cats

The primary purpose of this study was to determine whether commercially available latex agglutination and indirect hemagglutination kits for the detection of Toxoplasma gondii-specific antibodies were capable of detecting T. gondii-specific immunoglobulin M (IgM) in the serum of cats. Serum samples from 35 cats containing either T. gondii-specific IgM, T. gondii-specific immunoglobulin G (IgG), or both were collected. Each serum sample was assayed using a latex agglutination kit, an indirect hemagglutination kit, an enzyme-linked immunosorbent assay (ELISA) for the detection of T. gondii-specific IgG, and an ELISA for the detection of T. gondii-specific IgM. When serum samples containing only T. gondii-specific IgM as determined by ELISA were assayed, the latex agglutination kit and the indirect hemagglutination kit detected antibodies in 33.3% and 13.3%, respectively. When T. gondii-specific IgG was present in a serum sample, the results from the latex agglutination kit, the indirect hemagglutination kit, and the IgG-ELISA were similar; however, there was a wide variation in titer magnitude results between the three assays. It was concluded that the latex agglutination kit and the indirect hemagglutination kit did not adequately detect T. gondii-specific IgM in feline serum.     

A Brief Overview of Transgenic Farm Animals

A rapid recombinant antigen-based latex agglutination test (LAT) has been developed to detect specific anti-leptospiral antibodies from human and dog sera. The recombinant LipL32 antigen developed and used for detecting the antibodies is specific in detection of the pathogenic serovars of Leptospira as the expression of the LipL32 antigen is restricted only to the pathogenic leptospires. The sensitized latex beads are stable and could be stored at 4^°C for more than three months without showing loss of activity for both weakly and strongly positive samples. The test is found to be sensitive, specific and accurate as compared to the standard microscopic agglutination test (MAT). Moreover, the recombinant antigen-coated latex beads could detect the specific anti-leptospiral antibodies in the acute phase of the illness. The test is simple and inexpensive, and is rapid in the management of large numbers of patients.     

Evaluation of a recombinant MIC3 based latex agglutination test for the rapid serodiagnosis of Toxoplasma gondii infection in swines

The entire gene encoding microneme protein 3 (MIC3) from Toxoplasma gondii was cloned into the plasmid pGEX-KG and subsequently expressed in Escherichia coli as a glutathione-S-transferase (GST) fusion protein. The recombinant MIC3 (rMIC3) was purified and evaluated in a latex agglutination test (LAT) as the diagnostic antigen for the detection of antibodies to T. gondii in pig sera. The specificity, stability, and reproducibility of the test were examined. No agglutination was found when the sensitized latex beads were mixed with phosphate-buffered saline (PBS), borate-buffered saline (BBS), normal saline, and negative serum samples. There was no cross-reactivity with the standard positive sera of other pathogens. But intense agglutination occurred with T. gondii antibody positive serum samples. In our study, the coincidence rate of tested positive-sera of the LAT with rMIC3-sensitized latex particles and the ELISA with rSAG1 was up to 92.8%, T. gondii specific antibodies were detected by the LAT in all piglets that were experimentally infected with T. gondii tachyzoites from 8 to 42 days after infection. Our results indicated that the rMIC3 based latex agglutination test appears to be suitable for the detection of T. gondii antibodies at the early stage of infection.     

Comparative studies of the detection of rotavirus in fecal samples of calves with diarrhea with the latex test "Slidex Rota-Kit 2" and electron microscopy

Of 68 fecal samples from calves with diarrhoea which were tested for rotavirus with the latex agglutination test "Slidex Rota-Kit2" and by electron microscopy 33 samples were positive and 33 were negative with both tests respectively. Divergent results (latex test positive/EM negative and vice versa) were observed in one specimen only, respectively. Cross reactions with other viruses diagnosed by electron microscopy were not observed with the latex agglutination test. The "Slidex Rota-Kit2" is another suitable test for the diagnostic laboratory as well as for the veterinary practitioner for the detection of rotavirus in fecal samples of calves.     

Canine serum C-reactive protein detected by means of a near-patient test for human C-reactive protein

OBJECTIVES: The aim of the study was to evaluate the reliability of a rapid human C-reactive protein near-patient slide reversed passive latex agglutination test (Randox) for the semi-quantitative determination of canine serum C-reactive protein. METHODS: The concentration of C-reactive protein was determined in 244 canine serum samples by an established automated immunoturbidimetric method and in various predilutions by a commercially available reversed passive latex agglutination test for human C-reactive protein. The results were compared to assess if the reversed passive latex agglutination test reflected the results of the established method with special emphasis on the reversed passive latex agglutination test's ability to identify samples characterised as positive or negative by the established method. RESULTS: The reversed passive latex agglutination test reflected the C-reactive protein concentration in canine serum samples at all the tested predilutions (undiluted, 1:4, 1:8 and 1:16). When applying a predilution of 1:8, the positive and negative analytical predictive values for discriminating between positive and negative samples (according to the established quantitative method) were high (0.94 [0.82 to 0.99] and 0.97 [0.93 to 0.99], respectively). CLINICAL SIGNIFICANCE: In conclusion, this near-patient test was able to reflect the serum C-reactive protein concentration in canine samples in a reliable and clinically useful manner and could be applicable for general practice for evaluating C-reactive protein levels in canine serum.     

