Relentless increase of resistance to fluoroquinolones and expanded-spectrum cephalosporins in Escherichia coli: 20 years of surveillance in resource-limited settings from Latin America.

Previous studies on commensal Escherichia coli from healthy children in the Bolivian Chaco have shown remarkable resistance rates to the old antibiotics since the early 1990s, and the emergence of resistance to newer drugs (fluoroquinolones and expanded-spectrum cephalosporins) in the 2000s. Here we report the results of a new survey conducted in 2011 in the same setting. Rectal swabs were obtained from 482 healthy children (aged 6-72 months) from three urban areas of the Bolivian Chaco. Screening for antibiotic-resistant E. coli was performed by a direct plating method, as in the previous studies. The blaCTX-M genes were investigated by PCR/sequencing, and CTX-M-producing isolates were subjected to genotyping and detection of several plasmid-mediated quinolone resistance mechanisms. Results showed high rates of resistance to nalidixic acid (76%), ciprofloxacin (44%) and expanded-spectrum cephalosporins (12.4%), demonstrating a relentless increase of resistance to those drugs over the past two decades. CTX-M-type extended-spectrum beta-lactamases were found to be widespread (12%, 97% of extended-spectrum beta-lactamase producers). Compared with the previous studies, CTX-M-producing E. coli underwent a dramatic dissemination (120-fold increase since early 2000s) and a radical change of dominant CTX-M groups (CTX-M-1 and CTX-M-9 groups versus CTX-M-2 group). Most CTX-M producers were not susceptible to quinolones (91%), and 55% carried plasmid-mediated quinolone resistance genes (different combinations of aac(6')-Ib-cr, qnrB and qepA). This study shows the rapid and remarkable increasing trend for resistance to fluoroquinolones and expanded-spectrum cephalosporins in one of the poorest regions of Latin America, and underscores the need for urgent control strategies aimed at preserving the efficacy of those drugs in similar settings.

Chagas disease in Italy: breaking an epidemiological silence.

Chagas disease, a neglected tropical disease that due to population movements is no longer limited to Latin America, threatens a wide spectrum of people(travellers, migrants, blood or organ recipients,newborns, adoptees) also in non-endemic countries where it is generally underdiagnosed. In Italy, the available epidemiological data about Chagas disease have been very limited up to now, although the country is second in Europe only to Spain in the number of residents from Latin American. Among 867 at-risk subjectsscreened between 1998 and 2010, the Centre for Tropical Diseases in Negrar (Verona) and the Infectious and Tropical Diseases Unit, University of Florence found 4.2% patients with positive serology for Chagas disease (83.4% of them migrants, 13.8% adoptees).No cases of Chagas disease were identified in blood donors or HIV-positive patients of Latin American origin. Among 214 Latin American pregnant women,three were infected (resulting in abortion in one case).In 2005 a case of acute Chagas disease was recorded in an Italian traveller. Based on our observations, we believe that a wider assessment of the epidemiological situation is urgently required in our country and public health measures preventing transmission and improving access to diagnosis and treatment should be implemented.

QuantiFERON-TB Gold and the TST are both useful for latent tuberculosis infection screening in autoimmune diseases.

Screening for active tuberculosis (TB) and latent TB infection (LTBI) is mandatory prior to the initiation of tumour necrosis factor-alpha inhibitor therapy. However, no agreement exists on the best strategy for detecting LTBI in this population. The aim of the present study was to analyse the performance of the tuberculin skin test (TST) and QuantiFERON-TB Gold in-tube (QFT-GIT) on LTBI detection in subjects with immunomediated inflammatory diseases (IMID). The TST and QFT-GIT were prospectively performed in 398 consecutive IMID subjects, 310 (78%) on immunosuppressive therapy and only 16 (4%) had been bacillus Calmette-Guérin (BCG) vaccinated. Indeterminate results to QFT-GIT were found in five (1.2%) subjects. Overall, 74 (19%) out of 393 subjects were TST-positive and 52 (13%) were QFT-GIT-positive. Concordance between TST and QFT-GIT results was good (87.7%): 13 were QFT-GIT-positive/TST-negative and 35 QFT-GIT-negative/TST-positive. By multivariate analysis both tests were significantly associated with older age. Only the TST was associated with BCG vaccination and radiological lesions of past TB. Use of immunosuppressive drugs differently modulated QFT-GIT or TST scoring. Use of the QuantiFERON-TB Gold in-tube, as a screening tool for latent tuberculosis among immunomediated inflammatory disease subjects, is feasible. Until further data will elucidate discordant tuberculin skin test/QuantiFERON-TB Gold in-tube results, a strategy of simultaneous tuberculin skin and QuantiFERON-TB Gold in-tube testing in a low prevalence bacillus Calmette-Guérin vaccinated population, should maximise potentials of latent tuberculosis diagnosis.

