Escherichia coli ST131 clones harbouring AggR and AAF/V fimbriae causing bacteremia in Mozambican children: Emergence of new variant of fimH27subclone

Multidrug-resistant Escherichia coli ST131 fimH30 responsible for extra-intestinal pathogenic (ExPEC) infections is globally distributed. However, the occurrence of a subclone fimH27 of ST131 harboring both ExPEC and enteroaggregative E. coli (EAEC) related genes and belonging to commonly reported O25:H4 and other serotypes causing bacteremia in African children remain unknown. We characterized 325 E. coli isolates causing bacteremia in Mozambican children between 2001 and 2014 by conventional multiplex polymerase chain reaction and whole genome sequencing. Incidence rate of EAEC bacteremia was calculated among cases from the demographic surveillance study area. Approximately 17.5% (57/325) of isolates were EAEC, yielding an incidence rate of 45.3 episodes/105 children-years-at-risk among infants; and 44 of isolates were sequenced. 72.7% (32/44) of sequenced strains contained simultaneously genes associated with ExPEC (iutA, fyuA and traT); 88.6% (39/44) harbored the aggregative adherence fimbriae type V variant (AAF/V). Sequence type ST-131 accounted for 84.1% (37/44), predominantly belonging to serotype O25:H4 (59% of the 37); 95.6% (35/44) harbored fimH27. Approximately 15% (6/41) of the children died, and five of the six yielded ST131 strains (83.3%) mostly (60%; 3/5) due to serotypes other than O25:H4. We report the emergence of a new subclone of ST-131 E. coli strains belonging to O25:H4 and other serotypes harboring both ExPEC and EAEC virulence genes, including agg5A, associated with poor outcome in bacteremic Mozambican children, suggesting the need for prompt recognition for appropriate management

Community prevalence of carbapenemase-producing Gram-negative bacteria

Purpose: To raise awareness of carbapenemase-producing organisms, identify “at-risk” patients when admitted in a medical healthcare facility, and to outline effective infection prevention and control measures in order to halt the entry and spread of these organisms. Methods: A total of 1043 un-duplicated urine specimens of healthy volunteers who had no travel history or history of hospitalization were screened. The carbapenemase genotype of each imipenem-resistant strain was determined. Molecular typing and homology analysis of the main carbapenemase-producing strains were used to reveal the mode of transmission of resistance genes. Through transfer joint experiments, the potential risk of spread of carbapenemase genes was assessed. Results: A total of 19 carbapenemase-producing strains from 1,043 non-duplicated healthy volunteers (1.82 %) were identified. The main carbapenemase-producing organism was E. coli (42.1 %, 8/19). The main carbapenemase genotype of E. coli was blaKPC-2 (7 strains). Results from multi-locus sequence typing (MLST) indicated that 7 E. coli isolates belonged to ST-10, ST-101, ST-131, ST-405, ST-410 and ST-1193 and ST-2562. Homologous cluster analysis revealed that the sequence types among the 7 E. coli were high in diversity. The blaKPC-2 gene was successfully transferred from these isolates to 10.22-14 via conjugation. All recipient cells showed marked decreases in carbapenem sensitivity to imipenem (p < 0.05)). The degrees of conjugation were 2.10±0.12 ×10-4, 1.96±0.14×10-4, 2.72±0.18 ×10-4, 3.15±0.20 × 10-4, 2.92±0.23 ×10-4, 3.50±0.20 ×10-4 and 4.12±0.24 ×10-4 in recipient cells of TC7.23-51, TC8.9-42, TC8.15-11, TC8.23-59-3, TC8.23-83, TC9.08-47 and TC10.13-15, respectively. Conclusion: The findings demonstrate the pattern and features of carbapenemase-insensitive E. coli. The blaKPC-2 was the main community-prevalent gene of carbapenem-resistant E. coli. In view of increasing incidence of resistance to multi-drug therapy, surveillance of insensitivity to antibiotics is vital, especially urinary system infection due to carbapenem-insensitive E. coli

Detection of sequence type 131 in multi-drug resistant uropathogenic Escherichia coli isolates from two hospitals of Sabah

