Carbamoyl phosphate biosynthesis in Streptococci

Nine streptococcal strains belonging to different serological groups (A, B, C, D) were shown to synthesize carbamoyl phosphate from ammonium hydrocarbonate and ATP. The reaction was catalyzed by carbamate kinase (EC 2.7.2.2). The speed of the reaction was evaluated according to the increase of the content of citrullin (the combination of carbamate kinase and ornithine transcarbamoylase). The representatives of different serological groups were found to have quantitative differences in carbamate kinase activity: the highest specific activity (13 nmol of citrullin per minute in 1 mg of dried microbial biomass) was detected in group A streptococci, while group D streptococci showed the lowest specific activity (0.5 nmol).     

Streptococcus group B in meningitis and infection of newborn infants

Streptococci were isolated from the liquor or blood of 102 newborn infants and 16 infants in the first month of their life, suspected of having purulent meningitis, in 22 cases (18,5%). 5 isolated streptococcal strains were classified with group B on the basis of their cultural, biochemical and serological features. All of these strains were isolated from newborn infants during the first 3-4 days of their life. The occurrence of group B streptococci among all examined newborn infants was 4.8%; among the newborns with the positive results of bacteriological examination (73 infants) this figure was as high as 6.8%. The authors emphasize the necessity of producing, on an industrial scale, diagnostic preparations for the identification of group B streptococci playing a significant role in septic diseases and meningitides in newborns.     

Nucleic acid hybridization and cell wall composition studies of pyogenic streptococci

Abstract DNA-rRNA and DNA-DNA hybridization studies indicate that the classical pyogenic streptococci can be divided into five homology clusters. Based on these studies the term pyogenic streptococci should be confined to the first cluster consisting of serological groups A, A-variant, C, G ('large' colony, type II) and L.
Streptococci of serological groups B and M form the second cluster. The third cluster is composed of streptococci of serological groups R and S and serological groups U, V and P are found in the fourth cluster. The fifth cluster comprises strains of Streptococcus anginosus S. intermedius, Streptococcus MG and serological groups G ('minute' colony, type I) and F (type I). Most of the test strains contain the peptidoglycan type Lys-Ala_1–3. Only streptococci of serogroups R and S reveal a directly cross-linked peptidoglycan. Rhamnose was found as characteristic component of all cell wall polysaccharides. The impact of our results on the systematics of classical pyogenic streptococci will be discussed.     

Rapid Extraction Method with Pronase B for Grouping Beta-Hemolytic Streptococci

The enzyme, Pronase B, was used to extract the C carbohydrates of betahemolytic streptococci for serological grouping. Reference strains of streptococci, groups A through N, were accurately grouped by using this extraction method. Of the beta-hemolytic streptococci isolated from patients' cultures, 1,197 were classified as either group A or not group A by this method. There was 100% correlation with the reference Lancefield method. In contrast, false reactions occurred with the presumptive bacitracin disc method in 4.1% of the group A and 12.7% of the non-group A cultures. The Pronase method proved simple, accurate, and readily adaptable to the clinical laboratory routine.     

Incidence of aflatoxin producing strains and aflatoxin contamination in dry fruit slices of quinces (Cydonia oblonga Mill.) from the Indian state of Jammu and Kashmir

An investigation was undertaken to obtain data on the occurrence of aflatoxins and the aflatoxin producing potential of Aspergillus flavus strains isolated from dry fruit slices of quinces produced in jammu and Kashmir, India. A total of 147 A. flavus isolates recovered from dr fruit slices were grown in liquid rice flour medium and screened for the production of various aflatoxins by thin layer chromatography. The results showed that 23.14% of the tested isolates were aflatoxigenic, producing aflatoxins B1 and B2 in varying amounts. Aflatoxins G1 and G2 were not detected. All 25 of the investigated market samples were also found to be aflatoxin B1 positive and the level of contamination ranged from 96 to 8164 micrograms/kg of the dry fruit which is quite high in comparison to the permissible level of 30 ppb. As per these results biochemical composition of dry fruit slices of quinces, along with climatic conditions seem to be very favourable for aflatoxin production by the toxigenic A. flavus strains. Therefore, monitoring of aflatoxins in dry fruit slices of quinces is recommended for this region.     

