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Isolation and characterization of nutritionally variant streptococci from animal sources.

Ten nutritionally variant streptococci were recovered from clinical specimens submitted to the Kansas State University Veterinary Clinical Bacteriology laboratory over a 4-year period. Isolates were recognized visually on primary blood agar plates by their satellite growth around a previously overlaid Staphylococcus aureus culture. All isolates grew within 24 hours in Todd-Hewitt and heart infusion broths supplemented with 5% bovine fetal serum and 5% S aureus filtrate. They also grew anaerobically in supplemented broths within 48 hours. However, isolates did not grow aerobically or anaerobically in the absence of supplements up to a 7-day postinoculation period. As determined by the standard Kirby-Bauer technique, the isolates were highly susceptible to antimicrobial agents commonly recommended in veterinary medicine. The isolates did not react with the corresponding Lancefield group-specific antisera, as tested by the capillary precipitin test.

Anaerobiosis↗

[Diagnostic valve of ELISA-IgG for trichinosis using Melcher's antigen].

An enzyme-linked immunosorbent assay (ELISA) for trichinosis using a Melcher's antigen was developed for the detection of IgG antibodies in 41 serum samples from patients confirmed or suspected to have trichinosis by strong clinical and epidemiological evidences. ELISA- IgG was compared with a precipitin test (PT), a bentonite floculation test (BFT) and an indirect hemagglutination test (IHAT). The cut-off value was determined using serum samples from 67 apparently healthy persons employing two serum dilutions (1:100 and 1:500) with three standard deviations (SD). The sensitivity of ELISA- IgG was 97.6% and 95.2% using serum dilutions of 1:100 and 1:500 respectively, whereas the values for the other tests were: PT (92.7%), BFT (63.4%) and IHAT(85.4%). According to MacNemar test, ELISA-IgG did not present statistical significance (p > 0.05) compared with PT and IHAT, whereas all of them were different compared with BHT (p > 0.01). In order to find out the specificity of ELISA-IgG, additional 124 serum samples from individuals with other parasitoses, such as cysticercosis (31), fascioliasis (17), hydatidosis (51) and toxocariasis (25) were also tested. ELISA-IgG presented a specificity of 99.5% with both serum dilutions. The positive predictive values were 97.6 and 97.5%, whereas the negative one were 99.5 and 99.0% for 1:100 and 1:500 serum dilutions respectively. The use of ELISA-IgG and Melcher's antigen in the diagnosis of human trichinosis is discussed.

Adult↗

Goat serums for fluorescent antibody conjugates to chlamydial antigens.

Serums from goats hyperimmunized with Chlamydia psittaci consistently produce antichlamydial fluorescent antibody conjugate of high titer. The titer of the fluorescent antibody conjugate prepared from a given serum correlated well with the titer obtained by agar gel precipitin, but not with the complement fixation. The agar gel precipitin test can be used to predict whether a given serum is satisfactory for use in production of a conjugate for direct fluorescent antibody tests. Serums with an agar gel precipitin titer of 1/8 or higher generally produce a usable fluorescent antibody conjugate. Labeling gamma globulins with fluorescein isothiocyanate at a ratio of 1/150 resulted in satisfactory fluorescent antibody conjugates. Cultures of Vero cells infected with chlamydiae were found to be suitable for titration of the fluorescent antibody conjugates.

Animals↗

Marek's disease virus (MDV) antigens in the feather follicle epithelium: difference between oncogenic and nononcogenic MDV.

Feather tips from chickens infected with oncogenic or nononcogenic Marek's disease virus (MDV) were individually tested for MDV antigens by the agar-gel-precipitin test to compare antigen contents of feather follicles infected with oncogenic or nononcogenic MDV. There was a significant difference between oncogenic and nononcogenic MDV in the development of MDV antigens in the feather follicle epithelium of infected chickens. Significantly more feather tips from chickens infected with oncogenic MDV were positive for MDV antigens, particularly at 3 and 6 weeks postinoculation, whereas the feather tips from those infected with nononcogenic MDV were rarely positive, whether tested with the homologous or heterologous anti-MDV serum. Such a difference was consistent in separate trials employing three different strains of chickens. Infectious cell-free MDV was extracted more frequently from the feather tips of chickens infected with oncogenic MDV than from those of birds infected with nononcogenic MDV, and the virus titers of extracted MDV were always numerically or statistically higher in birds infected with oncogenic MDV.

