Extended-spectrum beta-lactamases among Enterobacteriaceae isolated in a public hospital in Brazil

Extended-spectrum β-lactamases (ESBL) in enterobacteria are recognized worldwide as a great hospital problem. In this study, 127 ESBL-producing Enterobacteriaceae isolated in one year from inpatients and outpatients at a public teaching hospital at São Paulo, Brazil, were submitted to analysis by PCR with specific primers for blaSHV, blaTEM and blaCTX-M genes. From the 127 isolates, 96 (75.6%) Klebsiella pneumoniae, 12 (9.3%) Escherichia coli, 8 (6.2%) Morganella morganii, 3 (2.3%) Proteus mirabilis, 2 (1.6%) Klebsiella oxytoca, 2 (1.6%) Providencia rettgeri, 2 (1.6%) Providencia stuartti, 1 (0.8%) Enterobacter aerogenes and 1 (0.8%) Enterobacter cloacae were identified as ESBL producers. BlaSHV, blaTEM and blaCTX-M were detected in 63%, 17.3% and 33.9% strains, respectively. Pulsed field gel eletrophoresis genotyping of K. pneumoniae revealed four main molecular patterns and 29 unrelated profiles. PCR results showed a high variety of ESBL groups among strains, in nine different species. The results suggest the spread of resistance genes among genetically different strains of ESBL-producing K. pneumoniae in some hospital wards, and also that some strongly related strains were identified in different hospital wards, suggesting clonal spread in the institutional environment.

Sensibilidade das enterobacteriáceas produtoras de beta-lactamases de espectro alargado à fosfomicina

Introdução: A emergência de enterobacteriáceas produtoras de beta-lactamases de espectro alargado (ESBL) nos últimos anos representa um problema de saúde pública, escasseando alternativas eficazes para o seu tratamento. Vários trabalhos internacionais têm demonstrado uma sensibilidade in vitro muito elevada destas bactérias à fosfomicina, havendo alguns que testaram a eficácia clínica do tratamento de cistites agudas não complicadas no subgrupo das Escherichia coli com resultados promissores. No Hospital Prof. Dr. Fernando Fonseca EPE (HFF) tem havido um aumento anual progressivo do isolamento destes patogéneos. Os autores pretenderam testar a sensibilidade das enterobacteriáceas produtoras de ESBL à fosfomicina no HFF e averiguar o eventual potencial terapêutico. Material e métodos: Estudo prospectivo, durante 6 meses, no qual foi testada a sensibilidade à fosfomicina das enterobacteriáceas produtoras de ESBL isoladas. Foi utilizado o equipamento VITEK 2® para identificação das estirpes. A susceptibilidade à fosfomicina foi determinada através do método de difusão de disco (Oxoid®). O tratamento estatístico foi realizado através do programa Microsoft Excel®. Resultados: Foram identificadas 150 enterobacteriáceas ESBL, das quais 52% corresponderam a Klebsiella pneumoniae e 44% a Escherichia coli. Cerca de 88% das Escherichia coli e 68% das Klebsiella pneumoniae apresentaram sensibilidade à fosfomicina. Conclusões: De acordo com os dados obtidos a nível internacional e no nosso hospital, os autores recomendam a utilização da fosfomicina para tratamento de cistites agudas não complicadas provocadas por Escherichia coli produtoras de ESBL, sugerindo, concomitantemente, a realização de trabalhos futuros de eficácia clínica para consubstanciar esta prática e recomendação.

