Diversity of metallo-b-lactamase-encoding genes found in distinct species of Acinetobacter isolated from the Brazilian Amazon Region

BACKGROUND The multidrug resistance (MDR) phenotype is frequently observed in Acinetobacter baumannii, the most clinically relevant pathogenic species of its genus; recently, other species belonging to the A. calcoaceticus-A. baumannii complex have emerged as important MDR nosocomial pathogens. OBJECTIVES The present study aimed to verify the occurrence of metallo-β-lactamase genes among distinct Acinetobacter species in a hospital located in the Brazilian Amazon Region. METHODS Antimicrobial susceptibility profiles were determined by broth microdilution. The genetic relationships among these isolates were assessed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Pyrosequencing reads of plasmids carrying the bla NDM-1 gene were generated using the Ion Torrent™ platform sequencing. FINDINGS A total of six isolates carried bla NDM-1: A. baumannii (n = 2), A. nosocomialis (n = 3), and A. pittii (n = 1); three carried bla IMP-1: A. baumannii, A. nosocomialis, and A. bereziniae. Resistance to colistin was observed for an NDM-1-producing A. nosocomialis isolate. Diverse PFGE patterns and sequence types were found among A. nosocomialis and A. baumannii isolates. The bla NDM-1 sequence was inserted in a Tn125 transposon, while the bla IMP-1 was found as a gene cassette of the class 1 integron In86. MAIN CONCLUSIONS To the best of our knowledge, this is the first report describing the dissemination of bla NDM-1 among distinct Acinetobacter species recovered from the same hospital in South America.

High rate of detection of OXA-23-producing Acinetobacter from two general hospitals in Brazil

Abstract INTRODUCTION In recent decades, the prevalence of carbapenem-resistant Acinetobacter isolates has increased, and the production of oxacillinase (OXA)-type carbapenemases is the main mechanism underlying resistance. We evaluated OXA production from 114 Acinetobacter isolates collected between March and December 2013 from different clinical specimens of patients in two hospitals (Hospital 1 [n = 61] and Hospital 2 [n = 53]) located in Niterói, Rio de Janeiro, Brazil. We also evaluated the genetic diversity of OXA-producing isolates. METHODS All the isolates were identified through the automated system Vitek II and matrix-assisted laser desorption ionization-time of flight mass spectrometry MALDI-TOF MS as belonging to the A. baumannii-A. calcoaceticuscomplex. Antimicrobial susceptibility profiles were verified through agar diffusion tests. The presence of OXA-encoding genes was confirmed by PCR. The genetic diversity of isolates positive for carbapenemase production was analyzed through pulsed-field gel electrophoresis. RESULTS There was a high rate of resistance to carbapenems in the isolates (imipenem: 96%; meropenem: 92%) from both hospitals. Moreover, a high percentage (95.6%) of OXA-23-positive isolates was observed for both hospitals, indicating that this was the main mechanism of carbapenem-resistance among the studied population. In addition, most isolates (96.5%) were positive for bla OXA-51. A high genetic diversity and a few major genotypes were found among the OXA-23-positive isolates analyzed. Only intra-hospital dissemination was observed. CONCLUSIONS The elevated dissemination of bla OXA-23-like observed among Acinetobacter isolates from both the studied hospitals highlights the need for continuous epidemiological surveillance in these institutions.

Characterization of multidrug-resistant Acinetobacter ssp. strains isolated from medical intensive care units in Cali - Colombia / Caracterización de cepas de Acinetobacter spp. resistentes a múltiples fármacos aisladas de unidades de cuidados intensivos en Cali - Colombia

