Po Xing Zheng

Complete Genome Sequence of emm1 Streptococcus pyogenes A20, a Strain with an Intact Two-Component System, CovRS, Isolated from a Patient with Necrotizing Fasciitis

ABSTRACT Here, we announce the complete sequence of Streptococcus pyogenes A20. This strain was isolated from a patient with necrotizing fasciitis. Given that A20 harbors an intact two-component system, CovRS, the discovery of its genome sequence provides more insight into the pathogenesis of a pandemic emm1 strain.

emm1/Sequence Type 28 Strains of Group A Streptococci That Express covR at Early Stationary Phase Are Associated with Increased Growth and Earlier SpeB Secretion

ABSTRACT Streptococcus pyogenes (group A streptococcus [GAS]) is a versatile human pathogen, and emm1/sequence type 28 (ST28) is the most frequently isolated type from GAS infections. The emm1/ST28 strain is associated with necrotizing fasciitis and streptococcal toxic shock syndrome. Growth-phase regulation is one of the important regulatory mechanisms in GAS, which controls gene expression at restricted phases of growth. CovRS, a two-component regulatory system, is considered the regulator of streptococcal pyrogenic exotoxin B (SpeB) and is thought to be activated in the exponential phase of growth. In the present study, Northern hybridization analysis showed that 52% of the analyzed GAS strains expressed covR at the exponential phase, but 48% of the strains expressed covR at the early stationary phase of growth. Strains transcribing covR at the early stationary phase showed better growth and earlier SpeB expression than the other group of strains. Multilocus sequence typing and pulsed-field gel electrophoresis analysis showed only emm1/ST28 strains (which comprise a clonal cluster) were expressing covR at the early stationary phase of growth, indicating that emm1/ST28 strains have special characteristics which may be related to their worldwide distribution.

Associations of the major international high-risk resistant clones and virulent clones with specific ompK36 allele groups in Klebsiella pneumoniae in Taiwan

This study was conducted to investigate the association between ompK36 variants and international high-risk clones in Klebsiella pneumoniae. Fifty-nine sequence types (STs) divided into four ompK36 allele groups (groups A to D) were identified among 185 K. pneumoniae isolates. The major high-risk clones (29 ST11, 13 ST15, 7 ST37 and 1 ST147 isolates) were assigned to group A, while 6 STs (15 ST23, 2 ST65, 3 ST86, 1 ST163, 1 ST373 and 2 ST375 isolates) associated with pyogenic liver abscess were assigned to group C. The genotyping assay developed in this study may be useful for screening of epidemic STs.

Environmental pH changes, but not the LuxS signalling pathway, regulate SpeB expression in M1 group A streptococci.

The autoinducer-2/LuxS signalling pathway participates in quorum sensing in diverse bacterial species. In group A streptococci (GAS), LuxS has been shown to be involved in regulating the expression of several important virulence factors. Streptococcal pyrogenic exotoxin B (SpeB), a cysteine protease that has important roles in GAS pathogenesis, is positively regulated by LuxS in M3 and M5 strains. In the present study, it was found that the supernatant harvested from an overnight culture stimulated M1 strains to express speB. However, mutation of the luxS gene in M1 strains or treating M1 strains with luxS mutant culture supernatant did not affect speB expression, indicating that the LuxS pathway is not involved in regulation of speB expression in M1 strains. In addition, the acid property of culture broth was found to be able to stimulate M1 strains to express speB in the same LuxS-independent manner. These results indicate that speB expression in M1 strains is induced by environmental pH changes but is not regulated by the LuxS signalling pathway.

Allocation of Klebsiella pneumoniae Bloodstream Isolates into Four Distinct Groups by ompK36 Typing in a Taiwanese University Hospital

