Evaluation of Conventional PCR for Detection of Toxigenic Corynebacterium diphtheriae Strains in Malaysia.

Diphtheria is an infectious but vaccine preventable disease caused by Corynebacterium diphtheriae and humans are the only reservoir. While toxigenic strains most frequently cause pharyngeal diphtheria, non-toxigenic strains commonly cause cutaneous infections. In 2016, there was a sudden increase in cases of C. diphtheriae reported in Malaysia. The toxigenic strains are currently determined using Elek's test and are carried out only in the reference laboratory. With the sudden increase in diphtheria cases in Malaysia, it is important for local laboratories in state hospitals to be able to perform a rapid, reliable diagnostic test for the detection of the exotoxin. In this study, we aimed to evaluate the application of conventional PCR method to detect toxigenic strains of C. diphtheriae compared to the Elek's test. Forty-eight C. diphtheriae strains were subjected to PCR detection of toxin gene A and B subunits, and also Elek's test. The A and B subunits of the toxin gene were detected in all C. diphtheriae strains except for one strain which was isolated from a foot ulcer. Elek's test was also positive for all the PCR positive strains. This study showed 100% corelation between the results of PCR and Elek's test assay. The conventional PCR can be used at the state laboratories for rapid detection of toxin genes in toxigenic C. diphtheriae cultures, thus early treatment can be given to the patients while waiting for Elek's test results.

Evaluation of Conventional PCR for Detection of Toxigenic Corynebacterium diphtheriae Strains in Malaysia

@#Diphtheria is an infectious but vaccine preventable disease caused by Corynebacterium diphtheriae and humans are the only reservoir. While toxigenic strains most frequently cause pharyngeal diphtheria, non-toxigenic strains commonly cause cutaneous infections. In 2016, there was a sudden increase in cases of C. diphtheriae reported in Malaysia. The toxigenic strains are currently determined using Elek’s test and are carried out only in the reference laboratory. With the sudden increase in diphtheria cases in Malaysia, it is important for local laboratories in state hospitals to be able to perform a rapid, reliable diagnostic test for the detection of the exotoxin. In this study, we aimed to evaluate the application of conventional PCR method to detect toxigenic strains of C. diphtheriae compared to the Elek’s test. Forty-eight C. diphtheriae strains were subjected to PCR detection of toxin gene A and B subunits, and also Elek’s test. The A and B subunits of the toxin gene were detected in all C. diphtheriae strains except for one strain which was isolated from a foot ulcer. Elek’s test was also positive for all the PCR positive strains. This study showed 100% corelation between the results of PCR and Elek’s test assay. The conventional PCR can be used at the state laboratories for rapid detection of toxin genes in toxigenic C. diphtheriae cultures, thus early treatment can be given to the patients while waiting for Elek’s test results.

Community-acquired necrotising pneumonia caused by Panton-Valentine leucocidin-producing methicillin-resistant Staphylococcus aureus.

A 61-year-old male presented with community-onset pneumonia not responding to treatment despite given appropriate antibiotics. Computed tomography scan of the thorax showed large multiloculated pleural effusion with multiple cavitating foci within collapsed segments; lesions which were suggestive of necrotising pneumonia. Drainage of the effusion and culture revealed methicillin-resistant Staphylococcus aureus, which had the same antibiotic profile with the blood isolate and PVL gene positive.

Surgical treatment of children with raised intra ocular pressures associated with sturge weber syndrome at hospital kuala lumpur, malaysia.

No abstract available.

Outbreak of melioidosis and leptospirosis co-infection following a rescue operation.