Animals↗

Studies of Candida serology in denture stomatitis patients.

The aim of the study was to determine the humoral antibody response to Candida by means of hemagglutinin and precipitin tests and to correlate these results to yeast colonization in the oral sites and feces and to the relapse tendency after treatment; and furthermore, to evaluate whether Candida serology can serve as a tool to predict a denture stomatitis risk group. The serologic tests were performed by means of passive hemagglutination (HA) and immunoelectroosmophoresis (counterimmunoelectrophoresis, CIE) tests in 51 denture stomatitis patients and compared with 25 individuals in the reference group. There were significantly more patients with increased hemagglutinin titers (pos. greater than or equal to 1:160) to Candida albicans polysaccharide (mannan) antigen and/or precipitin to crude C. albicans cytoplasmic antigen in serum (P less than 0.01) than in the reference group. In the denture stomatitis group there was no correlation between the presence of hemagglutinating and precipitating Candida antibody in serum and the parameters yeast colonization of the oral sites/feces, yeast score on the denture base and palatal erythema score. After local oral antimycotic treatment there was no significant change in the serum Candida hemagglutinin titers and serum precipitins. Serologic tests might be useful as a guidance in a selected group of patients for continued prosthetic treatment and as a prognostic instrument.

Adult↗

ORIGIN AND PROPERTIES OF NATURALLY OCCURRING HAPTEN FROM ESCHERICHIA COLI.

Anacker, R. L. (National Institute of Allergy and Infectious Diseases, Rocky Mountain Laboratory, Hamilton, Mont.), R. A. Finkelstein, W. T. Haskins, M. Landy, K. C. Milner, E. Ribi, and P. W. Stashak. Origin and properties of naturally occurring hapten from Escherichia coli. J. Bacteriol. 88:1705-1720. 1964.-Haptens found in preparations of endotoxin and in fractions of disrupted cells, particularly one termed "native hapten," which appeared to be associated with the protoplasm of cells rather than with cell walls, have been further investigated with a view to establishing their origin and composition as well as their host-reactive properties. For this purpose, cells from a smooth strain of Escherichia coli O111:B4 were either extracted directly or disrupted and separated into cell-wall and protoplasmic fractions. Haptens were obtained by gel filtration of endotoxins, by trichloroacetic acid extraction of protoplasm, and by a mild acid hydrolysis of endotoxin. Several lines of evidence indicated that native hapten originated in the protoplasm rather than by autolysis or degradation of cell-wall endotoxin during procedures employed in disruption. In gel diffusion and quantitative precipitin tests, no hapten was identical with endotoxin, but native hapten was serologically the most complex of the haptens and precipitated the most antibody. Native and acid haptens, on a weight basis, fixed about 1% of the quantity of complement fixed by homologous endotoxin. Haptens did not stimulate the production of antibodies in mice or rabbits and did not elicit endotoxic host reactions. Chemically, native hapten differed from endotoxin and from acid hapten in that it lacked phosphorus, heptose, long-chain fatty acids, and 2-keto-3-deoxyoctonate. These substances did not appear to be determinants of antigenic specificity, but they may provide necessary bonds for assembling hapten-like units into fully antigenic and toxic macromolecules.

Animals↗

Isolation and antigenic characteristics of axial filaments from the Reiter Treponeme.

Axial filaments were isolated and purified from Reiter treponemes after detergent solubilization of the cells" outer envelope. The axial filaments were separated from the spirochetal cells by shearing, purified by density gradient centrifugation, and fragmented by ultrasonication. Acrylamide gel electrophoresis of dissociated filaments revealed two major protein bands. Gel diffusion precipitin tests and immunoelectrophoresis between a purified axial filament suspension and anti-Reiter treponeme serum gave a single precipitin line. Checkerboard complement fixation tests also gave results consistent with a single antigen-antibody system. Tests with immune sera to other cultivable spirochetes were positive with some and negative with others. In addition, strongly positive reactions were obtained in complement fixation and precipitin tests with sera from rabbits and humans with syphilis and other treponematoses. However, both serological tests gave reactions of partial identity between the antigen(s) of Reiter treponeme axial filaments and those of the pathogenic treponemes. It was concluded from these studies that the axial filaments were probably the cellular locus of the so-called "Reiter protein" antigen of syphilis serology.