Abstract Introduction: The extensive use of antibiotics has led to the emergence of multi-resistant strains in some species of the genus Acinetobacter. Objective: To investigate the molecular characteristics of multidrug-resistant of Acinetobacter ssp. strains isolated from 52 patients collected between March 2009 and July 2010 in medical intensive care units in Cali - Colombia. Methods: The susceptibility to various classes of antibiotics was determined by disc diffusion method, and the determination of the genomic species was carried out using amplified ribosomal DNA restriction analysis (ARDRA) and by sequencing of the 16s rDNA gene. Also, the genes of beta-lactamases as well as, integrases IntI1 and IntI2 were analyzed by PCR method. Results: The phenotypic identification showed that the isolates belong mainly to A. calcoaceticus- A. baumannii complex. All of them were multi-resistant to almost the whole antibiotics except to tigecycline and sulperazon, and they were grouped into five (I to V) different antibiotypes, being the antibiotype I the most common (50.0%). The percent of beta-lactamases detected was: blaTEM (17.3%), blaCTX-M (9.6%), blaVIM (21.2%), blaIMP (7.7%), blaOXA-58 (21.2%), and blaOXA-51 (21.2%). The phylogenetic tree analysis showed that the isolates were clustering to A. baumannii (74.1%), A. nosocomialis (11.1%) and A. calcoaceticus (7.4 %). Besides, the integron class 1 and class 2 were detected in 23.1% and 17.3% respectively. Conclusion: The isolates were identified to species A. baumanii mainly, and they were multiresistant. The resistance to beta-lactams may be by for presence of beta-lactamases in the majority of the isolates.

Resumen Introducción: El uso extensivo de antibióticos ha llevado a la emergencia de cepas multirresistentes en algunas especies del género Acinetobacter. Objetivo: Investigar las características moleculares de resistencia a múltiples fármacos de cepas aisladas de Acinetobacter spp. colectadas entre marzo de 2009 y julio de 2010 en 52 pacientes de unidades de cuidados intensivos en Cali - Colombia. Métodos: La susceptibilidad a diversas clases de antibióticos se determinó mediante el método de difusión de disco, y la determinación de la especie genómica se llevó a cabo usando un análisis de restricción de ADN ribosómico amplificado (ARDRA) y mediante la secuenciación del gen 16s de ADNr. Además, se analizaron por el método de PCR los genes de las beta-lactamasas, como también, las integrasas IntI1 e IntI2. Resultados: La identificación fenotípica mostró que los aislamientos pertenecen principalmente al complejo A. calcoaceticus - A. baumannii. Todos ellos eran multirresistentes a casi todos los antibióticos excepto tigeciclina y sulperazón, y se agruparon en cinco (I a V) antibitipos diferentes, siendo el antibiotipo I el más común (50%). El porcentaje de betalactamasas detectadas fue: blaTEM (17,3%), blaCTX-M (9,6%), blaVIM (21,2%), blaIMP (7,7%), blaOXA-58 (21,2%), blaOXA- 51 (21.2%). El análisis del árbol filogenético mostró que los aislados se agrupaban en A. baumannii (74.1%), A. nosocomialis (11.1%) y A. calcoaceticus (7.4%). Además, el integrón clase 1 y clase 2 se detectaron en 23.1% y 17.3% respectivamente. Conclusión: los aislamientos se identificaron principalmente como la especie A. baumanii, y fueron multirresistentes. La resistencia a los betalactámicos puede deberse a la presencia de betalactamasas en la mayoría de los aislamientos.

Draft genome sequence of a multidrug-resistant beta-lactamase OXA-357-producing Acinetobacter pittii ST865 clinical isolate from China

Abstract Worldwide increasing emergence of carbapenem-resistant Acinetobacter spp. has rendered the limited availability of effective antimicrobial agents and has become a major public health concern. In this study, we report the draft genome sequence of A. pittii TCM156, a multidrug-resistant isolate that harbored the blaOXA-357 gene. The genome sequence was further analyzed by various bioinformatics methods. The genome size was estimated to be 3,807,313 bp with 3508 predicted coding regions and G + C content is 38.7%. These findings have raised awareness of the possible emergence of OXA-type enzyme-producing A. pittii isolate in China.