ABSTRACT The OmpK36 porin plays a role in carbapenem resistance and may contribute to bacterial virulence in Klebsiella pneumoniae. This study aimed to investigate the characteristics of different groups of K. pneumoniae separated by ompK36 typing. Among 226 nonduplicate K. pneumoniae bloodstream isolates collected at a Taiwanese hospital in 2011, four ompK36 types, designated types A, B, C, and D, were identified by PCR in 61, 28, 100, and 36 isolates, respectively; 1 isolate was untypeable. Statistical analysis showed significantly higher rates of antimicrobial resistance (all tested antibiotics except meropenem), extended-spectrum β-lactamases or DHA-1 (47.5% together), Qnr-type quinolone resistance determinants (50.8%), and IncFIIA-type plasmids (49.2%) in group A than in others. Seventeen isolates were identified as belonging to 3 international high-risk clones (4 sequence type 11 [ST11], 10 ST15, and 3 ST147 isolates); all isolates but 1 ST15 isolate were classified in group A. The significant characteristics of group C were hypermucoviscosity (62.0%) and a higher virulence gene content. This group included all serotype K1 (n = 30), K2 (n = 25), and K5 (n = 3) isolates, 6 of 7 K57 isolates, all isolates of major clones associated with pyogenic liver abscesses (29 ST23, 11 ST65, 5 ST86, 7 ST373, and 1 ST375 isolates), and 16 (94.1%) of 17 isolates causing bacteremic liver abscesses. Twelve (42.9%) of the group B isolates were responsible for bacteremic biliary tract infections. Group D was predominant (83.3%) among 12 K20 isolates. This study suggests that most clinical K. pneumoniae isolates can be allocated into four groups with distinct characteristics based on ompK36 types.

Peroxide Responsive Regulator PerR of group A Streptococcus Is Required for the Expression of Phage-Associated DNase Sda1 under Oxidative Stress

The peroxide regulator (PerR) is a ferric uptake repressor-like protein, which is involved in adaptation to oxidative stress and iron homeostasis in group A streptococcus. A perR mutant is attenuated in surviving in human blood, colonization of the pharynx, and resistance to phagocytic clearance, indicating that the PerR regulon affects both host environment adaptation and immune escape. Sda1 is a phage-associated DNase which promotes M1T1 group A streptococcus escaping from phagocytic cells by degrading DNA-based neutrophil extracellular traps. In the present study, we found that the expression of sda1 is up-regulated under oxidative conditions in the wild-type strain but not in the perR mutant. A gel mobility shift assay showed that the recombinant PerR protein binds the sda1 promoter. In addition, mutation of the conserved histidine residue in the metal binding site of PerR abolished sda1 expression under hydrogen peroxide treatment conditions, suggesting that PerR is directly responsible for the sda1 expression under oxidative stress. Our results reveal PerR-dependent sda1 expression under oxidative stress, which may aid innate immune escape of group A streptococcus.

Lactobacillus pentosus strain LPS16 produces lactic acid, inhibiting multidrug-resistant Helicobacter pylori

BACKGROUND/PURPOSE Helicobacter pylori is a human gastric pathogen. Antibiotic resistance of H. pylori has become a problem increasing the failure of H. pylori eradication. Therefore alternative approaches are required. The aim of this study was to evaluate the anti-H. pylori activity of Lactobacillus pentosus strain LPS16 and the mechanism of its killing effect. METHODS The anti-H. pylori activity of LPS16 was determined by the disc diffusion test and time killing assay. High-performance liquid chromatography analysis was used to analyze the secreted compounds of LPS16. Sixty H. pylori strains isolated from different gastric diseases, having different antibiotic susceptibility were collected to analyze the spectrum of anti-H. pylori activity of LPS16. Adhesion ability of LPS16 to gastric epithelial cell lines was assayed by flow cytometry. RESULTS The anti-H. pylori activity of LPS16 depended on the secreted component, and lactic acid mediated bactericidal activity against H. pylori. The bactericidal activity did not vary significantly among the strains isolated from different diseases having different antibiotic susceptibility. Moreover, LPS16 can adhere on gastric epithelial cell lines AKG and MKN45. CONCLUSION L. pentosus strain LPS16 had the broad-spectrum anti-H. pylori activity, suggesting that it can be used to prevent H. pylori infection.

Invasive hypermucoid variant of group A Streptococcus is defective in growth and susceptible to DNA-damaging treatments

Hyaluronic acid capsule is one of the most important virulence factors of group A Streptococcus (GAS). Over-production of capsule has been thought to enhance GAS virulence during infections. However, although the increased of capsule expression associates with increased bacterial virulence and invasive ability, over-production of capsule has not often been observed among clinical isolates. In the present study, we identified two mucoid emm12 type isolates that can convert to the hypermucoid morphology under both in vitro and in vivo conditions. Consistent with previous studies, hypermucoid variants are more invasive in the mouse air-pouch infection model. However, one of the hypermucoid variants showed a growth-defective phenotype in regular broth culture conditions and is significantly more susceptible to various DNA-damaging treatments when compared with the mucoid variant. These properties of the hypermucoid variant may be adverse factors inhibiting its adaptation to the host environment during infections.

