Neisseria meningitidis group B was subdivided into a number of different serotypes. A sensitive microbactericidal assay procedure was developed, and, with this method, over 10 different serotypes were provisionally established. The method by which distinct serotype designations were assigned is described. Almost half of the strains examined thus far contained a common serotype antigen which was distinct from the grouping antigen. Differentiation of group B meningococci into distinct types will aid in epidemiological studies and in the selection of appropriate strains for vaccines against group B meningococci. Neisseria meningitidis consists of seven major serological groups (2,13). Epidemiological study of meningococcal infection is hampered by a lack of suitable procedures for distinguishing different strains of the same serological group. Roberts (11) was one of the first to demonstrate that antigenic variation does exist between different strains of the same group. He showed that antisera against two strains of group B meningococci did not have reciprocal opsonic or bactericidal activity. Frisch (4) also found that sera from human cases of group B meningococcal infection showed marked differences in bactericidal activity when tested against various strains of group B meningococci.Gold and Wyle (5) have recently identified distinct serotypes of group C meningococci with a bactericidal assay. We have developed a microbactericidal procedure for typing group B meningococci and have used this procedure to establish over 10 distinct provisional serotypes within this group.MATERIALS AND METHODS Strains. Many of the group B meningococcal strains were kindly supplied by Neylan Vedros, Neisseria Repository (NAMRU-1), Berkeley, Calif. Several strains were obtained from Great Lakes Naval Training Station (NAMRU-4), and others were isolated locally in the Minneapolis-St. Paul area.Stock strains were maintained in the lyophilized state. Lyophilized cultures were regrown in Trypticase soy broth (TSB; BBL) and then frozen in vials in fetal calf serum at -20 C. All working cultures were taken as needed from this frozen stock.Antisera. Immune bactericidal sera were prepared against the various group B strains in young American Dutch rabbits weighing approximately 1.5 kg. The rabbits received graded multiple intravenous injections of Formalin-killed and washed group B cells according to the following immunization schedule:(i) week 1, 0.25 ml of vaccine (2 X 108 organisms/ml) given Monday, Wednesday, and Friday (MWF);(ii) week 2, 0.50 ml of vaccine, MWF; (iii) week 3, 1.0 ml of vaccine, MWF. On week 4, the animals were bled by cardiac puncture and then given a single intravenous injection of approximately 2 X 108 live organisms. They were bled again on week 5.The antisera were divided into 1-dr vials and promptly frozen and stored at -20 C until used.Complement sera. Complement required for the bactericidal assay was obtained by bleeding three or more 3-kg American Dutch rabbits via heart puncture. Sera so obtained were pooled an...

Serial observations including cultures of the upper respiratory tract and of infected skin lesions and streptococcal antibody determinations were made over a two-year period in a semi-closed population of children between 10 months and 15 years of age. There was a high prevalence of group A streptococci in nose and throat cultures and of skin lesions containing these organisms. Almost 90% of the study population developed streptococcal impetigo during the study period. A slightly higher proportion of males than females developed skin infection but there was no relationship to age. Impetigo was observed throughout the calendar year, exceeding 12% of child-visits in one winter month, but was generally more common in the summer and fall. Conversely, group A streptococci were more often isolated from nose and throat cultures in the winter months. The increase in recovery of streptococci from nose and throat cultures lagged behind the increase in streptococcal impetigo and continued into the winter months, when the prevalence of impetigo had declined. Calculation of ratios for individual streptococcal serotypes isolated from different body sites revealed a clear cut distinction between "respiratory" and "impetigo" serotypes, with respect to both prevalence and acquisition rates. Respiratory serotypes were more commonly isolated in the winter and impetigo serotypes in the summer and fall. Significant antibody responses to extracellular antigens of the streptococcus were documented for pharyngeal acquisitions of both impetigo and respiratory serotypes and for skin lesions associated with impetigo serotypes. Group A streptococcal serotypes may be divided into three categories on the basis of their human pathogenicity for body sites: some with the potential for respiratory infection, others with the potential for skin infection and a few unusual serotypes with the potential for infecting both sites.

A B S T R A C T The immune response after streptococcal infection of the skin and of the upper respiratory tract (URT) was studied prospectively in a group of normal children, ages 3-6 yr. The children were examined and cultures for group A streptococci were obtained weekly from the throat, nose, and skin lesions (when present). Paired sera were collected at the beginning and end of the study, and the changes in antibody titers were measured for three different streptococcal antigens: streptolysin 0, deoxyribonuclease B (DNAse B), and nicotinamide adenine dinucleotidase (NADase).The findings suggest that in contrast to infection of the URT antibody response to streptolysin 0 is relatively feeble after streptococcal infection which is limited to the skin. The response to NADase is also poor after cutaneous infection. Antibody responses to DNAse B are generally good regardless of the site of the infection. These and other studies indicate that anti-DNAse B is the antibody of choice in studying streptococcal infection of the skin and its complications.

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Over 10 distinct serotypes of group B Neisseria meningitidis have been found to date by using a sensitive microbactericidal assay developed by the authors. The serotype antigens have now been extracted by hot acid or saline extraction procedures. It was found that these extracted serotype antigens may be used in a simple capillary precipitin method. This method uses unadsorbed, undiluted rabbit antisera. In the capillary precipitin method a 3+ to 4+ reaction was considered significant. The microbactericidal assay and capillary precipitin methods for serotyping group B meningococci show excellent agreement. Group B meningococci were also serotyped by an agar gel double diffusion technique. The latter technique conserves reagents and has the further advantage that it does not show the minor cross-reactions observed in the capillary precipitin method. Thus two simple, reproducible methods for serological typing of group B meningococci have been developed. These methods developed for the serological typing of group B meningococci will aid in epidemiological studies of meningococcal disease. They may also be of value for selecting suitable strains for vaccine production.