Molecular epidemiology and virulence markers of Salmonella Infantis isolated over 25years in São Paulo State, Brazil

AbstractInfection of humans and animals caused by Salmonella is a major public health problem worldwide. Among the more than 2500 serovars, S. Infantis has been one of the 15 most isolated serovars in the world. Despite its clinical importance, little is known about the molecular characteristics of S. Infantis strains from Brazil. The aims of this study were to type S. Infantis isolates of this country and to assess their pathogenic potential. The molecular epidemiology of 35 S. Infantis strains, isolated from human sources (25) and food items (10) between 1984 and 2009 in São Paulo State, Brazil, were investigated using ERIC-PCR, PFGE and MLST. Furthermore, the presence of some virulence markers from Salmonella pathogenicity islands (SPIs) SPI-1 and SPI-2 and from the virulence plasmid was assessed by PCR. Using ERIC-PCR, 34 S. Infantis strains exhibited a high genetic similarity (⩾93.7%) and using PFGE, 32 strains exhibited a similarity ⩾80.6%. Additionally, MLST showed a high clonal similarity among strains that all presented the same ST32. Thirty-two isolates under investigation contained the virulence markers invA, sopB, sopD, sipA, sipD, ssaR, sifA, flgK, fljB and flgL. In conclusion, the S. Infantis strains studied were genetically similar, suggesting that a prevalent subtype has been causing disease and food contamination during a 25year period in São Paulo State, an important metropolitan region in Brazil. Furthermore, the contamination between strains from food items and sick humans indicates that better control measures for S. Infantis may be needed in this country

Antimicrobial activity of products based on potassium monopersulfate on bacteria associated with avian infections

O objetivo deste trabalho foi avaliar a atividade antimicrobiana de quatro novos produtos à base de monopersulfato de potássio em biofilmes bacterianos associados a infecções aviárias, para a desinfecção de bebedouros em granjas avícolas. Inicialmente, realizaram-se testes em células bacterianas planctônicas, para verificar a atividade antimicrobiana e a concentração inibitória mínima dos produtos, denominados PA, PB, PC e PD. Esses produtos foram testados em biofilmes maduros dos patógenos aviários Escherichia coli, Salmonella enterica subsp. enterica serovar Enteritidis e Listeria monocytogenes, cultivados em espécimes preservados em blocos de acrílico, por meio da contagem de unidades formadoras de colônias, por microscopia eletrônica de varredura e por microscopia de fluorescência confocal. Todos os produtos foram eficazes contra as espécies bacterianas avaliadas. Os produtos PA e PB inibiram o crescimento bacteriano em concentrações ≤ 0,13%, e PC e PD apresentaram o mesmo efeito em concentrações ≤ 0,25%. Além disso, o produto PA foi capaz de eliminar os biofilmes maduros de S. enterica subsp. enterica serovar Enteritidis e L. monocytogenes. Os produtos à base de monopersulfato de potássio avaliados, notadamente o PA, são eficazes contra biofilmes bacterianos associados a infecções aviárias e apresentam potencial como sanitizantes e desinfetantes em bebedouros de granjas avícolas.The objective of this work was to evaluate the antimicrobial activity of four new potassium monopersulfate-based products on bacterial biofilms associated with avian infections, in order to disinfect drinking fountains in poultry farms. Initially, tests were performed in planktonic bacterial cells, to verify the antimicrobial activity and the minimum inhibitory concentration of the products, named PA, PB, PC, and PD. These products were tested on mature biofilms of the avian pathogens Escherichia coli, Salmonella enterica subsp. enterica serovar Enteritidis, and Listeria monocytogenes, grown on specimens preserved in acrylic blocks, by counting colony-forming units, scanning electron microscopy, and confocal fluorescence microscopy. All products were effective against the evaluated bacterial species. The PA and PB products inhibited the bacterial growth at ≤ 0.13% concentrations, and PC and PD showed the same effect at ≤ 0.25% concentrations. Furthermore, the PA product was able to eliminate mature biofilms of S. enterica subsp. enterica serovar Enteritidis and L. monocytogenes. The evaluated monopersulfate-based products, notably PA, are effective against bacterial biofilms associated with avian infections and show potential as sanitizers and disinfectants for drinking fountains in poultry farms

High Prevalence of Multidrug-Resistant Klebsiella pneumoniae Harboring Several Virulence and β-Lactamase Encoding Genes in a Brazilian Intensive Care Unit

