Electrophoretic Patterns of Extracellular Deoxyribonuclease (DNase) and Their Correlation with T‐Type in Group A Streptococci

Miyakawa, Yozo; Yamada, Toshihiko; Shitara, Masatsugu; Yoshimura, Fuminobu

doi:10.1111/j.1348-0421.1985.tb00819.x

Cited by 6 publications

(13 citation statements)

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“…Hence, we designated Spy0747 as SpnA (S. pyogenes nuclease A). All S. pyogenes strains express one or more DNase genes (Wannamaker, 1958;Miyakawa et al, 1985;Ferreira et al, 1992), and recent genome studies have revealed that all S. pyogenes strains analysed contain several genes presumed to encode extracellular DNases. Extracellular streptococcal DNases have been considered as virulence factors.…”

Section: Discussionmentioning

confidence: 99%

“…These factors include M protein, streptococcal inhibitor of complement, streptococcal pyrogenic toxins, haemolysins, and several DNases (Cunningham, 2000). DNases have been extensively characterized (McCarty, 1949;Miyakawa et al, 1985;Ferreira et al, 1992). However, there was no direct evidence that DNases are virulence factors, until Sumby et al (2005a) provided such evidence.…”

Section: Introductionmentioning

confidence: 99%

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Characterization of a virulence-associated and cell-wall-located DNase of Streptococcus pyogenes

et al. 2010

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We investigated culture supernatant proteins from the M1 serotype of Streptococcus pyogenes by two-dimensional gel electrophoresis and peptide mass mapping analysis, and characterized the single protein spots. Among them, we analysed the Spy0747 protein. This protein is homologous to the SsnA protein, a cell-wall-located DNase expressed in Streptococcus suis serotype 2. We designated the Spy0747 protein as SpnA. SpnA protein was also detected in the insoluble fraction of whole-cell lysates using shotgun proteomic analysis, suggesting that SpnA is also located in the cell wall. SpnA was expressed as a glutathione S-transferase-fusion protein in Escherichia coli. We confirmed that the recombinant protein had DNase activity that was dependent on Ca 2+ and Mg 2+ , like SsnA. Blood bactericidal assays and mouse infection model experiments showed that the spnA knockout strain was less virulent than the parental strain, thus suggesting that SpnA could play an important role in virulence. Using PCR, we found that the spnA gene was present in all clinical S. pyogenes strains we examined. Our results, together with a previous report identifying Spy0747 as a surface-associated protein, suggest that SpnA is an important cell-wall-located DNase that is generally produced in S. pyogenes and is involved in virulence. INTRODUCTIONStreptococcus pyogenes is a Gram-positive bacterium infecting the upper respiratory tract, such as the tonsils and pharynx, and it also causes post-infection diseases such as rheumatic fever and glomerulonephritis. Furthermore, S. pyogenes causes many serious human diseases such as streptococcal toxic shock syndrome (STSS) (Cone et al., 1987;Cunningham, 2000). Because of the worldwide prevalence of STSS, a number of studies have been undertaken to identify relevant virulence factors (Hauser et al., 1991;Reichardt et al., 1992). These factors include M protein, streptococcal inhibitor of complement, streptococcal pyrogenic toxins, haemolysins, and several DNases (Cunningham, 2000). DNases have been extensively characterized (McCarty, 1949;Miyakawa et al., 1985;Ferreira et al., 1992). However, there was no direct evidence that DNases are virulence factors, until Sumby et al. (2005a) provided such evidence.We have investigated streptococcal exoproteins using 2-dimensional gel electrophoresis (2-DE) (Tanaka et al., 2005;Sawai et al., 2007). We have so far detected several DNases (Hasegawa et al., 2002a, b) and have analysed the influence of various culture conditions on production of these exoproteins (Nakamura et al., 2004). In these comprehensive analyses of culture supernatant proteins, we detected another putative DNase protein, Spy0747, in the culture supernatant of S. pyogenes SF370. The Spy0747 protein is homologous to the SsnA protein, which is a cell-wall-located DNase expressed in Streptococcus suis serotype 2 that requires Ca 2+ and Mg 2+ for its activity (Fontaine et al., 2004). Hence we designated the Spy0747 protein as SpnA. In the present study, we further characterized the SpnA protein an...

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Characterization of a virulence-associated and cell-wall-located DNase of Streptococcus pyogenes

et al. 2010

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“…Many indirect lines of evidence implicate DNases in GAS pathogenesis. First, all GAS strains produce at least one DNase, and most strains make several distinct enzymes (13,(15)(16)(17). Second, expression of DNase is up-regulated upon interaction of GAS with human epithelial cells, polymorphonuclear leukocytes (PMNs), and oropharynx (20,21,24,25).…”

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confidence: 99%

“…These factors include capsule, M protein, streptococcal inhibitor of complement, streptococcal pyrogenic toxin superantigens, hemolysins, and several DNases (8 -11). DNases were among the first secreted GAS proteins to be identified and extensively characterized (6,(12)(13)(14)(15)(16)(17). However, after almost 60 years of investigation, there is no direct evidence that DNases are virulence factors (18 -23).…”

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Extracellular deoxyribonuclease made by group A Streptococcus assists pathogenesis by enhancing evasion of the innate immune response