Potential use of serological methods in detecting cholera toxin

In the study of 50 Vibrio cholerae museum strains, 45 of them producing cholerigenic effect in suckling rabbits, cholera toxin, determined by means of the passive immune hemolysis (PIH) test, has been detected in the supernatant of the culture fluid of only two strains: V. cholerae 569 B, a well-known producer of cholera toxin, and V. cholerae (eltor) 1310, from whose population a toxigenic variant has been obtained by selection. To study the capacity of V. cholerae for producing toxin in vitro, in six cholerigenic strains, besides the supernatant of their culture fluids, also protein fractions, cell lysates and membrane fractions have been studied in the PIH test. In all these strains cholera toxin has been detected only in membrane fractions, which should be taken into consideration in the serological evaluation of the toxigenicity of V. cholerae.     

Septicaemic infections in pigs,caused by haemolytic streptococci of new Lancefield groups designated R,S and T

Pyogenic streptococci isolated from outbreaks and from sporadic infections in pigs and piglets were characterized by the almost unique combination of the properties of -haemolysis on horse blood agar and acid production from inulin. Three new serological groups were recognized, each with a single antigen different from those of any of the Lancefield groups. These antigens are polysaccharides located in the cell wall. About half the number of haemolytic streptococcal strains isolated from pigs were group R or group S streptococci, a few strains were group T streptococci, and the remaining strains were group L streptococci,S. equisimilis, group E streptococci, or group P streptococci, in this order of frequency. Only one out of about 150 haemolytic strains could not be identified serologically. Group R and group T streptococci differ from group S streptococci by acid production from raffinose. Infections with group R streptococci appeared to occur independently of age, whereas infections with group S streptococci were almost entirely confined to piglets.     

The Use of Standardized Reference Antigens in the Serogrouping of Streptococci

Streptococci isolated from the dental plaque of five animal species were identified by physiological and serological methods. Twenty-nine strains of streptococci considered to resemble Streptococcus mitior or Strep. bovis on the basis of physiological data reacted with Lancefield group B or group K antisera respectively in tube precipitation tests. Further serological studies with standardized antigens from known serogroup B and K streptococci revealed that only three of these 29 isolates had been serogrouped accurately and carried the appropriate group antigen. Comparisons were made between the reactivity of the antisera produced by Difco and Wellcome Reagents with acid and autoclaved extracts of the strains. It was shown that the accuracy of serogrouping such isolates could be improved if the tests were made in a gel diffusion system that included a reference antigen.     

Pheno- and genotypic properties of streptococci of serological group B of canine and feline origin

In the present study streptococci of serological group B isolated from canines (n=48) and felines (n=7) were comparatively investigated with group B streptococci from humans and bovines for cultural, biochemical and serological properties for antibiotic resistancies and by molecular analysis. An identification was performed with group B-specific antiserum, biochemical reactions, by PCR amplification and subsequent endonuclease digestion of the 16S rRNA gene and by amplification of species-specific parts of the 16S rDNA the 16S-23S rDNA intergenic spacer region and the CAMP factor gene cfb. Phenotypic similarities of group B streptococci of canine and feline origin with group B streptococci from humans and differences to group B streptococci of bovine origin could be observed in lactose fermentation, serotype patterns, pigmentation, growth properties of the bacteria in fluid medium and soft agar, hemagglutination reactions and in minocycline and tetracycline resistance. A negative hyaluronidase plate test, a hylB amplicon with a size of 4.6 kb and an insertion sequence 1548 could be observed among canine, feline and human group B streptococci of serotype III. The remaining hyaluronidase positive strains, also including all isolates of bovine origin, had a hylB gene with a size of 3.3 kb. Further genotypic differences could be observed in the occurrence of the genes lmb and scpB which appeared generally among canine, feline and human group B streptococci, but less pronounced among bovine isolates of this species. According to the presented data group B streptococci of canine and feline origin seemed to be more related to human than to bovine isolates of this species possibly indicating some epidemiological relation.     

A rapid method of grouping beta-hemolytic streptococci using extemporaneous coagglutination

Extemporaneous coagglutination procedure for the serological grouping of beta-hemolytic streptococci is reported. Streptococcal group antigens were extracted with nitrous acid. 250 strains of groups A, B, C, F and G streptococci were tested with this method. An agreement of 100% was found between this method and the Lancefield capillary precipitation procedure. Extemporaneous coagglutination method was found to be rapid, reliable, easy and economical and could be adopted in any routine diagnostic laboratory.     