Draft genome sequence of phenol degrading Acinetobacter sp. Strain V2, isolated from oil contaminated soil

Abstract We report here the draft genome sequence of Acinetobacter sp. Strain V2 isolated from the oil contaminated soil collected from ENGEN, Amanzimtoti, South Africa. Degradation of phenolic compounds such as phenol, toluene, aniline etc. at 400 ppm in 24 h and oil degrading capability makes this organism an efficient multifunctional bioremediator. Genome sequencing of Acinetobacter spp. V2 was carried out on Illumina HiSeq 2000 platform (performed by the Beijing Genomics Institute [BGI], Shenzhen, China). The data obtained revealed 643 contigs with genome size of 4.0 Mb and G + C content of 38.59%.

Draft genome sequence of Acinetobacter pittii ST643 shared by cystic fibrosis patients

Acinetobacter pittii has emerged as an important hospital pathogen that is associated with outbreaks and drug resistance. In cystic fibrosis (CF) patients, the detection of Acinetobacter spp. is rare; however, we isolated the A. pittii sequence type ST643 in several Brazilian CF patients treated in the same centre. The current study describes the draft genome of A. pittii ST643.

GPo1 alkB gene expression for improvement of the degradation of diesel oil by a bacterial consortium

To facilitate the biodegradation of diesel oil, an oil biodegradation bacterial consortium was constructed. The alkane hydroxylase (alkB) gene of Pseudomonas putida GPo1 was constructed in a pCom8 expression vector, and the pCom8-GPo1 alkB plasmid was transformed into Escherichia coli DH5α. The AlkB protein was expressed by diesel oil induction and detected through SDS-polyacrylamide gel electrophoresis. The culture of the recombinant (pCom8-GPo1 alkB/E. coli DH5α) with the oil biodegradation bacterial consortium increased the degradation ratio of diesel oil at 24 h from 31% to 50%, and the facilitation rates were increased as the proportion of pCom8-GPo1 alkB/E. coli DH5α to the consortium increased. The results suggested that the expression of the GPo1 gene in E. coli DH5α could enhance the function of diesel oil degradation by the bacterial consortium.

Excess body weight in children may increase the length of hospital stay

OBJECTIVES: To investigate the prevalence of excess body weight in the pediatric ward of University Hospital and to test both the association between initial nutritional diagnosis and the length of stay and the in-hospital variation in nutritional status. METHODS: Retrospective cohort study based on information entered in clinical records from University Hospital. The data were collected from a convenience sample of 91 cases among children aged one to 10 years admitted to the hospital in 2009. The data that characterize the sample are presented in a descriptive manner. Additionally, we performed a multivariate linear regression analysis adjusted for age and gender. RESULTS: Nutritional classification at baseline showed that 87.8% of the children had a normal weight and that 8.9% had excess weight. The linear regression models showed that the average weight loss z-score of the children with excess weight compared with the group with normal weight was −0.48 (p = 0.018) and that their length of stay was 2.37 days longer on average compared with that of the normal-weight group (p = 0.047). CONCLUSIONS: The length of stay and loss of weight at the hospital may be greater among children with excess weight than among children with normal weight. .

Evaluation of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry-Based VITEK MS System for the Identification of Acinetobacter Species from Blood Cultures: Comparison with VITEK 2 and MicroScan Systems

