Molecular typing and antibiotic resistance patterns of methicillin-resistant Staphylococcus aureus isolates from clinical samples in Malaysia: An update.

A total of 120 non-consecutive MRSA isolates were obtained from hospitalized patients at Hospital Kuala Lumpur, Malaysia. All isolates were subjected to antimicrobial susceptibility tests and genotyping based on staphylococcal cassette chromosome mec(SCCmec), Staphylococcus aureus protein A typing (spa) and multilocus sequence typing (MLST). Vast majority of MRSA isolates were resistant to more than three classes of antibiotics. Five antibiotic resistance profiles were observed among the MRSA isolates. All isolates tested were still susceptible to vancomycin. Genotyping revealed isolates are highly clonal, where all MRSA belonged to the predominant Asian clone ST239 comprising 4 spa types. Spa typing revealed four different spa types. Continuous monitoring and effective therapeutic options for Asian MRSA clone is recommended.

Nasal carriers are more likely to acquire exogenous Staphylococcus aureus strains than non-carriers.

We performed a prospective observational study in a clinical setting to test the hypothesis that prior colonization by a Staphylococcus aureus strain would protect, by colonization interference or other processes, against de novo colonization and, hence, possible endo-infections by newly acquired S. aureus strains. Three hundred and six patients hospitalized for >7 days were enrolled. For every patient, four nasal swabs (days 1, 3, 5, and 7) were taken, and patients were identified as carriers when a positive nasal culture for S. aureus was obtained on day 1 of hospitalization. For all patients who acquired methicillin-resistant S. aureus (MRSA) or methicillin-susceptible S. aureus via colonization and/or infection during hospitalization, strains were collected. We note that our study may suffer from false-negative cultures, local problems with infection control and hospital hygiene, or staphylococcal carriage at alternative anatomical sites. Among all patients, 22% were prior carriers of S. aureus, including 1.9% whom carried MRSA upon admission. The overall nasal staphylococcal carriage rate among dermatology patients was significantly higher than that among neurosurgery patients (n = 25 (55.5%) vs. n = 42 (16.1%), p 0.005). This conclusion held when the carriage definition included individuals who were nasal culture positive on day 1 and day 3 of hospitalization (p 0.0001). All MRSA carriers were dermatology patients. There was significantly less S. aureus acquisition among non-carriers than among carriers during hospitalization (p 0.005). The mean number of days spent in the hospital before experiencing MRSA acquisition in nasal carriers was 5.1, which was significantly lower than the score among non-carriers (22 days, p 0.012). In conclusion, we found that nasal carriage of S. aureus predisposes to rather than protects against staphylococcal acquisition in the nose, thereby refuting our null hypothesis.

Experimental infection of brown-marbled grouper, Epinephelus fuscoguttatus (Forskal), with Vibrio parahaemolyticus identifies parvalbumin beta-2 subunit I, alpha-2-macroglobulin, nattectin and immunoglobulin light chain, differentially expressed in resistant grouper.

The mechanisms through which brown-marbled grouper accomplishes resistance to infection, particularly against Vibrios, are not yet fully understood. In this study, brown-marbled grouper fingerlings were experimentally infected with Vibrio parahaemolyticus, to identify disease resistance grouper, and the serum proteome profiles were compared between resistant and susceptible candidates, via two-dimensional gel electrophoresis (2-DE). The results showed that putative parvalbumin beta-2 subunit I, alpha-2-macroglobulin, nattectin and immunoglobulin light chain proteins were among proteins that significantly overexpressed in the resistant fish as compared to the susceptible group of fish, whereas apolipoprotein E and immunoglobulin light chain proteins were observed to be differentially overexpressed in the susceptible fish. Further analysis by peptide sequencing revealed that the immunoglobulin light chain proteins identified in the resistant and susceptible groups differed in amino acid composition. Taken together, the results demonstrated for the first time that putative parvalbumin beta-2 subunit I, alpha-2-macroglobulin, nattectin and immunoglobulin light chain are among important proteins participating to effect disease resistance mechanism in fish and were overexpressed to function collectively to resist V. parahaemolyticus infection. Most of these molecules are mediators of immune response.

Putative apolipoprotein A-I, natural killer cell enhancement factor and lysozyme g are involved in the early immune response of brown-marbled grouper, Epinephelus fuscoguttatus, Forskal, to Vibrio alginolyticus.

