J. T. Poolman

Lipo-oligosaccharide immunotyping of Neisseria meningitidis by a whole-cell ELISA with monoclonal antibodies

To assess the applicability of a whole-cell ELISA (WCE) with monoclonal antibodies (MAbs) for lipo-oligosaccharide (LOS) immunotyping of Neisseria meningitidis, 675 meningococcal isolates obtained in 1989 and 1990 in the Netherlands and 57 isolates collected in 1974, of which the immunotype had been determined previously by microprecipitation, were analysed. Despite the lack of specific MAbs for L2 and L4, an algorithm was developed for the assignment of immunotypes on the basis of the reaction patterns of the reference strains and these isolates to a combination of 14 MAbs. The immunotypes found by WCE were in accordance with those obtained by microprecipitation and the results from WCE were reproducible. The distribution of immunotypes among isolates of the various serogroups in the Netherlands in 1989-1990 is presented. Based on the reaction patterns of the isolates, two main categories of related immunotypes could be distinguished among isolates of serogroups B and C: L2/L4 and L3/L1/L8. Some isolates of the latter category were of one immunotype, but many isolates expressed one or two additional immunotypes, either strongly or weakly, indicating that the differences in this category are quantitative rather than qualitative. The results of this study have demonstrated that the WCE method for LOS immunotyping is easily applicable and provides better definition of test strains for in-vitro bactericidal assays and research into pathogenesis.

Colony Variants of Neisseria Meningitidis Strain 2996 (B: 2b: P1.2): Influence of Class-5 Outer Membrane Proteins and Lipopolysaccharides

Different colonial morphologies were found among colonies of Neisseria meningitidis strain 2996 (B:2b:P1.2). Examination of cultures, selected on the basis of colony transparency or opacity, revealed that both lipopolysaccharides (LPS) and class-5 outer membrane proteins (OMP) are associated with differences in colonial morphology. Among 13 variants, four LPS variants and two class-5 OMP variants were recognised. All variants were non-fimbriate. The LPS variations were confirmed by immunoprecipitation. In addition to these qualitative variations of LPS, meningococci synthesise LPS of different molecular size depending upon growth phase; larger LPS molecules were found after analysis of stationary-phase cultures than with exponential-phase cultures. These changes did not cause a change in serotyping characteristics. The recognition in this study of intra-strain heterogeneity of meningococcal LPS and class-5 OMPs is important for the understanding of meningococcal pathogenicity. This heterogeneity was also detected in simultaneous isolates from different sites of a patient.

Meningococcal disease in The Netherlands, 1959–1981: The occurrence of serogroups and serotypes 2a and 2b of Neisseria meningitidis

By means of a filter radioimmunoassay and the use of monoclonal anti-2a and anti-2b antibodies, we have serotyped 3164 of 3688 strains of Neisseria meningitidis isolated from patients in The Netherlands between 1959 and 1981. Serotypes 2a and 2b were distributed differently among the major serogroups A, B, C, and W-135. Neither of the types was found among group A strains. Type 2b strains of serogroup B emerged in 1965, causing a country-wide epidemic which reached a peak incidence in March and April of 1966 and continued to predominate within group B until 1979. Type 2a strains of serogroup C were responsible for a substantial number of sporadic cases over a long period without any association with outbreaks or with a shift in the pattern of the serogroup. After the appearance of group W-135 in 1971, W-135 strains caused a small non-focal epidemic wave. The upsurge of disease due to virulent sub-populations of strains B:2b and C:2a appeared to be closely related to a basic pattern of regular cyclical waves with peak intervals which differed for serogroups A, B, and C. In recent years both serotype 2a and 2b strains within the different serogroups fell to insignificant numbers. Our results show that retrospective large-scale serotyping of collected strains provides insight into the epidemiological patterns of endemic meningococcal disease.

Recurrent localized outbreaks of group C meningococcal disease and selective vaccination programmes

Conclusions(1) The high attack rate indicates that one virulent strain was responsible for the outbreaks. (2) The first vaccination programme carried through was not efficient enough to irradicate group C meningococcal disease whereas extension of the programme to the whole high-risk group was successful.

Immunochemical characterization of meningococcal serotype antigens by immunodiffusion and SGIP

The electronic device (2), consisting of amplifiers, an analog digital (AD) converter, a timer (1) and a tube counter, steers the printing device (3). After each measurement, tube number, time of measurement and mV are printed out. The mV values are converted into absorbance expressed as Klettunits, corrected for initial mV, tube diameter and instrument factors with a Diehl Alphatronic (Diehl Datensystem Niirnberg, West-Germany) desk computer. The complete instrument fits in an incubator room of 40 • 40 • 100 cm. A linear relation between turbidity and viable counts was found between 2 x 107 and 8 x 109 c.f.u./ml (r > 0.99). The aeration resulted in a remarkable increase of bacteria (from 45 % in staphylococci up to 123 ~ in Pseudomonas aeruginosa as measured turbidimetrically. No surface growth or pellicle formation was observed with the latter strain because of the regular stirring. Mechanical damage of bacteria by the stirring rod was small. Because of its small size, the instrument can be used for anaerobic cultures by placing it in an anaerobic cabinet. Yields of 8 x 109 c.f.u./ml of Bacteroidesfragilis were obtained. In research laboratories the instrument is more versatile than the commercial types.