Serological Diagnosis of Pullorum Disease with the Microagglutination System

The application of a tetrazolium-stained Salmonella pullorum antigen in the microagglutination test is described and compared with the macroscopic tube agglutination test for detecting carriers of pullorum disease and fowl typhoid in chickens. Titers revealed by both testing procedures are similar; however, the microagglutination test is preferred because of the savings in time, space, and cost.     

Serological investigation of the fish pathogen Edwardsiella ictaluri, cause of enteric septicemia of catfish.

The serological relationships among 32 isolates of Edwardsiella ictaluri obtained from fish were studied. The strains were extremely homogeneous in protein and lipopolysaccharide preparations as observed by sodium-dodecyl-sulfate polyacrylamide gel electrophoresis. Only minor variations were observed in the structural O-side chain subunits in three isolates; however, such variation did not preclude antigenic recognition by two E. ictaluri antisera in either microagglutination or Western blot immunoassays. The antigenic homogeneity of E. ictaluri was further demonstrated by microagglutination assays with both formalin-killed and heat inactivated cellular antigens. The minimal degree of antigenic variability observed suggested that most isolates of E. ictaluri compose a single antigenic serotype.     

Avian Salmonella-Stained Microtest Antigens Produced on Solid Media

Procedures are described for the preparation of Salmonella pullorum and Salmonella typhimurium tetrazolium-stained microagglutination and microantiglobulin antigens with solid media for culture propagation. Cell suspensions were found to be more easily harvested from solid medium than from previously used liquid medium, and greater yields of antigen were obtained. Liquid medium microagglutination and microantiglobulin test antigens were found to be more sensitive than those prepared on solid medium. The microtest antigens produced on solid medium were easier to prepare and in serological tests gave titers more comparable to those demonstrated with the standard macroscopic tube agglutination test.     

The diagnosis of antileptospiral antibodies in human subjects by solid-phase immunoenzyme analysis

The possibility of using the enzyme-linked immunosorbent assay (ELISA) for the diagnosis of leptospirosis has been shown. This method has proved to be more simple and sensitive than the leptospiral microagglutination and lysis test. The data on obtaining genus-specific leptospiral antigens are presented. As revealed in this study, the antigens obtained by the complex treatment of microbial cells with ultrasound and detergents show the maximum activity in ELISA. The optimum parameters of the ELISA system for the diagnosis of leptospirosis have been established.     

Use of the immunofluorescent microagglutination reaction for the purpose of studying ornithosis infection. 1. Production and approbation of a corpuscular luminescent diagnosticum]

A method of obtaining the fluorescent ornithosis corpuscular diagnostic agent providing for the extraction of the corpuscular antigen with its subsequent conjugation with fluorochrome-fluoresceine isothiocyanate. The use of this preparation permits to stage the agglutination reaction on the basis of immunofluorescent analysis, this facilitating the recording of the reaction results and considerably decreasing consumption of the diagnostic agent. As shown, the suggested immunofluorescent microagglutination reaction was characterised by a sufficiently high sensitivity, specificity, and can be used for detection of antibodies to the causative agent of ornithosis.     

Use of the immunofluorescent microagglutination reaction for the intrageneric differentiation of the immune response in chlamydiosis

The possibility of using, on principle, the immunofluorescent microagglutination test with Chlamydia fluorescent corpuscular antigen for the strain-specific differentiation of immune response induced by the causative agents of ornithosis (strain B), enzootic abortus ovis (strain EAE) and lymphogranuloma venereum (strain LV) has been experimentally demonstrated. The calculation of indices, characterizing the specificity of differences between the systems under comparison, by the method of S. Frasser and D. Berman (1965) has confirmed the significance of such differentiation.     

Methodological aspects of serological diagnosis in leptospirosis

The results of serological study for leptospirosis of 515 blood serum specimens from patients coming to different clinics of the Republic of Bangladesh, are presented. The sera were tested in microagglutination test. To exclude intergroup reactions and to enhance reliability of results, immunoglobulin classes were determined with the use of cysteine as a reducing agent and immunoabsorption test. In 51 patients (9.9%) antileptospiral antibodies were detected in titers from 1:20 to 1:1600 against pathogenic leptospires of different serological groups.     

Properties of phase I purified cellular antigen of Coxiella burnetii

An attempt was made to purify phase I cell suspension of Coxiella burnetii used as an antigen in diagnostic serological tests. Homogenised suspension of chick embryos infected with phase I Henzerling and "Z" strains, after preliminary purification from host cell contaminants of chick embryos was subjected to consecutive centrifugation in sucrose/uropoline gradient and to continuous 20-45% uropoline gradient. The fractions obtained from uropoline gradient centrifugation were applied as phase I antigen C. burnetii in the following tests: complement fixation and microagglutination. Only fractions containing protein were serologically active. They proved to be of similar specificity and sensitivity as the antigens obtained by standard method. Moreover, it was found that after formalin treatment of C. burnetii cells no soluble antigens are liberated which could be detected by complement fixation test.     