Klebsiella pneumoniae is an important opportunistic pathogen that commonly causes nosocomial infections and contributes to substantial morbidity and mortality. We sought to investigate the antibiotic resistance profile, pathogenic potential and the clonal relationships between K. pneumoniae (n = 25) isolated from patients and sources at a tertiary care hospital’s intensive care units (ICUs) in the northern region of Brazil. Most of K. pneumoniae isolates (n = 21, 84%) were classified as multidrug resistant (MDR) with high-level resistance to β-lactams, aminoglycosides, quinolones, tigecycline, and colistin. All the 25 isolates presented extended-spectrum beta-lactamase-producing (ESBL), including carbapenemase producers, and carried the blaKPC (100%), blaTEM (100%), blaSHV variants (n = 24, 96%), blaOXA-1 group (n = 21, 84%) and blaCTX-M-1 group (n = 18, 72%) genes. The K2 serotype was found in 4% (n = 1) of the isolates, and the K1 was not detected. The virulence-associated genes found among the 25 isolates were mrkD (n = 24, 96%), fimH-1 (n = 22, 88%), entB (100%), iutA (n = 10, 40%), ybtS (n = 15, 60%). The genes related with efflux pumps and outer membrane porins found were AcrAB (100%), tolC (n = 24, 96%), mdtK (n = 22, 88%), OmpK35 (n = 15, 60%), and OmpK36 (n = 7, 28%). ERIC-PCR was employed to determine the clonal relationship between the different isolated strains. The obtained ERIC-PCR patterns revealed that the similarity between isolates was above 70%. To determine the sequence types (STs) a multilocus sequence typing (MLST) assay was used. The results indicated the presence of high-risk international clones among the isolates. In our study, the wide variety of MDR K. pneumoniae harboring β-lactams and virulence genes strongly suggest a necessity for the implementation of effective strategies to prevent and control the spread of antibiotic resistant infections

Whole genome sequencing of the multidrug-resistant Chryseobacterium indologenes isolated from a patient in Brazil

Chryseobacterium indologenes is a non-glucose-fermenting Gram-negative bacillus. This emerging multidrug resistant opportunistic nosocomial pathogen can cause severe infections in neonates and immunocompromised patients. This study aimed to present the first detailed draft genome sequence of a multidrug-resistant C. indologenes strain isolated from the cerebrospinal fluid of an infant hospitalized at the Neonatal Intensive Care Unit of Brazilian Tertiary Hospital. We first analyzed the susceptibility of C. indologenes strain to different antibiotics using the VITEK 2 system. The strain demonstrated an outstanding resistance to all the antibiotic classes tested, including β-lactams, aminoglycosides, glycylcycline, and polymyxin. Next, C. indologenes was whole-genome-sequenced, annotated using Prokka and Rapid Annotation using Subsystems Technology (RAST), and screened for orthologous groups (EggNOG), gene ontology (GO), resistance genes, virulence genes, and mobile genetic elements using different software tools. The draft genome contained one circular chromosome of 4,836,765 bp with 37.32% GC content. The genomic features of the chromosome present numerous genes related to cellular processes that are essential to bacteria. The MDR C. indologenes revealed the presence of genes that corresponded to the resistance phenotypes, including genes to β-lactamases (blaIND–13, blaCIA–3, blaTEM–116, blaOXA–209, blaVEB–15), quinolone (mcbG), tigecycline (tet(X6)), and genes encoding efflux pumps which confer resistance to aminoglycosides (RanA/RanB), and colistin (HlyD/TolC). Amino acid substitutions related to quinolone resistance were observed in GyrA (S83Y) and GyrB (L425I and K473R). A mutation that may play a role in the development of colistin resistance was detected in lpxA (G68D). Chryseobacterium indologenes isolate harbored 19 virulence factors, most of which were involved in infection pathways. We identified 13 Genomic Islands (GIs) and some elements associated with one integrative and conjugative element (ICEs). Other elements linked to mobile genetic elements (MGEs), such as insertion sequence (ISEIsp1), transposon (Tn5393), and integron (In31), were also present in the C. indologenes genome. Although plasmids were not detected, a ColRNAI replicon type and the most resistance genes detected in singletons were identified in unaligned scaffolds. We provided a wide range of information toward the understanding of the genomic diversity of C. indologenes, which can contribute to controlling the evolution and dissemination of this pathogen in healthcare settings