BACKGROUND: Acinetobacter species are the leading cause of bloodstream infection (BSI), but their correct identification is challenging. We evaluated the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)-based VITEK MS (bioMerieux, France), and two automated systems, VITEK 2 (bioMerieux) and MicroScan (Siemens, USA) for identification of Acinetobacter BSI isolates. METHODS: A total of 187 BSI isolates recovered at a university hospital in Korea between 2010 and 2012 were analyzed. The identification results obtained using VITEK MS and two automated systems were compared with those of rpoB sequencing. RESULTS: Of 187 isolates analyzed, 176 were identified to the species level by rpoB sequencing: the Acinetobacter baumannii group (ABG; 101 A. baumannii, 43 A. nosocomialis, 10 A. pittii isolates) was most commonly identified (82.4%), followed by Acinetobacter genomic species 13BJ/14TU (5.3%), A. ursingii (2.1%), A. soli (2.1%), A. bereziniae (1.1%), and A. junii (1.1%). Correct identification rates to the species group (ABG) level or the species level was comparable among the three systems (VITEK MS, 90.3%; VITEK 2, 89.2%; MicroScan, 86.9%). However, VITEK MS generated fewer misidentifications (0.6%) than VITEK 2 (10.8%) and MicroScan (13.1%) (P<0.001). In addition, VITEK MS demonstrated higher specificity (100%) for discrimination between ABG and non-ABG isolates than the other systems (both, 31.8%) (P<0.001). CONCLUSIONS: The VITEK MS system is superior to the VITEK 2 and MicroScan systems for identification of Acinetobacter BSI isolates, with fewer misidentifications and better discrimination between the ABG and non-ABG isolates.

Study of Acinetobacter Species with Special Reference to Carbapenem Resistance.

Bacteria of genus acinetobacter, invariably susceptible to many antibiotics earlier have emerged as a multidrug resistant opportunistic pathogen in recent years. Unwarranted and unrestricted usage of an antibiotic is associated with drug resistance in thereat in infections by acinetobacter species. Rapid detection of Metallo – Betalactamase starins can be done to prevent their dissemtination. The present study is undertaken to know antibiotic sensitivity pattern of acinetobacter species and screen the imipenem resistant starins for Metallo Beta Lactamase (MBL) production by various phenotypic methods.

Molecular characterization of clinical multiresistant isolates of Acinetobacter sp. from hospitals in Porto Alegre, State of Rio Grande do Sul, Brazil / Caracterização molecular de isolados clínicos Acinetobacter sp. multirresistentes em hospitais de Porto Alegre, Estado do Rio Grande do Sul, Brasil

INTRODUCTION: Hospitals around the world have presented multiresistant Acinetobacter sp. outbreaks. The spread of these isolates that harbor an increasing variety of resistance genes makes the treatment of these infections and their control within the hospital environment more difficult. This study aimed to evaluate the occurrence and dissemination of Acinetobacter sp. multiresistant isolates and to identify acquired resistance genes. METHODS: We analyzed 274 clinical isolates of Acinetobacter sp. from five hospitals in Porto Alegre, RS, Brazil. We evaluated the susceptibility to antimicrobial, acquired resistance genes from Ambler's classes B and D, and performed molecular typing of the isolates using enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) technique. RESULTS: A high (68 percent) percentage of multiresistant isolates of Acinetobacter sp. was observed, and 69 percent were resistant to carbapenems. We identified 84 percent of isolates belonging to species A. baumannii because they presented the gene blaOXA-51. The gene blaOXA-23 was detected in 62 percent of the isolates, and among these, 98 percent were resistant to carbapenems. Using the ERIC-PCR technique, we identified clones of Acinetobacter sp. spread among the four hospitals analyzed during the sampling period. CONCLUSIONS: The data indicate the dissemination of Acinetobacter sp. isolates among hospitals and their permanence in the hospital after one year.

INTRODUÇÃO: Hospitais no mundo todo têm apresentado surtos de Acinetobacter sp. multirresistentes. A disseminação destes isolados com uma variedade cada vez maior de genes de resistência torna difícil o tratamento destas infecções e seu controle dentro do ambiente hospitalar. Este trabalho teve como objetivo avaliar a ocorrência e disseminação de isolados de Acinetobacter sp. multirresistentes e identificar genes de resistência adquirida. MÉTODOS: Foram avaliados 274 isolados clínicos de Acinetobacter sp. obtidos de cinco hospitais da Cidade de Porto Alegre, RS, Brasil. Avaliamos o perfil de suscetibilidade a antimicrobianos, genes de resistência adquirida das classes B e D de Ambler e realizamos a tipificação molecular dos isolados utilizando a técnica de enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR). RESULTADOS: Encontramos uma alta (68 por cento) porcentagem de isolados de Acinetobacter sp. multirresistentes e 69 por cento dos isolados apresentaram resistência aos carbapenêmicos. Foram identificados 84 por cento de isolados pertencentes a espécie A. baumannii, pois apresentaram o gene blaOXA-51. Em 62 por cento dos isolados, foi detectado o gene blaOXA-23, sendo que 98 por cento destes isolados foram resistentes aos carbapenêmicos. Através da tipificação molecular pela técnica de ERIC-PCR identificamos clones de Acinetobacter sp. disseminados entre quatro dos hospitais analisados e nos anos de 2006 e 2007. CONCLUSÕES: Os dados obtidos indicam a disseminação de isolados de Acinetobacter sp. entre hospitais assim como sua permanência no ambiente hospitalar após um ano.