Isolation of a novel sequevar of Mycobacterium flavescens from the synovial fluid of an AIDS patient.

This report describes the characterisation of a mycobacterium involved in a case of septic arthritis in an AIDS patient that was treated successfully with specific anti-mycobacterial drugs. The biochemical and cultural features, and the mycolic acid pattern as assessed by high-performance liquid chromatography, were fully compatible with the isolate being Mycobacterium flavescens. However, the isolate's 16S rDNA sequence differed by five nucleotides from the two known sequevars of M. flavescens, thus indicating that this isolate belonged to a new 16S rDNA sequevar.

In-vitro activity of nisin against clinical isolates of Clostridium difficile.

Nisin is a cationic peptide produced by Lactococcus lactis. Its activity against clinical isolates of Clostridium difficile was compared to that of vancomycin and metronidazole by minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and time-kill studies. Nisin was more active than the other agents, with a MIC90 of 0.256 mg/L and strong bactericidal activity. Nisin may be a promising agent for the management of C. difficile associated diarrhea.

Mycobacterium elephantis: not an exceptional finding in clinical specimens.

Following the recent report of new 16S rDNA sequences of Mycobacterium elephantis, three clinical strains suspected to belong to such species were investigated using biochemical and cultural tests, high performance liquid chromatography of cell wall mycolic acids and genetic sequencing. Antimicrobial susceptibility was also determined. The findings confirmed recent data concerning human isolates of this new mycobacterium and identified a new 16S rDNA sequevar for this species.

Burden of unidentifiable mycobacteria in a reference laboratory.

Modern identification techniques at the genomic level have greatly improved the taxonomic knowledge of mycobacteria. In adjunct to nucleic acid sequences, mycobacterial identification has been endorsed by investigation of the lipidic patterns of unique mycolic acids in such organisms. In the present investigation, the routine use of high-performance liquid chromatography (HPLC) of mycolic acids, followed by the sequencing of the 16S rRNA, allowed us to select 72 mycobacterial strains, out of 1,035 screened, that do not belong to any of the officially recognized mycobacterial species. Most strains (i.e., 47) were isolated from humans, 13 were from the environment, 3 were from animals, and 9 were from unknown sources. The majority of human isolates were grown from the respiratory tract and were therefore most likely not clinically significant. Some, however, were isolated from sterile sites (blood, pleural biopsy, central venous catheter, or pus). Many isolates, including several clusters of two or more strains, mostly slow growers and scotochromogenic, presented unique genetic and lipidic features. We hope the data reported here, including the results of major conventional identification tests, the HPLC profiles of strains isolated several times, and the whole sequences of the 16S rRNA hypervariable regions of all 72 mycobacteria, may encourage reporting of new cases. The taxonomy of the genus Mycobacterium is, in our opinion, still far from being fully elucidated, and the reporting of unusual strains provides the best background for the recognition of new species. Our report also shows the usefulness of the integration of novel technology to routine diagnosis, especially in cases involving slow-growing microorganisms such as mycobacteria.

Prevalence of antibodies against hepatitis A and E viruses among rural populations of the Chaco region, south-eastern Bolivia.

