Study of susceptibility to antibiotics and molecular characterization of high virulence Staphylococcus aureus strains isolated from a rural hospital in Ethiopia.

We characterised 80 Staphylococcus aureus strains isolated from human patients with SSTIs at a rural hospital in Ethiopia. Susceptibility to antibiotic of all strains was tested. The MLST method was used to type and a phylogenetic analysis was conducted employing the sequences of 7 housekeeping genes. PCR amplification was used to investigate the presence of the following virulence genes in all strains: hla (α-haemolysin), tstH (toxic shock syndrome toxin), luk PV (Panton-Valentine leukocidin), fnbA (fibronectin binding protein A) and mecA (methicillin resistance). Most of the strains were resistant to penicillin and ampicillin, but only 3 strains were resistant to oxacillin, and 1 of them was a true MRSA. The MLST results showed a high diversity of sequence types (ST), 55% of which were new, and ST152 was the most prevalent. A phylogeny study showed that many of the new STs were phylogenetically related to other previously described STs, but bore little relationship to the only ST from Ethiopia described in the database. Virulence gene detection showed a high prevalence of strains encoding the hla, fnbA and pvl genes (98.77%, 96.3% and 72.84%, respectively), a low prevalence of the tst gene (13.58%) and a markedly low prevalence of MRSA (1.25%). S. aureus strains isolated from patients in a rural area in Ethiopia showed low levels of antibiotic resistance, except to penicillin. Moreover, this study reveals new STs in Eastern Africa that are phylogenetically related to other previously described STs, and confirm the high prevalence of the pvl gene and the low prevalence of MRSA on the continent.

Genetics and Pathogenicity Factors of Group C and G Streptococci.

Of the eight phylogenetic groups comprising the genus Streptococcus, Lancefield group C and G streptococci (GCS and GGS, resp.) occupy four of them, including the Pyogenic, Anginosus, and Mitis groups, and one Unnamed group so far. These organisms thrive as opportunistic commensals in both humans and animals but may also be associated with clinically serious infections, often resembling those due to their closest genetic relatives, the group A streptoccci (GAS). Advances in molecular genetics, taxonomic approaches and phylogenomic studies have led to the establishment of at least 12 species, several of which being subdivided into subspecies. This review summarizes these advances, citing 264 early and recent references. It focuses on the molecular structure and genetic regulation of clinically important proteins associated with the cell wall, cytoplasmic membrane and extracellular environment. The article also addresses the question of how, based on the current knowledge, basic research and translational medicine might proceed to further advance our understanding of these multifaceted organisms. Particular emphasis in this respect is placed on streptokinase as the protein determining the host specificity of infection and the Rsh-mediated stringent response with its potential for supporting bacterial survival under nutritional stress conditions.

Emergence of Streptococcus pyogenes emm102 causing toxic shock syndrome in Southern Taiwan during 2005-2012.

BACKGROUND: Streptococcal toxic shock syndrome (STSS) is an uncommon but life-threatening disease caused by Streptococcus pyogenes. METHODS: To understand the clinical and molecular characteristics of STSS, we analyzed clinical data and explored the emm types, superantigen genes, and pulsed-field gel electrophoresis of causative S. pyogenes isolates obtained between 2005 and 2012. RESULTS: In total, 53 patients with STSS were included in this study. The median age of the patients was 57 years (range: 9-83 years), and 81.1% were male. The most prevalent underlying disease was diabetes mellitus (45.3%). Skin and soft-tissue infection accounted for 86.8% of STSS. The overall mortality rate was 32.1%. Underlying diseases had no statistical impact on mortality. A total of 19 different emm types were identified. The most prevalent emm type was emm102 (18.9%), followed by emm11 (17%), emm1 (11.3%), emm87 (9.4%), and emm89 (7.5%). There was no statistically significant association between emm type and a fatal outcome. Among the superantigen genes, speB was the most frequently detected one (92.5%), followed by smeZ (90.6%), speG (81.1%), speC (39.6%), and speF (39.6%). The majority of emm102 strains were found to have speB, speC, speG, and smeZ. The presence of speG was negatively associated with a fatal outcome (P = 0.045). CONCLUSIONS: Our surveillance revealed the emergence of uncommon emm types, particularly emm102, causing STSS in southern Taiwan. Characterization of clinical, epidemiological, and molecular characteristics of STSS will improve our understanding of this life-threatening disease.