Animals↗

IMMUNOLOGICALLY ACTIVE POLYSACCHARIDES FROM NOCARDIA ASTEROIDES AND NOCARDIA BRASILIENSIS.

Zamora, A. (University of Mexico, Mexico, D.F.), L. F. Bojalil, and Fernando Bastarrachea. Immunologically active polysaccharides from Nocardia asteroides and Nocardia brasiliensis. J. Bacteriol. 85:549-555. 1963.-Two immunologically active polysaccharides were isolated from Nocardia asteroides (Poly I Na and Poly II Na) and N. brasiliensis (Poly I Nb and Poly II Nb). These polysaccharides were isolated from cell extracts and purified by methanol precipitation, chloroform extraction of extraneous material, and deproteinization with trichloroacetic acid. The crucial step used for separation of Poly I and Poly II from both nocardias was differential solubility. From dried preparations containing both polysaccharides, Poly I was solubilized at pH 10, whereas Poly II remained insoluble and was subsequently solubilized at pH 5. Poly I Na and Poly I Nb are apparently the same. Arabinose and galactose were the monosaccharide constituents of these polysaccharides, and their molar ratios were similar. Furthermore, Poly I Na and Poly I Nb cross-reacted in agar diffusion precipitin tests with rabbit antisera prepared against either N. asteroides or N. brasiliensis. Either polysaccharide absorbed serum antibodies against the other. These polysaccharides can be regarded as groupspecific. Poly II Na and Poly II Nb are different and species-specific. They are composed of arabinose, galactose, and mannose but exhibit different molar ratios of these sugars according to species. They reacted only with homologous antisera.

Allergy and Immunology↗

The efficacy of human schistosomicide treatment may depend on the rate of transmission.

The efficacy of different treatment protocols in humans infected with Schistosoma mansoni at sites with different transmission conditions was evaluated by the disappearance of anti-worm intestine IgM antibodies in an indirect fluorescence antibody test (IgM-IFT) and anti-egg antibodies in the circumoval precipitin test (COPT). Patient sera coming from sites of active low transmission (ALT), active high transmission (AHT) and low interrupted transmission (LIT) from Venezuela were studied. Chemotherapy protocols were (1) ALT, 60 mg/kg praziquantel (Pzq60); (2) AHT, one dose of 40 mg/kg Pzq followed by one dose of 20 mg/kg oxamniquine for one group and one dose of 40 mg/kg Pzq alone for the other group; (3) LIT, one dose of 40 mg/kg Pzq repeated every 3 months up to three doses. Cure rates occurred mostly between 3 and 12 months with the exception of Pzq60-ALT where it was evident before 3 months. Higher cure rates were evident in both places of low transmission (ALT and LIT) and the lowest in the AHT regardless of the treatment protocol. Cure was more evident with COPT compared to IgM-IFT. The rate of serological cure appears then to depend on the previous state of transmission. The differential cure rate evaluated by both techniques is probably due to the persistence of antibodies against antigens in different stages of the parasite.

Animals↗

[Cercaria dermatitis (swimmer's itch). Case report of cercaria dermatitis caused by Trichobilharzia (Digena, Schistosomatidae)].

A 63-year-old woman developed a pruritic exanthema on the extremities after one hour of work in her garden pond in Planegg, Southern Bavaria. As the appropriate vectors (ducks and snails) were present, we made the tentative diagnosis of cercarial dermatitis (swimmer's itch). By different serological methods (cercarial fluorescent antibody test, cercarial Hüllen reaction, circumoval precipitin test) antibodies against cercariae could be demonstrated in the serum of the patient 14 days later. Cercarial dermatitis appears worldwide, but in Central Europe the disease is often not recognized.

Animals↗

Immunochemical studies of antitoxin produced in normal and allergic individuals hyperimmunized with diphtheria toxoid. IV. Differences between human precipitating and non-precipitating skin-sensitizing diphtheria antitoxin as shown by electrophoresis.