Meningite Bacteriana no Recém-Nascido. Casuística de 12 Anos

Objectivo — Conhecer a epidemiologia da meningite bacteriana em recém-nascidos admitidos na Unidade de Cuidados Intensivos Neonatais do Hospital de Dona Estefânia. Doentes e métodos — Foi feita a revisão dos processos de recém-nascidos admitidos na Unidade de Cuidados Intensivos Neonatais do Hospital de Dona Estefânia de Janeiro de 1985 a Dezembro de 1996 — 12 anos, provenientes da maternidade do Hospital ou do exterior. Foram excluídas as infecções congénitas e as crianças com idade superior a 28 dias. Definiu-se como precoce a infecção com início nas primeiras 72 horas de vida. Resultados — Houve 36 casos de meningite bacteriana correspondendo a 1,1% das admissões. A incidência de meningite bacteriana precoce na Maternidade do Hospital foi 0,13 por mil nados-vivos. Vinte e quatro crianças eram do sexo masculino (66,7%), 7 eram pré-termo, 4 de baixo peso e 1 de muito baixo peso. Dez recém-nascidos tiveram meningite precoce (27,8%) e 26 (72,2%) meningite tardia. Houve isolamento do agente bacteriano no líquido cefalorraquidiano em 27 crianças (77,1%): E. coli (n=7); Streptococcus do grupo B(SGB) (n=6); Klebsiella pneumoniae (n=3); Proteus mirabilis (n=2), Listeria monocytogenes (n=1), Streptococcus bovis (n=1), Staphylococcus aureus (n=1), Neisseria meningitidis (n=2) e Salmonella tiphy (n=1). Houve ainda o isolamento de 3 Gram negativos não identificados. A hemocultura foi positiva em 19 de 32 colheitas (59,4%). Na ausência de terapêutica antibiótica, em 6 casos a cultura do líquor foi positiva e a hemocultura negativa e noutros 2 ambas as culturas foram negativas. Durante o internamento faleceram 9 recém-nascidos — mortalidade de 25% e em 11 foram detectadas sequelas. Conclusão — Houve um predomínio de casos de meningite tardia, em recém-nascidos de termo e do sexo masculino. Os agentes mais frequentemente encontrados foram a E. coli e a Streptococcus do grupo B.

Sepsis in burned patients

A prospective study was conducted from June 2001 to May 2002 at the Burns Unit of Hospital Regional da Asa Norte, Brasília, Brazil. During the period of the study, 252 patients were treated at the Burns Unit, 49 (19.4%) developed clinically and microbiologically proven sepsis. Twenty-six (53.1%) were males and 23 (46.9%) females with a mean age of 22 years (range one to 89 years) and mean burned body surface area of 37.7 ± 18.4% (range 7 to 84%). Forty-three patients had flame burns, five a scald and one an electric burn. These 49 patients had a total of 62 septic episodes. Forty (81.6%) patients had only one and nine (18.4%) had up to three episodes of sepsis. Thirty (61.2%) patients had their first septicemic episode either earlier or by one week postburn. Out of 62 septic episodes, 58 were due to bacteria and four due to Candida sp. The most common bacteria isolated from blood culture were Staphylococcus aureus, coagulase-negative Staphylococcus, Acinetobacter baumannii, Enterobacter cloacae and Klebsiella pneumoniae. Eleven (18.9%) episodes were due to oxacillin resistant Staphylococcus aureus. Acinetobacter baumannii was sensitive to ampicillin/sulbactam in 71.4% and to imipenem in 85.7% of the cases. The primary foci of sepsis were the burn wound in 15 ( 24.2% ) episodes. The most common clinical findings of sepsis in these patients were fever, dyspnea, hypotension and oliguria. The most common laboratory findings of these patients were anemia, leukocytosis, hypoalbuminemia and thrombocytopenia. Twelve (24.5%) patients died. The appropriate knowledge of clinical, epidemiological, laboratorial and microbiological aspects of sepsis in burned patients permits an adequate diagnosis and treatment of this complication.

Meningoencephalitis by Naegleria fowleri: epidemiological study in Anzoategui state, Venezuela

A case of primary amoebic meningoencephalitis produced by Naegleria fowleri was diagnosed in the Independencia county of Anzoategui State, Venezuela. This case motivated the realization of the present epidemiological study with the aim of identifying free-living amoebae in this area. Representative water samples were taken and physicochemical and microbiologic analyses were carried out. Trophozoites and cysts of Naegleria spp, were detected in 44.4% (n=4). An excellent concordance was found among the observations of free-living amoebae in smears and those of monoxenic cultures in non nourishing agar with Klebsiella pneumoniae (Kappa=1; p= 0.003). A variable load of aerobic mesophils was obtained. Moulds and yeast averages presented 3.0 CFU/ml (SD± 2.0) and 102.9 CFU/ml (SD± 32.2), respectively. One hundred per cent of the samples presented a most probable number of total and fecal coliforms of 240,000 NMP/100mL. Naegleria spp was present in waters of the Independence county of Anzoategui state, which constitutes a risk for people that use these sources.

