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Fulltext Human Plasmodium knowlesi infections in South-East Asian countries

Zaw MT, Lin Z

J Microbiol Immunol Infect, 2019 Oct;52(5):679-684.
PMID: 31320238 DOI: 10.1016/j.jmii.2019.05.012

Plasmodium knowlesi is now regarded as the fifth malaria parasite causing human malaria as it is widely distributed in South-East Asian countries especially east Malaysia where two Malaysian states namely Sabah and Sarawak are situated. In 2004, Polymerase Chain Reaction (PCR) was applied for diagnosing knowlesi malaria in the Kapit Division of Sarawak, Malaysia, so that human P. knowlesi infections could be detected correctly while blood film microscopy diagnosed incorrectly as Plasmodium malariae. This parasite is transmitted from simian hosts to humans via Anopheles vectors. Indonesia is the another country in South East Asia where knowlesi malaria is moderately prevalent. In the last decade, Sarawak and Sabah, the two states of east Malaysia became the target of P. knowlesi research due to prevalence of cases with occasional fatal infections. The host species of P. knowlesi are three macaque species namely Macaca fascicularis, Macaca nemestrina and Macaca leonina while the vector species are the Leucosphyrus Complex and the Dirus Complex of the Leucophyrus Group of Anopheles mosquitoes. Rapid diagnostic tests (RDT) are non-existent for knowlesi malaria although timely treatment is necessary for preventing complications, fatality and drug resistance. Development of RDT is essential in dealing with P. knowlesi infections in poor rural healthcare services. Genetic studies of the parasite on possibility of human-to-human transmission of P. knowlesi were recommended for further studies.
Fulltext Emergence of ST131 H30-Rx Subclones among Uropathogenic Escherichia coli Isolates from two large hospitals of Kota Kinabalu, Sabah, Malaysia

Yun, Mei Lai, Myo, Thura Zaw, Nor Amalina Emran, Lin, Zaw

Borneo Journal of Medical Sciences, 2017;11(3):19-24.
MyJurnal

Escherichia coli sequence type 131 (ST131) carries multiple drug resistance (MDR) genes as well as virulence genes. Drug resistant characteristics give a management problem to health care personnel. Four MDR Escherichia coli ST131 H30-Rx subclones were identified among 80 Uropathogenic E. coli (UPEC) isolates by using 4 allelic-specific Polymerase Chain Reactions (PCR) in two hospitals of Kota Kinabalu, Sabah, Malaysia. There is emergence of multidrug resistant E. coli in Kota Kinabalu.
Comparison of the Activities among Three SUL Genes Present in Uropathogenic Escherichia Coli

Yun, Mei Lai, Myo, Thura Zaw, Tin, Sabai Aung, Tin, Win, Lin, Zaw

Borneo Journal of Medical Sciences, 2019;13(1):17-22.
MyJurnal

The three plasmid borne alternative dihydropteroate synthetase (DHPS) genes namely sul1, sul2 and sul3 genes were heterologous in amino acid sequence and have about 40 – 50% identity. However, they have same DHPS activity with disc diffusion zone size of 6 mm with suphamethoxazole disc in our previous study. Sul1, sul2 and sul3 genes were observed in sulphamethoxazole resistant uropathogenic Escherichia coli (UPEC). In this study, all the three genes were cloned into E. coli host and minimum inhibitory concentration (MIC) was investigated for each sul gene to compare the activities of sul genes. The MIC values of E.coli containing sul2, sul1 and sul3 genes inserted recombinant plasmid were observed to have 18.5 mg/ml, 18 mg/ml and 17.5 mg/ml respectively as mean value of five experimental results. Although comparable MICs were obtained as a result, the MIC value was highest in E. coli carrying sul2 gene indicating that this DHPS enzyme activity of sul2 was strongest among three sul genes.
Fulltext Two sympatric types of Plasmodium ovale and discrimination by molecular methods

Zaw MT, Lin Z

J Microbiol Immunol Infect, 2017 Oct;50(5):559-564.
PMID: 28065415 DOI: 10.1016/j.jmii.2016.08.004