The gram-negative bacterium, Vibrio alginolyticus, has frequently been identified as the pathogen responsible for the infectious disease called vibriosis. This disease is one of the major challenges facing brown-marbled grouper aquaculture, causing fish farmers globally to suffer substantial economic losses. The objective of this study was to investigate the proteins involved in the immune response of brown-marbled grouper fingerlings during their initial encounter with pathogenic organisms. To achieve this objective, a challenge experiment was performed, in which healthy brown-marbled grouper fingerlings were divided into two groups. Fish in the treated group were subjected to intraperitoneal injection with an infectious dose of V. alginolyticus suspended in phosphate-buffered saline (PBS), and those in the control group were injected with an equal volume of PBS. Blood samples were collected from a replicate number of fish from both groups at 4 h post-challenge and analysed for immune response-related serum proteins via two-dimensional gel electrophoresis. The results showed that 14 protein spots were altered between the treated and control groups; these protein spots were further analysed to determine the identity of each protein via MALDI-TOF/TOF. Among the altered proteins, three were clearly overexpressed in the treated group compared with the control; these were identified as putative apolipoprotein A-I, natural killer cell enhancement factor and lysozyme g. Based on these results, these three highly expressed proteins participate in immune response-related reactions during the initial exposure (4 h) of brown-marbled grouper fingerling to V. alginolyticus infection.

Genetic variation among methicillin-resistant Staphylococcus aureus isolates from cancer patients in Saudi Arabia.

One hundred and twenty methicillin-resistant Staphylococcus aureus (MRSA) isolated from cancer and non-cancer patients in Saudi Arabia were investigated for antibiotic resistance, virulence determinants and genotypes. The majority of MRSA isolates from cancer (n = 44, 73.3 %) and non-cancer patients (n = 34, 56.7 %) were multi-resistant to more than four classes of antibiotics. Virulence gene profiling showed that all strains were commonly positive for adhesin genes, except ebps and bbp genes, which were not detected in any isolate. Although the presence of adhesin genes varied slightly among MRSA isolates from cancer and non-cancer patients, these variations were not found to be statistically significant. In contrast, the presence of the toxin genes seb, sec, seg and sei was significantly elevated in MRSA strains isolated from cancer patients. Multilocus sequence typing (MLST) detected six and nine sequence types (STs) among isolates from cancer and non-cancer patients, respectively. Using spa typing, 12 and 25 types were detected, including four new types. The ability of different MRSA clones to become multi-resistant and their ability to acquire different virulence factors may contribute to their success as pathogens in individual groups of patients.

Diversity in the antimicrobial susceptibility patterns of methicillin-resistant Staphylococcus aureus clones.

Methicillin-resistant Staphylococcus aureus (MRSA) is well known for its epidemicity, with the emergence of new clones on a daily basis. Diversity in the clonal types of MRSA challenges the success of treatment, as different clones respond to different sets of antibiotics. However, the antibiotic susceptibility among the isolates within the same clones is largely unexplored. In a previous study on MRSA epidemiology in Malaysia, we identified six major clonal complexes (ST-239-CC8, ST-1-CC1, ST-188-CC1, ST-22-CC22, ST-7-CC7 and ST-1283-CC8). In the present study, we investigated the antibiotic susceptibility patterns of isolates of different clones. Three hundred and eighty-nine MRSA isolates were subjected to the disc diffusion test, oxacillin minimum inhibitory concentration (MIC) determination and assessment of the distribution of macrolide, lincosamide and streptogramin B (MLS(B)) resistance genes. Thirty-six different antibiotic profiles were observed: 30 (83.3 %) among ST-239, 2 (5.6 %) among ST-1283 and 1 (2.8 %) each for ST-1, ST-7, ST-22 and ST-188. All ST-239 (362, 9 %) isolates were multiple drug-resistant (MDR; resistant to more than three classes of antibiotics) and had oxacillin MICs >256 mg/l. Among the 385 clindamycin-resistant isolates, 375 (96.4 %) illustrated inducible resistance (D-zone-positive), while 10 (2.6 %) showed constitutive resistance. The vast majority of the macrolide-resistant isolates carried the ermA gene (95.1 %), followed by ermC (12.9 %). Diversity in the antibiotic susceptibilities of isolates within the clones emphasises the need for continuous surveillance of MDR strains to prescribe the correct antibiotic rather than empirical treatment. This will likely reduce the emergence of new endemic or epidemic resistant MRSA clones.

High prevalence of qacA/B carriage among clinical isolates of meticillin-resistant Staphylococcus aureus in Malaysia.

The minimum inhibitory concentrations (MICs) of 60 meticillin-resistant Staphylococcus aureus (MRSA) isolates from Malaysia to three antiseptic agents - benzalkonium chloride (BZT), benzethonium chloride (BAC) and chlorhexidine digluconate (CHG) - were determined. All isolates had MICs ranging from 0.5 to 2 mg/L. Antiseptic resistance genes qacA/B and smr were detected in 83.3% and 1.6% of the isolates, respectively. Carriage of qacA/B correlated with reduced susceptibility to CHG and BAC. This is the first report of the prevalence of qacA/B and smr gene carriage in Malaysian MRSA isolates, with a high frequency of qacA/B carriage. The presence of these antiseptic resistance genes and associated reduced susceptibility to antiseptic agents may have clinical implications.

mec-associated dru typing in the epidemiological analysis of ST239 MRSA in Malaysia.