Prevalência e suscetibilidades bacterianas das infecções comunitárias do trato urinário, em Hospital Universitário de Brasília, no período de 2001 a 2005

A infecção do trato urinário é uma das afecções mais comuns da clínica médica, sendo mandatório o conhecimento epidemiológico da mesma e do perfil de sensibilidade dos agentes etiológicos. O estudo teve como objetivo identificar os agentes etiológicos mais freqüentes e o perfil de sensibilidade aos antimicrobianos das bactérias isoladas de uroculturas de pacientes ambulatoriais atendidos no Hospital Universitário de Brasília no período de 2001 a 2005. Foram analisadas 2.433 uroculturas positivas realizadas no laboratório de microbiologia do Hospital Universitário de Brasília. A Escherichia coli foi a bactéria mais isolada (62,4%), seguida de Klebsiella pneumoniae (6,8%) e Proteus mirabillis (4,7%). A Escherichia coli apresentou maior sensibilidade à amicacina (98,6%), gentamicina (96,2%), nitrofurantoína (96,3%), e às quinolonas ciprofloxacina (90,9%) e norfloxacina (89,8%), com baixa sensibilidade ao sulfametoxazol-trimetoprima (50,6%). As outras bactérias apresentaram similar padrão de sensibilidade. Em conclusão, a Escherichia coli foi a bactéria mais isolada, sendo altamente sensível aos amiglicosídeos, nitrofurantoína e quinolonas.

Nosocomial infection and characterization of extended-spectrum β-lactamases-producing Enterobacteriaceae in Northeast Brazil

INTRODUCTION: Extended spectrum β-lactamases (ESBLs) are enzymes that degrade β-lactam antibiotics and have been reported to be an important cause of nosocomial infection in worldwide. METHODS: During 2009, 659 enterobacteria strains were isolated from different clinical specimens and tested for ESBL production. The disk approximation test, combined disk method and addition of clavulanic acid were used for phenotypic detection of the ESBL-producing strains and PCR for detection of the blaTEM and blaCTX-M genes. RESULTS: Among the isolates, 125 were ESBL producers. The blaCTX-M and blaTEM genes were detected in 90.4% and 75% of the strains, respectively. Most strains were isolated from urine. Klebsiella pneumoniae was the most prevalent organism. Microorganisms presented high resistance to the antibiotics. CONCLUSIONS: These results support the need for extending ESBL detection methods to different pathogens of the Enterobacteriaceae family because these methods are only currently standardized by the CLSI for Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca and Proteus mirabilis. Carbapenems were the antibiotic class of choice for the treatment of infections caused by ESBL-producing Enterobacteriaceae.

Detection of bla KPC-2 in Proteus mirabilis in Brazil

INTRODUCTION : Infections caused by Klebsiella pneumoniae carbapenemase (KPC)-producing isolates pose a major worldwide public health problem today. METHODS : A carbapenem-resistant Proteus mirabilis clinical isolate was investigated for plasmid profiles and the occurrence of β-lactamase genes. RESULTS : The isolate exhibited resistance to ertapenem and imipenem and was susceptible to meropenem, polymyxin, and tigecycline. Five plasmids were identified in this isolate. DNA sequencing analysis revealed the presence of bla KPC-2 and bla TEM-1 genes. An additional PCR using plasmid DNA confirmed that bla KPC-2 was present in one of these plasmids. Conclusions: We report the detection of bla KPC-2 in P. mirabilis in Brazil for the first time. This finding highlights the continuous transfer of bla KPC between bacterial genera, which presents a serious challenge to the prevention of infection by multidrug-resistant bacteria.