Evaluation of an enzyme-labeled antiglobulin test for anti-Brucella immunoglobulin G among 3 cattle populations

Antibody to smooth Brucella abortus lipopolysaccharide antigen on the surface of polystyrene tubes was detected with peroxidase-labeled antibody against bovine immunoglobulin G. The enzyme-labeled antiglobulin test (ELAT) activity of samples was expressed in arbitrary units/0.01 ml by reference to a standard curve based on tests of dilutions of a positive serum pool. Reactions greater than 3.0 U/0.01 ml were classified positive because specificity at this level was 99.8% (417/418 samples correctly classified negative) with agglutination test-negative sera from 33 Brucella-free herds. Results of the ELAT were compared with results of agglutination tests and the complement-fixation test (CFT), using 430 sera from cattle in 7 infected herds. Activity of greater than 5.0 ELAT U/0.01 ml was detected in all 54 sera classified as positive (titer greater than 1:10) by the CFT, including 5 sera classified as negative by the tube agglutination test. Sera from 8 nonvaccinated cows in the infected herds reacted only by the ELAT, whereas reactions were obtained with 25 and 5 sera by only agglutination tests and the CFT, respectively. The ELAT and CFT results were in agreement for 25 of 26 sera from agglutination test-reactor cattle in herds of unknown status. Comparisons of milk ring and whey agglutination tests with the whey ELAT on 146 quarter samples from cows in an infected herd revealed no ELAT activity greater than or equal to 1.0 U/0.01 ml in the 73 samples considered negative by the 2 other tests. Samples (n = 47) that contained greater than or equal to 1.0 ELAT U/0.01 ml included all (n = 40) samples with milk ring or whey agglutination titers greater than or equal to 1:16 and greater than or equal to 32, respectively, and 7 samples that gave weaker reactions to the latter tests.     

Cryptococcosis of the central nervous system in a dog

A 3-year-old female German Shepherd Dog was evaluated for progressive mental obtundation and vestibular signs. Central nervous system cryptococcosis was diagnosed on the basis of growth of Cryptococcus neoformans in fungal culture of CSF, as well as detection of the organism in CSF via microscopy. Cryptococcal capsular latex antigen agglutination titer was 1:262,144 in CSF and 1:1,048,576 in serum samples. Imaging with magnetic resonance augmented diagnosis. The dog improved after long-term treatment with fluconazole. Fluconazole is useful in the treatment of CNS cryptococcosis, because it attains high concentration in the CNS. Long-term therapy is often required for resolution of clinical signs, and affected animals may require long-term follow-up with periodic evaluation of CSF via fungal culture and latex agglutination tests. Monitoring serum latex agglutination test results may provide a safe, less invasive means of monitoring response to treatment.     

Evaluation of a rapid specific test for detecting colostral IgG1 in the neonatal calf

A commercially available latex agglutination test was used to measure the concentration of IgG1 in bovine plasma and the results were compared with radial immunodiffusion and zinc sulphate turbidity methods. For concentrations of IgG1 up to 80 g/litre there were highly significant (P less than 0.001) correlation coefficients between the latex agglutination test and radial immunodiffusion, and between the latex agglutination test and zinc sulphate turbidity method (0.93 and 0.74 respectively). The coefficient of variation for the latex agglutination test ranged from 8.1 per cent to 17.9 per cent. IgG1 concentration was measured using the latex agglutination test in whole blood on a farm, in whole blood at a laboratory and in plasma at a laboratory. The correlation coefficients were highly significant (P less than 0.001) in all cases. The latex agglutination test is easy to use, rapid and specific. It is suitable for checking the colostral status of young calves on commercial farms.     

Serologic evaluation of cattle inoculated with Toxoplasma gondii: comparison of Sabin-Feldman dye test and other agglutination tests

Six cows and 6 calves were each inoculated with 100 or 100,000 Toxoplasma gondii oocysts. Serum samples were analyzed, using the Sabin-Feldman dye test (DT), indirect hemagglutination test, latex agglutination test, and the modified direct agglutination test (MAT). Antibody titers in cows were lower than in calves. In the cows, DT titers increased briefly during the first month after inoculation, after which the titers were negative; however, T gondii was isolated from the tissues of 4 cows. Indirect hemagglutination and latex agglutination titers were generally less than 1:256. The MAT titers increased to 1:1,024 during the first month after inoculation. In 5 of the 6 cows, the MAT titers persisted. The 6th cow had a preinoculation MAT titer of 1:2,000 for 3 to 6 months. Therefore, the DT was not useful in serologic surveys for T gondii in cattle; the MAT was the most sensitive test and may be useful in the diagnosis of T gondii infection in cattle.     

