In Malaysia, Shigella spp. was reported to be the third commonest bacterial agent responsible for childhood diarrhoea. Currently, isolation of the bacterium and confirmation of the disease by microbiological and biochemical methods remain as the "gold standard". This study aimed to detect the prevalence of four Shigella virulence genes present concurrently, in randomly selected Malaysian strains via a rapid multiplex PCR (mPCR) assay.
A report is presented of seven patients with acute septicaemic melioidosis seen at the University Hospital, Kuala Lumpur, Malaysia, during 1976-1979. All had associated disorders which rendered them more susceptible to infection. As prognosis depends on early diagnosis it is important that this disease be considered in the differential diagnosis of a septicaemic illness in such patients from endemic areas. The treatment of choice is a combination of tetracyclines and chloramphenicol, initially used in massive doses, and continued for at least six month to prevent relapses.
With the increasing recognition of the importance of H. pylori in gastrointestinal disease, there is a need for a reliable, efficient and yet inexpensive diagnostic test. The performance of the rapid urease test (RUT) as an endoscopy suite diagnostic test was compared to the established methods of culture, histology and Gram stain of tissue smear, in 274 gastric biopsy samples. Histology had the highest sensitivity of 99.3% followed by the RUT (96.6%). Culture and Gram stain of tissue smear had 100% specificity, while the rapid urease test had 99.2% specificity. The RUT had a positive predictive value of 99.3% and a negative predictive value of 96.2%. The RUT is an inexpensive, rapid and reliable diagnostic test of H. pylori infection.
Central nervous system melioidosis is an unusual infection in humans. This article reports a case of melioidosis presenting as an acute spinal epidural abscess. A discussion of this case and its management together with a brief review of melioidosis of the central nervous system is presented.
To evaluate the potential role of extracellular proteins in the pathogenicity and virulence of Burkholderia pseudomallei, the activities of several enzymes in the culture filtrates of nine clinical and six environmental isolates were investigated in vitro and in vivo in ICR strain of mice. The production of protease, phosphatase, phospholipase C, superoxide dismutase, catalase and peroxidase were detected in the culture filtrates of all the 15 isolates at different time points of growth 4-24h. Over time, activity of each enzyme at each time point varied. Profile of secretion was similar among the 15 isolates irrespective of source, that is clinical or environmental. Catalase, phosphatase and phospholipase C were found to be increased in 60-100% of the isolates post-passage in mice. In vivo inoculation studies in ICR mice demonstrated a wide difference in their ability to cause bacteraemia, splenic or external abscesses and mortality rate ranged from few days to several weeks.
We examined human tissues infected by Burkholderia (Pseudomonas) pseudomallei which is endemic in Malaysia to study the types of inflammation invoked, and to look for histopathological clues to its diagnosis. The lesions which varied from acute to chronic granulomatous inflammation were not tissue-specific. In five autopsy cases, the inflammation was usually a focal or diffuse, acute necrotising inflammation with varying numbers of neutrophils, macrophages, lymphocytes and 'giant cells'. The 'giant cells' probably represent giant macrophages with phagocytosed leukocytes. There were numerous gram-negative, non-acid-fast, intra- and extracellular bacilli, occurring either singly or in chains. Intracellular bacteria within macrophages and 'giant cells' were so numerous as to resemble globi. This feature has not been previously reported and may be a useful diagnostic clue in melioidosis. In 14 surgical cases biopsies showed acute inflammatory lesions that appeared no different from acute inflammation due to other causes. In many biopsies, however, the inflammation was either an acute-on-chronic inflammation with a focal granulomatous component, or was purely granulomatous in character. Bacilli were difficult to demonstrate in surgical biopsies even with the gram strain.
Pseudomonas aeruginosa is the third most common pathogen causing nosocomial infections. The objective of this study was to investigate the antimicrobial resistance profiles and genetic diversity of hospital isolates of P. aeruginosa and to investigate the presence of several resistance genes and integrons.
Subtyping of Salmonella Paratyphi A isolates from India, Pakistan, Indonesia and Malaysia was carried out by pulsed-field gel electrophoresis (PFGE) to assess the extent of genetic diversity of these isolates from different endemic countries.
A representative sample of 20 isolates of Salmonella weltevreden strains from stool cultures of patients admitted at the University Hospital, Kuala Lumpur, Malaysia were analyzed. All the strains were susceptible to ampicillin, ceftriaxone, ciprofloxacin, chloramphenicol, tetracycline, trimethoprim, gentamicin and co-trimoxazole. Ribosomal RNA gene restriction pattern analysis of PstI-digested DNA gave three ribotypes while pulsed-field gel electrophoresis (PFGE) analysis of XbaI-digested DNA gave ten distinct profiles. PFGE was more discriminative than ribotyping in distinguishing the strains. The majority of the strains analyzed were very closely related with similarity coefficient values ranging from 0.8 to 1.0. Both PFGE and ribotyping could distinguish one of the strains which was obtained from a patient following a bone marrow transplant for beta-thalassemia major, indicating that this particular strain was unrelated to the rest of the strains from patients with acute gastroenteritis.
The study was undertaken to determine clonal relationship and genetic diversity of the human strains of Salmonella enterica serovar Enteritidis isolated from 1995 to 2002 from different parts of Malaysia.
DNA fingerprinting of Salmonella enterica serotype Paratyphi B isolated in Malaysia during 1982-83, 1992 and 1996-2002 was carried out by pulsed-field gel electrophoresis (PFGE), antimicrobial susceptibility tests and D-tartrate utilization tests to assess the extent of genetic diversity of these isolates in Malaysia.
