25S rDNA genotyping and multi-locus sequence typing of Candida albicans of pathogenic and commensal origins in the Klang Valley, Malaysia.

Candida albicans is an important opportunistic fungal pathogen capable of causing fatal systemic infections in humans. Presently in Malaysia, there is little information available on the genetic diversity of this organism and trends in behavioural characteristics. In this project, three genotyping methods: 25S rDNA genotyping, Alternative Lengthening of Telomerase (ALT) sequence typing and Multi-Locus Sequence Typing (MLST) were applied to study the genetic diversity of strains from infected hospital in-patients and asymptomatic individuals in the community. The results showed that, with the 25S rDNA genotyping, as in other parts of the world, the most common genotype was type A which accounted for approximately 70% of the 111 isolates tested. Further typing with the ALT sequence showed type 3 to be the most common in the isolates tested. MLST analysis revealed many possibly novel sequence types, as well as a statistically significant association between pathogenicity and a group of closely related isolates, most of which were from hospital samples. Further work on genotypes associated with enhanced virulence will help to clarify the value of genotyping for clinical and epidemiological investigations.

Draft Genome Sequence of a Clinical Isolate of Mycobacterium tuberculosis Strain PR05.

We report the annotated genome sequence of a clinical isolate, Mycobacterium tuberculosis strain PR05, which was isolated from the human cerebrospinal fluid of a patient diagnosed with tuberculosis.

Hepatitis B seroprevalence among University of Malaya Students in the Post-universal Infant Vaccination Era.

AIM: A nationwide HBV vaccination for neonates in the Expanded Programme on Immunization (EPI) was implemented in Malaysia in 1989. The objective of this study was to investigate the prevalence of HBsAg, anti-HBs and anti-HBc among the new student intakes in the Faculties of Medicine and Dentistry, University of Malaya from 2005 to 2011. MATERIALS AND METHODS: All new students enrolled for undergraduate and postgraduate courses were screened for HBV infection. Serum samples collected were tested for the presence of HBsAg, anti-HBs and anti-HBc with the use of fully automated analysers. Statistical analyses were done using Open Epi version 2.3.1 RESULTS: The overall HBsAg prevalence among the 2923 new intakes was 0.62%. The HBsAg prevalence rate was 1.08% (15/1390) for those born before 1989 and only 0.20% (3/1533) among those born in or after 1989. By year of testing, HBsAg prevalence declined steadily from 1.27% (5/394) in 2005 to 1.20% (5/418) in 2006, 0.95% (4/421) in 2007, 0.49% (2/410) in 2008, 0.49% (2/407) in 2009 and finally 0% in both 2010 (0/445) and 2011 (0/428). Although 66.14% of those vaccinated during infancy had no demonstrable immunity at the time of screening, only 6 (0.39%) students were found to have anti-HBc, including the 3 who were HBsAg positive. CONCLUSION: These findings suggested effective prevention of HBV transmission with the universal and voluntary vaccination programs in Malaysia.

Tuberculosis in captive Asian elephants (Elephas maximus) in Peninsular Malaysia.

A cross-sectional study was conducted from 10 January to 9 April 2012, to determine the seroprevalence of tuberculosis (TB) of all captive Asian elephants and their handlers in six locations in Peninsular Malaysia. In addition, trunk-wash samples were examined for tubercle bacillus by culture and polymerase chain reaction (PCR). For 63 elephants and 149 elephant handlers, TB seroprevalence was estimated at 20.4% and 24.8%, respectively. From 151 trunkwash samples, 24 acid-fast isolates were obtained, 23 of which were identified by hsp65-based sequencing as non-tuberculous mycobacteria. The Mycobacterium tuberculosis-specific PCR was positive in the trunk-wash samples from three elephants which were also seropositive. Conversely, the trunk wash from seven seropositive elephants were PCR negative. Hence, there was evidence of active and latent TB in the elephants and the high seroprevalence in the elephants and their handlers suggests frequent, close contact, two-way transmission between animals and humans within confined workplaces.