Resistencia a los antibacterianos en América Latina: consecuencias para la infectología / Antibacterial drug resistance in Latin America: consequences for infectiousdisease control

Antibacterial drug resistance is a particularly significant issue in Latin America. This article explores antimicrobial resistance in three classes of clinically important bacteria: gram-positive bacteria, enterobacteria, and nonfermenting gram-negativebacilli. The gram-positive bacteria frequently responsible for infections in humans are for the most part cocci: staphylococci, streptococci (including pneumococci), and enterococci,in both community and hospital settings. This situation is no different in theRegion of the Americas. Among the gram-positive bacteria, the causative agents of bacteremia are most commonly strains of coagulase-negative Staphylococcus, followed by enterococci. This report explores the resistance of these species to different antimicrobial drugs, resistance mechanisms in community and hospital strains, and new drugs for treating infections caused by these bacteria. In Latin America, antimicrobialresistance in Enterococcus strains is still a minor problem compared to the situation in the United States. The strains of the genus Streptococcus isolated from respiratory infections are still sensitive to penicillin. Furthermore, the resistance of enterobacteriais extremely important in the Region, particularly because of the broad dissemination of CTX-M extended-spectrum beta-lactamases (ESBL), some of which originated in Latin America. This article analyzes the resistance of Streptococcus pneumoniae, betahemolytic streptococci, and viridans group streptococci. Among the nonfermentinggram-negative bacilli, while Pseudomonas aeruginosa strains remain the leading cause of bacteremia, infections caused by strains of Acinetobacter spp. have proliferatedextensively in some areas. With regard to antibiotics, several options are available for treating gram-positive bacterial infections...

La resistencia a los fármacos antibacterianos tiene particular importancia en América Latina. En este artículo se analiza la resistencia a los antimicrobianos de tres clases de bacterias de importancia clínica: bacterias grampositivas, enterobacterias y bacilos gramnegativos no fermentadores.Las bacterias grampositivas que producen infecciones humanas frecuentes son, en su mayoría, cocos: estafilococos, estreptococos (incluidos neumococos) y enterococos, tanto en elmedio comunitario como en el nosocomial. Esta situación no es diferente en la Región de las Américas. Entre las bacterias grampositivas, las que causan bacteriemia con mayor frecuencia corresponden a cepas de estafilococos coagulasa negativos, seguidas de las de enterococos. Eneste informe se analiza la resistencia de estas especies a distintos antimicrobianos, los mecanismosde resistencia para las cepas de origen hospitalario y comunitario y los nuevos medicamentos para tratar las infecciones por estas bacterias. La resistencia a los antimicrobianos delas cepas de Enterococcus en América Latina todavía es un problema menor en relación con la situación en los Estados Unidos de América. Las cepas del género Streptococcus aisladasde infecciones respiratorias aún son sensibles a penicilina. Por otra parte, la resistencia de las enterobacterias es de gran importancia en la Región, particularmente por la gran difusión debetalactamasas de espectro extendido (BLEE) de tipo CTX-M, algunas de las cuales se originaron en América Latina. En el presente artículo se analizan la situación de la resistencia de las cepas de Streptococcus pneumoniae, y de los estreptococos betahemolítico y del grupo viridans. Entre los bacilos gramnegativos no fermentadores, si bien las cepas de Pseudomonasaeruginosa siguen siendo la causa principal de bacteriemias, la proliferación de infecciones por cepas de Acinetobacter spp. tiene en algunas partes gran magnitud...