We analyzed the epidemiological data of all people who were involved in the search and rescue operation in Lubuk Yu, a natural recreational forest with waterfall and stream. The hospital admission records of the cases who fulfilled the case definition and the environmental samples result taken at Lubuk Yu recreational area were studied. 153 people were exposed to this outbreak, 85 (55.5%) were professional rescuers from various government agencies and 68 (44.5%) were villagers. 21 fulfilled the case definition. Ten cases were confirmed melioidosis, six melioidosis alone and four coinfected with leptospirosis. There were eight deaths in this outbreak, seven were villagers and one professional rescuer. Overall case fatality was 70%. All confirmed melioidosis cases and seven who died had diabetes mellitus. The morbidity rate were higher among the villagers, 23.5% compared to professional rescuers, 5.9%. The case fatality rate were also higher in this group which was 100% compared to 33.3% in professional rescuers. The soil and water samples in Lubuk Yu recreational area were positive for leptospira and Burkholderia pseudomallei. The presence of co-infection and co-morbidities especially diabetes mellitus among the exposed led to the high mortality in this outbreak hence a high index of suspicion is important among the healthcare professionals in the management of melioidosis cases. To avoid similar incident in future, search and rescue operation should be only conducted by professional rescuers with appropriate personal protective equipment. A register of rescuers should be maintained for surveillance and follow up if necessary.

Seroprevalance of brucellosis among suspected cases in Malaysia.

Brucellosis is a zoonotic disease which can be transmitted by direct or indirect contact with infected animal or their products. It is an important public health problem but little is known on brucellosis in the Malaysian population. The aim of this study was to determine the presence of Brucella antibodies using commercial Brucella IgG and IgM ELISA kits (Vircell, SL, Barcelona Spain). A total of 184 sera from suspected patients were received from 16 hospitals in Malaysia over the years 2004 to 2009. Only 10 serum samples (5.4%) were positive for Brucella antibodies in which 5 showed the presence of both IgM and IgG. Most of the positive patients were occupationally involved with animals. This study suggests the seroprevalance of brucellosis among individuals who have contact with infected animals in Malaysia is low.

DNA fingerprinting of septicemic and localized Burkholderia pseudomallei isolates from Malaysian patients.

We have analysed DNA fingerprinting patterns by pulsed-field gel electrophoresis (PFGE) of 52 unrelated Burkholderia pseudomallei strains isolated from septicemic and localized infections from Malaysian subjects. A total of 38 PFGE types were observed among 36 septicemic and 16 localized strains with no predominant pattern. Type 25 was seen in 2 epidemiologically related strains, suggesting human to human transmission. Twelve PFGE types were shared among 26 strains (21 septicemic and 5 localized) showing close genetic relatedness with coefficient of similarity of 0.81 to 1.0. The other 26 strains (15 septicemic and 11 localized) were unrelated as shown by the similarity coefficient of < 0.8. This study showed that our B. pseudomallei strains in Malaysia were mainly heterogenous with no predominant type both in septicemic or localized strains.

Globe avulsion secondary to maxillofacial trauma.

We report a case of globe avulsion secondary to maxillofacial trauma and propose potential mechanisms of injury. This case highlights the importance of wearing proper safety attire, especially during motorcycling.

Seroprevalance of brucellosis among suspected cases in Malaysia

Brucellosis is a zoonotic disease which can be transmitted by direct or indirect contact with infected animal or their products. It is an important public health problem but little is known on brucellosis in the Malaysian population. The aim of this study was to determine the presence of Brucella antibodies using commercial Brucella IgG and IgM ELISA kits (Vircell, SL, Barcelona Spain). A total of 184 sera from suspected patients were received from 16 hospitals in Malaysia over the years 2004 to 2009. Only 10 serum samples (5.4%) were positive for Brucella antibodies in which 5 showed the presence of both IgM and IgG. Most of the positive patients were occupationally involved with animals. This study suggests the seroprevalance of brucellosis among individuals who have contact with infected animals in Malaysia is low.

Analysis of PCR-RAPD DNA and antibiotic susceptibility profiles of antrum and corpus isolates of Helicobacter pylori from Malaysian patients.

