Utilization of serological methods in the retrospective diagnosis of cholera and in detecting vibrion carriers]

Sensitivity and specificity of the three serological methods were studied comparatively: the vibriocidal test, the reaction of bacterial agglutination and of indirect hemagglutination, with the use of erythrocytes sensitized with the vibrio lyzate, cholera species O-antigen and cholerogen. Investigations were conducted with the blood sera of cholera patients, vibrio carriers and contacts. Vibriocidal test proved to be the most sensitive; its data correlated with the results of bacterial agglutination and indirect hemagglutination with erythrocytes, sensitized with the lysate of the vibrios and the cholera O-antigen. None of the used serological methods provided a 100% coincidence with the results of bacteriological analysis. The frequency of detection of anticholera antibodies decreased in the following order: cholera patients, vibrio carriers, contacts.     

The evaluation of the vibriocidal antibody test in establishing a diagnosis of cholera or Vibrio carriage]

On the basis of the serological survey of cholera patients, vibrio carriers and persons having had contacts with the source or reservoir of Vibrio cholerae the conclusion has been made that the test for the presence of vibriocidal antibodies, together with the bacteriological study of the patient, is of diagnostic importance in the diagnosis of cholera or vibrio carriership. The detection of vibriocidal antibodies, especially in the study of paired sera, permits the detection of cholera cases which have not been bacteriologically confirmed due to various reasons; besides, it makes it possible to exclude the diagnosis of cholera made only on the basis of clinical data. Like bacteriological study, the determination of vibriocidal antibodies must be obligatory for persons hospitalized in a provisory hospital or an isolation ward; it will undoubtedly improve the quality of cholera diagnosis and permit taking timely antiepidemic measures in the focus of infection.     

The agglutinability of cholera O-monoclonal immunoglobulins]

A set of 10 monoclonal antibodies specific for vibrio species and of 5 monoclonal antibodies specific for serovar (Ogawa) was studied. These monoclonal antibodies were directed toward V. cholerae O1 antigen in agglutination reaction and on slide plates. Monoclonal antibodies agglutinating typical strains of cholera vibrios with titration range from 1: 6000 to 1: 25,000 were selected. MA were revealed to interact with cholera vibrio strains with reduced agglutinability. The set of agglutinating O monoclonal immunoglobulins was developed for laboratory identification of cholera O1 vibrios.     

Avian cholera exposure and carriers in greater white-fronted geese breeding in Alaska, USA

We conducted a 3-yr study (2001-03) on greater white-fronted geese (Anser albifrons frontalis) breeding in Alaska, USA, to determine the exposure of this population to Pasteurella multocida and the potential role of these birds as disease carriers. We tested sera from nearly 600 adult geese for antibodies to P. multocida serotype 1. We found a low prevalence (<5%) of positive antibodies in adult geese, and based on the short duration of detectable antibodies, these findings indicate recent infection with P. multocida. Prevalence was similar to serologic results from both breeding and wintering lesser snow geese. We also collected oral (n=1,035), nasal (n=102), and cloacal (n=90) swab samples to determine the presence of avian cholera carriers in this population. We were unable to isolate P. multocida serotype 1 from any of the birds sampled. Based on comparison with other waterfowl species, we concluded that these geese may be exposed to avian cholera during the winter or spring migration but are unlikely to play a significant role as carriers of the bacterium causing avian cholera.     

Characterization of Vibrio cholerae cultures isolated in foci of cholera in the city of Kazan

During the period of the registered outbreak of cholera in 2001 in Kazan 171 V. cholerae cultures were isolated in the focus of the infection (from patients, carriers and 7 environmental objects). The use of the basic and additional tests, including the polymerase chain reaction, made it possible to establish the circulation of V. cholerae, phagovar 15, in the focus of the infection. The strain isolated from the water reservoir Azino-1 in Kazan was identical in its properties to the epidemically dangerous strains isolated from patients. On the whole, the data obtained in the identification of the strains showed that the cultures isolated from patients, vibrio-carriers and environmental objects were identical.     

