Concentrated specimen smear microscopy utilising a polymer membrane sandwich filtration vessel for the detection of acid-fast bacilli in health facilities in Sabah, East Malaysia.

A simple, ready-to-use concentrated specimen smear microscopy method employing a nanometer silicon polyvinylidene fluoride (PVDF) polymer membrane sandwich filtration vessel to concentrate acid-fast bacilli (AFB) in samples (SFV-CSSM, Hunan-Tech New Medical System Co. Ltd. China) was compared with direct sputum smear microscopy (DSSM) to determine its performance using culture on modified Ogawa agar as reference. The results for 4114 clinical samples collected from health facilities in Sabah were interpreted with reference to culture results, sample collection-transportation conditions and clinical data including responses to anti-TB drug treatment. The SFV-CSSM showed higher sensitivity than DSSM (79.4% versus 60.5%) and less background interference. Its ability to detect low levels of AFB at an affordable cost makes it an excellent tool for the screening of pauci-bacillary samples as well as for active case finding in TB control programs.

Comparative evaluation of two commercial real-time PCR kits (QuantiFast™ and abTES™) for the detection of Plasmodium knowlesi and other Plasmodium species in Sabah, Malaysia.

BACKGROUND: The monkey parasite Plasmodium knowlesi is an emerging public health issue in Southeast Asia. In Sabah, Malaysia, P. knowlesi is now the dominant cause of human malaria. Molecular detection methods for P. knowlesi are essential for accurate diagnosis and in monitoring progress towards malaria elimination of other Plasmodium species. However, recent commercially available PCR malaria kits have unpublished P. knowlesi gene targets or have not been evaluated against clinical samples. METHODS: Two real-time PCR methods currently used in Sabah for confirmatory malaria diagnosis and surveillance reporting were evaluated: the QuantiFast™ Multiplex PCR kit (Qiagen, Germany) targeting the P. knowlesi 18S SSU rRNA; and the abTES™ Malaria 5 qPCR II kit (AITbiotech, Singapore), with an undisclosed P. knowlesi gene target. Diagnostic accuracy was evaluated using 52 P. knowlesi, 25 Plasmodium vivax, 21 Plasmodium falciparum, and 10 Plasmodium malariae clinical isolates, and 26 malaria negative controls, and compared against a validated reference nested PCR assay. The limit of detection (LOD) for each PCR method and Plasmodium species was also evaluated. RESULTS: The sensitivity of the QuantiFast™ and abTES™ assays for detecting P. knowlesi was comparable at 98.1% (95% CI 89.7-100) and 100% (95% CI 93.2-100), respectively. Specificity of the QuantiFast™ and abTES™ for P. knowlesi was high at 98.8% (95% CI 93.4-100) for both assays. The QuantiFast™ assay demonstrated falsely-positive mixed Plasmodium species at low parasitaemias in both the primary and LOD analysis. Diagnostic accuracy of both PCR kits for detecting P. vivax, P. falciparum, and P. malariae was comparable to P. knowlesi. The abTES™ assay demonstrated a lower LOD for P. knowlesi of ≤ 0.125 parasites/µL compared to QuantiFast™ with a LOD of 20 parasites/µL. Hospital microscopy demonstrated a sensitivity of 78.8% (95% CI 65.3-88.9) and specificity of 80.4% (95% CI 67.6-89.8) compared to reference PCR for detecting P. knowlesi. CONCLUSION: The QuantiFast™ and abTES™ commercial PCR kits performed well for the accurate detection of P. knowlesi infections. Although the QuantiFast™ kit is cheaper, the abTES™ kit demonstrated a lower LOD, supporting its use as a second-line referral-laboratory diagnostic tool in Sabah, Malaysia.

Parasitic infections in Malaysian aborigines with pulmonary tuberculosis: a comparative cross-sectional study.

The geographical distribution of tuberculosis (TB) overlaps with various parasitic infections. Uncovering the characteristics of coinfecting parasites that potentially affect the host susceptibility to TB is pertinent as it may provide input to current TB therapeutic and prophylactic measures. The present study was aimed at examining the types of parasitic infections in TB patients and healthy TB contacts (HC) in Orang Asli, Malaysian aborigines, who dwelled in the co-endemic areas. Stool and serum samples were collected from Orang Asli who fulfilled the selection criteria and provided written informed consents. Selected parasitic infections in the two study groups were determined by stool examination and commercial serum antibody immunoassays. The prevalence of parasitic infections in TB and HC participants were 100% (n = 82) and 94.6% (n = 55) respectively. The parasitic infections comprised toxocariasis, trichuriasis, amoebiasis, toxoplasmosis, hookworm infection, ascariasis, strongyloidiasis, and brugian filariasis, in decreasing order of prevalence. Overall, helminth or protozoa infection did not show any significant association with the study groups. However, when the species of the parasite was considered, individuals exposed to trichuriasis and toxoplasmosis showed significant odds reduction (odds ratio (OR) 0.338; 95% confidence interval (CI) 0.166, 0.688) and odds increment (OR 2.193; 95% CI 1.051, 4.576) to have active pulmonary TB, respectively. In conclusion, trichuriasis and toxoplasmosis may have distinct negative and positive associations respectively with the increase of host susceptibility to TB.