Study on pectinase and sclerotium producing abilities of two kinds ofAspergillus flavus isolates from Zhejiang

Pectinase and sclerotium production by strains ofAspergillus flavus were determined with a pectinase culture plate assay and a Cz 3% NaNO₃ medium plate assay. In theA. flavus population, 51% of isolates produced sclerotia, the toxigenic strains showing a tendency to have smaller sclerotia. Strains producing both abundant small sclerotia and a large quantity of aflatoxin were not found. There was no linear correlation between the amount of aflatoxin produced and the number of sclerotia. Levels of pectinase produced by the toxigenic strains were higher than that of the non-toxigenic strains, and this character was more obvious in the sclerotium-producing strains than in the non-sclerotium-prodcing strains. In theA. flavus population from Zhejiang in which the toxigenic strain rate was low, toxigenic strains may require higher levels of pectinase to compete with the non-toxigenic strains when infecting foodstuffs.     

New serotype of mutans streptococci isolated from pig oral cavity

Gram-positive streptococcal mutans-like strains, but with clearly different colony formation than S. orisuis on Mitis Salivarius agar, were isolated from the pig oral cavity and identified by 16S rRNA sequencing, G+C content, DNA-DNA homology and extensive biochemical and serological testing. The phenotypic data showed that the strains were similar to S. orisuis except for susceptibility to bacitracin. DNA-DNA homology between the isolates and S. orisuis was 72∼81%. However, serological data showed that they have a different sero-specific antigen from S. orisuis and other mutans streptococci. A new serotype, designated p, strains are classified in a serovar of S. orisuis, one of mutans streptococci.     

Evaluation of the usefulness of selected commercial systems for identification of Streptococcus species

The usefulness was assessed of three commercially available systems for rapid identification of streptococcal strains. The studied material comprised 68 strains of streptococci and enterococci (including 24 standard strains) belonging to serological groups: A (14 strains), B (10), C (11), D (10), F (3) and G (10), as well as 10 S. pneumoniae strains. The strains were isolated from throat, nasal, wound swabs, blood, pus of inpatients and throat and nasal swabs of outpatients. For the identification of streptococci 3 commercially available systems were used: API 20 STREP (bioMerieux, France), rapid ID 32 Strep (bioMerieux, France), Streptoplast PPL 18 (HTL, Poland). The determinations were done according to producer's instructions. The highest percent of correctly identified strains was obtained with the rapid ID 32 Strep--80.9%, with the API 20 STREP--76.4% strains were identified correctly and with the PPL 18--61.8%. The study showed that the API 20 STREP and rapid ID 32 STREP are suitable for the identification of streptococcal strains from groups: A, B, C, D, F and enterococci--group D. The proportions of correctly identified strains from these groups with the Streptoplast PPL 18 were lower than those determined with the bioMerieux systems. Using of three identification systems streptococci from group G and S. pneumoniae strains cannot be identified.     

Comparative biological characteristics of the L-form of Streptococcus group A and B

The comparative study of the biological and serological properties of the L-forms of streptococci, groups A and B, has been made. Their morphological similarity on the level of light microscopy has been demonstrated. The use of ring precipitation, gel diffusion, passive hemagglutination, aggregate hemagglutination, as well as the immunoferritin technique, has made it possible to establish the presence of specific antigens in the L-forms of streptococci, groups A and B. Serological cross reactions are negligible. The future development of a diagnosticum for the specific indication of these antigens is proposed. The fact of the presence of specific antigens in the L-forms of streptococci in comparison with the initial streptococcal strains has been confirmed.     

Serologic pattern of meningococci in a focus of infection]

The authors present the results of typing of meningococci (isolated from the patients and carriers) by the sera of serological groups A, B, C, D, X, Y, and Z made at the Leningrad Institute of Vaccine and Sera. The causative agents of serological group A were most frequently isolated from the patients with the generalized forms of meningococcus infection. The percentage of detection of meningococci of serological group A was the greatest in the patients and much less in the carriers in and outside the foci of this infection. Many dissociated cultures were revealed among the strains isolated from the carriers. Sera of the Leningrad Institute of Vaccine and Sera have permitted typing of meningococci cultures belonging to various serological groups in accordance with the international classification.     

