Siti Nursheena Mohd Zain

Pathogenic and Saprophytic Leptospira Species in Water and Soils from Selected Urban Sites in Peninsular Malaysia

Leptospira species were studied in water and soils from selected urban sites in Malaysia. A total of 151 water (n=121) and soil (n=30) samples were collected from 12 recreational lakes and wet markets. All samples were filtered and inoculated into semi-solid Ellinghausen and McCullough modified by Johnson and Harris (EMJH) media supplemented with additional 5-fluorouracil. The cultures were then incubated at 30°C and observed under a dark field microscope with intervals of 10 days. A PCR assay targeting the rrs gene was used to confirm the genus Leptospira among the isolates. Subsequently, the pathogenic status of the isolates was determined using primer sets G1/G2 and Sapro1/Sapro2, which target the secY and rrs genes, respectively. The isolates were identified at serogroup level using the microscopic agglutination test (MAT) while their genetic diversity was assessed by pulsed field gel electrophoresis (PFGE). Based on dark field microscopy, 23.1% (28/121) water and 23.3% (7/30) soil cultures were positive for Leptospira spp. Of the 35 positive cultures, only 8 were pure and confirmed as Leptospira genus by PCR assay. Two out of 8 isolates were confirmed as pathogenic, 5 were saprophytic and one was intermediate. These 8 isolates were negative for the 25 reference hyperimmune rabbit sera tested in the MAT. PFGE showed that all 8 of these environmental Leptospira spp. were genetically diverse. In conclusion, the presence of pathogenic Leptospira spp. in the urban Malaysian environment may indicate and highlight the importance of water screening, especially in recreational lakes, in order to minimize any chance of Leptospira infection.

Helminth communities from two urban rat populations in Kuala Lumpur, Malaysia.

BackgroundThe prevalence of parasitic infections among commensal animals such as black and brown rats in many tropical countries is high and in comparison with studies on rodents in temperate climates, little is known about the community structure of their parasites. Rodent borne parasites pose threats to human health since people living in close proximity to rodent populations can be exposed to infection.MethodsThe helminth community structures of two urban rat populations in Kuala Lumpur, Malaysia were investigated. The rats were from two contrasting sites in the city caught over a period of 21 months in 2000-2002.ResultsEleven species of helminth parasites comprising seven nematodes (Heterakis spumosum, Mastophorus muris, Nippostrongylus brasiliensis, Syphacia muris, Pterygodermatites tani/whartoni, Gongylonema neoplasticum, Angiostrongylus malaysiensis), three cestodes (Hymenolepis (Rodentolepis) nana, H. diminuta and Taenia taeniaeformis) and one acanthocephalan (Moniliformis moniliformis) were recovered from 346 Rattus rattus and 104 R. norvegicus from two urban sites, Bangsar and Chow Kit, during 2000-2002. Rattus rattus harboured over 60% of all helminths compared with R. norvegicus, although both host species played a dominant role in the different sites with, for example R. norvegicus at Bangsar and R. rattus at Chow Kit accounting for most of the nematodes. Overall 80% of rats carried at least one species of helminth, with the highest prevalences being shown by H. diminuta (35%), H. spumosum (29.8%) and H. nana (28.4%). Nevertheless, there were marked differences in prevalence rates between sites and hosts. The influence of extrinsic (year, season and site) and intrinsic (species, sex and age) factors affecting infracommunity structure (abundance and prevalence of infection) and measures of component community structure were analyzed.ConclusionsSince at least two species of rat borne helminths in Kuala Lumpur have the potential to infect humans, and these showed high prevalences in the rats, the assessment and regular monitoring of infections carried by wild rodents have important roles to play in public health.

Isolation and molecular characterization of leptospira interrogans and leptospira borgpetersenii isolates from the urban rat populations of Kuala Lumpur, Malaysia

Rats are considered the principal maintenance hosts of Leptospira. The objectives of this study were isolation and identification of Leptospira serovars circulating among urban rat populations in Kuala Lumpur. Three hundred urban rats (73% Rattus rattus and 27% R. norvegicus) from three different sites were trapped. Twenty cultures were positive for Leptospira using dark-field microscopy. R. rattus was the dominant carrier (70%). Polymerase chain reaction (PCR) confirmed that all isolates were pathogenic Leptospira species. Two Leptospira serogroups, Javanica and Bataviae, were identified using microscopic agglutination test (MAT). Pulsed-field gel electrophoresis (PFGE) identified two serovars in the urban rat populations: L. borgpetersenii serovar Javanica (85%) and L. interrogans serovar Bataviae (15%). We conclude that these two serovars are the major serovars circulating among the urban rat populations in Kuala Lumpur. Despite the low infection rate reported, the high pathogenicity of these serovars raises concern of public health risks caused by rodent transmission of leptospirosis.