Distribution of extended-spectrum β-lactamase types in a Brazilian tertiary hospital

INTRODUCTION: Epidemiological data on the prevalence of extended-spectrum β-lactamases (ESBLs) are scarce in Brazil despite the fact that these data are essential for empirical treatment and control measures. The objective of this study was to evaluate the prevalence of different ESBLs by type and distribution in a tertiary hospital in southern Brazil. METHODS: We evaluated 1,827 enterobacterial isolates between August 2003 and March 2008 isolated from patients at a tertiary hospital. Samples were identified using a Vitek automated system and were confirmed by biochemical testing. The identified ESBL strains were characterized by phenotypic methods, polymerase chain reaction (PCR), and sequencing. Genetic similarities were evaluated by pulsed-field gel electrophoresis. RESULTS: It was 390 (21.3%) ESBL-producing strains, which expressed the ESBLs CTX-M (292), SHV (84), CTX and SHV (10), TEM (2), and PER (2). CONCLUSIONS: The prevalence of ESBL-expressing strains was high, especially in Klebsiella pneumoniae and Enterobacter spp. CTX-M was the predominant type of ESBL observed, and its genetic variability indicates a polyclonal distribution.

Study of the risk factors related to acquisition of urinary tract infections in patients submitted to renal transplant

INTRODUCTION: Urinary tract infections (UTI) among transplant recipients are usually caused by gram-negative microorganisms and can provoke a high incidence of morbidity and mortality. The aim of this study was to evaluate the risk factors associated with the acquisition of UTIs during the first year after renal transplantation. METHODS: Here, we report a single-center retrospective cohort study of 99 renal transplant patients followed for the first year after surgery. The definition of a UTI episode was a urine culture showing bacterial growth and leucocyturia when patients presented with urinary symptoms. The absence of infection (asymptomatic bacteriuria) was defined as an absence of symptoms with negative urine culture or bacterial growth with any number of colonies. RESULTS: Ninety-nine patients were included in the study. During the study, 1,847 urine cultures were collected, and 320 (17.3%) tested positive for bacterial growth. Twenty-six (26.2%) patients developed a UTI. The most frequent microorganisms isolated from patients with UTIs were Klebsiella pneumoniae (36%), with 33% of the strains resistant to carbapenems, followed by Escherichia coli (20%). There were no deaths or graft losses associated with UTI episodes. CONCLUSIONS: Among the UTI risk factors studied, the only one that was associated with a higher incidence of infection was female sex. Moreover, the identification of drug-resistant strains is worrisome, as these infections have become widespread globally and represent a challenge in the control and management of infections, especially in solid organ transplantation.

Successful treatment of lower urinary tract infections with oral fosfomycin: a report of three cases

Infections due to multidrug-resistant organisms continue to increase, and therapeutic options remain scarce. Given this challenge, it has become necessary to use older antimicrobials for treatment of these pathogens. We report three patients with lower urinary tract infections caused by Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae who were successfully treated with a seven-day course of oral fosfomycin monotherapy.

Pharmacokinetic/pharmacodynamic target attainment of intravenous β-lactam regimens against Gram-negative bacteria isolated in a Brazilian teaching hospital

