Three open reading frames (ORFs) were identified by a genome walking strategy in the genomes of serotype M49 group A streptococcal (GAS) strains CS101 and 591. These ORFs were located between themgacore regulon and the dipeptide permease operon. The deduced amino acid (aa) sequences contained signature sequences indicative of a lipoprotein (306 aa), an intracellular protein (823 aa), and a secreted peptide (66 aa), respectively. ORF1 (named Lsp for lipoprotein ofStreptococcus pyogenes) and ORF2 exhibited a high degree of homology to thelmb/ORF2 genes ofS. agalactiae(B. Spellerberg et al., Infect. Immun. 67:871-878, 1999). The three ORFs were found to be present in each of the 27 GAS serotype strains tested. Transcription analysis revealed a polycistroniclsp/ORF2 and a monocistronic ORF3 message that were detected primarily at the transition from exponential to stationary growth phase.lspand ORF2 mutants, ORF2- and ORF3-luciferase reporter fusions, and antiserum against recombinant Lsp were produced to examine the biological role of these genes. Although high Zn²⁺and Cu²⁺ion concentrations decreasedlspoperon expression, Lsp did not transport divalent cations as described for other LraI-type operons. Thelspmutant had reduced fibronectin binding. Although no direct binding of Lsp to fibronectin could be demonstrated, thelspmutant showed decreased transcription ofprtF2encoding the fibronectin-binding protein F2. Both thelspand ORF2 mutants showed decreased laminin binding. Adherence to and internalization into A549 epithelial cells of both mutants was reduced without a detectable effect on eukaryotic cell viability. The transcription of a number of virulence factors was altered in thelspmutants and ORF2 mutants. The changes in laminin binding and eukaryotic cell internalization could be explained by changes in transcription ofspeB(cysteine protease) and/or the global regulatorsmga,csrRS, andnra.

The serotype M6 group A streptococcal RofA regulator was previously shown to exert a direct positive control of protein F1 expression and, concomitantly, fibronectin binding. Using a serotype M6 rofA mutant, we demonstrate here that this regulator has a potentially indirect negative influence on the expression of the mga , emm 6, pel-sagA , and speA virulence genes. Additionally, the rofA mutant exhibited reduced eukaryotic cell internalization rates in combination with decreased host cell viability.

Background: The Streptococcus pyogenes (group A streptococci, GAS) two-component signal transduction system CovRS has been described to be important for pathogenesis of this exclusively human bacterial species. If this system acts uniquely in all serotypes is currently unclear. Presence of serotype- or strain-dependent regulatory circuits and polarity is an emerging scheme in Streptococcus pyogenes pathogenesis. Thus, the contribution of the sensor kinase (CovS) of the global regulatory two-component signal transduction system CovRS on pathogenesis of several M serotypes was investigated. Results: CovS mutation uniformly repressed capsule expression and hampered keratinocyte adherence in all tested serotypes. However, a serotype- and even strain-dependent contribution on survival in whole human blood and biofilm formation was noted, respectively. Conclusions: These data provide new information on the action of the CovS sensor kinase and revealed that its activity on capsule expression and keratinocyte adherence is uniform across serotypes, whereas the influence on biofilm formation and blood survival is serotype or even strain dependent. This adds the CovRS system to a growing list of serotype-specific acting regulatory loci in S. pyogenes.

Recently, a total of 32 carbapenem- and fluoroquinolone-resistant Acinetobacter baumannii (Ab) isolates was isolated from war-injured patients who were treated at German Bundeswehr Hospitals, and preliminarily typed by “DiversiLab” repetitive elements sequence-based (rep-) PCR. Core genome-based sequence typing was also used to provide more detailed epidemiological information. From the clusters observed by rep-PCR, selected Ab strains were subjected to Next Generation Sequencing (NGS) in order to compare them with international outbreak-associated Ab strains and to identify MLST (multi-locus sequence type) lineages, as well as to identify known resistance genes. Accordingly, NGS indicated higher diversity than rep-PCR, but also confirmed likely transmission events. The identified acquired carbapenem-resistant genes comprised bla_OXA-23, bla_OXA-72 and bla_GES-12, as well as various other intrinsic and acquired resistance-associated genetic elements. All isolates clustered with the previously identified international clonal lineages IC1, IC2, IC6 and IC7, with corresponding Pasteur sequence types ST1, ST2, ST78 and ST25, respectively. In conclusion, the assessment confirmed a broad spectrum of resistance-associated genes in Ab isolated from war-injured patients from the Eastern Ukraine, and provided the first insights into locally abundant clonal lineages.

At the Bundeswehr Hospitals of Hamburg and Westerstede, patients repatriated from subtropical war and crisis zones of Northern Africa and the Middle East were medically treated, including microbiological assessment. Within a six-year interval, 16 Acinetobacter spp. strains, including 14 Acinetobacter baumannii (Ab) isolates with resistance against carbapenems and origins in Afghanistan (n = 4), Iraq (n = 2), Libya (n = 2), and Syria (n = 8) were collected. While clonal relationships of Libyan and Syrian strains had been assessed by superficial next generation sequencing (NGS) and “DiversiLab” repetitive elements sequence-based (rep-)PCR so far, this study provides core genome-based sequence typing and thus more detailed epidemiological information. In detail, sequencing allowed a definitive species identification and comparison with international outbreak-associated Ab strains by core genome multi locus sequence typing (cgMLST) and the identification of MLST lineages, as well as the identification of known resistance genes. The sequence analysis allowed for the confirmation of outbreak-associated clonal clusters among the Syrian and Afghan Ab isolates, indicating likely transmission events. The identified acquired carbapenem resistance genes comprised bla_OXA-23, bla_OXA-58, bla_NDM-1, and bla_GES-11, next to other intrinsic and acquired, partly mobile resistance-associated genes. Eleven out of 14 Ab isolates clustered with the previously described international clonal lineages IC1 (4 Afghan strains), IC2 (6 Syrian strains), and IC7 (1 Syrian strain). Identified Pasteur sequence types of the 14 Ab strains comprised ST2 (Syrian), ST25 (Libyan), ST32 (Iraqi), ST81 (Afghan), ST85 (Libyan), and ST1112 (Syrian), respectively. In conclusion, the study revealed a broad spectrum of resistance genes in Ab isolated from war-injured patients from Northern Africa and the Middle East, thereby broadening the scarcely available data on locally abundant clonal lineages and resistance mechanisms.