Macroparasite communities in stray cat populations from urban cities in Peninsular Malaysia

The occurrence of macroparasites was studied from 543 stray cats in four urban cities from the west (Kuala Lumpur), east (Kuantan), north (Georgetown) and south (Malacca) of Peninsular Malaysia from May 2007 to August 2010. Five ectoparasites species were recovered namely, Ctenocephalides felis, Felicola subrostratus, Haemaphysalis bispinosa, Heterodoxus spiniger and Lynxacarus radovskyi. Two cats from Georgetown were infested with the dog louse, H. spiniger and this represented the first host record for this species in Malaysia. Up to nine species of helminths were recovered with overall high prevalences of infection of 83% in Kuantan, followed by 75.1% in Kuala Lumpur, 71.6% in Georgetown and 68% in Malacca. The helminth species comprised five nematodes, Toxocara malaysiensis, Toxocara cati, Ancylostoma braziliensis, Ancylostoma ceylanicum, Physaloptera praeputialis, two cestodes Taenia taeniaeformis, Dipylidium caninum and one trematode, Playtnosomum fastosum. The majority of helminths were present in the four study sites except for the absence of P. praeputialis in Kuala Lumpur. The prevalence and abundance of infections were analysed taking intrinsic (host age and sex) and extrinsic (season) factors into consideration. Levels of infection and infestation were mainly influenced by host age and to a lesser extent sex and season, whereas four nematode species exhibited significant interactions within the intestine of the cat host. The potential for transmission of some macroparasite species from stray cats to the human population in urban areas is discussed.

Human Leptospirosis in Malaysia: Reviewing the Challenges After 8 Decades (1925-2012).

The history and epidemiology of human leptospirosis in Malaysia from 1925 to 2012 are described. Previous studies have demonstrated that leptospirosis is an endemic disease in Malaysia occurring in both urban and rural locations. The number of cases has risen dramatically since the Ministry of Health Malaysia highlighted leptospirosis as a notifiable disease in 2010, with reported cases increasing from 248 cases in 2004 to 3604 in 2012. The incidence of infection among the population suggests that occupation, sex, age, ethnic background, water recreational activities, and sporting events are risk factors. A robust surveillance system is now in place to monitor temporal and spatial changes in the incidence and prevalence of infection and to identify risk areas and disease behavior. Despite extensive studies over the past decade, there is a still a need to describe local serovars in host carriers and the human population, with the view to develop an effective vaccine against leptospirosis.

Epidemiology of human leptospirosis in Malaysia, 2004-2012

Leptospirosis is an emerging disease, especially in countries with a tropical climate such as Malaysia. A dramatic increase in the number of cases has been reported over the last decade; however, information on the epidemiological trends of this disease is lacking. The objective of this study is to provide an epidemiological description of human leptospirosis cases over a 9-year period (2004-2012) and disease relationship with meteorological, geographical, and demographical information. A retrospective study was undertaken to describe the patterns of human leptospirosis cases and their association with intrinsic (sex, age, and ethnicity) and extrinsic (location, rainfall, and temperature) factors. Data was grouped according to age, sex, ethnicity, seasonality and geographical distribution, and analyzed using statistical tools to understand the influence of all the different factors on disease incidence. A total of 12,325 cases of leptospirosis were reported between 2004 and 2012 with an upward trend in disease incidence, with the highest in 2012. Three hundred thirty-eight deaths were reported with an overall case fatality rate of 2.74%, with higher incidence in males (9696; 78.7%) compared with female patients (2629; 21.3%), and overall male to female ratio of 3.69:1. Patients aged cohorts between 30-39 years old (16.22 per 100,000 population) had the highest disease incidence while the lowest incidence occurred between <1 to 9 years old (3.44 per 100,000 population). The average incidence was highest amongst Malays (10.97 per 100,000 population), followed by Indians (7.95 per 100,000 population). Stratification according to geographical distribution showed that the state of Malacca had the highest average disease incidence (11.12 per 100,000 population) followed by Pahang (10.08 per 100,000 population). The states of Terengganu, Kelantan, and Perak recorded similar rates of incidence (≈8.00 per 100,000 population), while Johor with the least number of reported cases (1.80 per 100,000 population). Positive relationships were recorded between the number of reported cases with the number of raining days per month and monthly average temperature (p-value<0.05). However, no significant association was noted between rainfall volume and number of reported Leptospirosis cases. This collaborative efforts between medical, academic and governmental institutions has enabled the construction of this comprehensive database that is essential to understand the disease trends in Malaysia and add insights into the prevention and control of this disease.

Migrant Workers in Malaysia: Current Implications of Sociodemographic and Environmental Characteristics in the Transmission of Intestinal Parasitic Infections.