Salmonella enterica serovar Paratyphi A is a causative agent of paratyphoid fever. The clinical syndrome caused by paratyphoid fever overlaps with other febrile illnesses and cannot be distinguished from typhoid fever. Conventional methods used for diagnosis are time consuming, costly, and labor-intensive. We evaluated the specificity, sensitivity, and application of a multiplex polymerase chain reaction (PCR) previously developed by the method (Ou, H.Y., Teh, C.S.J., Thong, K.L., et al., J. Mol. Diagn., 9, 624-630, 2007) using 6 S. Paratyphi A, 22 S. Typhi, and 85 other Salmonella serovars as well as 36 non-Salmonella strains. The detection limit of the multiplex PCR was 4 x 10(4) cfu ml(-1). In a blind test of the other 50 strains, this multiplex PCR correctly identified the only S. Paratyphi A in the panel of strains. The sensitivity of this PCR using spiked blood and stool samples was 1 x 10(5) cfu ml(-1) and 2 x 10(5) cfu ml(-1), respectively, but increased to 1 x 10(4) cfu ml(-1) and 2 x 10(3) cfu ml(-1) after 5-h enrichment. We believe that this multiplex PCR is a promising technique for the specific and sensitive detection of S. Paratyphi A in clinical, environmental, and food samples.
The suitability of a PCR procedure using a pair of primers targeting the hilA gene was evaluated as a means of detecting Salmonella species. A total of 33 Salmonella strains from 27 serovars and 15 non-Salmonella strains from eight different genera were included. PCR with all the Salmonella strains produced a 784 bp DNA fragment that was absent from all the non-Salmonella strains tested. The detection limit of the PCR was 100 pg with genomic DNA and 3 x 10(4) c.f.u. ml(-1) with serial dilutions of bacterial culture. An enrichment-PCR method was further developed to test the sensitivity of the hilA primers for the detection of Salmonella in faecal samples spiked with different concentrations of Salmonella choleraesuis subsp. choleraesuis serovar Typhimurium. The method described allowed the detection of Salmonella Typhimurium in faecal samples at a concentration of 3 x 10(2) c.f.u. ml(-1). In conclusion, the hilA primers are specific for Salmonella species and the PCR method presented may be suitable for the detection of Salmonella in faeces.
An immunofluorescent assay (IFAT) using whole cell antigen derived from Burkholderia thailandensis used for detection of total antibodies to Burkholderia pseudomallei, was found to compare favorably with a previous published report on a B. pseudomallei IFAT assay. At a 1:20 cut-off titer, the assay had high sensitivity (98.9%) and satisfactory specificity (92.3%), when tested against sera from 94 patients suspected of melioidosis. Sera from 12 patients with culture proven melioidosis gave absolute concordance with the 2 test antigens. No sera from 50 blood donors had a titer of > or =20. Cross-reactivity with patients' sera positive for Chlamydia, Mycoplasma, Legionella and typhoid was not observed, except for 3 sera from typhus patients and one from a patient with leptospirosis. The major advantage of this assay is that the cultivation and preparation of B. thailandensis as antigen can be carried out in any laboratory with basic microbiological set-up. The serodiagnosis of melioidosis can be made safe for medical laboratory personnel, particularly in B. pseudomallei endemic regions.
In order to assess the usefulness of immunohistochemistry in the diagnosis of melioidosis, an infection by Burkholderia pseudomallei, polyclonal antibodies were applied to tissues from known cases of melioidosis and to other infected tissues. Formalin-fixed, paraffin-embedded tissues were stained by a modified immunoperoxidase technique. In autopsy tissues with inflammatory lesions of melioidosis, the cytoplasm of phagocytes and intact bacilli, both intra- and extracellular, were stained very strongly positive. Relatively more focal positive staining was observed in some but not all surgical biopsies from proven cases of melioidosis. In granulomas staining was mainly found in the central necrotic areas, with little staining of peripheral phagocytes. All control materials stained negative. Immunohistochemistry appears to be a useful diagnostic tool in melioidosis.
A 13-year old boy with acute lymphoblastic leukemia on chemotherapy developed neutropenia and acute cellulitis progressing to fulminating septicemia due to Pseudomonas pseudomallei. Septicemic melioidosis should be considered in the differential diagnosis of a febrile illness in children who are susceptible to infections.
The predictors of severe disease or death were determined for 85 melioidosis patients in Kuala Lumpur, Malaysia. Most of the patients were male, > 40 years old, and diabetic. Severe disease or death occurred in 28 (32.9%) cases. Lower lymphocyte counts and positive blood cultures were significant independent predictors of severe disease, but age, presentations with pneumonia, inappropriate empirical antibiotics, or flagellin types of the infecting isolates were not. Knowledge of local predictors of severe disease is useful for clinical management.
Routine use of selective media improves diagnosis of Burkholderia pseudomallei, but resources may be limited in endemic developing countries. To maximise yield in the relatively low-prevalence setting of Kuala Lumpur, Malaysia, B. pseudomallei selective agar and broth were compared with routine media for 154 respiratory specimens from patients with community-acquired disease. Selective media detected three additional culture-positive specimens and one additional melioidosis patient, at a consumables cost of US$75. Burkholderia pseudomallei was not isolated from 74 diabetic foot ulcer samples. Following careful local evaluation, focused use of selective media may be cost-effective.
We have developed a multiplex PCR assay for rapid identification and differentiation of cultures for Burkholderia pseudomallei, Burkholderia thailandensis, Burkholderia mallei and Burkholderia cepacia complex. The assay is valuable for use in clinical and veterinary laboratories, and in a deployable laboratory during outbreaks.