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Possible dual function of M protein: resistance to bacteriophage A25 and resistance to phagocytosis by human
Abstract:
Spontaneous phage A25-resistant (A25(R)) mutants of group A streptococci, strain K56, were isolated. The mutant cultures were unable to adsorb phage particles and hyperproduced M protein. Trypsin-digested A25(R) cells regained the ability to adsorb phage particles, but failed to become infectious centers. This failure indicated that the mutation created a double barrier to phage growth: (i) receptors were masked by M protein; (ii) irreversibly adsorbed phage were unable to multiply. Spontaneous variants of one A25(R) mutant, shown to be M negative (M(-)) by electron microscopy, serological tests, and sensitivity to phagocytosis, rapidly adsorbed phage and were able to become infectious centers. Therefore, it was concluded that the mutant phenotype, A25(R), arose by a single mutation and genes coding for this trait and M protein synthesis were either genetically linked, controlled by a common gene or were biochemically interdependent. The A25(R) phenotype was unstable and, as expected for plasmid-coded properties, acridine orange induced segregation of this phenotype. The parental M(+), A25-sensitive (A25(S)) cultures proved to be a mixed population. Infection at various multiplicities indicated that this culture was composed of phage A25(S) cells and cells more resistant to infection. Morphological comparison of thin sections of A25(R) and A25(S) cells by electron microscopy demonstrated striking differences. The A25(R) culture was composed entirely of cells uniformly covered with M protein, whereas the A25(S)M(+) wild-type culture was a mixed population, the majority of cells devoid of M protein. Phagocytosis by human blood enriched the culture for the latter cell type, suggesting that differences in phage sensitivity in the wild-type culture were also determined by the presence or absence of M protein. Thus M protein can serve a dual function for the streptococcal cell by allowing it to avoid infection by bacteriophage and ingestion by human leukocytes.
Insights
Group A streptococci mutants resistant to phage A25 hyperproduced M protein, blocking phage adsorption. Loss of M protein restored phage sensitivity, indicating M protein
Area of Science:
- Microbiology
- Bacteriology
- Virology
Background:
- Group A streptococci (GAS) are significant human pathogens.
- Bacteriophages are viruses that infect bacteria and can be used to study bacterial genetics and physiology.
- M protein is a major virulence factor of GAS, contributing to host immune evasion.
Purpose of the Study:
- To investigate the genetic and biochemical basis of phage A25 resistance in group A streptococci.
- To elucidate the role of M protein in phage resistance and bacterial virulence.
- To characterize the phenotype of spontaneous phage-resistant mutants.
Main Methods:
- Isolation and characterization of spontaneous phage A25-resistant (A25(R)) mutants of GAS strain K56.
- Assessment of phage adsorption, infectivity, and M protein production in wild-type and mutant strains.
- Electron microscopy to visualize M protein expression and cell morphology.
- Phagocytosis assays using human blood to assess the role of M protein in host immune evasion.
Main Results:
- A25(R) mutants exhibited hyperproduction of M protein, which masked receptors and prevented phage adsorption.
- Treatment with trypsin restored phage adsorption in A25(R) mutants but did not restore infectivity, indicating a second block to phage growth.
- Spontaneous M-negative (M(-)) variants of A25(R) mutants regained phage sensitivity and became infectious centers.
- The A25(R) phenotype was unstable and segregatable, suggesting plasmid-mediated inheritance.
- Wild-type cultures were a mixed population with varying M protein expression, influencing phage sensitivity and phagocytosis.
Conclusions:
- The A25(R) phenotype arises from a single mutation affecting M protein synthesis or regulation.
- M protein plays a dual role in GAS, mediating resistance to bacteriophage infection and evasion of phagocytosis.
- Genetic linkage or biochemical interdependence exists between phage resistance and M protein synthesis.
- M protein expression levels significantly impact GAS-phage interactions and host immune evasion strategies.