This study was conducted to determine whether there was any genetic heterogeneity among Helicobacter pylori strains isolated from the antrum and corpus of the same individual in a Malaysian population and to determine the presence of heterogeneous susceptibility of the isolates by comparing PCR-RAPD and antibiotic profiles. Forty-four H. pylori isolates cultured from the antrum and corpus of 22 patients were analyzed. Antibiotic susceptibility testing was carried out by minimum inhibitory concentration determination, using E-Test method strips. PCR-RAPD was carried out on all the strains and the profiles generated were analysed for cluster analysis. Twenty-nine different PCR-RAPD profiles were observed in the 44 isolates. Fifteen pairs of the isolates from the same patients had the same PCR-RAPD patterns while in 7 pairs, the profiles were different. The strains were clustered into 2 separate clusters at a low coefficient of similarity, where most of the strains were in cluster 1. The degree of similarity was very low among most of the isolates. Most of the patients (16 of 22) were infected with strains that have the same antibiotic susceptibility profiles. Out of these, only 10 pairs shared the same PCR-RAPD and antibiotic profiles. Five pairs of isolates with similar PCR-RAPD profiles differed in their antibiotic profiles due to metronidazole resistance in one of the sites. A large degree of genetic heterogeneity was observed among H. pylori strains circulating among Malaysian patients. An individual patient can be infected with multiple strains and the strains can be antibiotic resistant.

Vancomycin treatment failure in a vancomycin susceptible methicillin-resistant Staphylococcus aureus (MRSA) infected patient.

A 64-year old patient, who had bacteraemia, did not respond to vancomycin despite the MRSA isolate being sensitive to the antibiotic at MIC 2 microg/mL. Electron microscopy of the MRSA isolate showed thickening of the cell wall, which was not observed in MRSA with lower vancomycin MIC.

Subdural collections due to non-typhi Salmonella infections in two Malaysian children.

Subdural collections caused by Salmonella infection are rarely encountered in children. We present two cases caused by non-typhi Salmonella, one a four-and-a-half-month-old boy presenting with subdural effusion, and the other, a 16-month-old boy with empyema. The diagnosis was confirmed on blood and subdural pus cultures. One patient had status epilepticus following focal fit, and the other had prolonged fever without any localising signs of infection on admission. They responded well to prompt surgical drainage and prolonged systemic antibiotic therapy. Contrary to previous reports, both patients showed favourable outcome in terms of neurological sequelae.

Serogroups and antibiotic susceptibility patterns of Neisseria meningitidis isolated from army recruits in a training camp.

Invasive Neisseria meningitidis infection is rare but carries a high mortality rate. The carriage rate in the normal population is around 10% and can be higher in confined populations. A study on the prevalence of carriage of N. meningitidis was conducted among 3195 army recruits after 2 months of intensive training in an army camp. N. meningitidis was isolated from 37.0% of these recruits. Two hundred and ten of N. meningitidis isolates were subjected to serogrouping and 100 to antibiotic sensitivity testing by the disc diffusion method and E-test for penicillin. Ten (4.8%) of 210 Neisseria meningitidis serogrouped belonged to serogroup W135, 3.33% serogroup A and 81.4% belonged to either serogroup X, Y or Z. With the agar disc diffusion method, all the N. meningitidis showed susceptiblity to chloramphenicol, rifampicin, cefotaxime and levofloxacin; 85% of the strains were resistant to cotrimoxazole and 12.5% resistant to penicillin. However, based on minimum inhibitory concentration, none of the Neisseria meningitidis tested was resistant to penicillin.

In vitro screening of five local medicinal plants for antibacterial activity using disc diffusion method.

