Dynamics of clonal and plasmid backgrounds of Enterobacteriaceae producing acquired AmpC in Portuguese clinical settings over time.

OBJECTIVES: The objective of this work was to provide detailed molecular data on clinically acquired AmpC (qAmpC)-producing Enterobacteriaceae from two different periods (2002-2008 and 2010-2013) in order to clarify the contribution of clonal and plasmid genetic platforms for the current epidemiological scenario concerning extended-spectrum beta-lactams resistance. METHODS: We analysed 1246 Enterobacteriaceae non-susceptible to third-generation cephalosporins from two hospitals and one community laboratory between 2010 and 2013. Bacterial identification, antibiotic susceptibility, identification of qAmpC and plasmid-mediated quinolone resistance genes, clonal (pulsed-field gel electrophoresis (PFGE), Multilocus sequence typing (MLST)) and plasmid (S1-/I-CeuI-PFGE, replicon typing, hybridization) analysis were performed by standard methods. Whole-genome sequencing (WGS) was performed in two ST11-Klebsiella pneumoniae isolates harbouring DHA-1. RESULTS: The occurrence of qAmpC was lower (2.6%) than that observed in a previous survey (7.4%), and varied slightly over time. Isolates produced DHA-1 (53%), CMY-2 (44%) or DHA-6 (3%), but significant epidemiological changes were observed in the two surveys. While DHA-1 persisted in different institutions by selection of a worldwide epidemic IncR plasmid in an ST11 harbouring KL105, CMY-2 rates increased over time linked to IncI1 plasmids (instead of IncK or IncA/C2) in multiple Escherichia coli clones. CONCLUSIONS: The higher frequency of DHA-1 qAmpC in these species contrasts with the scenario in most European countries. Furthermore, the different genetic backgrounds associated with either extended-spectrum ß-lactamases (ESBLs) or acquired AmpC ß-lactamases (qAmpC) in our country might have contributed to their differential expansion.

High occurrence and unusual serotype diversity of non-typhoidal Salmonella in non-clinical niches, Angola.

Non-typhoidal Salmonella is an important burden, particularly in developing countries of the African region. We report for the first time in Angola, a sub-Saharan African country with commercial/travel relationships with Europe, an unexpectedly high occurrence of Salmonella (n = 12/63, 19%) from a high diversity of sources, particularly farm and wild animals. The detection of diverse serotypes (n = 12), involving putative new S. enterica subsp. salamae serotypes, is also of note, reinforcing the need for a comprehensive surveillance in Angola critical to identify animal/food/environmental sources of salmonellosis with impact on animal health, local people, tourists and exported products.

Atypical epidemiology of CTX-M-15 among Enterobacteriaceae from a high diversity of non-clinical niches in Angola.

OBJECTIVES: The objective of this study was to investigate the distribution and molecular epidemiology of ESBLs, acquired AmpCs and carbapenemases in Enterobacteriaceae from non-clinical niches in Angola, an under-researched sub-Saharan country. METHODS: Eighty-one samples were recovered from healthy persons (n = 18), healthy animals (n = 33) and their environments (n = 10) or aquatic settings (n = 20) in south Angola (2013). Samples were plated onto CHROMagar™ Orientation with/without antibiotics. Standard methods were used for bacterial identification, characterization of bla genes, antibiotic susceptibility testing and conjugation assays. Clonal analysis (XbaI-PFGE, MLST and Escherichia coli phylogroups), location of bla and plasmid characterization (S1-PFGE, I-CeuI-PFGE, replicon typing and hybridization) were also performed. RESULTS: ESBLs (almost exclusively CTX-M-15, 98%) were detected in 21% (45/216) of the isolates, recovered from diverse non-clinical niches and belonging to different Enterobacteriaceae species (mainly E. coli). Acquired AmpCs or carbapenemases were not found. The pandemic B2-ST131 E. coli clone was not identified, but some widespread clonal complexes (CCs) from A (CC10 and CC168), B1 (CC156) or D (CC38) phylogroups were detected. blaCTX-M-15 was variably identified on typeable (29%; 100-335 kb; IncFII, IncFIIK6, IncHI2 and IncY) or non-typeable (16%; 70-330 kb) plasmids or on the chromosome (14%), while for 41% of the isolates its specific location was not determined. CONCLUSIONS: This study reports, for the first time in Angola, an unexpected high occurrence of CTX-M-15 in diverse non-clinical niches and Enterobacteriaceae species, and uncovers novel plasmid replicons in under-researched geographical regions. The diffusion of blaCTX-M-15 through such a high diversity of genetic backgrounds (clones, typeable/non-typeable plasmids and genetic environments) unveils an extraordinary ability for blaCTX-M-15 acquisition and mobilization favoured by unrecognized ecological factors.

Long-term dissemination of acquired AmpC ß-lactamases among Klebsiella spp. and Escherichia coli in Portuguese clinical settings.

We investigated the occurrence, diversity and molecular epidemiology of genes coding for acquired AmpC ß-lactamases (qAmpC) among clinical isolates of Enterobacteriaceae lacking inducible chromosomal AmpCs in Portugal. A total of 675 isolates non-susceptible to broad-spectrum cephalosporins obtained from four hospitals and three community laboratories during a 7-year period (2002-2008) were analysed. The presence of genes coding for qAmpC was investigated by phenotypic criteria, polymerase chain reaction (PCR) and sequencing. Bacterial identification, antibiotic susceptibility testing, conjugation assays and clonal analysis were performed by standard procedures. The presence of bla(qAmpC) genes was detected in 50 % (50/100; 41 Klebsiella pneumoniae, 5 Escherichia coli, 4 Klebsiella oxytoca) of the presumptive qAmpC producers. DHA-1, detected in those species, was the most prevalent qAmpC (94 %, 47/50), being identified since 2003 and throughout the studied period in different institutions. Despite the high clonal diversity observed, three DHA-1-producing Klebsiella spp. clones were more frequently identified. CMY-2 (6 %, 3/50) was observed in B1-E. coli clones. Conjugative transfer was only observed in one (2 %) CMY-2-producing isolate. Most qAmpC producers (94 %, 47/50) co-expressed SHV-type and/or OXA-1 or CTX-M-32 extended-spectrum ß-lactamases (ESBLs). To the authors' knowledge, this is the first description of the molecular epidemiology and the long-term dissemination of qAmpC-producing Enterobacteriaceae in Portuguese clinical settings, highlighting an evolution towards a more complex epidemiological situation regarding cephalosporin resistance in Portugal.