Molecular typing of Mycobacterium abscessus based on tandem-repeat polymorphism.

A variable-number tandem-repeat (VNTR) typing assay for the differentiation of Mycobacterium abscessus strains was developed. This assay showed complete reproducibility, locus stability, and a discriminatory power (Hunter-Gaston discriminatory index [HGDI] of 0.9563) that is superior to that of multilocus sequencing. It is a promising tool for the investigation of Mycobacterium abscessus epidemiology and nosocomial outbreaks.

Multiplex PCR for the detection of urogenital pathogens in mothers and newborns.

Two duplex PCR assays were established for the detection of C. trachomatis (Ct), N. gonorrhoeae (GC), M. hominis (Mh), and U. urealyticum (Uu). These assays were used on clinical specimens obtained from women with Premature Rupture of Membrane or Post Partum Fever, from preterm infants, as well as from women with uneventful pregnancies and their babies delivered vaginally at term. The analytical sensitivity of the duplex PCR assays with internal controls incorporated is 7.0, 19.0, 5x10(3) and 7x10(2) genome copies per reaction for Ct, GC, Mh and Uu respectively. Specificity was demonstrated by the amplification of only target DNA in the presence of other organisms. Among 40 women with normal, at term, deliveries, there were 6 positives for Ct, 2 for GC and 1 for Uu. None of these women had signs of genital tract infection. The Mh/Uu PCR was positive in 11 of 40 PROM cases, with 7 women positive for Uu, 2 for Mh and 2 others for both organisms. Of 40 blood cultures taken from post-partum maternal infections, 6 were positive for Ct and 1 for Mh. Respiratory secretions from 30 premature neonates yielded 5 positives for Uu and one each for Mh and Ct. In contrast, there was only 1 positive result (for Mh) in 30 mature neonates. With 1 exception, all mycoplasma and ureaplasma positives were confirmed by culture and the concordance between paired tracheal aspirates and nasopharyngeal swabs from neonates was 96.7%. These results show the potential use of the duplex PCR assays for the diagnosis of maternal and neonatal disease caused by the four urogenital pathogens.

Real-time PCR detection of male-specific coliphages.

Male-specific coliphages are often used as indicators of contamination by enteric viruses. These phages can be detected in water samples by plaque assays and by polymerase chain reaction. In this study, the M13 coliphage was used to develop a real-time PCR assay for the detection of male-specific DNA coliphages. The real-time PCR was found to have a reaction efficiency of 1.45 and detection limit of 10(-3) plaque forming units per reaction mix. Repeated amplification and melting curve analyses demonstrated high specificity and reproducibility of the real-time assay. Quantitative detection with the real-time PCR should allow rapid assessment of the level of viral contamination in water.

Community-acquired vancomycin-resistant Enterococcus faecium: a case report from Malaysia.

Vancomycin-resistant enterococci (VRE) are formidable organisms renowned for their ability to cause infections with limited treatment options and their potential for transferring resistance genes to other Gram-positive bacteria. Usually associated with nosocomial infections, VRE are rarely reported as a cause of community-acquired infection. Presented here is a case of community-acquired infection due to vancomycin-resistant Enterococcus faecium. The patient had been applying herbal leaves topically to his cheek to treat a buccal space abscess, resulting in a burn of the overlying skin. From pus aspirated via the skin a pure culture of E. faecium was grown that was resistant to vancomycin with a MIC of >256 microg ml-1 by the E test and resistant to teicoplanin by disc diffusion, consistent with the VanA phenotype. The organism was suspected of contaminating the leaf and infecting the patient via the burnt skin. This case highlights the need for further studies on the community prevalence of VRE among humans and animals to define unrecognized silent reservoirs for VRE, which may pose a threat to public health.

