- Volume 60, Issue 10, 2011

We performed genotyping of Mycobacterium leprae present in skin biopsy samples that were collected during the first and the second disease occurrences from eight leprosy patients, seven of whom were diagnosed as suffering from disease relapse. Sequence analysis of part of the M. leprae rpoB, folP1, gyrB and gyrA genes did not show genetic change that supported the presence of drug-resistant bacilli. However, we observed a synonymous nucleotide change at position 297 of gyrA among five of these patients, one presenting C to T (CgyrAT) and four presenting T to C (TgyrAC) at this position. Additional genotyping by analysis of the four short tandem repeats GAA, GTA9, AT17 and TA18 showed that the gyrA single nucleotide polymorphism change was accompanied by a change in short tandem repeat genotype. Our data suggest that leprosy relapse in these patients, living in an area endemic for leprosy, could be caused by M. leprae with a genotype different from the one that caused initial disease.

Rapid diagnosis of multidrug-resistant tuberculosis (MDR-TB) is essential for the prompt initiation of effective second-line therapy to improve treatment outcome and limit transmission of this obstinate disease. A variety of molecular methods that enable the rapid detection of mutations implicated in MDR-TB have been developed. The sensitivity of the methods is dependent, in principle, on the repertoire of mutations being detected, which is typically limited to mutations in the genes rpoB, katG and the promoter region of inhA. In this study, a new reverse hybridization assay, REBA MTB-MDR (M&D), that probes mutations in the oxyR–ahpC intergenic region, in addition to those in rpoB, katG and the inhA promoter region, was evaluated. A set of 240 Mycobacterium tuberculosis clinical isolates from patients receiving retreatment regimens was subjected to conventional phenotypic drug-susceptibility testing (DST) and the REBA MTB-MDR assay. The nucleotide sequences of the loci known to be involved in drug resistance were determined for comparison. In brief, the results showed that the REBA MTB-MDR assay efficiently recognized nucleotide changes in the oxyR–ahpC intergenic region as well as those in rpoB, katG and the inhA promoter region with higher sensitivity, resulting in an 81.0 % detection rate for isoniazid resistance. Inclusion of the oxyR–ahpC intergenic region in the REBA MTB-MDR assay improved the overall sensitivity of molecular DST for MDR-TB from 73.1 to 79.9 %.

Pneumococcal nasopharyngeal carriage isolates recovered from Brazilian children attending day-care centres in 2005 were assessed for serotype, genotype and penicillin susceptibility phenotype. As 124 of the 253 isolates (49 %) were characterized previously with respect to serotype and penicillin susceptibility, the primary objectives were to examine clonal associations and penicillin susceptibility within major serotypes and to assess the suitability of conventional multiplex PCR for deducing carriage serotypes within this population. Using a combination of PCR-based serotyping and the Quellung reaction, serotypes were identified for 81 % (205/253) of the isolates, with serogroups or types 14, 6, 23F, 19F and 18 being predominant. Included within the 205 isolates successfully serotyped by PCR were 28 isolates that had become non-viable. Forty-eight isolates were non-typable using both the PCR method and the Quellung reaction. Penicillin non-susceptibility was observed within 16 of the 18 multilocus sequence types detected. Thus, this study provides further evidence from a diverse collection of pneumococcal clones that PCR-based serotype deduction is useful for providing supportive evidence for pneumococcal conjugate vaccine implementation.

To study the prevalence pattern and trends in the phenotypic and genetic characteristics of shigellae, we tested 212 isolates isolated from diarrhoeal patients admitted to the Infectious Diseases Hospital, Kolkata, India, from November 2007 to October 2010. Prevalence of Shigella spp. was higher in the >5 years age group (69 %) than in children in the <5 years age group (31 %). Serotypes 2a, 3a and untypable isolates of Shigella flexneri were frequently detected. An increase in the isolation of Shigella sonnei (15 %) is a novel trend in this region. Fluoroquinolone resistance among S. flexneri serotypes 2a, 3a and other serogroups of shigellae is another evolving trend. The set gene was exclusively present in S. flexneri 2a, and the sen gene was detected in all serogroups. PFGE revealed the grouping of S. flexneri isolates according to their serotypes with approximately 80–100 % similarity, whilst Shigella dysenteriae type 2 and S. sonnei were clonal in nature. There was no demarcation in the prevalence of serotypes, antimicrobial resistance or clonality between the two age groups.

Invasive aspergillosis is associated with high morbidity and mortality rates; nevertheless, blood cultures almost invariably yield a negative result. The recovery and detection time of Aspergillus fumigatus, Aspergillus flavus and Aspergillus terreus were studied in BACTEC Plus Aerobic/F, Mycosis-IC/F and Myco/F Lytic vials, incubated in the BACTEC 9240 and 9000 MB automated systems. Two different approaches were used for subculture in solid medium: (i) the routine method, using a sterile airway needle/subculture unit, and (ii) a novel procedure, using instead a tuberculin disposable syringe and collecting a larger aliquot (100 µl), following vigorous agitation of the vials. A. fumigatus was detected at inoculum concentrations of >3 conidia per 10 ml after 21–40 h, in both BACTEC Plus Aerobic/F and BACTEC Mycosis-IC/F vials. A few more hours were needed to detect A. flavus and A. terreus. The novel subculture procedure of BACTEC culture vials on solid medium resulted in several positive results that were not detected by the routine sampling procedure. BACTEC Plus Aerobic/F vials show an advantage particularly in patients under antifungal treatment. In cases of polymicrobial bloodstream infections (concurrent bacterial growth), the inoculation of blood samples into a BACTEC Mycosis-IC/F vial achieved the best results. Further multicentre studies are needed to validate this improved automated detection of Aspergillus spp. from blood cultures in clinical laboratories, as this diagnostic procedure allows antifungal susceptibility testing of moulds.