A microagglutination test for blood typing rhesus monkeys, Macaca mulatta

We have developed a microagglutination test for typing rhesus monkey erythrocytes that is sensitive, accurate and easy to perform. The technique requires only microliter quantities of antiserum and cells, and agglutination is easily detected using an inverted microscope. An advantage of this technique is that the typing plates can be stored at -70 degrees C without loss of activity. The results of typing over 400 rhesus blood samples with this technique were 95% concordant with results using the standard microtitre agglutination technique. Preliminary results indicate that this test is also adaptable to typing human blood.     

Comparison of tests for the diagnosis of spontaneous encephalitozoonosis in rabbits

The effectiveness of immunofluorescence, complement fixation, microagglutination serologic tests, intradermal skin test, and detection of histologic lesions were compared for use in diagnosis of spontaneous encephalitozoonosis in rabbits. The India ink and microbead agglutination reactions were compared with immunofluorescence and complement fixation by testing 11 single or pooled sera. Serologic tests correlated best with each other and less well with intradermal tests or presence of lesions. Immunofluorescence, India ink reaction and microbead agglutination were equally useful in detecting antibodies to Encephalitozoon cuniculi. The intradermal test correlated best with the presence of detectable lesions.     

土拉弗朗西斯菌检测研究进展

土拉弗朗西斯菌（Francisella tularensis）是土拉菌病（Tularemia）的致病菌,是最具传染性的致病菌之一,在自然界中已发现一百种以上的动物感染此菌。因其传播途径多样,易扩散、毒性强而被美国疾病控制预防中心列入A类生物恐怖制剂。土拉菌病是一种人畜共患病,致死率高,及时、准确的检测土拉菌对于土拉菌病患者及时治疗和防止扩散具有重要的意义。土拉菌检测方法很多,如菌培养,微凝集实验、酶联免疫吸附、快速检测试纸条、生物传感器、PCR、核酸杂交检测、质谱分析、基因芯片等。但到目前为止还没有一种成熟的用于土拉菌检测方法,其主要原因在于土拉菌致病性强,且不易分离培养。本文综述了土拉菌细菌学、免疫学、分子生物学方法检测的最新研究进展。     

Nephelometric density adjustment of leptospiral antigen.

In testing leptospirosis by microagglutination, reproducibility of the results is affected by population age and germ concentration. These undesirable factors can be avoided if antibody identification is made with cultures incubated for 7 to 21 days and adjusted to 50 X 10(6) leptospires per mililitre. Most suitable for the purpose is an analogue mode based on nephelometry and the probability theory. The obtained statistical data furnish calculated and tabulated apparatus values which are used for standardizing with the aid of a newly developed nepheloflask with compression-spring stirrer.     

Application of PCR for Specific Diagnosis of Leptospirosis in Humans in Ukraine

Leptospirosis remains one of the most widespread zoonotic diseases in the world and Ukraine, in particular. Ukrainian clinicians have been faced with early detection of the disease due to the availability of only a serological method for routine diagnostics in Ukraine, namely the microscopic agglutination test (MAT). This paper demonstrates the first results of the complex application of MAT and polymerase chain reaction (PCR) for routine verification of leptospirosis, which were first applied simultaneously in Lviv Oblast of Ukraine in 2016. We examined the sera of 150 patients clinically suspected of leptospirosis, 31 of whom were treated at the Lviv Oblast Clinical Hospital for Infectious Diseases (LOCHID). The application of PCR during the first seven days of the disease allowed increasing the share of confirmed leptospirosis cases by 16,1% in patients that were treated in LOCHID during 2016–2017.Key words: Leptospirosis, diagnostics, microagglutination test, polymerase chain reaction     

A simple method for the detection of antibodies to Encephalitozoon cuniculi in rabbits.

Rabbit antibodies against Encephalitozoon cuniculi were detected in an indirect microagglutination test using a bead substrate to which anti-rabbit immunoglobin G light and heavy chain antibodies were coupled. The test was positive using immune whole serum or F(ab)' and F(ab)'2 fragments of immunoglobin G but negative using the F(c) fragment. The reaction was blocked by saturating the beads with rabbit serum or by absorbing positive sera with excess Encephalitozoon cuniculi. The test provided a simple method to detect antibodies to Encephalitozoon cuniculi, did not require elaborate equipment and could be performed using frozen antigen.     