Ethnic distribution of Chlamydophila pneumoniae antibodies in a Malaysian population and possible correlation with coronary heart disease.

Chlamydophila pneumoniae, an important respiratory pathogen causing lower respiratory tract infections, has also been implicated in coronary heart disease (CHD). This study reports a cross-sectional, demographic, serological analysis of the prevalence of Chlamydophila pneumoniae antibodies in a multiracial Malaysian population. Generally, Malaysian Indians had the highest degree of seropositivity (58%) followed by the Chinese (54%) and the Malays (32%), results which were statistically significant (CI: 95%; p < 0.01). Interestingly, this trend was also reflected in the study group consisting of patients with acute myocardial infarctions (AMI) and chronic CHD. Again, the Indians were more frequently seropositive (65%), with more than 50% having IgG titres > or = 128. Comparatively, the Chinese and Malays showed 51 and 17% seropositivity respectively. These results were also statistically significant (CI: 95%; p < 0.01). Malaysian Indians are more commonly afflicted with CHD. A variety of factors have been suggested to explain this prevalence including diet, social habits, genetics and the possible role of infectious agents. This study notes an interesting association between this pattern of racial prevalence and the possible role of C. pneumoniae infections as a contributory/predisposing factor in the development of cardiovascular disease.

Infrequent occurrence of vancomycin-resistant enterococci in poultry from Malaysian wet markets.

Fifty samples of chicken, duck and geese faeces were obtained from 13 wet markets in Kuala Lumpur to study the prevalence of vancomycin-resistant enterococci (VRE) among local market poultry. Biotyping of colonies grown on azide agar incubated at 45 degrees C yielded E. pseudoavium, E. faecalis, E. faecium and E. gallinarum from chicken faeces and E. malodoratus, E. faecalis, E. faecium, E. gallinarum, E. hirae/dispar, and E. durans from goose and duck faeces. On agar containing 6 mg/ l of vancomycin, one strain of E. flavescens was identified, giving a VRE detection rate of 2.0%. This isolate had a vancomycin M.I.C. of 8 mg/l as determined by the Etest, and the van C-3 gene that was identified by PCR followed by sequence analysis. The prevalence of VRE among poultry sold in local markets appears to be low, and may reflect the infrequent use of antimicrobials in our poultry farms. Nevertheless, the possibility of human acquisition of microbes via the food chain cautions against the use of antimicrobials in animal husbandry that may encourage the emergence and spread of multi-drug resistant organisms like the VRE among animal microbial flora.

Mycoplasma Pneumoniae infection in Malaysian children admitted with community acquired pneumonia.

Mycoplasma pneumoniae is increasingly recognized as an important cause of community acquired pneumonia (CAP) in children. We determined the importance of M. pneumoniae as a causative agent in 170 children aged 1 month to 15 years who were hospitalized with CAP over a 6-month period. The diagnosis of M. pneumoniae infection was based on serological evidence obtained by a particle agglutination test (SERODIA-MYCO II). A positive serological diagnosis was made if the acute phase serum titer was more than 1:160 or paired samples taken 2-4 weeks apart showed a four-fold or greater rise in the serum titer. M. pneumoniae was identified as the causative agent in 40 (23.5%) children. Children with M. pneumoniae infection were more likely to be older than 3 years (OR 4.0 95%CI 1.8-9.1, p<0.001), Chinese (OR 4.3 95%CI 2.0-8.9, p<0.001), have a duration of illness longer than 7 days prior to admission (OR 6.0 95%CI 2.7-13.5, p<0.001) and have perihilar interstitial changes on chest X-ray (OR 4.6 95%CI 2.2-9.9, p<0.001). A significant number of hospital admissions for CAP in Malaysian children can be attributed to M. pneumoniae. It is important to identify these children so as to administer the most appropriate antibiotic treatment.

Colonization and transmission of Ureaplasma urealyticum and Mycoplasma hominis from mothers to full and preterm babies by normal vaginal delivery.