Resistencia a los medicamentos del Mycobacterium Leprae detectados en cultivo de almohadilla plantar de ratón entre 1997 y 2013 en Malasia / No disponible

Antecedentes: Después de tres décadas de implementación de la multiterapia (MDT), consistente en una combinación de rifampicina, dapsona y clofazimina, en Malasia la aparición de resistencia farmacológica del Mycobacterium leprae constituye una preocupación, ya que puede llevar al fracaso del tratamiento y la recidiva de la enfermedad. Objetivos: Determinar el modelo de resistencia farmacológica del M. leprae en Malasia. Métodos: Se analizaron los cultivos en almohadilla plantar de ratón (MFP) de todas las biopsias cutáneas de pacientes con lepra borderline lepromatosa y lepra lepromatosa enviados a la Unidad de la Lepra, Laboratorio Nacional de Salud Publica, Sungai Buloh, Malasia, entre 1997-2013. Resultados: Se realizaron 651 cultivos MFP. La edad media de los pacientes fue de 41 anos (rango: 6-88). La proporción varón/hembra era de 3·8:1. Cuatrocientos cuarenta y cuatro pacientes (69·1%) eran malayos. La proporción de M. leprae positivo en cultivo era del 66·6% (433 of 651). El Índice Bacteriologico (IB) y el Índice Morfológico (IM) promedios para los cultivos positivos fue de 3·7 and 2·8 respectivamente. El IB y el IM de los que no crecieron en la MFP eran significativamente menores que los que presentaban cultivos positivos (P < 0·001). La dapsona presento el mayor índice de resistencia del 55% (238 of 433). Sin embargo, el elevado grado de resistencia a la dapsona (0·01%) fue de 6·24%. Hubo 407 MFP con rifampicina 0·003% y 12 (2·9%) resultaron resistentes a la misma. La clofazimina presento el menor grado de resistencia intermedia (0·001%) que fue del 0·2% (1 of 429). No había diferencias significativas entre el patrón de resistencia y género o nacionalidad de los pacientes. Conclusiones: Mas de la mitad de los cultivos MFP presentaron resistencia de baja intensidad a la dapsona; menos del 3% eran resistentes a la rifampicina y la resistencia a la clofazimina resulto muy baja

Background: After three decades of implementing multidrug therapy (MDT) consisting of rifampicin, dapsone and clofazimine in Malaysia, the drug resistance pattern of Mycobacterium leprae is a growing concern as it may lead to failure of treatment and relapse of disease. Objective: To determine the drug resistance patterns of M. leprae in Malaysia. Methods: Mouse footpad (MFP) culture of all skin biopsy samples from patients with borderline lepromatous and lepromatous leprosy sent to the Leprosy Unit, National Public Health Laboratory, Sungai Buloh, Malaysia between 1997-2013 were retrospectively studied. Results: There were 651 MFP cultures performed. The mean age of patients was 41 years old (range: 6-88). The male: female ratio was 3·8:1. Four hundred and forty four patients (69·1%) were Malaysian. The rate of positive M. leprae culture was 66·6% (433 of 651). The mean Bacteriological Index (BI) and median Morphological Index (MI) for those with positive culture were 3·7 and 2·8 respectively. The mean BI and MI of those which failed to grow in the MFP were significantly lower than those with positive cultures (P < 0·001). Dapsone has the highest resistance rate of 55% (238 of 433). Nevertheless, high degree dapsone resistance (0·01%) was 6·24%. There were 407 MFP tests using rifampicin 0·003% and 12 (2·9%) were resistant to it. Clofazimine has the lowest intermediate degree (0·001%) resistance rate of 0·2% (1 of 429). There were no significant differences between the drug resistance pattern and the gender or the nationality of the patients. Conclusion: More than half of our positive MFP cultures showed low-level resistance to dapsone; less than 3% were resistant to rifampicin, and clofazimine resistance remained very low

High prevalence of pfcrt K76t mutants among Plasmodium falciparum isolates from Sabah, Malaysia.