Panton-Valentine leukocidin is not the primary determinant of outcome for Staphylococcus aureus skin infections: evaluation from the CANVAS studies.

The impact of Panton-Valentine leukocidin (PVL) on the severity of complicated skin and skin structure infections (cSSSI) caused by Staphylococcus aureus is controversial. We evaluated potential associations between clinical outcome and PVL presence in both methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) isolates from patients enrolled in two large, multinational phase three clinical trials assessing ceftaroline fosamil for the treatment of cSSSI (the CANVAS 1 and 2 programs). Isolates from all microbiologically evaluable patients with monomicrobial MRSA or MSSA infections (n = 473) were genotyped by PCR for pvl and underwent pulsed-field gel electrophoresis (PFGE). Genes encoding pvl were present in 266/473 (56.2%) isolates. Infections caused by pvl-positive S. aureus were associated with younger patient age, North American acquisition, and presence of major abscesses (P<0.001 for each). Cure rates of patients infected with pvl-positive and pvl-negative S. aureus were similar overall (93.6% versus 92.8%; P = 0.72), and within MRSA-infected (94.5% vs. 93.1%; P = 0.67) and MSSA-infected patients (92.2% vs. 92.7%; P = 1.00). This finding persisted after adjustment for multiple patient characteristics. Outcomes were also similar when USA300 PVL+ and non-USA300 PVL+ infections were compared. The results of this contemporary, international study suggest that pvl presence was not the primary determinant of outcome in patients with cSSSI due to either MRSA or MSSA.

Increasing resistance in multiresistant methicillin-resistant Staphylococcus aureus clones isolated from a Chinese hospital over a 5-year period.

BACKGROUND: The aim was to study the changes in the antimicrobial resistance of methicillin-resistant Staphylococcus aureus (MRSA) clones over a 5-year period (from 2000 to 2005) at a representative hospital in Beijing, China. METHODS: A total of 100 randomly selected MRSA strains were analyzed using antimicrobial susceptibility testing, pulsed-field gel electrophoresis, spa typing, multilocus sequence typing, SCCmec typing, and PCR for the Panton-Valentine leukocidin virulence factor. RESULTS: Resistance to rifampin greatly increased from 32% (16/50) to 68% (34/50). High-level mupirocin-resistant isolates were found only in 2005, when four were identified. Intermediate susceptibly to quinupristin-dalfopristin increased from 22% (11/50) to 52% (26/50) between 2000 and 2005. The main antimicrobial resistance profiles changed from TC-GM-CI-EM-CM in 2000 to TC-GM-CI-EM-CM-RI in 2005. The main pulsed-field gel electrophoresis type changed from types C, L, and E in 2000 to types J, F, and N, respectively, in 2005. ST239-MRSA-III was the most predominant clone in 2000 and 2005, whereas ST5-MRSA-II was found only in 2005. CONCLUSIONS: There were increasing levels of antimicrobial resistance and epidemiological changes in the hospital-associated MRSA strains isolated in this facility between 2000 and 2005.

Distribution of methicillin-resistant Staphylococcus aureus in a low-prevalence area.