Medicinal plants have many traditional claims including the treatment of ailments of infectious origin. In the evaluation of traditional claims, scientific research is important. The objective of the study was to determine the presence of antibacterial activity in the crude extracts of some of the commonly used medicinal plants in Malaysia, Andrographis paniculata, Vitex negundo, Morinda citrifolia, Piper sarmentosum, and Centella asiatica. In this preliminary investigation, the leaves were used and the crude extracts were subjected to screening against five strains of bacteria species, Methicillin Resistant Staphylococcus aureus (MRSA), Staphylococcus aureus, Klebsiella pneumoniae, Pseudomonas aeruginosa and Escherichia coli, using standard protocol of Disc Diffusion Method (DDM). The antibacterial activities were assessed by the presence or absence of inhibition zones and MIC values. M. citrifolia, P. sarmentosum and C. asiatica methanol extract and A. paniculata (water extract) have potential antibacterial activities to both gram positive S. aureus and Methicillin Resistant S. aureus (MRSA). None of the five plant extracts tested showed antibacterial activities to gram negative E. coli and K. pneumoniae, except for A. paniculata and P. sarmentosum which showed activity towards P. aeruginosa. A. paniculata being the most potent at MIC of 2 g/disc. This finding forms a basis for further studies on screening of local medicinal plant extracts for antibacteria properties.

In-vitro activity of quinupristin/dalfopristin, levofloxacin and moxifloxacin against fusidic acid and rifampicin-resistant strains of methicillin-resistant Staphylococcus aureus (MRSA) from Malaysian hospitals.

The in-vitro susceptibility of quinupristin/dalfopristin, levofloxacin and moxifloxacin against methicillin-resistant Staphylococcus aureus (MRSA) strains, which are also resistant to fusidic acid and rifampicin were carried out to determine whether these antibiotics can be used as an alternative treatment for multiply resistant MRSA strains. The minimum inhibitory concentrations (MIC) of these antibiotics were determined by E-test. Quinupristin/dalfopristin had good activity (MIC90 = 1 mg/L) against these strains while most of the strains showed intermediate resistance to moxifloxacin with MIC90 = 2 mg/L). However, more than 90% of these strains were resistant to levofloxacin with the MICs that ranged from 8 mg/L to 16 mg/L with the majority inhibited at 8 mg/L.

A case of trauma related Acanthamoeba keratitis.

Acanthamoeba is an uncommon cause of keratitis but one of the most severe because of the prolonged and painful course of the disease and poor visual outcome. Although contact lens use is the principal risk factor, about 10% of cases occur following trauma and exposure to contaminated soil or water. Two cases of Acanthamoeba keratitis involving women contact lens wearers have previously been reported in Malaysia but this is the first time, a non contact lens related Acanthamoeba keratitis is reported. The case involved a 28 year old Indonesian male construction worker who had a trauma of the right eye during work. His eye was struck by sand and dust particles after which he quickly washed with water from an open tank at the construction site. He experienced pain, redness, glaring and blurring of vision of the right eye three days later. The diagnosis was missed at initial presentation but culture of the corneal scraping had proven Acanthamoeba as the aetiological agent. The history and clinical findings of this trauma related Acanthamoeba keratitis are briefly discussed.

Staphylococcus aureus carriage in selected communities and their antibiotic susceptibility patterns.

The carriage and antibiotic susceptibility patterns of Staphylococcus aureus in the community were determined. Nasal, throat and axillary swabs were taken from 100 healthy adults and 90 disabled nursing home inmates. Antibiotic disc susceptibility testing was conducted following the NCCLS method. Staphylococcus aureus carriage was noted in 29% of healthy adults and 47.7% of nursing home inmates. Out of 79 strains, resistance to antibiotics were as follows; penicillin (92.4%), genetamicin (2.5%), tetracycline (6.3%), fusidic acid (11.3%), erythromycin (3.8%), pefloxacin (5.1%), mupirocin (3.8%), amikacin (3.8%), ciprofloxacin (2.5%) and chloramphenicol (2.5%). Methicillin-resistant Staphylococcus aureus was not isolated. Multiple colonizations and multi-antibiotic resistant Staphylococcus aureus were shown to occur in healthy individuals without risk factors and not previously hospitalized.

A major methicillin-resistant Staphylococcus aureus clone predominates in Malaysian hospitals.

