Physiological and serological variation in Streptococcus mitis biovar 1 from the human oral cavity during the first year of life

Abstract

Objective: The purpose of the study was to explore the physiological and antigenic diversity of a large number of Streptococcus mitis biovar 1 isolates in order to begin to determine whether these properties contribute to species persistence.

Design: S. mitis biovar 1 was collected from four infants from birth to the first year of age. At each of eight to nine visits, 60 isolates each were obtained from the cheeks, tongue and incisors (once erupted) yielding 4440 in total. These were tested for production of neuraminidase, beta1-N-acetylglucosaminidase, beta1-N-acetylgalactosaminidase, IgA1 protease and amylase-binding. Antigenic diversity was examined by ELISA and Western immunoblotting using antisera raised against S. mitis biovar 1 NCTC 12261(T) and SK145.

Results: Three thousand three hundred and thirty (75%) of the isolates were identified as S. mitis biovar 1 and 3144 (94.4%) could be divided into four large phenotypic groups based on glycosidase production. Fifty-four percent of the isolates produced IgA1 protease, but production was disproportionate among the phenotypes. Between one-third and one-half of the strains of each phenotype bound salivary alpha-amylase. Antisera against strains NCTC 12261(T) and SK145 displayed different patterns of reactivity with randomly selected representatives of the four phenotypes.

Conclusions: S. mitis biovar 1 is physiologically and antigenically diverse, properties which could aid strains in avoiding host immunity and promote re-colonization of a habitat or transfer to a new habitat.

FIGURE 1

A sampling frame was created by assigning a unique number to each isolate and then, within each phenotypes, 12 isolates were randomly chosen using a computer generated random numbers list. Wells of microtiter plates were coated with washed whole cells of the selected isolates and an ELISA was performed as described in Materials and Methods using serial twofold dilutions (1:500 to 1:32,000) of rabbit antisera raised against S. oralis, SK100 and S. mitis biovar 1, SK145. Individual plates were used for each antiserum which always included the homologous strain. Pre-immune serum served to control for natural antibodies reactive with the antigen. For each isolate tested the sum of the absorbance value at each dilution was expressed as a percentage of the sum of the absorbance value at each dilution of the homologous strain. Filled bars represent SK145 antiserum and open bars represent SK100 antiserum. The interrupted horizontal lines are set at 25%, 50%, and 75% of the antibody binding of the homologous control. In the text isolates of the respective phenotypes are assigned the prefix A, B, C or D.

FIGURE 2

Western immunoblots were run on cell wall extracts of the same isolates tested by ELISA. The blots were developed using antisera against S. mitis biovar 1 strains NCTC 12261^T (Figure 2A) and SK 145 (Figure 2B). Antisera to S. oralis strains ATCC 35037^T and SK100 were non-reactive in Western blotting. Digital images of the blot strips were compared by cluster analysis using UPGMA. To the right of the strips is shown the identity of the isolate and its phenotype.

FIGURE 2

Western immunoblots were run on cell wall extracts of the same isolates tested by ELISA. The blots were developed using antisera against S. mitis biovar 1 strains NCTC 12261^T (Figure 2A) and SK 145 (Figure 2B). Antisera to S. oralis strains ATCC 35037^T and SK100 were non-reactive in Western blotting. Digital images of the blot strips were compared by cluster analysis using UPGMA. To the right of the strips is shown the identity of the isolate and its phenotype.