Investigating circulating methicillin-resistant Staphylococcus aureus (MRSA) strains and identifying their accumulations in society are important in the search for strategies for eradicating the pathogen. The aim of this study was to describe the distribution of MRSA in a low-prevalence area where MRSA could be establishing endemicity. MRSA isolates from 802 patients (803 isolates) were included and placed into a timeline (1991-2006) under different categories: hospital (n=270), long-term care facility (LTCF) (n=175) and general practitioner (GP) (n=358). MRSA isolates had been characterized using multilocus sequence-typing, staphylococcal cassette chromosome mec-typing and detection of Panton-Valentine leukocidin-encoding genes (lukS/F-PVL), and were placed in exotoxin-encoding gene clusters. The GP category increased mainly in a cluster with few exotoxin-encoding genes (r=0.760), the LTCF (r=0.804) and the hospital category (r=0.876) mainly in clusters with more exotoxin-encoding genes. ST8-IV, lukS/F-PVL present, increased in the community (1-41 isolates) in the time period 2002-2006, later in the hospital (1-8 isolates, 2004-2006), and finally reached the LTCF (1 isolate, 2006). ST8-IV, lukS/F-PVL absent, could have attained endemicity in LTCFs, where 51 isolates were isolated in 2006. ST125-IV, lukS/F-PVL absent, showing epidemic qualities abroad, caused outbreaks at five LTCFs.

Presence of streptococcal pyrogenic exotoxin A and C genes in human isolates of group G streptococci.

The bacteriophage-associated genes speA and speC encode streptococcal pyrogenic exotoxins of group A streptococci (GAS). Human isolates of group C and G streptococci (GCS and GGS) are commensals and the closest known genetic relatives of GAS; on occasion, GCS-GGS can cause infection that is clinically similar to GAS disease. Thirty-four human isolates of GCS-GGS were tested for speA and speC. Two GGS isolates harbored speA only, whereas a third GGS had both genes. All spe alleles found in GGS were identical to known spe alleles of GAS, except for one speA allele, which was unique. The presence of shared speA and speC alleles in GAS and GGS is highly suggestive of recent interspecies transfer. Acquisition of GAS-like virulence genes by GGS may lead to enhanced pathogenicity in this usually commensal-like organism.

Rapid isolation and identification of staphylococcal exoproteins by reverse phase capillary high performance liquid chromatography-electrospray ionization mass spectrometry.

The isolation of staphylococcal extracellular toxins and enzymes (exoproteins) usually requires time-consuming purification steps such as repeated chromatographic separations and isoelectric focusing. We performed rapid isolation, quantification and identification of staphylococcal exoproteins by reverse phase capillary high performance liquid chromatography-electrospray ionization mass spectrometry (LC-ESI/MS) followed by the determination of N-terminal amino acid sequences of separated peaks. We identified two novel exoproteins as well as previously reported antigens ORF-1 and ORF-2, glutamyl endopeptidase in Staphylococcus aureus NCTC8325 and protein A, staphylococcal enterotoxin C3 (SEC3), toxic shock syndrome toxin-1 (TSST-1) and alpha-toxin in a clinical isolate methicillin-resistant S. aureus (MRSA) 3543. MRSA3543 secreted 5.33 and 1.45 microg of SEC3 and TSST-1 per 20 microg total exoproteins ml(-1), respectively. The capillary LC treatment of the exoprotein fraction separated at least 12 peaks, indicating its high-resolution power. We found that when a protein was once determined by its N-terminal sequence, its mass spectrum and the obtained molecular mass was applicable for the assignment of the protein.

Fatal group a streptococcal toxic shock-like syndrome in a child with varicella: report of the first well documented case with detection of the genetic sequences that code for exotoxins SPE A and B, in Sao Paulo, Brazil

Crianca de sete anos, previamente higida, foi admitida na unidade de terapia intensiva por quadro de toxemia associado a varicela. Evoluiu rapidamente para choque e insuficiencia de multiplos orgaos e sistemas e, apesar do tratamento intensivo, morreu no quarto dia apos a admissao. A cultura de secrecao colhida por puncao profunda de partes moles em regiao toracica foi positiva para Streptococcus pyogenes, proteina-M nao tipavel e carreador dos genes codificadores da producao de exotoxinas pirogenicas estreptococicas A e B, preenchendo os criterios para definicao de Sindrome do choque toxico estreptococico. Os autores discutem aspectos clinicos e fisiopatologicos desta sindrome, bem como alguns aspectos incomuns relacionados a este caso

Are bacterial exotoxins cytokine network regulators?