Hemolytic Vibrio cholerae O1 that is sensitive to Mukerjee's cholera phage IV and the phage produced by the hemolytic vibrio lysogenized with the infection of Mukerjee's cholera phage IV

Strains of hemolytic Vibrio cholerae O1 (El Tor vibrio) which are sensitive to Mukerjee's cholera phage group IV were isolated from cholera patients in North-East Thailand in 1986. Plaques of the phage on these hemolytic V. cholerae O1 were usually translucent but almost transparent on some strains, just like the plaques on non-hemolytic V. cholerae O1 (classical vibrio). These hemolytic V. cholerae O1 were lysogenized with the infection of cholera phage IV, and the lysogenized strains produced phage different from cholera phage IV. These hemolytic strains were classified into Cured type in prophage typing of V. cholerae O1, El Tor, because they were also lysogenized with Kappa phage and were hemolytic. When Cured-type V. cholerae O1, El Tor previously isolated in various countries were examined for the sensitivity to cholera phage IV, some of the isolates were sensitive.     

Titration of Cholera Antitoxin in Human Sera by Microhemagglutination with Formalinized Erythrocytes

Microtiter hemagglutination tests employing formalinized sheep erythrocytes sensitized with either crude or purified cholera toxin were used to assay the cholera antitoxin content of human sera. Comparable results were obtained with either crude or purified toxin-sensitized cells with the exception of two sera that gave unusually high hemagglutination titers with the crude toxin. Sera from 13 convalescent cholera patients showed a high degree of correlation between antitoxin levels as determined in vitro by the hemagglutination test and in vivo by the skin permeability factor neutralization test. Fourfold or greater rises in antitoxin levels between acute and convalescent sera were detected in 9 of 15 patients with bacteriologically proven cholera. No significant increases in titer were observed in 14 cases of noncholera diarrhea. Cholera antitoxin was detected by hemagglutination in only 1 of 33 sera, obtained from eight countries, containing vibriocidal antibodies. Formalinized sheep erythrocytes sensitized with toxin and stored at 4 C in the presence of 1:10,000 thimerosal were stable and sensitive for at least 6 months (the longest time tested).     

Trial of the toxins of the Sonne microbe and cholera vibrio in an isolated loop of mouse small intestine]

Enterotoxic properties of neuro-and endotoxins of Sonne dysentery bacillus, of cholerogen, and Bowen's cholera vibrio endotoxin were determined on a model of an isolated loop of the mouse small intestine. In definite doses all the mentioned preparations could induce dilatation of the isolated loop of the mouse small intestine. In case of Sonne bacillus neurotoxin this property correlated with the toxicity of the preparation for the animals and failed to depend on the virulence and biochemical reference of the strain from which it was obtained. Marked variations in the sensitivity of mongrel and linear mice to the mentioned preparations were noted.     

Characteristics of the heat-stable enterotoxin of NAG vibrios

Cell-free culture filtrate boiled for 15 minutes has been found to retain its biological activity in various experimental models used for the determination of the toxicogenicity of cholera vibrio filtrates. During gel filtration of the concentrated filrate o. NAG vibrio, strain NO. 9852, through Sephadex G-75 toxic activity could be detected in the free volume of the column, which was indicative of the fact that the molecular weight of the thermostable enterotoxin was about 70,000 daltons and greater. The methods of gel diffusion and aggregated hemagglutination have been used to show that the thermostable enterotoxin of NAG vibrio No. 9852 is immunologically unrelated to cholerogen. Some data obtained in experimental models suggest that the thermostable enterotoxin probably differs from cholera enterotoxin in the mechanism of its action.     