Background: Escherichia coli sequence type 131 (ST131) has emerged among bacteria causing urinary tract infection (UTI) in the previous decade. This ST contains multiple drug resistant (MDR) genes together with genes encoding many virulence factors. As a result, this strain of uropathogenic E. coli (UPEC) gives rise to treatment failure with consequent prolonged stay in a hospital. Therefore, earlier identification of this strain in the hospital has advantage in combating severe type of UTI. Objective: To detect ST 131 strains in MDR UPEC isolates from two hospitals of Sabah. Materials and Methods: Antibiotic susceptibility tests were performed to detect MDR isolates. Two polymerase chain reactions (PCRs) including mdh and gyrB allelic-specific PCR were performed on these MDR to detect ST131 strains. Results: The results showed four isolates were resistant to TMP-SMX, gentamycin, ciprofloxacin, and cefotaxime, and three isolates of these were investigated to be ST131 clones by two PCR reactions. Conclusion: There is the presence of ST131 strains in hospitals of Sabah. This information will be a guideline for the clinician in the management of UTI in the clinical settings

Extended-spectrum β-lactamases, transferable quinolone resistance, and virulotyping in extra-intestinal E. coli in Uruguay

Introduction: To characterize extended-spectrum β-lactamases (ESBLs) and plasmid-mediated quinolone resistance (PMQR) genes in Escherichia coli isolates obtained from extra-intestinal samples in three Uruguayan hospitals. Methodology: Fifty-five ESBL-producing E. coli isolates were studied. Virulence genes, ESBLs, and PMQR genes were detected by polymerase chain reaction. ESBL-producing isolates were compared by pulsed-field gel electrophoresis. Multi-locus sequence typing was also performed on 13 selected isolates. Results: Thirty-seven isolates harbored blaCTX-M-15 (67.3%), eight blaCTX-M-2 (14.6%), five blaCTX-M-14 (9.1%), three carried both blaCTX-M-2 and blaCTX-M-14, one blaCTX-M-9, and one blaCTX-M-8. Among the CTX-M-15 producers, 92% belonged to sequence types ST131 and ST405, and carried aac(6’)Ib-cr as well. Isolates harboring blaCTX-M-2, blaCTX-M-14, blaCTX-M-9, or blaCTX-M-8 were found to be genetically unrelated. Conclusions: The successful dissemination of CTX-M-15-producing E.coli isolates seems to be linked to the spreading of high-risk clones and horizontal gene transfer. A trade-off between carrying more antibiotic resistance and less virulence-related genes could partially account for the evolutionary advantages featured by successful clones.Fil: Vignoli, Rafael. Universidad de la República; UruguayFil: García Fulgueiras, Virginia. Universidad de la República; UruguayFil: Cordeiro, Nicolás F.. Universidad de la República; UruguayFil: Bado, Inés. Universidad de la República; UruguayFil: Seija, Verónica. Universidad de la República; Uruguay. Hospital Pasteur de Montevideo; UruguayFil: Aguerrebere, Paula. Universidad de la República; UruguayFil: Laguna, Gabriel. Universidad de la República; UruguayFil: Araújo, Lucía. Universidad de la República; UruguayFil: Bazet, Cristina. Universidad de la República; UruguayFil: Gutkind, Gabriel Osvaldo. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Chabalgoity Rodríguez, José Alejandro. Universidad de la República; Urugua

Survival of Staphylococcus aureus ST398 in the human nose after artificial inoculation.

There is evidence that MRSA ST398 of animal origin is only capable of temporarily occupying the human nose, and it is therefore, often considered a poor human colonizer.We inoculated 16 healthy human volunteers with a mixture of the human MSSA strain 1036 (ST931, CC8) and the bovine MSSA strain 5062 (ST398, CC398), 7 weeks after a treatment with mupirocin and chlorhexidine-containing soap. Bacterial survival was studied by follow-up cultures over 21 days. The human strain 1036 was eliminated faster (median 14 days; range 2-21 days) than the bovine strain 5062 (median 21 days; range 7-21 days) but this difference was not significant (p = 0.065). The bacterial loads were significantly higher for the bovine strain on day 7 and day 21. 4/14 volunteers (28.6%) showed elimination of both strains within 21 days. Of the 10 remaining volunteers, 5 showed no differences in bacterial counts between both strains, and in the other 5 the ST398 strain far outnumbered the human S. aureus strain. Within the 21 days of follow-up, neither human strain 1036 nor bovine strain 5062 appeared to acquire or lose any mobile genetic elements. In conclusion, S. aureus ST398 strain 5062 is capable of adequately competing for a niche with a human strain and survives in the human nose for at least 21 days

Massive Increase, Spread, and Exchange of Extended Spectrum {beta}-Lactamase-Encoding Genes Among Intestinal Enterobacteriaceae in Hospitalized Children With Severe Acute Malnutrition in Niger.

