Mariana Pagano

High endemic levels of multidrug-resistant Acinetobacter baumannii among hospitals in southern Brazil

BACKGROUND Most published data on multidrug-resistant Acinetobacter baumanii (MDR Ab) are derived from outbreaks. We report incidence trends on health care-acquired infections due to MDR Ab over a 12-month period in the city of Porto Alegre in southern Brazil. METHODS Clinical and epidemiologic data were obtained from the local health care information system of the municipal health department. Polymerase chain reaction was used to detect the presence of the genes bla(OXA-23-like), bla(OXA-24-like), bla(OXA-51), and bla(OXA-58), and repetitive sequence-based polymerase chain reaction and pulsed-field gel electrophoresis were performed for molecular typing. RESULTS The highest rate of infection (9.0/1,000 inpatient-days) was identified in a trauma hospital. The gene bla(OXA-23-like) was identified in 99.0% of MDR Ab isolates. Eight main clonal groups were identified by molecular typing, and 3 of these were found in all hospitals. CONCLUSION The presence of 3 clones in all hospitals demonstrates the ability of MDR Ab to spread among hospitals. Moreover, the occurrence of one particular clone (clone 4) throughout the study period suggests its increased ability to cause outbreaks and to remain in the environment. The monitoring of epidemic strains by molecular methods is of paramount importance to prevent or reduce the spread of MDR Ab.

Emergence of NDM-1-producing Enterobacteriaceae in Porto Alegre, Brazil

OBJECTIVES To evaluate the emergence of New Delhi metallo-β-lactamase 1 (NDM-1)-producing Enterobacteriaceae isolates in Brazil. METHODS From April to October 2013, following the detection of the first NDM-1-producing isolate, a surveillance study was performed for the detection of blaNDM-1 among Enterobacteriaceae isolates with reduced susceptibility to carbapenems in 17 hospitals of Porto Alegre, Brazil. Real-time PCR was used to determine the presence of carbapenemase genes, which were further sequenced. Clonal relatedness was assessed by pulsed-field gel electrophoresis (PFGE). RESULTS A total of 1134 isolates were evaluated. blaNDM-1 was detected in 11 (0.97%) isolates: nine Enterobacter cloacae complex (eight belonging to a single clone recovered from two distinct hospitals and the other strain from a third hospital) and two Morganella morganii (belonging to a single clone recovered from one hospital). Most isolates presented high-level resistance to carbapenems. CONCLUSIONS NDM-1-producing Enterobacteriaceae have emerged rapidly in the hospitals of the Brazilian city where they were first detected. The emergence of NDM-1 in Brazil is of great concern, since it is a severe threat to antimicrobial therapy against Enterobacteriaceae in this country.