A prospective study was carried out among pregnant women and their newborn babies in the University Hospital, Kuala Lumpur from January 1996 to June 1997. The maternal cervical colonization rates of Ureaplasma urealyticum (UU) and Mycoplasma hominis (MH) were found to be 57.5% and 15.8% respectively while the isolation rates from nasopharyngeal secretions of the newborns were 50.8% for UU and 6.6% for MH. The overall transmission rates were 88.4% for UU and 42.1% for MH. There was no significant difference in the transmission rates of either organism from mothers to their respective newborn babies by the maturity of pregnancy. In preterm babies, the nasopharyngeal isolation rates of UU and MH were not influenced by the babies' gestational age and birth weight nor by the maternal history of abortion or parity. However, there was a tendency for UU to persist in the nasopharyngeal secretion of preterm babies especially those of birth weight below 2 kg. None of the babies contaminated with mycoplasmas at birth developed respiratory symptoms during six to eight weeks of follow-up.

An immunogenic epitope of Chlamydia pneumoniae from a random phage display peptide library is reactive with both monoclonal antibody and patients sera.

Random 15-mer peptides displayed on filamentous phages were screened in binding studies using a Chlamydia pneumoniae-specific monoclonal antibody (RR-402) and affinity-purified, polyclonal sera from patients seropositive for C. pneumoniae infections by the microimmunofluorescence (MIF) test. One 15-mer epitope, epitope Cpnl5A (LASLCNPKPSDAPVT) was identified in both the monoclonal and polyclonal screenings, and showed higher ELISA reactivity with C. pneumoniae MIF-positive sera compared to patients with other chlamydial infections, non-chlamydial respiratory infections and normal healthy sera (MIF-negative). Interestingly, epitope Cpnl5A also showed significant (52%) amino acid sequence homology to the 56 kDa type-specific antigen of Rickettsia tsutsugamushi, a protein implicated in the virulence of this organism.

Ureaplasma urealyticum and Mycoplasma hominis isolation from cervical secretions of pregnant women and nasopharyngeal secretions of their babies at delivery.

BACKGROUND: A prospective study was carried out at the University Hospital, Kuala Lumpur to determine the cervical carriage rate of Ureaplasma urealyticum and Mycoplasma hominis among healthy pregnant women at delivery and the incidence of nasopharyngeal colonisation among their infants. PATIENTS: Sixty mother and baby pairs were examined. RESULTS: Cervical colonisation among the mothers was found to be 56.7% for U.urealyticum and 17.7% for M.hominis. The transmission rate to their infants was 88.2% and 30% for U.urealyticum and M.hominis respectively. CONCLUSION: There was no statistically significant difference in the maternal colonisation rates according to ethnic group, parity and past history of abortion. All U.urealyticum isolates in our study were sensitive to erythromycin but about one-third were resistant to tetracycline and ciprofloxacin and 26.5% were resistant to minocycline.

Bacterial infection of central venous catheters in short-term total parenteral nutrition.

Fourteen severely ill ventilated patients in an intensive care unit, requiring short-term total parenteral nutrition, were examined for catheter-related infection. Microbiological analysis using Maki's SQ technique was carried out on catheter exit site, catheter hub, proximal subcutaneous segment of catheter and catheter up. Qualitative cultures were carried out on total parenteral nutrition and peripheral blood samples. Twenty six of 29 catheters removed (90%) were culture positive but only 7 catheters were related to positive blood cultures, giving a catheter-related bacteremia (CRB) rate of 24%. Haematogenous seeding was strongly implicated in 7/29 (24%) of catheters. Patients' skin flora appeared to be the main source of catheter-related infection. The organisms isolated for patients with CRB included coagulase-negative staphylococci, Acinetobacter and Klebsiella. It is suggested that to control infective complications of central venous catheters, emphasis should be focused on specialised intravenous therapy teams and the use of strict protocols for insertion and care of central lines.