Chloroquine (CQ) remains the first line drug for the prevention and treatment of malaria in Malaysia in spite of the fact that resistance to CQ has been observed in Malaysia since the 1960s. CQ-resistance is associated with various mutations in pfcrt, which encodes a putative transporter located in the digestive vacuolar membrane of P. falciparum. Substitution of lysine (K) to threonine (T) at amino acid 76 (K76T) in pfcrt is the primary genetic marker conferring resistance to CQ. To determine the presence of T76 mutation in pfcrt from selected areas of Kalabakan, Malaysia 619 blood samples were screened for P. falciparum, out of which 31 were positive. Blood samples were collected on 3 MM Whatman filter papers and DNA was extracted using QIAmp DNA mini kit. RFLP-PCR for the detection of the CQ-resistant T76 and sensitive K76 genotype was carried out. Twenty-five samples were shown to have the point mutation in pfcrt whereas the remaining samples were classified as CQ-sensitive (wild-type). In view of the fact that CQ is the first line anti-malarial drug in Malaysia, this finding could be an important indication that treatment with CQ may no longer be effective in the future.

Epidemiology of tuberculosis and leprosy, Sabah, Malaysia.

The objectives in this epidemiology review are to measure and report the extent of morbidity and mortality due to tuberculosis (TB), the proportion of new sputum smear positive cases in districts and the status of cohort analysis as of 1999. As for leprosy, the main objective is to determine morbidity and the treatment outcomes of Multiple Drug Therapy (MDT). Based on the results obtained, a comprehensive action plan for prevention, control and monitoring of tuberculosis and leprosy cases and patients is being produced and implemented throughout the state. The analysis concentrated on patients diagnosed at all out-patient units and admitted in all of the state's hospitals. The patient particulars were recorded using a standardized format based on TB and Leprosy Health Management Information System (TB HMIS). TB was the second highest by notification of communicable diseases in Malaysia in 2001. 29% or about one-third of the national TB cases are from Sabah. However, it has been noted that there was an average decline of 2.6% in annual notification since 10 years ago to date. There was also a reduction of 11.4% in 2001 as compared to annual notification in 2000. Immigrants contribute more than 24% in detection of new cases since 1990. Treatment success rate in term of completion of treatment to date is 82%. Mortality rate has steadily declined from 14 deaths to 7 deaths per 100,000 population. Leprosy in Sabah also contributes to 30% of the yearly total caseload of Malaysia and has the highest notification rate of 2 per every 100,000 population as compared to other states. The average registered leprosy cases over the past 5 years are 239 cases and the prevalence rate is 0.7/10,000 population. The state has successfully achieved its goal to decrease leprosy as per the World Health Organization (WHO) goal of yearly overall prevalence rate of less than 1 case for every 10,000 population. However, the districts of Kudat, Tawau, Lahad Datu, Kota Kinabalu and Semporna are still within the prevalence rate of more than one per 10,000 population. This review highlights some interesting findings which can be incorporated into the State and Districts action plans and strategies. It is also noted that in order to translate National Plans and Strategies into effective action at the community level, health workers need relevant up-to-date knowledge of the pattern of health and disease, and of their determinants, in each district. The Sabah Health Department continues to organize and support programs related to management and control of tuberculosis and leprosy to progressively reduce the incidence of these diseases in the community by breaking the chain of transmission of Mycobacterium tuberculosis and M. leprae, respectively.

Risk factors of tuberculosis among health care workers in Sabah, Malaysia.

Tuberculosis (TB) is one of the main public health problems in Sabah; 30% of the total number of TB cases reported in Malaysia every year occur in Sabah. The average incidence of TB among health care workers over the past 5 years is 280.4 per 100,000 population (1, Annual Report of Sabah State TB Control Programme, 1998). At present, there are no specific measures for the prevention of TB transmission in health care facilities. A case-control study was conducted among health care workers in Sabah in 2000-2001. Cases were health care workers with TB diagnosed between January 1990 and June 2000. Controls were health care workers without TB and working in the same facility as cases during the disease episode. The study attempted to identify risk factors for TB among the study population. Data were collected through structured interviews and review of patients' records. The notification rate of TB among health care workers was significantly higher than that to the general population (Z=4.893, p<0.01). The average notification rate of TB among health care workers over the last 5 years was two times higher than in the general population (280.4/100,000 compared to 153.9/100,000). Regression results showed that ethnicity, designation, family contact and TB related knowledge did not significantly contribute to the risk of contracting TB in this study. However, after controlling for the above factors, age, gender, history of TB contact outside the workplace (other than family contact), duration of service and failure to use respiratory protection when performing high-risk procedures, were the main risk factors of TB among health care workers. This study succeeded in identifying some of the risk factors of TB among health care workers. We managed to include the large ratio of controls to case (3:1) and those cases spanned over a period of 10 years. However, the findings from the study have to be applied with caution due to the limitations of this study, which include recall bias, dropouts, and small sample size. Based on the study findings, we recommend that health care workers in the first 10 years of service should take extra precautions, such as using respiratory protection when performing procedures that are considered to be of high risk with respect to TB infection. They should also undergo TB screening at least once every 2 years and, if symptomatic, offered prophylactic treatment. The Respiratory Protection Programme should be fully implemented to help reduce the risk of TB among health care workers in Sabah.