Galectin-3 inhibits galectin-8/parkin- mediated ubiquitination of group A streptococcus

ABSTRACT Group A streptococcus (GAS) is an important human pathogen that causes a wide variety of cutaneous and systemic infections. Although originally thought to be an extracellular bacterium, numerous studies have demonstrated that GAS can trigger internalization into nonimmune cells to escape from immune surveillance or antibiotic-mediated killing. Epithelial cells possess a defense mechanism involving autophagy-mediated targeting and killing of GAS within lysosome-fused autophagosomes. In endothelial cells, in contrast, we previously showed that autophagy is not sufficient for GAS killing. In the present study, we showed higher galectin-3 (Gal-3) expression and lower Gal-8 expression in endothelial cells than in epithelial cells. The recruitment of Gal-3 to GAS is higher and the recruitment of Gal-8 to GAS is lower in endothelial cells than in epithelial cells. We further showed that Gal-3 promotes GAS replication and diminishes the recruitment of Gal-8 and ubiquitin, the latter of which is a critical protein for autophagy sequestration. After knockdown of Gal-3 in endothelial cells, the colocalization of Gal-8, parkin, and ubiquitin-decorated GAS is significantly increased, as is the interaction of Gal-8 and parkin, an E3 ligase. Furthermore, inhibition of Gal-8 in epithelial cells attenuates recruitment of parkin; both Gal-8 and parkin contribute to ubiquitin recruitment and GAS elimination. Animal studies confirmed that Gal-3-knockout mice develop less-severe skin damage and that GAS replication can be detected only in the air pouch and not in organs and endothelial cells. These results demonstrate that Gal-3 inhibits ubiquitin recruitment by blocking Gal-8 and parkin recruitment, resulting in GAS replication in endothelial cells. IMPORTANCE In epithelial cells, GAS can be efficiently killed within the lysosome-fused autophaosome compartment. However, we previously showed that, in spite of LC-3 recruitment, the autophagic machinery is not sufficient for GAS killing in endothelial cells. In this report, we provide the first evidence that Gal-3, highly expressed in endothelial cells, blocks the tagging of ubiquitin to GAS by inhibiting recruitment of Gal-8 and parkin, leading to an enhancement of GAS replication. We also provide the first demonstration that Gal-8 can interact with parkin, the critical E3 ligase, for resistance to intracellular bacteria by facilitating the decoration of bacteria with ubiquitin chains. Our findings reveal that differential levels of Gal-3 and Gal-8 expression and recruitment to GAS between epithelial cells and endothelial cells may contribute to the different outcomes of GAS elimination or survival and growth of GAS in these two types of cells. In epithelial cells, GAS can be efficiently killed within the lysosome-fused autophaosome compartment. However, we previously showed that, in spite of LC-3 recruitment, the autophagic machinery is not sufficient for GAS killing in endothelial cells. In this report, we provide the first evidence that Gal-3, highly expressed in endothelial cells, blocks the tagging of ubiquitin to GAS by inhibiting recruitment of Gal-8 and parkin, leading to an enhancement of GAS replication. We also provide the first demonstration that Gal-8 can interact with parkin, the critical E3 ligase, for resistance to intracellular bacteria by facilitating the decoration of bacteria with ubiquitin chains. Our findings reveal that differential levels of Gal-3 and Gal-8 expression and recruitment to GAS between epithelial cells and endothelial cells may contribute to the different outcomes of GAS elimination or survival and growth of GAS in these two types of cells.

Complete Genome Sequence of Community-Acquired Klebsiella pneumoniae KP36, a Strain Isolated from a Patient with an Upper Urinary Tract Infection

ABSTRACT Here, we announce the complete genome sequence of Klebsiella pneumoniae KP36, a strain isolated from a patient with a severe community-acquired urinary tract infection. This genome provides insights into the pathogenesis of a pandemic K. pneumoniae strain from a community-acquired urinary tract infection.