[Show abstract][Hide abstract] ABSTRACT: Variable number of tandem repeat (VNTR) typing with a panel of 24 loci is the current gold standard in the molecular typing of Mycobacterium tuberculosis complex. However, due to technical problems a part of the loci can often not be amplified in the multiplex PCRs. Therefore, a considerable number of single-locus PCR reactions have to be repeated for the loci with missing results, which impairs the laboratory workflow. Therefore, the original in-house method described by Supply et al. in 2006 was re-evaluated. We modified 7 primers and the PCR mastermix, resulting in a strongly optimized in-house 24-locus VNTR typing method. The percentage of instantly complete 24-locus VNTR patterns, detected in the routine flow of typing activities increased to 84.7% compared to 72.3% for the typing conducted with the commercially available MIRU-VNTR typing kit by GenoScreen. The analytical sensitivity of the optimized in-house method was assessed by serial dilutions of M. tuberculosis in broncho-alveolar lavage fluid (BALF). The 1:10 dilution of the different strains tested was the lowest dilution for the detection of a complete 24-locus VNTR pattern.The optimized in-house 24-locus VNTR typing method will reduce the turn-around-time of typing significantly and also the financial burden of these activities.
Journal of clinical microbiology 02/2014; · 4.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A 43-year-old female developed an Epstein-Barr virus (EBV)-positive post-transplant lymphoproliferative disorder (PTLD) in the central nervous system (CNS), 14 years after renal transplantation. One year prior to presentation, the patients' treatment regimen was altered from cyclosporine, azathioprine, and prednisone to mycophenolate mofetil and prednisone. Magnetic resonance imaging of the brain revealed lesions suspect for malignant lymphoma. The EBV real-time polymerase chain reaction (PCR) on peripheral blood was negative, but highly positive on cerebrospinal fluid. EBV-positive PTLD was confirmed using histological analysis of cerebral biopsies. Despite tapering of immune suppressive medication and treatment with rituximab and chemotherapy, the patient deceased 50 days after presentation. This case illustrates that vigilance is required when presented with a negative EBV PCR result on peripheral blood when PTLD of the CNS is suspected. This late presentation suggests a relation to the switch in immunosuppressive regimen 1 year earlier.
Transplant International 08/2012; 25(11):e113-6. · 3.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A young female health professional was diagnosed with pulmonary tuberculosis caused by Mycobacterium bovis. Source finding and contact tracing was initiated by the regional municipal health service using both tuberculin skin test and QuantiFERON(®)-TB Gold (QFT-GIT (IGRA). The strain appeared near-identical to that of an elderly Dutch patient.
Medical Microbiology and Immunology 03/2012; 201(3):397-400. · 3.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Indonesia has a concentrated but rapidly growing HIV epidemic. We examined the effect of HIV on causative organisms, clinical features and prognosis of adult meningitis.
A prospective cohort study.
All adult patients at a referral hospital who underwent cerebrospinal fluid examination for suspected meningitis were examined for HIV and included in a prospective cohort study. Microbiological testing was done for common bacterial pathogens, mycobacteria and fungi. Patients were followed for at least 6 months, and logistic regression models were used to identify risk factors for mortality.
Among 185 patients who mostly presented with subacute meningitis, 60% were male and the median age was 30 years. HIV infection was present in 25% of the patients; almost two-thirds were newly confirmed, and all presented with severe immunosuppression (median CD4 cell count 13/microl, range 2-98). One-third of HIV-infected patients had cryptococcal meningitis whereas two-thirds suffered from tuberculosis. After 1 month, 41% of patients had died. HIV infection was strongly associated with 1-month mortality (adjusted odds ratio 12.15; 95% confidence interval 3.04-15.72) and death during extended follow-up (hazard ratio 2.48; 95% confidence interval 1.97-5.74).
Although HIV is still uncommon in the general population in Indonesia, its prevalence among adult meningitis cases already seems high. Mycobacterium tuberculosis and Cryptococcus neoformans are the main causes of meningitis in this setting, and mortality is very high, especially in HIV-infected patients. Our data suggest that adult meningitis cases in Indonesia should be screened routinely for HIV infection. Further studies are needed to address the high mortality.