We conducted a cross-sectional study to determine the seroprevalence of antibodies against hepatitis A and hepatitis E viruses (HAV and HEV) in the population of two rural areas, Camiri and Villa Montes, of the Chaco region, south-eastern Bolivia. HAV antibodies were detected in 461 (94.1%) of 490 serum samples tested, not differing significantly between sexes and study areas. The HAV seropositivity rate (64.7%) was high even in the youngest age group (1-5 years). The prevalence of HEV was 7.3%, with no significant differences between sexes. The prevalence of HEV antibodies in the population of the Camiri area (10.4%) was significantly higher than in the Villa Montes area (4.4%), possibly due to the better quality of drinking water in the Villa Montes area. In the population /= 31 year-old group. This is consistent with findings in other countries. This is the first report of the prevalence of HEV infection in Bolivia.

Patterns of antimicrobial use and antimicrobial resistance among healthy children in Bolivia.

OBJECTIVE: To determine the incidence of antimicrobial-resistant, nonpathogenic Escherichia coli among healthy children aged 6-72 months in Camiri town and a rural village, Javillo, in south-eastern Bolivia. METHOD: A community-based survey: stool samples were obtained from 296 healthy children selected by modified cluster sampling in Camiri and all 25 eligible children in Javillo. E. coli isolates were tested for antimicrobial susceptibility according to the standard disc diffusion method. By a questionnaire survey of 12 pharmacies and by using simulated patients, we investigated the antimicrobial availability and the usage patterns in Camiri town. RESULTS: In Camiri, over 90%, and in Javillo over 70% of children carried E. coli resistant to ampicillin, trimethoprim-sulphamethoxazole (TMP/SMX) or tetracycline. Overall, 63% of children carried E. coli with multiple resistance to ampicillin, TMP/SMX, tetracycline and chloramphenicol. In the simulated patients study, antimicrobials were dispensed inappropriately for 92% of adults and 40% of children with watery diarrhoea, and were under-prescribed for males with urethral discharge (67%) or females with fever and dysuria (58%). The dose and/or duration of antimicrobials dispensed was almost always too low. CONCLUSION: Our study showed a disturbingly high prevalence of carriage of nonpathogenic E. coli resistant to antimicrobials. The prevalence of resistance to ampicillin and TMP/SMX was higher than that previously reported in developing countries. The existence of a large reservoir of resistance genes in healthy individuals in developing countries represents a threat to the success of antimicrobial therapy throughout the world. Programmes to improve rational and effective drug use in developing countries are urgently needed.

Cervical lymphadenitis due to an unusual mycobacterium.

A scotochromogenic acid-fast bacillus was isolated from a lymph node of a 2-year-old female. On the basis of conventional testing, the mycobacterium appeared to be Mycobacterium scrofulaceum. Its mycolic acid profile, however, was not identical to that of Mycobacterium scrofulaceum but was similar to that of Mycobacterium interjectum. Direct sequencing of the 16S rRNA gene revealed a unique nucleic acid sequence, suggesting that the isolate represents a previously undescribed pathogenic species.

Mycobacterium malmoense in Italy: the modern Norman invasion?

The isolation of Mycobacterium malmoense has for a long time been restricted to few countries of Northern Europe; reports from countries other than Sweden, Great Britain and Finland are rare and the first Italian case report has been published in 1995. Since 1988, however, fifteen strains of M. malmoense have been isolated in Italy, eleven of which in the last two years; of these, ten appeared clinically significant on the basis of medical records. The susceptibility of the strains and the role of high performance liquid chromatography of cell wall mycolic acids for a reliable identification are discussed.

Characterization of mycobacterial isolates phylogenetically related to, but different from Mycobacterium simiae.

The use of high-performance liquid chromatography (HPLC) revealed four previously unreported profiles within a group of mycobacteria consisting of 14 clinical isolates. These mycobacteria, whose identification by conventional tests appeared problematic, mostly resembled Mycobacterium avium complex or Mycobacterium simiae. Genetic analysis revealed, within this group, six different nucleic acid sequences in a hypervariable 16S rRNA segment, but all the isolates appeared to be phylogenetically related to M. simiae. Six isolates representing the largest of groups defined by means of genetic sequencing turned out to belong to the newly described species Mycobacterium lentiflavum. Furthermore, three such clusters precisely coincided with three of those defined by HPLC, while the three remaining clusters shared almost identical HPLC profiles. All but one strain (which, although clearly not belonging to the M. avium complex, hybridized with specific commercial DNA probes) showed high-grade resistance to the majority of antimycobacterial drugs. Three of the isolates were clinically significant according to stringent criteria. Sophisticated techniques, like genetic sequencing or HPLC, by now seem indispensable for differentiating unusual and new mycobacteria from well-established ones.