The aim of this work was to study the antibacterial effect of coriander (Coriandrum sativum) essential oil against Gram-positive and Gram-negative bacteria. Antibacterial susceptibility was evaluated using classical microbiological techniques concomitantly with the use of flow cytometry for the evaluation of cellular physiology. Our results showed that coriander oil has an effective antimicrobial activity against all bacteria tested. Also, coriander oil exhibited bactericidal activity against almost all bacteria tested, with the exception of Bacillus cereus and Enterococcus faecalis. Propidium iodide incorporation and concomitant loss of all other cellular functions such as efflux activity, respiratory activity and membrane potential seem to suggest that the primary mechanism of action of coriander oil is membrane damage, which leads to cell death. The results obtained herein further encourage the use of coriander oil in antibacterial formulations due to the fact that coriander oil effectively kills pathogenic bacteria related to foodborne diseases and hospital infections.

The genetic characteristics of mumps virus (MuV) strains isolated from sporadic mumps cases between 2007 and 2009 in three provinces of south-west China were investigated. MuV detection was carried out by nested RT-PCR on 117 cases. The small hydrophobic gene of 33 isolated strains was identified, and sequence analysis revealed that all the isolates belonged to the lineage genotype F with slight nucleotide variation. Furthermore, the virus haemagglutinin–neuraminidase and nucleoprotein antigens exhibited a high degree of nucleotide and amino acid similarity (>99 %) within all isolates, whilst the fusion protein appeared to have certain geographical differences. This study on molecular surveillance will help to monitor the circulation of MuV in China.

Infection of the central nervous system (CNS) is a rare but devastating complication of invasive aspergillosis. We report a case of invasive aspergillosis with spinal involvement in a human immunodeficiency virus (HIV)-infected patient without neutropenia. A 42-year-old, antiretroviral-naïve, HIV-infected man presented with progressive weakness in the lower limbs and urinary and faecal incontinence for 2 weeks. The patient had been prescribed broad-spectrum antibiotics and prednisone. He had upper motor neuron signs and a sensory level at T1, with accompanying neck stiffness on flexion. Magnetic resonance imaging revealed diffuse abnormal signals of the vertebral bodies in the lower cervical and thoracic areas, with cord compression in the C2 and C3 region and signal distortions of the T2 and T3 vertebral bodies. Chest X-ray and computerized tomography demonstrated post-tuberculous apical cavities with suspected fungal colonization. Histopathology of an extradural spinal lesion at T1/T2 suggested invasive aspergillosis. The patient was started on fluconazole in response to the histopathological evidence of Aspergillus infection, but died within 3 weeks. Post-mortem analysis of the biopsy sample by PCR identified the infectious agent as Aspergillus fumigatus. Atypically, his CD4+ T-cell count was 239 cells mm−3 and he had no evidence of neutropenia. Invasive aspergillosis should be considered as part of the differential diagnosis among HIV-infected patients with non-specific, focal CNS symptoms, even among those without classical risk factors such as neutropenia, and aggressive antifungal therapy should be instituted as early as possible.

We describe a case of bacteraemia caused by Weissella confusa in a 48-year-old male who was operated on for adenocarcinoma of the gastro-oesophageal junction and maintained on total parenteral nutrition. Blood cultures were positive for a vancomycin-resistant streptococcus-like organism which was identified as W. confusa by 16S rRNA gene sequencing.

Invasive candidiasis is rare in children after the neonatal period, but can occur in children with (secondary) immunodeficiency with a damaged gastrointestinal or skin barrier, or when receiving antibiotics. A 10-month-old girl was diagnosed as suffering from cystic fibrosis (CF) when showing failure to thrive, pulmonary symptoms and hypoproteinaemia. At that time, Candida albicans was identified from blood culture and treated intravenously with liposomal amphotericin B for 13 days. Six weeks later, the girl presented with osteoarticular infection of the left knee caused by C. albicans. The infection showed insufficient response to therapy with liposomal amphotericin B, but the patient recovered after therapy with fluconazole and flucytosine. Follow-up over 4 years revealed no sequelae. In conclusion, invasive Candida infections may occur in patients with CF, and preventive measures might be considered in patients at risk. In the case of an invasive infection, prolonged treatment with a combination of antifungal drugs may be required.

Despite high vaccination coverage rates, there has been a gradual increase in reported pertussis cases. Although whooping cough affects all ages, young infants continue to suffer the greatest pertussis disease burden. Adolescents and adults are the primary source of infection for young babies. In this paper, we report two cases involving the likely transmission of pertussis from mothers to infants in Tunisia.