Lysis of the cellular walls of Streptococcus group A by enzymes produced by actinomycetes]

More than 80 cultures of actinomycetes belonging to different taxanomic groups were studied with a purpose of screening actinomycetes actively producing enzymes lyzing the cell walls of group A streptococci. 31 strains of the actinomycetes producing enzymes which lyzed the cell walls by 20-50 and 60-80 per cent within 1 and 4 hours respectively were selected. The proteolytic activity of the enzymes produced by these strains was also studied. It was shown that 4 cultures, i.e. Actinomyces albus, strains 6 and 9, Actinomyces levoris, strain 29 and Actinomyces gibsonii, strain 42 were of interest as organisms producing enzymes which lyzed the streptococcal cell wall without impairing its antigenic components.     

Characteristics of beta-hemolytic streptococci secreted in children in various climatic zones]

A study was made of the level of carrier-state, serological characteristics, virulence and toxigenicity of beta-hemolytic streptococci isolated from children in two towns with a different incidence of scarlet fever located in the contrast climatic zones. It was found that the level of carrier-state of beta-hemolytic streptococci and biological properties of the circulating strains were important factors associated with the incidence of scarlet fever.     

Identification with the aid of various serological reactions of L forms of streptococci isolated from the live organism]

Growth inhibition, agglutination, precipitation, and passive hemagglutination tests were used for the identification of the L-forms of streptococci isolated from the organism of experimental rabbits both after the infection with the L-forms of streptococci and with the streptococci of group A. The tests were positive not only with the antiserum of homologous, but also of heterologous strains of the L-form of streptococcus, group A. The L-form cultures isolated from the experimental animals failed to differ from the laboratory strain of the L-forms of streptococcus, group A, by serological properties.     

Serological Studies of Clostridium botulinum Type E and Related Organisms II. Serology of Spores

Pure spore antigens for the immunization of rabbits were prepared by enzymic digestion of vegetative components and separation of the cleaned spores in polyethylene glycol. Spore antisera were prepared to strains representative of toxigenic Clostridium botulinum type E; nontoxigenic boticin E-producing variants; nontoxigenic nonproducers of boticin E; nontoxigenic "atypical" strains, which differ somewhat from C. botulinum type E in their physiology; C. botulinum types A and B; and C. bifermentans. They were tested against these and additional strains representative of the above groups, other types of C. botulinum, and other Clostridium species. There was no evidence of agglutination of flagellar or somatic antigens of vegetative cells by these antisera. Agglutination and agglutinin absorption tests showed common antigens among toxigenic type E strains and nontoxigenic variants, both producers and nonproducers of boticin E. Some nontoxigenic "atypical" strains varied in their ability to be agglutinated by type E antisera, and others did not agglutinate at all. Of those atypical strains that were not agglutinated, one was agglutinated by C. bifermentans antiserum. Antisera prepared against C. botulinum types A and B and C. bifermentans did not agglutinate the spores of type E or its variants nor share antigens common to each other. Similarly, antisera to type E, its nontoxigenic variants, and nontoxigenic atypical strains did not agglutinate other C. botulinum types or any other Clostridium species investigated.     

Erythrogenic toxins A, B and C: Occurrence of the genes and exotoxin formation from clinical Streptococcus pyogenes strains associated with streptococcal toxic shock-like syndrome

We report the study of 53 clinical isolates of group A streptococci, all from patients with streptococcal toxic shock-like syndrome. The strains were analysed for the occurrence of the genes of erythrogenic toxins (pyrogenic exotoxins) types A, B and C and in vitro production of these toxins. In contrast to reports indicating that 85% of the toxic shock-like syndrome-associated isolates contained the erythrogenic toxin A gene, only 58.5% of our strains harboured this gene. The erythrogenic toxin C gene was detected in 22.6% of the isolates. Erythrogenic toxin A and erythrogenic toxin B were produced by 68.7% and 58.3% of the strains containing either gene. For all group A streptococci, irrespective of clinical association, the erythrogenic toxin B gene was detected in all the isolates tested. Thus, it is difficult to define a specific role for erythrogenic toxin B in toxic shock-like syndrome as there was no clear correlation between this disease and the presence of toxin genes. Our results suggest the existence of other pathogenic factor(s) produced by group A streptococci which may stimulate human peripheral T lymphocytes in a manner similar to that of erythrogenic toxins, thus explaining different observations in previous epidemiological genetic studies.