Background. From the time of CTX-M emergence, extended-spectrum β-lactamase-producing enterobacteria (ESBL-E) have spread worldwide in community settings as well as in hospitals, particularly in developing countries. Although their dissemination appears linked to Escherichia coli intestinal carriage, precise paths of this dynamic are largely unknown. Methods. Children from a pediatric renutrition center were prospectively enrolled in a fecal carriage study. Antibiotic exposure was recorded. ESBL-E strains were isolated using selective media from fecal samples obtained at admission and, when negative, also at discharge. ESBL-encoding genes were identified, their environments and plasmids were characterized, and clonality was assessed with polymerase chain reaction-based methods and pulsed-field gel electrophoresis for E. coli and Klebsiella pneumoniae. E. coli strains were subjected to multilocus sequence typing. Results. The ESBL-E carriage rate was 31% at admission in the 55 children enrolled. All children enrolled received antibiotics during hospitalization. Among the ESBL-E-negative children, 16 were resampled at discharge, and the acquisition rate was 94%. The bla(CTX-M-15) gene was found in >90% of the carriers. Genetic environments and plasmid characterization evidenced the roles of a worldwide, previously described, multidrug-resistant region and of IncF plasmids in CTX-M-15 E. coli dissemination. Diversity of CTX-M-15-carrying genetic structures and clonality of acquired ESBL E. coli suggested horizontal genetic transfer and underlined the potential of some ST types for nosocomial cross-transmission. Conclusions. Cross-transmission and high selective pressure lead to very high acquisition of ESBL-E carriage, contributing to dissemination in the community. Strict hygiene measures as well as careful balancing of benefit-risk ratio of current antibiotic policies need to be reevaluated

Saturation Mutagenesis of Lysine 12 Leads to the Identification of Derivatives of Nisin A with Enhanced Antimicrobial Activity

peer-reviewedIt is becoming increasingly apparent that innovations from the “golden age” of antibiotics are becoming ineffective, resulting in a pressing need for novel therapeutics. The bacteriocin family of antimicrobial peptides has attracted much attention in recent years as a source of potential alternatives. The most intensively studied bacteriocin is nisin, a broad spectrum lantibiotic that inhibits Gram-positive bacteria including important food pathogens and clinically relevant antibiotic resistant bacteria. Nisin is gene-encoded and, as such, is amenable to peptide bioengineering, facilitating the generation of novel derivatives that can be screened for desirable properties. It was to this end that we used a site-saturation mutagenesis approach to create a bank of producers of nisin A derivatives that differ with respect to the identity of residue 12 (normally lysine; K12). A number of these producers exhibited enhanced bioactivity and the nisin A K12A producer was deemed of greatest interest. Subsequent investigations with the purified antimicrobial highlighted the enhanced specific activity of this modified nisin against representative target strains from the genera Streptococcus, Bacillus, Lactococcus, Enterococcus and Staphylococcus.This work was supported by the Irish Government under the National Development Plan; by the Irish Research Council for Science Engineering and Technology (IRCSET); by Enterprise Ireland; and by Science Foundation Ireland (SFI), through the Alimentary Pharmabiotic Centre (APC) at University College Cork, Ireland, which is supported by the SFI-funded Centre for Science, Engineering and Technology (SFI-CSET) and provided P.D.C., C.H. and R.P.R. with SFI Principal Investigator funding

Phenotypic microarrays suggest Escherichia coli ST131 is not a metabolically distinct lineage of extra-intestinal pathogenic E. coli