Isolation of an unusual mycobacterium from an AIDS patient.

A mycobacterium isolated from a clinical sample of an AIDS patient was identified as Mycobacterium interjectum by direct 16S rRNA sequence determination. High-performance liquid chromatography, however, revealed a mycolic acid pattern which was different from the one shared by the previously analyzed strains of this species.

Characterization of an isolate of the newly described species Mycobacterium interjectum.

The phenotypic features of a clinical isolate of the new species Mycobacterium interjectum, identified on the basis of its 16S rRNA gene sequence, are compared with those of the type strain. The differentiation of M. interjectum from Mycobacterium gordonae or Mycobacterium scrofulaceum is not achievable on the basis of phenotypic traits usually tested for mycobacterial speciation, but it can be reached by 16S rRNA gene sequencing or by high performance liquid chromatography (HPLC) of cell wall mycolic acids. The former reveals sequence identity with the signature region of the type species, and the latter yields a profile which is easily differentiated from those of the other two species. The unique HPLC profile of M. interjectum is reported here for the first time and so are the MICs of a wide spectrum of drugs.

Identification of the newly described Mycobacterium poriferae from tuberculous lesions of snakehead fish (Channa striatus).

A mycobacterium isolated form a cultured snakehead with nodular lesions was identified on the basis of high performance liquid chromatography (HPLC) profile of cell wall mycolic acids, and confirmed by conventional tests, as Mycobacterium poriferae, a species previously isolated only from a marine sponge. The profiles of M. poriferae, Mycobacterium aurum and Mycobacterium parafortuitum are here reported for the first time.

In vitro activity of MDL 62,879 against gram-positive bacteria and Bacteroides species.

The new thiazolyl peptide antibiotic MDL 62,879 (GE2270 A) showed excellent in vitro activity in testing against staphylococci and streptococci, with MIC90s ranging from 0.23 to 0.9 mg/l. It was very active against Clostridium difficile and Propionibacterium acnes (MIC90 0.06 mg/l in each case) and had variable activity against Bacteroides spp. MDL 62,879 had exceptionally good activity against Enterococcus faecalis, including against a collection of high-level aminoglycoside-resistant isolates where it had an MIC90 of 0.047. The antibiotic was bacteriostatic for enterococcal isolates but bactericidal for a methicillin-resistant isolate of Staphylococcus aureus.

Utility of high-performance liquid chromatography for identification of mycobacterial species rarely encountered in clinical laboratories.

High-performance liquid chromatography (HPLC) has been demonstrated to be a suitable technique for determining the species of mycobacteria on the basis of their mycolic acid pattern. Representative HPLC profiles, which are needed for the visual recognition of chromatograms, have been published for the most frequently encountered mycobacterial species. No extensive study has been reported for less common species, and only a few, scattered chromatographic patterns are available in literature. This study evaluates the utility of this technique for the identification of several rare species. Mycobacterium celatum, Mycobacterium genavense and Mycobacterium simiae chromatographic profiles have been verified, and previously unreported profiles of other species investigated. The chromatographic pattern of Mycobacterium malmoense is presented for the first time.

Isolation of the newly described species Mycobacterium celatum from AIDS patients.

Mycobacterium celatum is a recently described species which, on the basis of conventional tests, may be misidentified as Mycobacterium xenopi or as belonging to the Mycobacterium avium complex. Only genomic sequencing or high-performance liquid chromatography of cell wall mycolic acids can presently allow a correct identification of this mycobacterium. Two cases of infection due to M. celatum, in AIDS patients, are described here. The quantitative susceptibility pattern of the isolates to a wide spectrum of drugs is also reported.