Electrophoresis on a starch-supporting medium was used to fractionate sera containing human diphtheria antitoxin of the following varieties (a) precipitating antitoxin, (b) non-precipitating skin-sensitizing antitoxin, and (c) mixtures containing precipitating and skin-sensitizing antitoxins. Aliquots of the protein fractions thus separated were tested for activity using the rabbit toxin neutralization test, precipitin techniques, and passive transfer tests in human skin. Non-precipitating, skin-sensitizing diphtheria antitoxin migrated largely as a fast moving gamma (gamma(1)) globulin. Passive transfer studies of isolated antitoxic fractions showed that they were as potent as whole serum in the ability to cause immediate wheal reactions. These fractions were not precipitable using appropriate quantities of purified toxoid. Precipitating diphtheria antitoxin migrated largely as a slow moving gamma (gamma(2)) globulin. Isolated antitoxic fractions of appropriate strength obtained from representative sera were precipitable by toxin and were unable to cause immediate wheal reactions upon toxoid challenge in human recipients. Mixtures of skin-sensitizing and precipitating antitoxins were separable by the technique of starch electrophoresis. The individual components removed from mixtures by this method retained the properties by which they could be characterized in whole serum.

Antibodies↗

Antigenic relationship between the common antigen (OEP) of Pseudomonas aeruginosa and Vibrio cholerae.

Antibodies were found by the OEP-passive hemagglutination test to cross-react with the common antigen (OEP) of Pseudomonas aeruginosa in sera of rabbits immunized with two serotype (Inaba and Ogawa) strains of Vibrio cholerae. The titer in the OEP-passive hemagglutination reaction rose later than did the agglutinin titer and reached a peak of 640 to 1,280. The titers of OEP antibody formation in rabbits immunized with V. cholerae were almost the same as that of P. aeruginosa. The common antigen of P. aeruginosa was confirmed to exist serologically in both strains of V. cholerae as determined by the indirect fluorescent antibody test and the agar gel precipitin test. Passive immunization with the V. cholerae immune rabbit serum significantly protected mice against P. aeruginosa infection. Purified antibodies cross-reacting with the OEP of P. aeruginosa derived from the V. cholerae immune rabbit sera by OEP-coupled affinity chromatography protected mice against P. aeruginosa infection as compared with the control group, which was injected with 100 microgram of immunoglobin G not containing OEP antibody. The purified antibodies (2.5 microgram per mouse) protected animals challenged with approximately 10,000 50% lethal doses in the control group. Consequently, the common antigen (OEP) of P. aeruginosa proved to be a common antigen of V. cholerae both serologically and in possessing infection protective properties.

Animals↗

Serological response of rhesus monkeys to histoplasma, Blastomyces, and Coccidioides antigens.

Fifteen adult rhesus monkeys were inoculated with nonviable Histoplasma capsulatum, Blastomyces dermatitidis, or Coccidioides immitis. Antibody assays were made periodically during a 2-year period by use of a complement-fixation (CF) test employing four antigens and a latex-agglutination test. Selected sera were also studied in an immunodiffusion test and a coccidioidin-precipitin test. The serological patterns obtained with the anti-Histoplasma, anti-Blastomyces, and anti-Coccidioides monkey sera were comparable to those of sera from patients with diseases caused by the respective organisms. Rhesus monkeys should provide a good laboratory model for additional studies, including the influence of multiple antigenic stimuli on serologic response and patterns. Monkeys could also be used for the production of antisera required for studies to improve the specificity of the currently available CF antigens.

Animals↗

CAMP-disk test for presumptive identification of group B streptococci.

The modification of the CAMP test for goup B streptococci involved substituting a paper disk impregnated with partially purified beta-hemolysin for the staphylococcal culture that was the source of beta-hemolysin in the original test. The disk is placed onto a sheep blood agar plate beside the streak of Streptococcus being tested. The plate is then incubated aerobically at 35 degrees C. A positive reaction consists of a lunar-shaped clear zone that appears within 24 h in the dark beta-hemolysin zone surrounding the disk. A double-blind study of 135 randomly coded streptococcal isolates showed complete agreement between the CAMP-disk test and the standard Lancefield precipitin test. All group B streptococci tested had positive reactions, and all strains tested from streptococcal groups A, C, D, and G were negative. The CAMP-disk test is a simple and convenient way to identify presumptively group B streptococci.

Bacteriological Techniques↗

[Prevalence of extrinsic allergic alveolitis in cattle breeders from the province of Reggio Emilia].