Antibody response to preS1 in hepatitis-B-virus-induced liver disease and after immunization

Antibodies to the preS1-encoded sequence of hepatitis B virus (HBV) envelope were detected by ELISA using a synthetic peptide analogue of preS1 proteins, in different groups of HBV-infected subjects and also in hepatitis B vaccine recipients. Such antibodies were specifically found in only 1 % of HBsAg chronic carriers including patients with cirrhosis and primary liver cancer. Anti-preS1 were detected in patients with acute hepatitis; in 13 % of the HBsAg⁺ sera obtained before recovery and in 37 % of the sera obtained after recovery.Anti-preS1 antibodies were detected in recipients of a plasma-derived vaccine, but not in those receiving a recombinant vaccine. The results indicate that anti-preS1 is an earlier serum marker of HBV clearance than anti-preS2 and anti-S antibodies.     

The isolation of Vibrio fluvialis on the territory of the USSR]

For the first time V. fluvialis strains were detected on the territory of the USSR. The taxonomic position of these vibrios was determined by their nucleotide DNA composition (the content of guanine + cytosine was 49.3-51.0 mole%) and the characteristic features of their phenotype. The individual features of the strains consisted in their capacity for agglutination with cholera antisera, groups 01 and Inaba, in diagnostic dilutions in the presence of differences in genomes and phenotypes with cholera vibrios. Molecular hybridization DNA-DNA also gave no confirmation of their relationship to cholera vibrios (23-26% homology). The comparative study of V. fluvialis strains from the USSR and other countries by a broader set of their phenotypical signs confirmed their identity.     

Basophil-sensitizing antibody response to herpes simplex viruses in rabbits.

Antibodies to herpes simplex virus type 1 and type 2 were detected in the sera of rabbits by release of histamine from basophils sensitized in vitro with the sera. The time course of the appearance of the antibodies, the dose-response curve of the release of histamine in relation to antigen concentration, the sedimentation characteristics of the antibodies in sucrose gradients, and the ability to destroy the sensitizing capacity of the sera with heat suggest that the antibodies being assessed were of the IgE class. These antibodies were induced in animals injected intradermally, intramuscularly, and i.p. with live virus. The antibodies were detected 1 week after primary injection and a similar time course of antibody appearance was observed after a second or third injection. The same cross-reactivity between type 1 and type 2 virus observed with IgG antibody was also observed with IgE antibody.     

Inhibiting effect of cholera vibrio neuraminidase in Rauscher mouse leukemia]

A study was made of the inhibitory effect of neuraminidase of cholera vibrio in mouse Rauscher leukemia. It was shown that the processing of cells of the spleen of mice suffering from leukemia with neuraminidase (in a dose of 50 units/ml and more) significantly inhibited the capacity of these cells to induce leukemia in their injection to BALB/c mice. In the mentioned concentration neuraminidase injected repeatedly parenterally produced no therapeutic effect in Rauscher leukemia.     

Antibody-dependent cellular cytotoxicity-inducing antibodies against human immunodeficiency virus. Presence at different clinical stages

The presence of antibodies mediating antibody-dependent cellular cytotoxicity (ADCC) against human immunodeficiency virus (HIV)-infected target cells was investigated with 170 sera from patients with varying severity of HIV infection. Approximately 40% of sera from individuals representing all stages of infection were ADCC-positive when tested against HTLV-IIIB infected 0937 clone 2 target cells. The positive sera had higher HIV antibody titers as measured by enzyme-linked immunosorbent assay compared with ADCC-negative sera. ADCC titers were lower in patients with acquired immune deficiency syndrome than in asymptomatic carriers. This decline in ADCC titer was not correlated with a general decrease of HIV antibodies. No correlation between the CD4:CD8 lymphocyte ratio and ADCC activity was found. The possible beneficial effect of ADCC-inducing antibodies early in infection is discussed in relation to the effect of ADCC-inducing antibodies in other retrovirus systems and to the nature of lentivirus infections.     