A cross-sectional study of intestinal parasitic infections amongst migrant workers in Malaysia was conducted. A total of 388 workers were recruited from five sectors including manufacturing, construction, plantation, domestic and food services. The majority were recruited from Indonesia (n = 167, 43.3%), followed by Nepal (n = 81, 20.9%), Bangladesh (n = 70, 18%), India (n = 47, 12.1%) and Myanmar (n = 23, 5.9.2%). A total of four nematode species (Ascaris lumbricoides, Trichuris trichiura, Enterobius vermicularis and hookworms), one cestode (Hymenolepis nana) and three protozoan species (Entamoeba histolytica/dispar, Giardia sp. and Cryptosporidium spp.) were identified. High prevalence of infections with A. lumbricoides (43.3%) was recorded followed by hookworms (13.1%), E. histolytica/dispar (11.6%), Giardia sp. (10.8%), T. trichura (9.5%), Cryptosporodium spp. (3.1%), H. nana (1.8%) and E. vermicularis (0.5%). Infections were significantly influenced by socio-demographic (nationality), and environmental characteristics (length of working years in the country, employment sector and educational level). Up to 84.0% of migrant workers from Nepal and 83.0% from India were infected with intestinal parasites, with the ascarid nematode A. lumbricoides occurring in 72.8% of the Nepalese and 68.1% of the Indian population. In addition, workers with an employment history of less than a year or newly arrived in Malaysia were most likely to show high levels of infection as prevalence of workers infected with A. lumbricoides was reduced from 58.2% to 35.4% following a year’s residence. These findings suggest that improvement is warranted in public health and should include mandatory medical screening upon entry into the country.

Antimicrobial susceptibility of Leptospira spp. isolated from environmental, human and animal sources in Malaysia

Leptospirosis is a zoonosis with worldwide distribution caused by pathogenic spirochetes of the genus Leptospira. The aim of this study was to evaluate the susceptibility of isolates obtained from different hosts. A total of 65 Leptospira isolates from humans (n = 1), zoonoses (rat, n = 60; dog, n = 1; swine, n = 1) and environment (n = 2) were tested against six antibiotics. All the isolates were resistant to trimethoprim and sulphamethoxazole and had high MIC toward chloramphenicol (MIC90: 6.25 μg/ml). All except one environment isolate were sensitive to ampicillin, doxycycline and penicillin G.

Isolation and Molecular Identification of Bartonellae from Wild Rats (Rattus Species) in Malaysia

This study describes our investigation on the prevalence and molecular identification of bartonellae from Rattus diardii and R. norvegicus in the urban areas of Malaysia. Of 95 rats investigated, Bartonella tribocorum, B. rattimassiliensis, B. coopersplainsensis, B. elizabethae, and B. queenslandensis were isolated from kidney and spleen homogenates of four rats. Bartonellae DNA was amplified from the rat organ tissues by using primers specific for the bartonellae RNA polymerase beta subunit (rpoB) gene in nine other rats. Sequence analysis of the rpoB gene fragments shows the identification of B. queenslandensis in five rats, B. elizabethae in three rats, and B. tribocorum in one rat. Combining the results of isolation and molecular detection of bartonellae, we found that the prevalence of Bartonella infection in the Rattus spp. investigated in this study was 13.7%. Implementation of effective rat control program in the urban areas is necessary to prevent the spillover of bartonellosis from rats to humans.

Characterization of Nontyphoidal Salmonella Isolates from Asymptomatic Migrant Food Handlers in Peninsular Malaysia

Asymptomatic Salmonella carriers who work as food handlers pose food safety and public health risks, particularly during food preparation, and this has serious implications for the disease burden in society. Therefore, we conducted a study to determine the number of Salmonella carriers in a migrant cohort in several food establishments in three major cities in Peninsular Malaysia. Sociodemographic data and stool samples were collected and analyzed using standard methods of detection and isolation. Antimicrobial susceptibility tests of the positive samples were also performed. A total of 317 migrant food handlers, originating from South and Southeast Asian countries, were recruited voluntarily. Nine (2.8%) stool samples were confirmed to be Salmonella positive. PCR serotyping and pulsed-field gel electrophoresis identified four serotypes as Typhimurium (n = 3), Corvallis (n = 2), Hadar (n = 1), Agona (n = 1) and two unknown serovars. Antimicrobial susceptibility tests revealed that all nine isolates were susceptible to amoxicillin-clavulanic acid, cefotaxime, ceftazidime, ceftriaxone, and gentamycin. However, seven isolates were found to be multidrug resistant to ampicillin, chloramphenicol, trimethoprim-sulfamethoxazole, sulfonamides, streptomycin, and tetracycline. This study highlights that carriers of nontyphoidal Salmonella exist among migrant food handlers, which poses a health risk to consumers through food contamination. Our results indicate a need for authorities to enhance food safety awareness in the migrant workers and to reevaluate current health screening methods to include preventive measure such as mandatory stool screening as part of the preemployment and routine health examinations.