Sumby

Barbian

Gardner

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

298

279

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Many pathogenic bacteria produce extracellular DNase, but the benefit of this enzymatic activity is not understood. For example, all strains of the human bacterial pathogen group A Streptococcus (GAS) produce at least one extracellular DNase, and most strains make several distinct enzymes. Despite six decades of study, it is not known whether production of DNase by GAS enhances virulence. To test the hypothesis that extracellular DNase is required for normal progression of GAS infection, we generated seven isogenic mutant strains in which the three chromosomal-and prophage-encoded DNases made by a contemporary serotype M1 GAS strain were inactivated. Compared to the wild-type parental strain, the isogenic triple-mutant strain was significantly less virulent in two mouse models of invasive infection. The triple-mutant strain was cleared from the skin injection site significantly faster than the wild-type strain. Preferential clearance of the mutant strain was related to the differential extracellular killing of the mutant and wild-type strains, possibly through degradation of neutrophil extracellular traps, innate immune structures composed of chromatin and granule proteins. The triple-mutant strain was also significantly compromised in its ability to cause experimental pharyngeal disease in cynomolgus macaques. Comparative analysis of the seven DNase mutant strains strongly suggested that the prophage-encoded SdaD2 enzyme is the major DNase that contributes to virulence in this clone. We conclude that extracellular DNase activity made by GAS contributes to disease progression, thereby resolving a long-standing question in bacterial pathogenesis research.virulence factor ͉ Streptococcus pyogenes ͉ neutrophil extracellular traps M any pathogenic bacteria produce extracellular DNase, but the benefit of this enzymatic activity is not understood. It has been hypothesized that secretion of DNase provides a growth advantage by enlarging the pool of available nucleotides by DNA hydrolysis (1). Extracellular DNase activity also has been hypothesized to play a role in the dissemination and spread of infecting bacteria by liquifying pus (2-6). In addition, a recent study implied a role for extracellular DNases in the evasion of the innate immune response by degrading neutrophil extracellular traps (NETs). NETs trap and kill bacteria extracellularly, are composed of chromatin and granule proteins, and their structure is dissipated by DNase activity (7). The extracellular bactericidal activity of neutrophils directed against Shigella flexneri and Staphylococcus aureus was reduced greatly when incubated with exogenously added DNase (7).Group A Streptococcus (GAS) is a bacterial pathogen responsible for many serious human diseases (8, 9). The wide diversity of disease manifestations caused by GAS infection is thought to be due in part to the variable number of secreted and cell-wall anchored virulence factors produced by this pathogen. These factors include capsule, M protein, streptococcal inhibitor of complement, streptococc...

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“…S. pyogenes strains commonly express DNase B, but the other three DNases are not frequently expressed and thus DNase B has a major share of total DNase activity in most strains (5,7,8). Determination of antibody titers to DNase B has been used to confirm a clinical diagnosis of a previous S. pyogenes infection and the level of anti-DNase B antibody in invasive cases was significantly lower than those in healthy individuals (1).…”

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Close Correlation of Streptococcal DNase B (sdaB) Alleles with emm Genotypes in Streptococcus pyogenes

Matsumoto

Sakae

Hashikawa

et al. 2005

Microbiology and Immunology

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Streptococcal DNases are major exoenzymes and possible virulence factors in Streptococcus pyogenes and are classified into four types: DNase A, B, C and D. Among these DNases, DNase B, which is also designated as mitogenic factor (MF) or streptococcal pyrogenic exotoxin F (speF) (3,4), is dominant, although its role in the pathogenesis of S. pyogenes has not been well elucidated. S. pyogenes strains commonly express DNase B, but the other three DNases are not frequently expressed and thus DNase B has a major share of total DNase activity in most strains (5,7,8). Determination of antibody titers to DNase B has been used to confirm a clinical diagnosis of a previous S. pyogenes infection and the level of anti-DNase B antibody in invasive cases was significantly lower than those in healthy individuals (1).The present study was undertaken to determine Abstract: DNase B is a major nuclease and a possible virulence factor in Streptococcus pyogenes. The allelic diversity of streptococcal DNase B (sdaB) gene was investigated in 83 strains with 14 emm genotypes. Of the 15 alleles identified, 11 alleles carried only synonymous nucleotide substitutions. On the other hand, 4 alleles had a non-synonymous substitution other than synonymous substitutions, resulting in the substitution of a single amino acid. The distribution of each allele was generally emm genotype-specific. Only sdaB7 was found in both emm2 and emm4. The promoter region was highly conserved and DNase B protein was similarly expressed in all alleles.

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Electrophoretic Patterns of Extracellular Deoxyribonuclease (DNase) and Their Correlation with T‐Type in Group A Streptococci

Cited by 6 publications

References 15 publications

Characterization of a virulence-associated and cell-wall-located DNase of Streptococcus pyogenes

Characterization of a virulence-associated and cell-wall-located DNase of Streptococcus pyogenes

Extracellular deoxyribonuclease made by group A Streptococcus assists pathogenesis by enhancing evasion of the innate immune response

Close Correlation of Streptococcal DNase B (sdaB) Alleles with emm Genotypes in Streptococcus pyogenes

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