Molecular epidemiology of aminoglycosides resistance in Acinetobacter SPP. with emergence of multidrug-resistant strains

Acinetobacter spp. is characterized as an important nosocomial pathogen and increasing antimicrobial resistance. Our aim was to evaluate antimicrobial susceptibility and aminoglycosides resistance genes of Acinetobacter spp. isolated from hospitalized patients. Sixty isolates were identified as Acinetobacter species. The isolates were tested for antibiotic resistance by disc diffusion method for 12 antimicrobials. The presence of aphA6, aacC1 aadA1, and aadB genes were detected using PCR. From the isolated Acinetobacter spp. the highest resistance rate showed against amikacin, tobramycin, and ceftazidim, respectively; while isolated bacteria were more sensitive to ampicillic/subactam. More than 66% of the isolates were resistant to at least three classes of antibiotics, and 27.5% of MDR strains were resistant to all seven tested classes of antimicrobials. The higher MDR rate presented in bacteria isolated from the ICU and blood samples. More than 60% of the MDR bacteria were resistance to amikacin, ceftazidim, ciprofloxacin, piperacillin/tazobactam, doxycycline, tobramycin and levofloxacin. Also, more than 60% of the isolates contained phosphotransferase aphA6, and acetyltransferase genes aacC1, but adenylyltransferase genes aadA1 [41.7%], and aadB [3.3%] were less prominent. 21.7% of the strains contain three aminoglycoside resistance genes [aphA6, aacC1 and aadA1]. The rising trend of resistance to aminoglycosides poses an alarming threat to treatment of such infections. The findings showed that clinical isolates of Acinetobacter spp. in our hospital carrying various kinds of aminoglycoside resistance genes

Quorum sensing signal profile of Acinetobacter strains from nosocomial and environmental sources / Perfil de sensores de quórum en cepas nosocomiales y ambientales de Acinetobacter

A set of 43 strains corresponding to 20 classified and unclassified genomic Acinetobacter species was analyzed for the production of typical N-acyl homoserine lactone quorum sensing molecules in culture broths. A large percentage of the strains (74%) displayed quorum sensing signals that could be separated into three statistically significantly different chromatographic groups (p < 0.001) based on their retention factor in TLC, i.e. Rf1 (0.22 ± 0.02); Rf2 (0.40 ± 0.02) and Rf3 (0.54 ± 0.02). Noteworthy, 63% of the strains tested produced more than one quorum signal. The frequency of signal appearance was Rf3 > Rf2 > Rf1. None of the three signals could be specifically assigned to a particular species in the genus; furthermore, no distinction could be made between the quorum sensing signals secreted by typical opportunistic strains of the A. calcoaceticus-A. baumannii complex, isolated from patients, with respect to the other species of the genus, except for the Rf1 signal which was present in all the QS positive strains belonging to this complex and DNA group 13 TU. In conclusion, quorum sensors in Acinetobacter are not homogenously distributed among species and one of them is present in most of the A. calcoaceticus-baumannii complex.

Se analizó la producción de moléculas típicas de N-acil homoserina lactona con actividad de quorum sensing en cultivos líquidos de un grupo de 43 cepas correspondientes a 20 especies genómicas clasificadas y no clasificadas de Acinetobacter. Un porcentaje alto de las cepas (74%) mostraron señales de quorum sensing que pudieron ser separadas en tres grupos cromatográficos significativamente diferentes entre sí (p < 0,001) sobre la base de sus factores de retención en TLC, a saber: Rf1 (0.22 ± 0.02); Rf2 (0.40 ± 0.02) y Rf3 (0.54 ± 0.02). Es de notar que 63% de las cepas ensayadas produjeron más de una señal de quorum. La frecuencia de aparición de las señales fue Rf3 > Rf2 > Rf1. Ninguna de las tres señales pudo ser asignada a una especie en particular dentro del género; es más, no se encontró diferencia entre las señales producidas por las cepas típicamente oportunistas (complejo A. calcoaceticus-A. baumannii) aisladas de pacientes respecto de las producidas por otras cepas del mismo género, excepto para el caso de Rf1, que se encontró presente en todos los aislamientos quorum sensing positivos del mencionado complejo y en las cepas del grupo de DNA 13TU. En conclusión, los sensores de quórum en Acinetobacter no están homogéneamente distribuidos entre especies y uno de ellos (Rf1) está presente en la mayoría de los miembros del complejo calcoaceticus-baumannii.

