Escherichia coli producing SHV-type extended-spectrum beta-lactamase is a significant cause of community-acquired infection.

OBJECTIVES: Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (ESBLEC) is an increasingly significant cause of community-acquired infection worldwide. The epidemiological features of CTX-M- and SHV-producing ESBLEC causing community-acquired infections are compared. METHODS: A multicentre cohort study including all community-acquired infections caused by ESBLEC in four geographical areas of Spain was carried out. ESBL characterization was by isoelectric focusing, PCR and sequencing. Demographics, previous healthcare contact, co-morbidity, use of antimicrobials, invasive procedures and type of infection were collected for all patients. Patients with CTX-M- and SHV-producing isolates were compared using logistic regression. RESULTS: One hundred and twenty-two cases (95% urinary tract infections) were included. ESBLs were characterized in 112 isolates; 77 isolates (69%) produced CTX-M, 36 (32%) produced SHV and 7 (6%) produced TEM enzymes (8 produced >1 ESBL). Patients with isolates producing CTX-M enzymes only (CTX-M group, n = 70) and SHV enzymes only (SHV group, n = 31) were compared. There were no differences in terms of underlying disease, previous healthcare contact, invasive procedures, antibiotic use or type of infection. Multivariate analysis including geographical area showed that a Charlson Index score of >2 (OR = 4.0; 95% CI = 1.2-12.6) was associated with SHV isolates, while age >60 (4.7; 1.7-12.5) was associated with CTX-M isolates. CONCLUSIONS: SHV-producing ESBLEC is a significant cause of community-acquired infection in Spain; the clinical epidemiology of such isolates seems very similar to that of CTX-M-producing E. coli.

Community infections caused by extended-spectrum beta-lactamase-producing Escherichia coli.

BACKGROUND: Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli is an increasingly important group of community pathogens worldwide. These organisms are frequently resistant to many of the antimicrobial agents usually recommended for the treatment of infections caused by E coli, such as penicillins, cephalosporins, fluoroquinolones, and trimethoprim-sulfamethoxazole. Data concerning risk factors, clinical features, and therapeutic options for such infections are scarce. METHODS: A case-control study was performed to investigate the risk factors for all types of community-acquired infections caused by ESBL-producing E coli in 11 Spanish hospitals from February 2002 to May 2003. Controls were randomly chosen from among outpatients with a clinical sample not yielding ESBL-producing E coli. The clinical features of these infections were investigated in the case patients. The efficacy of fosfomycin tromethamine and amoxicillin-clavulanate potassium was observationally studied in patients with cystitis. RESULTS: A total of 122 cases were included. Risk factors selected by multivariate analysis included the following: age older than 60 years; female sex; diabetes mellitus; recurrent urinary tract infections (UTIs); previous invasive procedures of the urinary tract; follow-up in outpatient clinic; and previous receipt of aminopenicillins, cephalosporins, and fluoroquinolones. Urinary tract infections accounted for 93% of the cases; 6% of the patients were bacteremic and 10% needed hospitalization. The cure rate of patients with cystitis was 93% with fosfomycin therapy (all isolates were susceptible); among patients treated with amoxicillin-clavulanate, cure rates were 93% for those with susceptible isolates (minimum inhibitory concentration < or =8 microg/mL) and 56% for those with intermediate or resistant isolates (minimum inhibitory concentration > or =16 microg/mL) (P = .02). CONCLUSIONS: In predisposed patients, ESBL-producing E coli is a notable cause of community-acquired infection, and particularly UTI. Fosfomycin and amoxicillin-clavulanate appear to be effective for cystitis caused by susceptible isolates.

Direct detection of rifampin- and isoniazid-resistant Mycobacterium tuberculosis in auramine-rhodamine-positive sputum specimens by real-time PCR.

Our objective was to evaluate the feasibility of a molecular assay based on a real-time PCR technique, carried out with a LightCycler instrument (Roche Biochemicals), to identify Mycobacterium tuberculosis bacilli and to detect rifampin and isoniazid resistance in DNA extracts from sputum samples. We studied three genes: rpoB, which is associated with rifampin resistance, and katG and inhA, which are associated with isoniazid resistance. A total of 205 sputum samples collected from 108 patients diagnosed with pulmonary tuberculosis with positive auramine-rhodamine-staining (AR) sputum samples, were tested. The sensitivities of the LightCycler PCR assay for the positive AR specimens was 97.5% (200 of 205) for rpoB and inhA genes and 96.5% (198 of 205) for the katG gene. For the total number of patients tested, the sensitivity was 100% (108 of 108 patients) for rifampin, whereas the sensitivity was 98.1% (106 of 108 patients) for isoniazid. Full agreement was found with the Bactec MGIT 960 method and the genotype inferred from the LightCycler data for rifampin. The phenotypic method for isoniazid reported 13 resistant strains (> or = 0.1 microg/ml). In seven (53.8%) strains there was a concordance between both methods, but we found that six (46.2%) strains reported as resistant by the phenotypic method were determined to be susceptible by real-time PCR. For the 75 strains reported as susceptible by the phenotypic method, the concordance with the LightCycler data was 100%. Our results demonstrate that rifampin-resistant M. tuberculosis could be detected in DNA extracted from auramine-rhodamine-positive sputum samples in a single-tube assay that took less than 3 h to perform for a collection of auramine-rhodamine-positive specimens obtained from patients with culture-documented pulmonary tuberculosis. Similarly, this occurs in half of the isoniazid-resistant M. tuberculosis DNA extracted from auramine-rhodamine-positive specimens.

Improved real-time PCR for rapid detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis clinical isolates.

In this study we designed two pairs of probes for the detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis with real-time PCR procedures. One pair of probes spans the region between codon 510 and 528 of the rpoB gene, and the other one screens for mutation at the regulatory region of the inhA gene. We have evaluated these probes in combination with two other pairs of probes previously described to detect mutations in 20 susceptible and 53 unique resistant M. tuberculosis clinical isolates. We were able to detect nine different mutations affecting five codons of the rpoB gene, two different mutations at codon 315 of the katG gene and a nucleotide substitution (C209T) in the regulatory region of the inhA gene within two hours turnaround.