BACKGROUND AND OBJECTIVES: Several new cases of Extrinsic Allergic Alveolitis or Farmer's Lung (FL) in farm workers were reported to Occupational Health Services in the province of Reggio Emilia (Italy). This prompted the Public Health Department to study the prevalence of the disease among milk-cow breeders involved in Parmigiano-Reggiano cheese production: who are the biggest hay users. METHODS: A suitable questionnaire was sent to 1875 farmers in three of the six districts of the province. Half of them (935: 841 males, 94 females) answered; further contacts and medical history research revealed 33 case of "likely FL". Twenty-three (2 females) (10 "missing"), underwent pulmonary function tests, chest X-rays, precipitins tests against Saccharopolyspora Rectivirgula and other fungal antigens and (22 farmers) bronchoalveolar lavage (BAL). RESULTS: According to the "Società Italiana di Medicina del Lavoro e di Igiene Industriale" diagnostic standards, we found 20 subjects suffering from FL among farmers collecting hay in large cylindrical (round) bales, dried on field (2.6%) and among others still using small (traditional), prismatic bales (0.5%). CONCLUSIONS: The prevalence on the whole exposed population (6000-9000 people) was estimated between 1.5% and 3.0% (90-270 people); no difference was found in FL prevalence between flat and hilly or mountain areas; the method of collecting hay in big "round" bales, dried on field, seems to produce higher frequencies of FL cases if compared with the traditional ones (more frequent in mountain areas). The new hay packing methods, using forced air driers, are suggested as a possible solution.

Adult↗

Comparisons of serologic responses of white Leghorn and New Hampshire red chickens to purified lipopolysaccharides of Pasteurella multocida.

White leghorn and New Hampshire red chickens were inoculated with purified lipopolysaccharides of 14 serotypes of Pasteurella multocida to determine their ability to produce serotype-specific antisera for somatic antigen typing. Specific antisera were made by both breeds of chicken to lipopolysaccharides of serotypes 1, 3, 4, 6, 8, and 16. No specific antisera were made against lipopolysaccharides of serotypes 2, 5, 7, 12, and 14. Lipopolysaccharides of serotypes 10 and 11 failed to stimulate antibody production. White leghorns were more responsive than New Hampshire red chickens. White leghorn antisera had higher titers to lipopolysaccharides in passive hemagglutination tests and produced more intense precipitin reactions with heat-stable antigens in the gel-diffusion-precipitin test.

Animals↗

An enzyme-linked immunosorbent assay for the detection of antibody against avian influenza virus.

An enzyme-linked immunosorbent assay (ELISA) was developed for detecting antibody to type A avian influenza (AI) virus. The sensitivity and group specificity of the AI-ELISA were compared with those of the agar-gel-precipitin test (AGPT) and the hemagglutination-inhibition (HI) test under conditions of both controlled and field exposure. During the course of temporal experimental infection (0-76 days) of specific-pathogen-free (SPF) chickens with AI subtype Hav9N2, the AI-ELISA was able to detect specific AI antibody as early as 8 days postinoculation (PI), and it measured rising levels of antibody through 35 days PI, at which time the chickens were re-exposed to AI virus. Conversely, AGP tests were negative through 35 days PI, and HI tests began to detect low levels of AI antibody only at 21 days PI. Following a secondary infection at 35 days PI with the same AI subtype, all tests measured rising levels of AI-specific antibody (35-76 days PI). However, the AGP test was positive at only the 7- and 14-day samplings postsecondary immunization. Under field conditions, the AI-ELISA was able to detect serum AI antibody in flocks from which highly pathogenic AI was isolated, but the AGP tests of these sera were negative.

Animals↗

ISOLATION OF L FORMS FROM GROUP A STREPTOCOCCI EXPOSED TO BACITRACIN.

Rotta, Jiri (Washington University School of Medicine, St. Louis, Mo.), Walter W. Karakawa, and Richard M. Krause. Isolation of L forms from group A streptococci exposed to bacitracin. J. Bacteriol. 89:1581-1585. 1965.-L forms were obtained from group A streptococci by exposure to bacitracin on gradient plates, although novobiocin was ineffective in this respect. Subcultures of these L forms had morphological and bacteriological properties similar to those obtained with penicillin. M protein was detected in L-form colony smears by immunofluorescent staining with type-specific conjugate. The L forms were not stained with group-specific conjugate. Parallel precipitin tests performed with extracts from a heavy growth of L forms on agar confirmed these findings. Thus, the L forms obtained with bacitracin continue to produce M protein but are devoid of the group-specific carbohydrate which is a major component of cell-wall structure.

Bacitracin↗