Emergence of NDM-1-producing Acinetobacter pittii in Brazil

Sir, The New Delhi metallo-lactamase (NDM), initially reported in Klebsiella pneumoniae and Escherichia coli, is now disseminated worldwide mostly among Enterobacteriaceae [1]. The NDM carbapenemase has also been described in Acinetobacter baumannii, but only in sporadic cases in countries such as China, India, Egypt, Germany, Israel and, more recently, Brazil [1,2]. Noteworthy, recent studies reported NDM-producers among non-baumannii Acinetobacter spp., which may also be human pathogens. Here we report the first case of NDM-1-producing Acinetobacter pittii in Brazil. A 66-year-old male patient with bladder carcinoma was admitted for radical cystectomy to a 900-bed tertiary care hospital in Porto Alegre, Southern Brazil, on 25 February 2013. Fifteen days later he presented an intestinal subocclusion and fever. Computerised tomography (CT) of the abdomen showed the presence of a collection in pelvis, which was drained surgically. This purulent secretion was cultured and a K. pneumoniae was identified (VITEK® 2 system; bioMérieux, La Balme-les-Grottes, France). Urine was also cultured and revealed the presence of Candida sp. (50 000 CFU/mL) and Acinetobacter sp. (>100 000 CFU/mL). The patient was treated with intravenous meropenem 500 mg every 12 h for 7 days, followed by cefepime 1 g every 24 h (doses adjusted to impaired renal function). Three subsequent urine cultures obtained 11, 28 and 44 days after the first culture were negative for Acinetobacter sp. The patient was therefore considered colonised by Acinetobacter sp. After 90 days the patient improved and was discharged from the hospital. The Acinetobacter sp. isolate MP was identified as A. pittii by matrix-assisted laser desorption/ionisation time-of-flight (MALDITOF) (Bruker Daltonik, Bremen, Germany), gyrB multiplex PCR and 16S rRNA gene sequencing. Minimum inhibitory concentrations (MICs) of -lactams, aminoglycosides, ciprofloxacin, fosfomycin, chloramphenicol, tigecycline, colistin and polymyxin B were determined (Etest® and microdilution method) and showed that the isolate was resistant to all -lactams (with the exception of aztreonam), including carbapenems (MICs of imipenem, ertapenem, doripenem and meropenem >32 g/mL). The isolate remained susceptible to amikacin, gentamicin, tigecycline, colistin, polymyxin B, ciprofloxacin and chloramphenicol. Carbapenemase genes were searched by real-time PCR (blaOXA-48, blaKPC, blaIMP, blaVIM and blaGES) and multiplex PCR (blaOXA-23-like, blaOXA-40-like, blaOXA-58-like and blaOXA-143). A positive signal was obtained only for the blaNDM gene, and sequencing identified the blaNDM-1 gene. To identify the location of this gene, electrotransformation assays were attempted using plasmid DNA extracts from A. pittii isolate MP using A. baumannii CIP7010 and E. coli TOP10 as recipients. Transfer of the blaNDM-1 gene by electrotransformation into these two recipient strains remained unsuccessful, suggesting that the gene might be chromosomally located in A. pittii MP, as reported in A. baumannii [3]. The genetic environment of the blaNDM-1 gene was determined by PCR mapping as described [3] and insertion sequence ISAba125 was identified upstream of the blaNDM-1 gene. However, attempts to identify another copy of ISAba125 downstream of blaNDM-1 remained unsuccessful, suggesting that the blaNDM-1 gene might be part of a truncated Tn125 transposon, as previously reported in A. baumannii [3]. Multilocus sequence typing (MLST) was performed according to the Institute Pasteur scheme (http://www.pasteur.fr) and A. pittii isolate MP was identified as ST119. Interestingly, two blaNDM-positive A. pittii isolates were recently identified in Paraguay [4], a neighbouring country of Brazil, but those isolates belonged to ST320 and ST321. The only reports of A. pittii ST119 isolates are from Japan, with isolates producing the carbapenemase IMP-19 [1]. Identification of blaNDM-positive non-baumannii Acinetobacter spp. is now increasingly reported worldwide, concomitantly with those of blaNDM-positive A. baumannii isolates. There are few reports of NDM-producing A. pittii, being from China, Turkey and recently Paraguay. This is of particular concern considering that Acinetobacter sp. may (i) act as reservoirs for blaNDM genes in non-human settings, as recently shown in several Chinese studies with identification of NDM-1-producers among Acinetobacter calcoaceticus and Acinetobacter junii from environmental samples from livestock farms [1], Acinetobacter johnsonii from hospital sewage [1] and Acinetobacter lwoffii from chickens [1], but also (ii) act as a source of blaNDM genes then horizontally transferred to enterobacterial species as evidenced [5].

Detection of blaGES-5 in Carbapenem-Resistant Kluyvera intermedia Isolates Recovered from the Hospital Environment

Vanessa B. Ribeiro, Alexandre P. Zavascki, Franciéli P. Rozales, Mariana Pagano, Cibele M. Magagnin, Carolina S. Nodari, Renato Cassol Ferreira da Silva, Micheline G. Dalarosa, Diego R. Falci, Afonso L. Barth ‹Laboratório de Pesquisa em Resistência Bacteriana, Centro de Pesquisa Experimental, Hospital de Clínicas de Porto Alegre (HCPA), Porto Alegre, Brazil; Infectious Diseases Service, Hospital de Clínicas de Porto Alegre, Porto Alegre, Brazil; Programa de Pós-Graduação em Ciências Farmacêuticas; Universidade Federal do Rio Grande do Sul (UFRGS), Porto Alegre, Brazil; Programa de Pós-Graduação em Ciências Médicas, UFRGS, Porto Alegre, Brazil; Hospital Nossa Senhora da Conceição, Porto Alegre, Brazil

Mobile genetic elements related to carbapenem resistance in Acinetobacter baumannii.

Acinetobacter baumannii is widely recognized as an important pathogen associated with nosocomial infections. The treatment of these infections is often difficult due to the acquisition of resistance genes. A. baumannii presents a high genetic plasticity which allows the accumulation of these resistance determinants leading to multidrug resistance. It is highlighted the importance of the horizontal transfer of resistance genes, through mobile genetic elements and its relationship with increased incidence of multidrug resistant A. baumannii in hospitals. Considering that resistance to carbapenems is very important from the clinical and epidemiological point of view, the aim of this article is to present an overview of the current knowledge about genetic elements related to carbapenem resistance in A. baumannii such as integrons, transposons, resistance islands and insertion sequences.

Emergence of OXA-72-producing Acinetobacter baumannii Belonging to High-Risk Clones (CC15 and CC79) in Different Brazilian States.

hand hygiene compliance observations and consecutive training efforts is important, given that <10% of all hand disinfections were performed correctly in an observational study by Tschudin-Sutter et al, who observed the 6-step technique. Appropriate hand-surface coverage was reached in only 7.9% of hand hygiene procedures observed by Park et al, despite a high rate of compliance with the correct indications. Shah et al performed a video observation of hand washing. Of 1,081 recordings, 403 (37.3%) were excellent, 521 (48.2%) were acceptable, and 157 (14.5%) were unacceptable. A limitation of our study is the lack of bacterial counts, but the results of Riley et al, who showed no correlation between hand coverage and bacterial counts with a 6-step technique compared to a 3-step approach, had not been published at the time of our experiment. Another limitation is the small number of participants and the experimental setting of this proof-of-principle study. However, we believe that based on our results, the addition of dichotomous subjective quality assessment using the parameters time and skin coverage during live observation by experienced infection control staff is feasible and could be a valuable addition to conventional hand hygiene observation.