乳胶凝集试验检测鸡传染性法氏囊病毒血清抗体的研究

以鸡传染性法氏囊病病毒(IBDV)VP2基因工程抗原致敏乳胶微粒,用IBDV阳性血清进行方阵滴定,以最佳致敏系件制成乳胶抗源,建立乳胶凝集试验(LAT),用来检测血清中IBD抗体。对467份待测血清分别、同时作LAT和双向琼琼脂免疫扩散试验(DATA)。结果,LAT阳性419份,阴性48份;DAGT阳性425份,阴性42份.试验表明乳胶凝集试验操作简便、快速、敏感性高、特异性强,可用于现场检测,适合基层单位检测IBDV血清抗体。     

Evaluation of quantitative latex agglutination for detection of Cryptosporidium parvum, E. coli K99, and rotavirus in calf feces.

A new methodology for detection of rotavirus, Escherichia coli K99, and Cryptosporidium parvum in bovine fecal samples was developed based on a quantitative latex agglutination technique (QLAT). Calibrated microspheres coated with specific antibodies to 1 of the enteric pathogens are quantitatively agglutinated by the antigens present in diluted fecal sample. The test is performed in a 96-well flat-bottom plate. The samples were tested with a commercial enzyme-linked immunosorbent assay kit prior to being analyzed by QLAT. The calculated sensitivity and specificity are adequate for field conditions, because the amount of the pathogenic agents is generally high. The overall time to perform the test was about 20 minutes.     

A slide latex agglutination test for the rapid detection of antibodies in serum against porcine parvovirus

A slide latex agglutination test (LAT) was developed and evaluated to detect serum antibodies against porcine parvovirus. Porcine parvovirus antigen was obtained by 10% PEG-6000 and 0.5 mol/l sodium chloride precipitation, and inactivated by 0.1% methanal. Two per cent suspensions of latex particles (0.5-0.8 microm) were coated by adding an equal volume of porcine parvovirus antigen at 0.34 microg/ml. Repeatability of latex agglutination test was evaluated with a panel of 100 sera using the same and different antigen lots. A good agreement between LAT and haemagglutination inhibit assay was observed. Because of convenience and speed of performance, this method would be used widely in clinic examination.     

Conjugation and Characterization of Latex Particles with Toxoplasma gondii–specific Immunoglobulin Y Antibodies for Diagnostic Aim and Evaluation Efficiency in In Vitro Culture

Toxoplasma gondii is a parasite that causes severe health problems in the world. Toxoplasmosis, an infection caused by T. gondii, leads to high risk of mortality in patients with immunodeficiency, transplantation, and cancer. Besides that, it causes miscarriages in pregnancy, various abnormalities such as hydrocephalus in infants and congenital diseases. Because the clinical indication of the disease is not specific, it is confused with many diseases, and this leads to the necessity of directly detecting the presence of the toxoplasmosis. Therefore, various diagnostic assays are needed for the diagnosis of the disease. Amongs them, latex agglutination assay is widely used for the detection of specific antibodies or antigens in samples. Latex particles are coated with immunogenic molecules (antigens) to detect antibodies in the blood or used to identify antigens when coated with specific antibodies. In both, aggregation of latex particles results in agglutination. Monoclonal antibodies are often used in latex agglutination assay as in other diagnostic methods. However, monoclonal antibodies can be produced in low quantities at a high cost. Besides, to produce monoclonal antibodies, an experienced staff, a well-equipped cell culture laboratory, a long period of time, and a burdened budget are needed. In recent years, as an alternative to monoclonal antibodies, immunoglobulin Y (IgY) antibodies, which are obtained from chicken eggs, and specifically produced against desired antigenic constructs, have become quite attractive in terms of both low cost and abundant production without requiring infrastructure. In contrast, the latex assay based on IgY antibodies for use in the diagnosis of T. gondii has not been developed. This study aimed to conjugate T. gondii-specific IgY antibodies to latex particles, characterize the particles by Fourier transform infrared spectroscopy, scanning electron microscopy, and spectroscopic methods, and finally demonstrate the interaction with T.gondii parasites in culture with scanning electron microscopy analysis.     

Sensitivity and specificity of latex agglutination tests used to identify Streptococcus agalactiae and Staphylococcus aureus isolated from bulk tank milk

Comparisons were made among rapid latex agglutination tests and conventional biochemical tests used to identify Streptococcus agalactiae and Staphylococcus aureus. Ninety-eight streptococci and 149 staphylococci isolated from bulk tank milk were tested. Sensitivity and specificity for the latex agglutination test used for identification of Str agalactiae were 97.6 and 98.2%, respectively. Sensitivity and specificity for the latex agglutination test used for identification of S aureus were 90.2 and 67.5%, respectively. Of 25 staphylococci considered false-positive by the latex agglutination test, 14 (56%) were considered tube coagulase-positive. Fifteen staphylococci considered false-positive by latex agglutination test had biotypes representative of S hyicus of S xylosus.