This study was conducted to determine the molecular epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) in Malaysian hospitals. A total of 264 MRSA isolates from eight hospitals were subjected to typing by pulsed-field gel electrophoresis (PFGE) of SmaI restricted DNA. Antibiotic disk susceptibility testing was also carried out to determine their resistance patterns. Thirty-one PFGE pattern types were identified. Three major pattern types A, ZC and K were found with type A the predominant profile in c. 80% of strains and present in all hospitals. Unlike type A, other DNA pattern types were unique to the hospitals in which they were isolated. PFGE type A also consisted of strains that were multiply antibiotic resistant. The presence of a single predominant PFGE type in Malaysian hospitals is an important finding which suggests that inter-hospital spread of MRSA had occurred frequently and regularly.

Molecular fingerprinting of fusidic acid- and rifampicin-resistant strains of methicillin-resistant Staphylococcus aureus (MRSA) from Malaysian hospitals.

The emergence and spread of multiresistant methicillin-resistant Staphylococcus aureus (MRSA) strains, especially those resistant to fusidic acid and rifampicin, in Malaysian hospitals is of concern. In this study DNA fingerprinting by PFGE was performed on fusidic acid- and rifampicin-resistant isolates from Malaysian hospitals to determine the genetic relatedness of these isolates and their relationship with the endemic MRSA strains. In all, 32 of 640 MRSA isolates from 9 Malaysian hospitals were resistant to fusidic acid and rifampicin. Seven PFGE types (A, ZC, ZI, ZJ, ZK, ZL and ZM) were observed. The commonest type was type ZC, seen in 72% of isolates followed by type A, seen in 13%. Each of the other types (ZI, ZJ, ZK, ZL and ZM) was observed in a single isolate. Each type, even the commonest, was found in only one hospital. This suggests that the resistant strains had arisen from individual MRSA strains in each hospital and not as a result of the transmission of a common clone.

DNA fingerprinting of methicillin-resistant Staphylococcus aureus (MRSA) by pulsed-field gel electrophoresis (PFGE) in a teaching hospital in Malaysia.

Methicillin-resistant Staphylococcus aureus (MRSA) has been prevalent in our hospital over the last three years. Differentiation among MRSA strains by DNA typing in addition to antibiotic resistance pattern surveillance is crucial in order to implement infection control measures. The aim of this study was to characterize MRSA isolates from patients admitted to Hospital Universiti Kebangsaan Malaysia (HUKM) by phenotypic (analyses of antibiotic susceptibility pattern) and genotypic (PFGE) techniques to determine the genetic relatedness of the MRSA involved and to identify endemic clonal profiles of MRSA circulating in HUKM. Seventy one MRSA strains collected between January to March 2000 from patients from various wards in HUKM were tested for antimicrobial resistance and typed by pulsed-field gel electrophoresis (PFGE). Four major types of PFGE patterns were identified (A, B, C and D) among MRSA strains. Two predominant PFGE types were recognised, Type A (59.2%) and Type B (33.8%). Most of these strains were isolated from ICU, Surgical wards and Medical wards. MRSA strains with different PFGE patterns appeared to be widespread among wards. Strains with the same antibiotype could be of different PFGE types. Most of isolates were resistant to ciprofloxacin, erythromycin, gentamicin and penicillin. One isolate with a unique PFGE pattern Type D and susceptible to gentamicin was identified as a different clone. Some isolates obtained from the same patient showed different PFGE subtypes suggesting that these patients were infected/colonized with multiple MRSA strains. PFGE analysis suggests that MRSA strains with different PFGE types was propagated within our hospital. The relationship between antibiotic susceptibility and PFGE patterns was independent. The ability of PFGE technique in differentiating our MRSA strains make it a method of choice for investigating the source, transmission and spread of nosocomial MRSA infection, and thus an appropriate control programme can be implemented to prevent the spread of MRSA infection.