Extraintestinal pathogenic E. coli (ExPEC) are the major aetiological agent of urinary tract infections (UTIs) in humans. The emergence of the CTX-M producing clone E. coli ST131 represents a major challenge to public health worldwide. A recent study on the metabolic potential of E. coli isolates demonstrated an association between the E. coli ST131 clone and enhanced utilisation of a panel of metabolic substrates. The studies presented here investigated the metabolic potential of ST131 and other major ExPEC ST isolates using 120 API test reagents and found that ST131 isolates demonstrated a lower metabolic activity for 5 of 120 biochemical tests in comparison to non-ST131 ExPEC isolates. Furthermore, comparative phenotypic microarray analysis showed a lack of specific metabolic profile for ST131 isolates countering the suggestion that these bacteria are metabolically fitter and therefore more successful human pathogens

PinR mediates the generation of reversible population diversity in Streptococcus zooepidemicus

Opportunistic pathogens must adapt to and survive in a wide range of complex ecosystems. Streptococcus zooepidemicus is an opportunistic pathogen of horses and many other animals, including humans. The assembly of different surface architecture phenotypes from one genotype is likely to be crucial to the successful exploitation of such an opportunistic lifestyle. Construction of a series of mutants revealed that a serine recombinase, PinR, inverts 114 bp of the promoter of SZO_08560, which is bordered by GTAGACTTTA and TAAAGTCTAC inverted repeats. Inversion acts as a switch, controlling the transcription of this sortase-processed protein, which may enhance the attachment of S. zooepidemicus to equine trachea. The genome of a recently sequenced strain of S. zooepidemicus, 2329 (Sz2329), was found to contain a disruptive internal inversion of 7 kb of the FimIV pilus locus, which is bordered by TAGAAA and TTTCTA inverted repeats. This strain lacks pinR and this inversion may have become irreversible following the loss of this recombinase. Active inversion of FimIV was detected in three strains of S. zooepidemicus, 1770 (Sz1770), B260863 (SzB260863) and H050840501 (SzH050840501), all of which encoded pinR. A deletion mutant of Sz1770 that lacked pinR was no longer capable of inverting its internal region of FimIV. The data highlight redundancy in the PinR sequence recognition motif around a short TAGA consensus and suggest that PinR can reversibly influence the wider surface architecture of S. zooepidemicus, providing this organism with a bet-hedging solution to survival in fluctuating environments

Characterization and Comparison of 2 Distinct Epidemic Community-Associated Methicillin-Resistant Staphylococcus aureus Clones of ST59 Lineage.

Sequence type (ST) 59 is an epidemic lineage of community-associated (CA) methicillin-resistant Staphylococcus aureus (MRSA) isolates. Taiwanese CA-MRSA isolates belong to ST59 and can be grouped into 2 distinct clones, a virulent Taiwan clone and a commensal Asian-Pacific clone. The Taiwan clone carries the Panton-Valentine leukocidin (PVL) genes and the staphylococcal chromosomal cassette mec (SCCmec) VT, and is frequently isolated from patients with severe disease. The Asian-Pacific clone is PVL-negative, carries SCCmec IV, and a frequent colonizer of healthy children. Isolates of both clones were characterized by their ability to adhere to respiratory A549 cells, cytotoxicity to human neutrophils, and nasal colonization of a murine and murine sepsis models. Genome variation was determined by polymerase chain reaction of selected virulence factors and by multi-strain whole genome microarray. Additionally, the expression of selected factors was compared between the 2 clones. The Taiwan clone showed a much higher cytotoxicity to the human neutrophils and caused more severe septic infections with a high mortality rate in the murine model. The clones were indistinguishable in their adhesion to A549 cells and persistence of murine nasal colonization. The microarray data revealed that the Taiwan clone had lost the ø3-prophage that integrates into the β-hemolysin gene and includes staphylokinase- and enterotoxin P-encoding genes, but had retained the genes for human immune evasion, scn and chps. Production of the virulence factors did not differ significantly in the 2 clonal groups, although more α-toxin was expressed in Taiwan clone isolates from pneumonia patients. In conclusion, the Taiwan CA-MRSA clone was distinguished by enhanced virulence in both humans and an animal infection model. The evolutionary acquisition of PVL, the higher expression of α-toxin, and possibly the loss of a large portion of the β-hemolysin-converting prophage likely contribute to its higher pathogenic potential than the Asian-Pacific clone