Plasmodium ovale is widely distributed in tropical countries, whereas it has not been reported in the Americas. It is not a problem globally because it is rarely detected by microscopy owing to low parasite density, which is a feature of clinical ovale malaria. P.o. curtisi and P.o. wallikeri are widespread in both Africa and Asia, and were known to be sympatric in many African countries and in southeast Asian countries. Small subunit ribosomal RNA (SSUrRNA) gene, cytochrome b (cytb) gene, and merozoite surface protein-1 (msp-1) gene were initially studied for molecular discrimination of P.o. curtisi and P.o. wallikeri using polymerase chain reaction (PCR) and DNA sequencing. DNA sequences of other genes from P. ovale in Southeast Asia and the southwestern Pacific regions were also targeted to differentiate the two sympatric types. In terms of clinical manifestations, P.o. wallikeri tended to produce higher parasitemia levels and more severe symptoms. To date, there have been a few studies that used the quantitative PCR method for discrimination of the two distinct P. ovale types. Conventional PCR with consequent DNA sequencing is the common method used to differentiate these two types. It is necessary to identify these two types because relapse periodicity, drug susceptibility, and mosquito species preference need to be studied to reduce ovale malaria. In this article, an easier method of molecular-level discrimination of P.o. curtisi and P.o. wallikeri is proposed.
Fulltext Importance of kelch 13 C580Y mutation in the studies of artemisinin resistance in Plasmodium falciparum in Greater Mekong Subregion

Zaw MT, Lin Z, Emran NA

J Microbiol Immunol Infect, 2020 Oct;53(5):676-681.
PMID: 31563454 DOI: 10.1016/j.jmii.2019.07.006

The mortality caused by Plasmodium falciparum was reduced by Artemisinin (ART) and ART combination therapy (ACT). However, Artemisinin resistance (ART-R) emerge during 2008 in Cambodia and spread to Greater Mekong Subregion (GMS). ART-R was confirmed not to spread to India, a gateway to whole Africa. The whole genome sequencing approach of P. falciparum assumed the k13 gene encoded Kelch protein was discovered to be associated with ART-R. Of the single nucleotide polymorphisms (SNPs) of k13 gene, C580Y mutant was commonly dominant in Cambodia, Myanmar, Thailand, Laos and Vietnam and assumed to be one of strong molecular markers for ART-R in P. falciparum isolates in GMS. Literatures published between 2017 and 2018 were reviewed in this work. F446I is observed to be doubtful molecular marker as ART-R marker. Transgenic experiment showed that parasite with F446I mutation displayed prolonged clearance in respond to ART while C580Y was applied as positive control mutant. Furthermore, study of C580Y allele in four countries Cambodia, Thailand, Laos resulted in single origin whereas the parasite with this allele showed multi-origin in three Provinces of Vietnam. As artemisinin was short acting drug, the role of long acting partner drug was studied by using transgenic C580Y mutant and C580 to leave recrudescent P. falciparum. Recently, there was treatment failure with ACT in some countries in GMS. In this review, the importance of C580Y mutation in the study of ART-R was discussed.
Fulltext Mutations inside rifampicin-resistance determining region of rpoB gene associated with rifampicin-resistance in Mycobacterium tuberculosis

Zaw MT, Emran NA, Lin Z

J Infect Public Health, 2018 04 26;11(5):605-610.
PMID: 29706316 DOI: 10.1016/j.jiph.2018.04.005

BACKGROUND: Rifampicin (RIF) plays a pivotal role in the treatment of tuberculosis due to its bactericidal effects. Because the action of RIF is on rpoB gene encoding RNA polymerase β subunit, 95% of RIF resistant mutations are present in rpoB gene. The majority of the mutations in rpoB gene are found within an 81bp RIF-resistance determining region (RRDR).
METHODOLOGY: Literatures on RIF resistant mutations published between 2010 and 2016 were thoroughly reviewed.
RESULTS: The most commonly mutated codons in RRDR of rpoB gene are 531, 526 and 516. The possibilities of absence of mutation in RRDR of rpoB gene in MDR-TB isolates in few studies was due to existence of other rare rpoB mutations outside RRDR or different mechanism of rifampicin resistance.
CONCLUSION: Molecular methods which can identify extensive mutations associated with multiple anti-tuberculous drugs are in urgent need so that the research on drug resistant mutations should be extended.
Fulltext Updates on k13 mutant alleles for artemisinin resistance in Plasmodium falciparum