ABSTRACTINTRODUCTION: Monte Carlo simulations have been used for selecting optimal antibiotic regimens for treatment of bacterial infections. The aim of this study was to assess the pharmacokinetic and pharmacodynamic target attainment of intravenous β-lactam regimens commonly used to treat bloodstream infections (BSIs) caused by Gram-negative rod-shaped organisms in a Brazilian teaching hospital.METHODS: In total, 5,000 patients were included in the Monte Carlo simulations of distinct antimicrobial regimens to estimate the likelihood of achieving free drug concentrations above the minimum inhibitory concentration (MIC; fT > MIC) for the requisite periods to clear distinct target organisms. Microbiological data were obtained from blood culture isolates harvested in our hospital from 2008 to 2010.RESULTS: In total, 614 bacterial isolates, including Escherichia coli, Enterobacterspp., Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa, were analyzed Piperacillin/tazobactam failed to achieve a cumulative fraction of response (CFR) > 90% for any of the isolates. While standard dosing (short infusion) of β-lactams achieved target attainment for BSIs caused by E. coliand Enterobacterspp., pharmacodynamic target attainment against K. pneumoniaeisolates was only achieved with ceftazidime and meropenem (prolonged infusion). Lastly, only prolonged infusion of high-dose meropenem approached an ideal CFR against P. aeruginosa; however, no antimicrobial regimen achieved an ideal CFR against A. baumannii.CONCLUSIONS:These data reinforce the use of prolonged infusions of high-dose β-lactam antimicrobials as a reasonable strategy for the treatment of BSIs caused by multidrug resistant Gram-negative bacteria in Brazil.

Antimicrobial activity of bergenin from Endopleura uchi (Huber) Cuatrec

Endopleura uchi (Huber) Cuatrec. is an Amazon species traditionally used as treatment for inflammations and female disorders. Bergenin was isolated from ethyl acetate fraction of bark of E. uchi by using column chromatography over sephadex LH-20 and then silica gel 60 flash. Its structure was identified on the basis of its NMR spectra. The antimicrobial activity of bergenin and fractions of methanol extract of E. uchi were evaluated against ATCC microorganisms (Escherichia coli, Salmonella enteritidis, Pseudomonas aeruginosa, Enterococcus faecalis, Staphylococcus aureus, Candida albicans, C. guilliermondii, Aspergillus flavus, A. nidulans). Clinically isolated strains of all of these microorganisms, along with C. tropicalis, A. niger, Shigella sonnei, Serratia marcenses and Klebsiella pneumoniae were also evaluated. The growth inhibition caused by bergenin, extracts and fractions of E. uchi against ATCC microorganisms were similar to the inhibition to microorganisms clinically isolated. The ethyl acetate fraction and the isolate bergenin inhibit the growth of the yeasts C. albicans, C. tropicalis, and C. guilliermondii, but present lower activity against filamentous fungi Aspergillus flavus, A. nidulans, A. niger, and did not inhibit the Gram positive and Gram negative bacteria. The activity of the ethyl acetate fraction and bergenin are in agreement wit its high concentration found in bark extract of E. uchi. Moreover, the selective activity against three Candida species helps to understand its traditional use against infections that affect women.

PEGylated ofloxacin nanoparticles render strong antibacterial activity against many clinically important human pathogens.

The rise of bacterial resistance against important drugs threatens their clinical utility. Fluoroquinones, one of the most important classes of contemporary antibiotics has also reported to suffer bacterial resistance. Since the general mechanism of bacterial resistance against fluoroquinone antibiotics (e.g. ofloxacin) consists of target mutations resulting in reduced membrane permeability and increased efflux by the bacteria, strategies that could increase bacterial uptake and reduce efflux of the drug would provide effective treatment. In the present study, we have compared the efficiencies of ofloxacin delivered in the form of free drug (OFX) and as nanoparticles on bacterial uptake and antibacterial activity. Although both poly(lactic-co-glycolic acid) (OFX-PLGA) and methoxy poly(ethylene glycol)-b-poly(lactic-co-glycolic acid) (OFX-mPEG-PLGA) nanoformulations presented improved bacterial uptake and antibacterial activity against all the tested human bacterial pathogens, namely, Escherichia coli, Proteus vulgaris, Salmonella typhimurium, Pseudomonas aeruginosa, Klebsiella pneumoniae and Staphylococcus aureus, OFX-mPEG-PLGA showed significantly higher bacterial uptake and antibacterial activity compared to OFX-PLGA. We have also found that mPEG-PLGA nanoencapsulation could significantly inhibit Bacillus subtilis resistance development against OFX.

Influence of oxygen conditions on bacterial interactions within biofilms related with cystic fibrosis

Dissertação de mestrado em Bioengineering