Acinetobacter baumannii resistente aos carbapenêmicos no Brasil: perfil de suscetibilidade e diversidade de oxacilinases

Introduction: The Acinetobacter calcoaceticus-baumannii (ABC) complex includes five species, and the A. baumannii is the most important of them because it carries mechanisms of carbapenems resistance, especially the oxacillinases. Objectives: The objectives of this study were to identify the species of the ABC complex, to evaluate the susceptibility profile and to investigate the presence of oxacillinases in carbapenems-resistant isolates from four Brazilian States. Methods: In the study period, 92 isolates from Rio Grande do Sul (RS), Rio de Janeiro (RJ), Paraná (PR) and São Paulo (SP) were collected. The isolates were identified by matrixassisted laser desorption ionization-time of fight mass spectrometry (MALDI-TOF MS) and sequencing of gyrB gene. Evaluation of susceptibility was performed by disk diffusion and broth microdilution. The presence of oxacillinases was performed by in-house multiplex polymerase chain reaction (PCR). Results: Ninety-one (99%) isolates were identified as A. baumannii by MALDI-TOF and sequencing. The majority of isolates (56; 61%) showed resistance to the six antimicrobial agents tested. Three isolates were resistant to polymyxin B [minimum inhibitory concentration (MIC) ≥ 4 μg/ml). Eighty (87%) isolates were positive to OXA-23-like, and twelve (13%) isolates to OXA-24-like. Conclusion: Our findings confirm the knowledge about the dissemination of the bla OXA-23 gene in Brazil and suggest the recent emergence and spread of bla OXA-24 gene, since it was identified in three of the four sampled states.

Emergence of Acinetobacter baumannii ST730 carrying the blaOXA-72 gene in Brazil

Over the last decade, Acinetobacter baumannii resistant to carbapenems has emerged in many medical centres and has been commonly associated with high morbimortality. In Brazil, this resistance is mainly attributed to the spread of OXA-23-producing clones and, to a lesser extent, to OXA-143-producing clones. Here, we describe, for the first time, two OXA-72-producing A. baumannii isolates in southern Brazil to a broad spectrum of antibiotics, except polymyxin B and tigecycline. Molecular typing by multilocus sequence typing (MLST) demonstrated that both OXA-72-producing isolates belong to a new sequence type (ST), ST730, which was recently identified in OXA-23-producing A. baumannii isolates in São Paulo, Brazil. We demonstrate that the two A. baumannii ST730 isolates carrying blaOXA-72share a common ancestral origin with the blaOXA-23producers in Brazil. This observation reinforces the importance of strain-typing methods in order to clarify the dynamics of the emergence of new clones in a geographic region.

False-positive results in screening for metallo-β-lactamase are observed in isolates of Acinetobacter baumannii due to production of oxacilinases

Carbapenemases production, either metallo-lactamases (MBLs), KPC or oxacillinases (OXA), is the main resistance mechanism responsible for resistance phenotype to carbapenems in Acinetobacter.1 While for MBLs and KPCs there are some screening tests for their detection,2,3 the same is not true for oxacillinases. During the investigation of a large outbreak, we analyzed 584 carbapenem-resistant Acinetobacter spp. isolates from seven hospitals. Isolates were identified using the API 20NE system (Biomerieux, Basingstoke, United Kingdom). PCR for the blaOXA-51 gene was performed as a marker of Acinetobacter baumannii at species level.1 The MIC for imipenem was performed by Etest® (AB BIODISK, Solna, Sweden) and was ≥8 g/mL in all isolates. A total of 562 (96.3%) and 553 (95%) isolates proved to be OXA-51 and OXA-23 producers respectively, by a multiplex PCR, which included primers for the blaOXA-23-like, blaOXA-24-like, blaOXA-51, blaOXA-58, blaOXA-143 genes. 4 Among the 553 OXA-23-like A. baumannii producing isolates, we observed 86 (15.5%) positive isolates in the screening test for MBLs either by the disk-approximation test using a ceftazidime (CAZ) and a 2-mercaptopropionic acid (MPA) or by the Etest MBL (Imipenem/Imipenem + EDTA − AB BIODISK, Solna, Sweden).2,5 The modified Hodge Test was performed in these isolates and only nine (1.5%) were positive.6 PCR using blaIMP-1-like, blaVIM-2-like blaSPM-1, blaNDM-1, blaKPC-1,2 primers 2–4,7