Zaw MT, Emran NA, Lin Z

J Microbiol Immunol Infect, 2018 Apr;51(2):159-165.
PMID: 28711439 DOI: 10.1016/j.jmii.2017.06.009

BACKGROUND: In the fight against malaria caused by Plasmodium falciparum, the successes achieved by artemisinin were endangered by resistance of the parasites to the drug. Whole genome sequencing approach on artemisinin resistant parasite line discovered k13 gene associated with drug resistance. In vitro and in vivo studies indicated mutations in the k13 gene were linked to the artemisinin resistance.
METHODOLOGY: The literatures published after April, 2015 up to December, 2016 on k13 mutant alleles for artemisinin resistance in Plasmodium falciparum and relevant literatures were comprehensively reviewed.
RESULTS: To date, 13 non-synonymous mutations of k13 gene have been observed to have slow parasite clearance. Worldwide mapping of k13 mutant alleles have shown mutants associated with artemisinin resistance were confined to southeast Asia and China and did not invade to African countries. Although in vitro ring stage survival assay of 0-3 h was a recently developed assay, it was useful for rapid detection of artemisinin resistance associated k13 allelic marker in the parasite. Recently, dissemination of k13 mutant alleles was recommended to be investigated by identity of haplotypes. Significant characteristics of well described alleles in the reports were mentioned in this review for the benefit of future studies.
CONCLUSION: According to the updates in the review, it can be concluded artemisinin resistance does not disseminate to India and African countries within short period whereas regular tracking of these mutants is necessary.
Fulltext Polymerase chain reaction-restriction fragment length polymorphism method for differentiation of uropathogenic specific protein gene types

Lai YM, Zaw MT, Shamsudin SB, Lin Z

J Microbiol Immunol Infect, 2016 Aug;49(4):591-4.
PMID: 26212311 DOI: 10.1016/j.jmii.2015.06.002

The putative pathogenicity island (PAI) containing the uropathogenic specific protein (usp) gene and three small open reading frames (orfU1, orfU2, and orfU3) encoding 98, 97, and 96 amino acid proteins is widely distributed among uropathogenic Escherichia coli (UPEC) strains. This PAI was designated as PAIusp. Sequencing analysis of PAIusp has revealed that the usp gene can be divided into two types - uspI and uspII - based on sequence variation at the 3' terminal region and the number and position of orfUs differ from strain to strain. Based on usp gene types and orfU sequential patterns, PAIusp can be divided into four subtypes. Subtyping of PAIusp is a useful method to characterize UPEC strains. In this study, we developed a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method to differentiate usp gene types. This method could correctly identify the usp gene type in usp-positive UPEC strains in our laboratory.
Fulltext Evaluation of PAIusp subtyping to characterize uropathogenic E. coli isolates

Lai YM, Zaw MT, Shamsudin SB, Lin Z

J Infect Dev Ctries, 2016 Oct 31;10(10):1053-1058.
PMID: 27801366 DOI: 10.3855/jidc.6944