Determinación de carbapenemasas y su relación con estructuras genéticas en aislamientos clínicos de Acinetobacter baumannii de hospitales de la ciudad de Cochabamba / Determinig carbapenemases and its relationship with genetic structures in clinical isolates of Acinetobacter baumannii at hospitals in the city of Cochabamba

En este estudio se analizaron un toal de 34 aislamientos clínicos de Acinetobacter baumannii con el fin de determinar la presencia de carbapenemasas y su relación con estructuras genéticas.

Rapid detection of non-enterobacteriaceae directly from positive blood culture using fluorescent in situ hybridization.

Fluorescent in situ hybridization (FISH) was carried out using two different oligonucleotide probes specific for Pseudomonas spp. and Acinetobacter spp. These probes were tested against different organisms and were found to be highly specific. Sensitivity testing showed that the probes were able to detect as low as 10 3 CFU/mL. In addition, FISH was carried out directly on positive blood culture samples and the detection of microorganisms took less than 2 h. We believe that FISH is a rapid method that can be used as a routine laboratory diagnostic technique for the detection of Acinetobacter spp. and Pseudomonas spp. in clinical samples.

Epidemiological investigation of nosocomial Acinetobacter infections using arbitrarily primed PCR & pulse field gel electrophoresis.

BACKGROUND & OBJECTIVE: Nosocomial infections caused by Acinetobacter spp. are a significant problem worldwide. Information on epidemiological investigation of outbreaks caused by Acinetobacter species in India is lacking. The present investigation was carried out to elucidate molecular epidemiology of Acinetobacter species isolated from nosocomial infections in a tertiary care hospital in south India using two DNA-based typing methods. METHODS: The medical records of 43 patients with Acinetobacter infection during a period of 24 months were reviewed and Acinetobacter isolates obtained from these patients were characterized phenotypically by assimilation tests and genotypically by arbitrarily primed PCR (AP-PCR) and pulse field gel electrophoresis (PFGE). Susceptibility testing results of the Acinetobacter isolates were also analysed. RESULTS: Most of the infections were nosocomial, and the majority of these were acquired in intensive care units (ICUs). A. baumannii accounted for 41.8 per cent (n=18) of all pneumonia acquired in the ICU. AP-PCR with M13 primer distinguished 8 different PCR patterns comprising of 2 to 6 DNA fragments of 0.1 to 1.0 kb. PFGE identified 9 distinct profiles with five subvarients. By APPCR, epidemiologically unrelated strains could not be differentiated and often differences within biotypes of A. baumannii were not detectable. ApaI macrorestriction (PFGE) identified at least 4 outbreaks caused by 3 clones of A. baumannii and one clone of DNA group 13TU, one replacing the other in a well-defined temporal order. INTERPRETATION & CONCLUSION: Most of A. baumannii isolates were multidrug resistant. PFGE was more discriminatory [Discriminatory Index (DI)=0.96 than AP-PCR fingerprinting (DI=0.88)] in the present study. However, AP-PCR fingerprinting is more useful as a simple and rapid identification technique for epidemiological investigation of nosocomial Acinetobacter infections.

Identification of Acinetobacter species isolated from clinical specimens by amplified ribosomal DNA restriction analysis.