Bacterial exotoxins are generally thought to act by damaging cells or altering cell metabolism. However, recent work has established that many exotoxins modulate eukaryotic cell cytokine synthesis. Cytokine induction may play a significant role in exotoxin action, and therapeutic targeting of cytokines could be beneficial in infectious diseases involving bacterial exotoxins.

Channel-forming leucotoxins from Staphylococcus aureus cause severe inflammatory reactions in a rabbit eye model.

Panton-Valentine leucocidin arises from the combination of one S component (LukS-PV) with one F component (LukF-PV), whereas gamma-haemolysin comprises two S components (HlgA and HlgC) with one F component HlgB. The intravitreal injection of rabbit eye with the six combinations (S + F) of channel-forming leucotoxins produced by Staphylococcus aureus ATCC 49775 induced acute inflammatory reactions depending on time and doses of toxins. These reactions involved posterior chamber as well as anterior chamber and conjunctiva, eyelids and annexes. Histological examination confirmed the involvement of eye tissues and the disruption of the retinal barrier. The lesions began only 4 h after injections and persisted for at least 5 days. Clinical and biological effects of each leucotoxin were modulated by the speed of onset and intensity of inflammation and necrosis, leading to a functional classification according to the severity of the lesions (HlgA + LukF-PV > HlgA + HlgB > or = LukS-PV + HlgB > or = LukS-PV + LukF-PV > HlgC + HlgB > or = HlgC + LukF-PV). Moreover, N-acetyl beta-D glucosaminidase assays on crude extracts of vitreous revealed granules and granule secretions from polymorphonuclear cells with levels according the above classification. These results show that channel-forming leucotoxins have a very significant inflammatory activity. As most S. aureus strains produce two or even six leucotoxins depending on the production of Panton-Valentine leucocidin, these compounds could be considered to be virulence factors.

Prevalence of the speA2 and speA3 alleles in Streptococcus pyogenes isolated from TSLS patients in Japan.

More than 100 cases have been identified as streptococcal toxic shock-like syndrome (TSLS) in Japan, since the first case was reported in 1993. Of 26 S. pyogenes isolates associated with TSLS, 69% were M-types 1 and 3. This study focused on speA alleles which were carried by all M-type 1 and 3 isolates and isolates of some other M-types, irrespective of sources. We sequenced the nucleotides of the relevant region of the speA allele in all of these isolates. Consistent with other reports, these M-type 1 and 3 isolates carried the speA2 and speA3 alleles, respectively. Our results indicate that these two speA alleles, which were prevalent world-wide have been disseminating in Japan.

Streptococcal toxic shock syndrome from a puncture wound to the foot.

Puncture wounds to the foot are a common occurrence. If treated properly, the majority will be resolved without major complications. Toxic shock syndrome and streptococcal toxic shock-like syndrome are devastating complications of some staphylococcal and streptococcal infections. This paper discusses the similarities and differences between the two toxic states, reviews the pathophysiology, and presents a case report of near-fatal streptococcal toxic shock-like syndrome secondary to a puncture wound of the foot.

Exotoxins of Aeromonas hydrophila.

Eighty of 103 strains of Aeromonas hydrophila cultured at 100 rev./min produced heat-labile enterotoxins detected using the suckling mouse assay. Results in intestinal perfusion agreed with the suckling mouse test in all strains tested by both methods. Enterotoxic activity correlated with haemolysin and cytotoxin production, but 4% of strains would have been wrongly classified using haemolysin assay in place of the suckling mouse test and 11% misclassified on the basis of cytotoxin assay. There was a significant association between haemolytic and cytotoxic activity, but 15% of strains produced only one of these toxins. Haemolysin, cytotoxin and enterotoxin were not always associated in a given isolate. The time of appearance of exotoxins during bacterial growth and the effects of dialysis, heating and proteolytic enzymes also suggest that haemolysins, cytotoxins and enterotoxins of Aeromonas hydrophila are separate toxins and not different manifestations of the same toxin.