The use of microdot immunoenzyme analysis with visual detection for the determination of tularemia antibodies

The possibility of using the micropoint enzyme immunoassay (EIA) on a nitrocellulose membrane with the visual evaluation of results for the detection of tularemia IgG antibodies in hamadryas baboons at the postvaccinal period has been studied. The sensitivity of this assay has been compared with that of the passive hemagglutination (PHA) test, the microagglutination (MA) test and EIA with the spectrophotometric evaluation of results in plates. As shown in this study, EIA in the above-mentioned modification can be successfully used for the detection of tularemia antibodies in the blood serum. The sensitivity of micropoint EIA has proved to be not inferior to that of EIA in plates, while exceeding the sensitivity of the PHA test 10- to 20-fold and the sensitivity of the MA test 10- to 1,000-fold. This method is simple, reliable, highly sensitive, economic and requires no special equipment, which makes it highly promising for the diagnosis of tularemia and the evaluation of humoral immunity at the postvaccinal period.     

A microagglutination test for blood typing rhesus monkeys, Macaca mulatta

Summary. We have developed a microagglutination test for typing rhesus monkey erythrocytes that is sensitive, accurate and easy to perform. The technique requires only μ1 quantities of antiserum and cells, and agglutination is easily detected using an inverted microscope. An advantage of this technique is that the typing plates can be stored at -70°C without loss of activity. The results of typing over 400 rhesus blood samples with this technique were 95% concordant with results using the standard microtitre agglutination technique. Preliminary results indicate that this test is also adaptable to typing human blood.     

The immunochemical and biological properties of Leptospira membranes]

The immunochemical and biological properties of purified membrane fractions obtained from Leptospira interrogans, serovar copenhageni, strain Rat 2, and Leptospira biflexa, strain Patoc 1, were studied. The presence of genus-specific and group-specific antigens in leptospiral membranes was established by the methods of immunodiffusion analysis, the microagglutination (MA) and lysis tests. In animal experiments cell membrane preparations produced no toxic and allergic effects. Leptospiral membranes obtained from strain Rat 2 ensured the protection of golden hamsters infected with Leptospira virulent culture and induced antibody production in high titers, detected with the use of the MA test, the lysis test and the enzyme immunoassay, in rabbits immunized in two injections.     

Prevalence of Leptospira and Brucella antibodies in wild boars (Sus scrofa) in Tuscany, Italy

Five hundred sixty-two blood samples were collected from wild boars (Sus scrofa) shot in six districts of Tuscany, central Italy, between 1997 and 2000. Sera were examined for antibodies specific for Leptospira interrogans by microagglutination test and Brucella spp. by the Rose Bengal test and indirect enzyme-linked immunosorbent assay. Thirty-four (6.0%) samples tested positive for anti-Leptospira antibodies, 29 (5.1%) sera were positive for anti-L. interrogans serovar bratislava antibodies (titres ranging from 1:100-1:400), and 5 (0.9%) sera were positive for anti-L. interrogans serovar icterohaemorrhagiae antibodies (titres 1:100). All the examined sera were negative for anti-Brucella antibodies.     

Microantiglobulin Test for Detecting Salmonella typhimurium Agglutinins

A sensitive antiglobulin (AG) test procedure for the demonstration and experimental study of the agglutinin response of chickens infected orally with Salmonella typhimurium is described. A tetrazolium-stained S. typhimurium antigen was employed with microagglutination techniques and equipment for the first time in conducting the AG test. Results with the conventional macroscopy tube agglutination test for S. typhimurium and the 24-hr microtest were comparable; however, the AG test enhanced titers as much as 16 times, and these persisted at a significant level for as long as 4 months. This study is being extended to other Salmonella serotypes and possible field applications of the AG test procedure.     

The effect of selected viruses on Corynebacterium kutscheri infection in rats

The influence of selected viral pathogens on rats that were previously infected with Corynebacterium kutscheri was investigated. A series of three separate experiments were performed to test the effect of sialodacryoadenitis virus, Sendai virus and rat virus. In each experiment, weanling rats were divided into three groups (C. kutscheri-inoculated, virus-inoculated and C. kutscheri plus virus-inoculated). Two groups were inoculated oronasally with C. kutscheri to establish subclinical infections. Two weeks later, two groups were inoculated intranasally with virus. At 5 weeks, the prevalence of C. kutscheri recovery from oral cavity and submaxillary lymph node and the prevalence of overt pseudotuberculosis was compared between treatment groups. Seroconversion of rats to C. kutscheri was measured by microagglutination and viruses by indirect immunofluorescence assays. Infection of rats with sialodacryoadenitis virus, Sendai virus or rat virus had no discernable effect on C. kutscheri-infected rats.