The usefulness of mec-associated dru typing in the epidemiological analysis of methicillin-resistant Staphylococcus aureus (MRSA) isolated in Malaysia was investigated and compared with pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and spa and SCCmec typing. The isolates studied included all MRSA types in Malaysia. Multilocus sequence type ST188 and ST1 isolates were highly clonal by all typing methods. However, the dru typing of ST239 isolates produced the clearest discrimination between SCCmec IIIa and III isolates, yielding more subtypes than any other method. Evaluation of the discriminatory power for each method identified dru typing and PFGE as the most discriminatory, with Simpson's index of diversity (SID) values over 89%, including an isolate which was non-typeable by spa, but dru-typed as dt13j. The discriminatory ability of dru typing, especially with closely related MRSA ST239 strains (e.g., Brazilian and Hungarian), underscores its utility as a tool for the epidemiological investigation of MRSA.

Cytotoxicity and scanning electron microscopy study of gentamycin-coated HA effect on biofilm.

Biofilms are adherent, multi-layered colonies of bacteria that are typically more resistant to the host immune response and routine antibiotic therapy. HA biomaterial comprises of a single-phased hydroxyapatite scaffold with interconnected pore structure. The device is designed as osteoconductive space filler to be gently packed into bony voids or gaps following tooth extraction or any surgical procedure. Gentamycin-coated biomaterial (locally made hydroxyapatite) was evaluated to reduce or eradicate the biofilm on the implant materials. The results indicated that the HA coated with gentamycin was biocompatible to human osteoblast cell line and the biofilm has been reduced after being treated with different concentrations of gentamycin-coated hydroxyapatite (HA).

Adult-to-adult living donor liver transplant: UK experience.

BACKGROUND: Adult-to-adult living donor liver transplantation (ALDLT) is being adopted widely in the USA and mainland Europe, fueled by the increasing waiting lists for cadaver organs. The present report describes the first UK experience with the procedure in patients from overseas who have the lowest priority for cadaver organ allocation. METHODS: The 16 patients seen over the period November 1998 to March 2002 had end-stage cirrhosis from chronic hepatitis C virus (HCV) or hepatitis B virus (HBV) infection (13 cases), with single instances of cryptogenic cirrhosis, secondary biliary cirrhosis and alcoholic liver disease. Grafts were left lobe in the first two recipients and right lobe in the subsequent 14 recipients, donated by nine sons/daughters and seven brothers/sisters. RESULTS: Twelve of the 16 recipients did well. The four recipients who died had recurrent sepsis; two of these died following hepatic arterial occlusion, and in three major surgical factors were present before transplantation. Serial computed tomography (CT) measurements in the survivors showed regeneration of the grafted lobe with final volumes reaching in each case the calculated standard liver volume for body size. In the donors, liver function tests had returned to normal by day 7-14, with rapid regeneration of the remaining lobe, although the final size attained that estimated before donation in only four donors. CONCLUSIONS: ALDLT, although requiring considerable facilities and organization, can give good results for both recipient and donor. As with cadaver grafts, outcome in the recipient if the larger right lobe is used is dependent on surgical risk factors and the severity of clinical decompensation before transplantation. Measures to ensure the safety of the donors remain the main concern.

Detection and molecular characterization of the zot gene in Vibrio cholerae and V. alginolyticus isolates.

A total of 11 Vibrio cholerae isolates from 1996-1998 outbreaks in Malaysia and 4 V. alginolyticus were analyzed. Isolates were characterized by polymerase chain reaction (PCR) and Southern hybridization for the presence of the gene encoding zonula occludens toxin (zot). Screening of zot gene by PCR revealed the presence of this gene in V. cholerae and V. alginolyticus. The zot gene from one V. cholerae Ogawa isolate that was cloned in a pCR 2.1 TOPO vector was sequenced. The sequences obtained were 99% homologous to the zot gene sequence from the Gene Bank.

Prevalence of pulmonary tuberculosis on the roof of the world.

SETTING: The Shimshal Valley, a remote village in Northern Pakistan, is one of the seven Pamirs of Central Asia, widely known as the roof of the world. OBJECTIVE: To investigate the prevalence of pulmonary tuberculosis (TB) in the Shimshal Valley. DESIGN: The Rapid Village Survey Method (RVS) was used to investigate the prevalence of pulmonary tuberculosis. The selection criteria were chronic cough, hemoptysis, past history of TB and close contact with a tuberculous patient. After clinical examination, a chest radiograph was done and a single spot sputum sample was obtained for smear examination. RESULTS: The total population of the village was 1077, of whom 231 cases were studied. Overcrowding affected 75% of the study population. The prevalence of smear positive pulmonary TB in the village studied was 554 per 100000 population, and the prevalence of active smear-negative TB was estimated at 1949/100 000. The prevalence of active pulmonary TB increased with age and the only risk factor for active TB was age over 45 years. Of the 21 cases with a past history of pulmonary TB, only 38% had completed a full course of chemotherapy. CONCLUSION: Pulmonary TB is a very serious health issue in the rural community (Shimshal Valley) of Pakistan. This study highlights the lack of efficacy of national tuberculosis control programs in the country.