The prevalence of Chlamydia trachomatis in patients with pelvic inflammatory disease.

Chlamydia trachomatis is recognized as the most prevalent sexually transmitted organism in many parts of the world. Most complications associated with chlamydial infection in women and their infants can be avoided by appropriate treatment. However, treatment is often not initiated because infections are frequently asymptomatic. The identification of at risk patients and treatment of these patients is a practical clinical approach in the reduction of transmission and prevention of complications. The prevalence of chlamydial infection among patients with pelvic inflammatory disease admitted to Seremban General Hospital was 22.7%. The difference in seropositivity between PID patients (20.5%) and antenatal controls (2.3%) was statistically significant. The corresponding cervical antigen detection rates were 6.8% and 2.3% respectively. Chlamydial infection should be screened for in gynaecological patients and antibiotic policies should take cognizance of the aetiological role played by this organism in pelvic inflammatory disease.

Young Malaysian children with lower respiratory tract infections show low incidence of chlamydial infection.

OBJECTIVE: The incidence of Chlamydia pneumoniae and Chlamydia trachomatis infection was studied among infants and young children admitted to hospital for the management of lower respiratory tract infections, over a 12 month period. METHODOLOGY: Respiratory secretions were examined for chlamydiae by cell culture, enzyme-linked immunosorbent assay and polymerase chain reaction-enzyme immunoassay. Sera were tested by micro-immunofluorescence for chlamydial IgG, IgM and IgA. Other bacterial and viral pathogens were also looked for by standard cultural and serological methods. RESULTS: Of 87 patients aged 2 months-3 years, an aetiologic diagnosis was made in 41 (47.1%). C. pneumoniae and C. trachomatis were each detected in 1 (1.2%) of the patients. Among common bacterial pathogens, Haemophilus influenzae (13.8%) and Streptococcus pneumoniae (8.1%) were the most frequently identified. Respiratory viruses and elevated Mycoplasma pneumoniae antibodies were found in 10.3% and 9.1% of patients, respectively. CONCLUSION: Chlamydiae are infrequent causes of community-acquired acute lower respiratory tract infections in infants and very young children in Malaysia.

Group B streptococcus: maternal carriage rate and early neonatal septicaemia.

Between January 1984 and December 1994, 30 cases of early neonatal group B streptococcus (GBS) septicaemia were managed in the Neonatal Unit, University Hospital, Kuala Lumpur. Two neonates were outborn and 28 were inborn, giving an average annual incidence of neonatal GBS septicaemia of 0.4/1000 livebirths among inborn babies. In a separate survey over a three-month period, GBS genital carriage rate among 196 parturients was found to be 9.7%. Of the infants with GBS septicaemia, the mean gestational age was 37.5 +/- 3.8 weeks and the mean birthweight was 2540 +/- 716 g. Twelve (40%) were preterm infants and 14 (47%) were low birthweight infants. Male and female infants were almost equally affected. Prolonged rupture of membranes and maternal pyrexia accounted for only 5 (17%) and 3 (10%) of the cases respectively. Twenty-four (80%) neonates had onset of symptoms within 6 hours of life and respiratory symptoms were observed in 24 (80%) of the cases, while meningitis was uncommon. Six (20%) neonates died. Preterm and low birthweight infants had higher mortality than their term counterparts: 42% versus 6% and 36% versus 6% respectively. Of those who died, 4 (67%) required respiratory support right from birth and the mean time of onset of symptoms was 4 hours (range 0 to 21 hours) and the duration of survival was only 28.8 hours (range 12 to 38 hours). As the incidence of neonatal GBS septicaemia was low, mass screening and chemoprophylaxis for GBS were not recommended. All the GBS isolates were sensitive to penicillin and ampicillin, thus one of these antibiotics should be included in the antimicrobial therapy of septic neonates.