BACKGROUND & OBJECTIVES: Taxonomy of Acinetobacter has been changing ever since it was recognized to be associated with human infections. Many biochemical schemes and molecular methods have been used for the species identification of this bacterium. Recently a simple molecular method called amplified ribosomal DNA restriction analysis (ARDRA) has been used to determine the genomospecies of ACINETOBACTER: An attempt is made in the present study to identify the Acinetobacter genomospecies isolated from clinical specimens using ARDRA and to see whether the environmental isolates are similar to those obtained from clinical specimens. METHODS: A total of 142 consecutive isolates of Acinetobacter sp. obtained from different clinical specimens (125) and environmental samples (17) of postoperative neurosurgery-intensive care unit were studied using ARDRA. Amplification was done using primers of 16S rRNA gene followed by restriction with Alu I, Cfo I and Mbo I enzymes separately to obtain a profile of patterns specific for a species. RESULTS: Of the 125 clinical isolates, 107 were Acinetobacter baumannii (genomospecies 2) and 18 were A. calcoaceticus (genomospecies 1); while 11 of the 17 environmental isolates were A. baumannii and 6 had unidentifiable patterns which were not found in the clinical isolates. INTERPRETATION & CONCLUSION: We found that ARDRA was a simple and reproducible method to be used in a clinical laboratory for identification of Acinetobacter species. A. baumannii was found to be the commonest species isolated from the patients and environment in our hospital. The presence of the same species of Acinetobacter in the environment suggests the role of environment as a source of infection to the patients in high risk units.

Biotyping, serotyping and ribotyping as epidemiological tools in the evaluation of Acinetobacter baumannii dissemination in hospital units, Sorocaba, Sao Paulo, Brazil

Dissemination of Acinetobacter baumannii strains in different units of a hospital in Sorocaba, Sao Paulo, Brazil was evaluated over a period of two years. By using biotyping, serotyping and ribotyping, 27 distinct clones were differentiated among 76 strains isolated between 1993-94, from clinical specimens of hospitalized patients. Two clones, 2:O4:A (biotype:serotype:ribotype) and 2:O29:A accounted for the majority of strains widely disseminated in the units during 1993. The introduction in the hospital setting, of a new clone, 6:O13:B, at the end of 1993 and its predominance through 1994 is discussed. Among 15 strains isolated from neonates, 6 (40 percent) belonged to the same clone, 2:O4:A. Interestingly, this clone was almost all recovered in neonatal intensive care unit, nursery and in pediatric unit. All strains were susceptible to imipenem and polymyxcin B. Multiresistant strains (up to 12 antimicrobial agents) accounted for 66.7 percent and 84.8 percent of the strains isolated in 1993 and in 1994, respectively

Presencia de integrones y su relación con la resistencia a cefalosporinas de tercera generación en cepas de acinetobacter baumannii de origen nosocomial / Presence of integrons and their relationship with the resistance to third generation cephalosporins among nosocomial isolates of acinetobacter baumannii

Background: Acinetobacter baumannii is an important etiological agent causing nosocomial infections. High level of resistance for different kind of antimicrobials has been observed, including ß-lactam antibiotics. This feature, chromosomal or plasmid encoded, has been associated to integrons harbouring antibiotic resistance gene cassettes. Aims: To investigate the presence of integrons among clinical isolates resistant to third generation cephalosporins (3GC). Material and methods: One hundred A. baumannii strains isolated from several Chilean hospitals were included in this study. Minimal inhibitory concentrations (MIC) of 3GC by an agar dilution method were carried out. Integrons class 1, 2 and 3 were investigated by colony blot hybridisation and confirmed by PCR. Results: High level of resistance to all assayed 3GC was observed. On the other hand, integrón class 2 was the most prevalent (77 percent of isolates) followed by integron class 1 (52 percent). Forty six percent of isolates hybridised with probes for both of them. However, no positive hybridisation was detected for integron class 3. Conclusions: Nevertheless, most isolates harboured one or both class of integron; there was no direct relationship between the presence of these genetic structures and the resistance to this kind of ß-lactam antibiotics