A Fatal Bacteremia Caused by Hypermucousviscous KPC-2 Producing Extensively Drug-Resistant K64-ST11 Klebsiella pneumoniae in Brazil

We report a fatal bacteremia caused by Klebsiella pneumoniae in a 60–70-year-old patient from Brazil. The genomic analysis of three isolates (from blood culture, nasal and anal swabs) showed that the bacteremia was caused by a KPC-2 producing extensively drug-resistant K64-ST11 hypermucousviscous K. pneumoniae (hmKP) harboring several virulence and antimicrobial resistance genes. Although the isolates did not present virulence markers associated with hypervirulent K. pneumoniae (hvKP), they showed invasion and toxicity to epithelial Hep-2 cells; resistance to cell microbicidal mechanisms; and blood and human serum survival, evidencing their pathogenic potential. This study highlights the risk of infection caused by hmKp strains not characterized as hvKP as well as the clinical implications and difficulty of treatment, especially in elderly or immunocompromised patients

New sequence type in multidrug-resistant Klebsiella pneumoniae harboring the blaNDM-1-encoding gene in Brazil

Objectives: The aim of this study was to investigate the blaNDM gene, pathogenic potential, and antimicrobial resistance of clinical isolates of carbapenem-resistant Klebsiella pneumoniae isolated from patients admitted to the University Hospital of Londrina between January 2014 and March 2017. Methods: blaNDM-1 and virulence genes were investigated using conventional PCR methods Antimicrobial susceptibility testing was performed by disk diffusion method according to the Clinical and Laboratory Standards Institute guidelines of 2017. Clonal relationships of the New Delhi metallo-β-lactamase (NDM)-positive isolates were determined by enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus sequence typing (MLST). Results: A total of 825 K. pneumoniae were identified, with four isolates (Kp6408, Kp6410, Kp6411, and Kp6715) presenting the blaNDM-1 gene. All NDM-1-producing isolates showed co-production of blaKPC-2 and blaTEM genes and also the virulence genes kfu, entB, mrkD, and fimH. Three isolates (Kp6408, Kp6410, and Kp6715) were classified as multidrug-resistant (MDR) and one (Kp6411) as extensively drug-resistant (XDR). ERIC-PCR analyses demonstrated that the isolates shared about 60% genetic similarity. MLST revealed four different sequence types (STs), described for the first time in Brazil, with two novel STs described in this study: ST3371 and ST3372. Conclusion: This study reports the identification of NDM-1 associated with KPC and virulence genes in four MDR K. pneumoniae with STs first described in Brazil. Keywords: MLST, Carbapenemases, Klebsiella pneumoniae, ND

Changes in vancomycin-resistant Enterococcus faecium causing outbreaks in Brazil

Enterococci have been implicated in severe human infections as a consequence of associated determinants of virulence and antimicrobial resistance. The majority of vancomycin-resistant Enterococcus faecium (VRE(fm)) connected to outbreaks worldwide pertains to the clonal complex 17 (CC17). In Brazil, the majority of VRE(fm) involved in outbreaks reported so far are not related to CC17. VRE(fm) strains responsible for an outbreak and sporadic cases in hospitals located in the city of Campinas, Brazil, were compared to other VRE(fm) strains in the country. Twenty-two out of 23 E. faecium were vancomycin-resistant and harboured the vanA gene. One vancomycin-susceptible E. faecium (VSE(fm)) strain was included in this study because it was isolated from a patient who one week later harboured a VRE(fm). All strains, except VSE, showed the same alteration in the VanA element characterised by deletion of the left extremity of the transposon and insertion of IS1251 between the vanS and vanH genes. Genes codifying virulence factors such as collageneadhesin protein, enterococcal surface protein and hyaluronidase were detected in the VRE(fm) and VSE(fm) studied. Both pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) revealed that VRE(fm) and VSE(fm) strains have a clonal relationship. New sequence types (STs) were identified by MLST as ST447, ST448, ST478 and ST412 but all belonged to the CC17. The present study revealed that VRE(fm) outbreaks in Brazil were caused by strains that did not share a common evolutionary history, and that VRE(fm) strains belonging to CC17 could be predominant in Brazil as in other countries. (C) 2011 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)[FAPESP 2008/56379-0