No abstract

SummaryGroup A streptococcus (GAS) causes diseases ranging from benign to severe infections such as necrotizing fasciitis (NF). The reasons for the differences in severity of streptococcal infections are unexplained. We developed the polymorphic-tag-lengthstransposon-mutagenesis (PTTM) method to identify virulence genes in vivo . We applied PTTM on an emm 14 strain isolated from a patient with NF and screened for mutants of decreased virulence, using a mouse model of human soft-tissue infection. A mutant that survived in the skin but was attenuated in its ability to reach the spleen and to cause a lethal infection was identified. The transposon was inserted into a small open reading frame (ORF) in a locus termed sil , s treptococcal i nvasion l ocus. sil contains at least five genes ( sil A-E) and is highly homologous to the quorum-sensing competence regulons of Streptococcus pneumoniae . sil A and sil B encode a putative two-component system whereas sil D and sil E encode two putative ABC transporters. sil C is a small ORF of unknown function preceded by a combox promoter. Insertion and deletion mutants of sil had a diminished lethality in the animal model. Virulence of a deletion mutant of sil C was restored when injected together with the avirulent emm 14-deletion mutant, but not when these mutants were injected into opposite flanks of a mouse. DNA transfer between these mutants occurred in vivo but could not account for the complementation of virulence. DNA exchange between the emm 14-deletion mutant and mutants of sil occurred also in vitro, at a frequency of ~ 10 ----8 for a single antibiotic marker. Whereas sil C and sil D mutants exchanged markers with the emm 14 mutant, sil B mutant did not. Thus, we identified a novel locus, which controls GAS spreading into deeper tissues and could be involved in DNA transfer.

Binding of the group A streptococcus (GAS) to respiratory epithelium is mediated by the fibronectin (Fn)-binding adhesin, protein F1. Previous studies have suggested that certain GAS strains express Fn-binding proteins that are different from protein F1. In this study, we have cloned, sequenced, and characterized a gene (prtF2) from GAS strain 100076 encoding a novel Fn-binding protein, termed protein F2. Insertional inactivation of prtF2 in strain 100076 abolishes its high-affinity Fn binding. prtF2-related genes exist in most GAS strains that lack prtF1 (encoding protein F1) but bind Fn with high affinity. These observations suggest that protein F2 is a major Fn-binding protein in GAS. Protein F2 is highly homologous to Fn-binding proteins from Streptococcus dysgalactiae and Streptococcus equisimilis, particularly in its carboxy-terminal portion. Two domains are responsible for Fn binding by protein F2. One domains (FBRD) consists of three consecutive repeats, whereas the other domain (UFBD) resides on a non-repeated stretch of approximately 100 amino acids and is located 100 amino acids aminoterminal of FBRD. Each of these domains is capable of binding Fn when expressed as a separate protein. In strain 100076, protein F2 activity is regulated in response to alterations in the concentration of atmospheric oxygen.

Recurrent group G Steptococcus bacteremia, associated with lymphatic disorders and possibly emm stG 840.0, is described.

To determine whether rapid emergence of Salmonella enterica serovar Infantis in Israel resulted from an increase in different biotypes or spread of 1 clone, we characterized 87 serovar Infantis isolates on the genotypic and phenotypic levels. The emerging strain comprised 1 genetic clone with a distinct pulsed-field gel electrophoresis profile and a common antimicrobial drug resistance pattern.

Our survey emphasizes the high rate of pregnancy-related listeriosis in Israel and shows that specific clones might account for this.

JRS4(HE), a highly encapsulated, mouse-passaged variant of group A streptococcal strain JRS4, was characterized. The mucoid phenotype of JRS4(HE) was preserved after extensive passage in vitro. The level and size of csrRS transcript in JRS4(HE) was similar to that of JRS4, yet JRS4(HE) expressed high levels of has and sagA and exhibited an increased activity of streptolysin S. These findings indicate that the CsrRS repressor system was inactive in JRS4(HE). JRS4(HE) adhered to HEp-2 cells at the stationary phase but did not internalize these cells. At midlogarithmic phase, JRS4(HE) neither adhered to nor internalized cells, because of an increased amount of hyaluronic acid. Mice injected subcutaneously with JRS4(HE) developed large, deep necrotic lesions. In contrast, mice challenged with JRS4 developed small, superficial lesions. Despite the use of a high inoculum, mice challenged with JRS4(HE) did not develop a lethal bacteremic infection. It is concluded that inactivation of CsrRS in vivo is insufficient to cause a spreading necrotic disease.

We performed emm typing of M nontypeable invasive group A streptococcal (GAS) isolates collected in a prospective population-based study in Israel. One hundred twenty of 131 isolates (92%) had emm sequences compatible with GAS, consisting of 51 different emm types. Eleven isolates were found to be group G streptococcus. Of the 120 isolates, 55 (46%) belonged to 32 types for which there were no typing sera available in the Streptococcal Reference Laboratory in Israel. The other 65 (64%) isolates, consisting of 19 types, had sera available and therefore could have been serotyped. Forty-three isolates had T and emm types which were not correlated according to standard M-typing protocols and were therefore missed. The principal effect of emm typing was the addition of 32 types not previously identified in Israel and the discovery of new associations between emm and T types. emm typing did not significantly change the proportion of M types; the five most common types were 3, 28, 2, 62, and 41. Twenty different types comprised 80% of all isolates. No new emm sequences were discovered. emm typing emphasized the unusually low incidence of M1 strains causing severe disease in Israel. As serological typing of GAS becomes more problematic due to lack of sera and the appearance of new emm types, reference laboratories should replace M typing with emm sequence typing. Development of a GAS vaccine relies on the emm type distributions in different geographical locations. In our study, 7% of isolates (types 41 and 62) are not included in a 26-valent vaccine that is being studied.Group A streptococcus (GAS) causes a variety of human infections. These range from mild, self-limited diseases like pharyngitis and impetigo to severe, sometimes life-threatening illnesses such as bacteremia, necrotizing fasciitis, and toxic shock syndrome. Typing of GAS has long been the hallmark of both epidemiological studies and the understanding of diseases caused by different strains (2). M protein is a major virulence determinant of GAS that is associated with resistance to phagocytosis, adherence to cells, and virulence in a mouse model of necrotizing fasciitis (1, 24). Serological M typing was developed many years ago and was the only means for typing GAS. Initially, only 50 serotypes were described (12), but later several reference laboratories added some 30 more serotypes (12). This laborious method is becoming obsolete because it is time-consuming and expensive. Many centers have stopped producing specific antisera in rabbits. Sequence analysis of the hypervariable portion of the emm gene encoding M protein (emm typing) has simplified GAS typing and has recently expanded the number of known GAS types from ϳ80 to 124 (12).In various regions of the world, the percentage of M nontypeable strains varies from Ͼ90% (20) to Ͻ20% (15). The reasons for this variation include technical difficulties and a high prevalence of new emm types, for which serum is not available for M typing (15). In Israel, 67% of 21,517 GAS isolates (mostly from pharyngeal s...