INTRODUCTION: Uropathogenic virulence factors have been identified by comparing the prevalence of these among urinary tract isolates and environmental strains. The uropathogenic-specific protein (USP) gene is present on the pathogenicity island (PAI) of uropathogenic Escherichia coli (UPEC) and, depending on its two diverse gene types and the sequential patterns of three open reading frame units (orfUs) following it, there is a method to characterize UPEC epidemiologically called PAIusp subtyping.
METHODOLOGY: A total of 162 UPEC isolates from Sabah, Malaysia, were tested for the presence of the usp gene and the sequential patterns of three orfUs following it using polymerase chain reaction (PCR). In addition, by means of triplex PCR, the prevalence of the usp gene was compared with other two VFs of UPEC, namely alpha hemolysin (α-hly) and cytotoxic necrotizing factor (cnf-1) genes encoding two toxins.
RESULTS: The results showed that the usp gene was found in 78.40% of UPEC isolates, indicating that its prevalence was comparable to that found in a previous study in Japan. The two or three orfUs were also associated with the usp gene in this study. All the PAIusp subtypes observed in Japan were present in this study, while subtype IIa was the most common in both studies. The usp gene was observed in a higher percentage of isolates when compared with α-hly and cnf-1 genes.
CONCLUSIONS: The findings in Japan and Sabah, East Malaysia, were similar, indicating that PAIusp subtyping is applicable to the characterization of UPEC strains epidemiologically elsewhere in the world.
Fulltext The study on the survival of Escherichia coli in water at room temperature

Aung TS, Masandid H, Oo KS, Lin Z, Rayaji SJ

Borneo Journal of Medical Sciences, 2016;10(1):14-18.
MyJurnal

Escherichia coli (E. coli) is one of the most frequent causes of many bacterial infections especially
gastroenteritis in developing countries. It is also used as an indicator for faecal pollution in the
surveillance of bacteriological quality of drinking water. This study was conducted to determine the
survival of E. coli in water at room temperature (27oC). E. coli which is cultured in Lactose Peptone Broth
was inoculated into 8 bottles each containing 10 millilitres of distilled water. They were kept at 27oC.
Starting from the day 1, ten-fold dilutions were made from each bottle number and E. coli count was
done from each dilutions by using pour plate method. The colony forming unit/ millilitre (CFU/ml) was
calculated. The same procedure was carried out from bottles number 2 to 8 from day 2 to day 8
consecutively. CFU/ml of E. coli in dilution 10-5was markedly decreased from 3.9 x 106
in day 1to 0 in
day 8. The findings suggest that if the water is contaminated with low number of E. coli, it can be
eliminated by keeping water at room temperature for only few days.
Fulltext Genetic diversity of toxigenic Vibrio cholerae O1 from Sabah, Malaysia 2015

Zaw MT, Emran NA, Ibrahim MY, Suleiman M, Awang Mohd TA, Yusuff AS, et al.

J Microbiol Immunol Infect, 2019 Aug;52(4):563-570.
PMID: 29428381 DOI: 10.1016/j.jmii.2018.01.003

BACKGROUND: Cholera is an important health problem in Sabah, a Malaysian state in northern Borneo; however, Vibrio cholerae in Sabah have never been characterized. Since 2002, serogroup O1 strains having the traits of both classical and El Tor biotype, designated as atypical El Tor biotype, have been increasingly reported as the cause of cholera worldwide. These variants are believed to produce clinically more severe disease like classical strains.
PURPOSE: The purpose of this study is to investigate the genetic diversity of V.cholerae in Sabah and whether V.cholerae in Sabah belong to atypical El Tor biotype.
METHODS: ERIC-PCR, a DNA fingerprinting method for bacterial pathogens based on the enterobacterial repetitive intergenic consensus sequence, was used to study the genetic diversity of 65 clinical V.cholerae O1 isolates from 3 districts (Kudat, Beluran, Sandakan) in Sabah and one environmental isolate from coastal sea water in Kudat district. In addition, we studied the biotype-specific genetic traits in these isolates to establish their biotype.
RESULTS: Different fingerprint patterns were seen in isolates from these three districts but one of the patterns was seen in more than one district. Clinical isolates and environmental isolate have different patterns. In addition, Sabah isolates harbor genetic traits specific to both classical biotype (ctxB-1, rstRCla) and El Tor biotype (rstRET, rstC, tcpAET, rtxC, VC2346).
CONCLUSION: This study revealed that V.cholerae in Sabah were genetically diverse and were atypical El Tor strains. Fingerprint patterns of these isolates will be useful in tracing the origin of this pathogen in the future.

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