Systemic immunity of experimental animals immunized with cholera vaccines

The levels of antitoxic and vibriocidal antibodies in the sera of suckling rabbits after their parenteral immunization with cholera vaccine, cholera toxoid and a combination of cholera vaccine and toxoid were examined. Cholera vaccine induces intensive production of vibriocidal antibodies, and cholera toxoid, of antitoxic antibodies. The parenteral administration of the serum of rabbits immunized with cholera toxoid neutralized the action of cholera toxin in the small intestine of suckling rabbits. The complex preparation combines the properties of the corpuscular vaccine and the toxoid, inducing the production of both vibriocidal and antitoxic antibodies.     

Studies of immunological activity of Aspergillus flavus Link. III. Presence of antibodies against fungal antigens in the sera of persons from selected groups

The aim of this study was to determine the occurrence of antibodies against antigens of A. flavus (APP, AEM, AS, API), A. fumigatus and A. candidus. One hundred and fifty two sera of individuals connected with industrial environment were tested, in which A. flavus was permanently isolated: 339 sera of healthy controls-blood donors of city of Poznań, and 24 sera of patients with confirmed or suspected Aspergillosis were also included in the study. The sera were tested for a presence of specific antibodies by immunoprecipitation in 1% agar gel, by using inactivated sera and above mentioned antigens. In a group of people having permanent contact with A. flavus, antibodies to antigens derived from this genus were present in 4.6% of individuals while against A. fumigatus antigens in 0% and A. candidus 0.7%. In blood donors group 5 times lower percentage of sera having anti-A. flavus antibodies was found and a complete lack of detectable antibodies for other two genera. The results of the studies of patient sera indicate a necessity of broadening a set of fungal antigens used for an investigation of this type of sera. Antibodies against A. flavus were found in three patients and for A. fumigatus in 7 patients. One patient had antibodies for both genera and two patients had antibodies against A. flavus lacking antibodies against A. fumigatus. The results of this study indicate that antigens of A. flavus should be included into serodiagnosis of Aspergillosis.     

Hepatitis B virus markers, beta 2 microglobulin and anti HTLV in a population of blood donors from a prison environment

212 male prisoners were collected at the prison in March 1983. Anti-HBc and HBs Ag were detected by a combined test (AUSRIA-CORE, Abbott Laboratories). 67 sera (31,5%) were anti-HBc positive, 7 of them HBs Ag positive. Screening for anti-HTLV (anti-p24) was negative for all the donors. Beta 2 microglobulin levels were determined on 115 sera (B2-micro RIA 100, Pharmacia Laboratories). 8 had levels above 2,6 mg/l (greater than 2 SD). These 8 sera were anti-HBc positive and one of them HBs Ag positive 3 HBs Ag positive donors had non elevated Beta 2 microglobulin levels. This survey confirms that prisoners as blood donors should be regarded as carrying a high risk of transmission of HBV and probably other infectious agents with similar epidemiology. The signification of elevated Beta 2 microglobulin levels deserves further investigations since this determination could be of value as an additive test to increase the safety of blood products.     

Cholera and NAG vibrio utilization of different C-containing substrates in Hiss media and a medium with a limited mineral content

A total of 55 V. cholerae strains and 175 NAG vibrio strains were studied with a view to establish their capacity for utilizing citrate in Simmons citrate agar or for growing in it in the absence of the source of carbon. The strains were divided into 3 groups, each containing approximately an equal number of cholera and NAG vibrios irrespective of their origin or serovars. None of 50 signs used in this investigation permitted the reliable differentiation of the cholera and NAG vibrio groups due to considerable differences between the strains within each group. The use of Hiss medium with starch instead of Kodam medium is proposed for the determination of the diastatic activity of cholera and NAG vibrios.     

Construction of a genetic map of the chromosome of Vibrio cholerae based on conjugational crossings

The results of cholera vibrio chromosomal mapping using Vibrio cholerae classica and V. cholerae eltor donor strains obtained with the help of various R. plasmids, are summarized in the paper. A genetic map of V. cholerae chromosome was established showing the order of 35 gene markers. The relationship between the genetic structures of cholera eltor and classical vibrio biotypes is discussed.