AIDS (London, England) 09/2009; 23(17):2309-16. · 4.91 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Comparison of Mycobacterium tuberculosis genotype distributions in different areas might help to find determinants of the emergence of certain genotypes, such as the Beijing family. In this study, M. tuberculosis isolates originating from patients from two Indonesian islands were genotyped, and possible associations with patients' characteristics and drug resistance were explored. A high degree of genetic diversity was observed among the M. tuberculosis strains, and a significant difference was found in the geographical distribution of genotype families. The predominant Beijing genotype family was isolated from 268 of 813 patients from West Java (33.0%) versus 12 of 84 patients from Timor (14.3%) (P = 0.002). Family F (East African-Indian) (33.3%) and family D (Latin American and Mediterranean) (20.0%) were more prevalent in Timor. No significant associations were found between genotype families and age, vaccination with Mycobacterium bovis BCG, previous treatment, disease localization, or drug resistance. Possible explanations for the differences in the geographical distribution of the M. tuberculosis genotypes are discussed.
Journal of clinical microbiology 11/2008; 46(11):3639-45. · 4.16 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Tuberculosis (TB) in Africa is increasing because of the human immunodeficiency virus (HIV) epidemic, and in HIV/AIDS patients it presents atypically. Pulmonary tuberculosis (PTB) in Africa is mainly diagnosed clinically, by chest radiograph or by sputum smear for acid fast bacilli (AFB).
We evaluated in 120 HIV-infected patients with chest infection the diagnostic accuracy of AFB smear of sputum and bronchoalveolar lavage (BAL) fluid, sputum Mycobacterium tuberculosis (MTB) culture, real-time PCR and MycoDot serological test, using MTB culture of BAL fluid as gold standard. We correlated PCR cycle threshold values (C(T)) to the culture results. Retrospectively, we evaluated the development of active TB in patients with positive PCR but negative culture.
Culture of BAL fluid identified 28 patients with PTB. Fifty-six patients could not produce adequate sputum. Sputum AFB smear and the serological test had sensitivities of 66.7% and 0%, respectively. PCR with C(T) 40 was positive in 73 patients, 27 of whom were also TB culture positive (96.4% sensitivity and 52.3% specificity of PCR). PCR with C(T) 32 had sensitivity of 85.7% and specificity of 90.9% to diagnose PTB in BAL. No patients with positive PCR but negative culture developed active TB during 18 months follow-up.
In these HIV-infected patients, AFB smear and serology had very low sensitivities. PCR of BAL with C(T) value 32 had improved specificity to diagnose active PTB. A prospective follow-up study is warranted in TB/HIV endemic settings, applying real time PCR to both sputum and BAL.
Tropical Medicine & International Health 11/2007; 12(10):1210-7. · 2.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The etiology of Crohn's disease in humans is largely unknown. Clinical signs of Crohn's disease partly resemble the clinical picture of Johne's disease in ruminants caused by Mycobacterium avium subsp. paratuberculosis. Because of the high prevalence of these bacteria in (products of) ruminants and their remarkable thermostability, concern has been raised about the possible role of these bacteria in the pathogenesis of Crohn's disease. In an attempt to develop a molecular typing method to facilitate meaningful comparative DNA fingerprinting of M. avium subsp. paratuberculosis isolates from the human and animal reservoirs, multilocus variable-number tandem-repeat analysis (MLVA) was explored and compared to IS900 restriction fragment length polymorphism (RFLP) typing. MLVA typing subdivided the most predominant RFLP type, R01, into six subtypes and thus provides a promising molecular subtyping approach to study the diversity of M. avium subsp. paratuberculosis.
Journal of Clinical Microbiology 12/2004; 42(11):5022-8. · 4.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mycobacterium heckeshornense is a rare isolate in clinical specimens. We performed simultaneous 16S rRNA sequence analysis of a mycobacterium culture and a histopathology specimen to determine the relevance of M. heckeshornense infection in an immunocompetent patient initially presenting with pneumothorax.
Journal of Clinical Microbiology 10/2004; 42(9):4386-9. · 4.07 Impact Factor