Multi-Laboratory Evaluation of a Lateral Flow Rapid Test for Detection of Amebic Liver Abscess.

Independent evaluations of XEh Rapid®, an IgG4-based rapid dipstick test, were performed to assess its diagnostic performance to detect amebic liver abscess (ALA) using 405 samples at seven laboratories in four countries. The test showed high diagnostic specificity (97-100%) when tested with samples from healthy individuals (n = 100) and patients with other diseases (n = 151). The diagnostic sensitivity was tested with a total of 154 samples, and the results were variable. It was high in three laboratories (89-94%), and moderate (72%) and low (38%) in two other laboratories. Challenges and issues faced in the evaluation process are discussed. Nevertheless, XEh Rapid is promising to be developed into a point-of-care test in particular for resource-limited settings, and thus merits further confirmation of its diagnostic sensitivity.

A holistic approach is needed to control the perpetual burden of soil-transmitted helminth infections among indigenous schoolchildren in Malaysia.

A cross-sectional survey was conducted among 1,142 Orang Ali schoolchildren in six states of Peninsular Malaysia to investigate the current prevalence and risk factors of STH infections. Faecal samples were examined using direct smear, formalin-ether sedimentation, Kato-Katz, and Harada-Mori methods. A pre-tested questionnaire was used to collect information on the demographic, socioeconomic, personal hygiene, and health status of the participants. Overall, 70.1% (95% CI = 67.4, 72.7) of the participants were infected with at least one of the STH species. The prevalence of Ascaris lumbricoides, Trichuris trichiura, and hookworm infections was 63.1%, 61.8% and 11.5%, respectively. Moderate-to-heavy STH infections accounted for 61.3% of the total infections. Univariate and logistic regression analyses revealed different sets of risk factors, with age (> 10 years) being the significant risk factor of all three STH species. Moreover, other species-specific risk factors were identified including being a member of the Senoi tribe, family size (≥ 7 members), school size (150-250 pupils), maternal unemployment, unimproved source of drinking water, lacking improved toilet in the house, inadequate WASH facilities at school, not washing hands before eating, and not washing fruits before eating; presence of domestic animals, and not wearing shoes when outside. The high prevalence of STH infections found in the study population exceeds the WHO policy intervention threshold (20% prevalence). Thus, an innovative holistic approach should be adopted to control STH infections among these children as part of the efforts to improve the quality of life of the entire Orang Asli population. .

Effect of Brugia pahangi co-infection with Plasmodium berghei ANKA in gerbils (Meriones unguiculatus).

Malaria and lymphatic filariasis (LF) are two leading and common mosquito-borne parasitic diseases worldwide. These two diseases are co-endemic in many tropical and sub-tropical regions and are known to share vectors. The interactions between malaria and filarial parasites are poorly understood. Thus, this study aimed at establishing the interactions that occur between Brugia pahangi and Plasmodium berghei ANKA (PbA) co-infection in gerbils. Briefly, the gerbils were matched according to age, sex, and weight and grouped into filarial-only infection, PbA-only infection, co-infection, and control group. The parasitemia, survival and clinical assessment of the gerbils were monitored for a period of 30 days post Plasmodium infection. The immune responses of gerbils to both mono and co-infection were monitored. Findings show that co-infected gerbils have higher survival rate than PbA-infected gerbils. Food and water consumption were significantly reduced in both PbA-infected and co-infected gerbils, although loss of body weight, hypothermia, and anemia were less severe in co-infected gerbils. Plasmodium-infected gerbils also suffered hypoglycemia, which was not observed in co-infected gerbils. Furthermore, gerbil cytokine responses to co-infection were significantly higher than PbA-only-infected gerbils, which is being suggested as a factor for their increased longevity. Co-infected gerbils had significantly elicited interleukin-4, interferon-gamma, and tumor necrotic factor at early stage of infection than PbA-infected gerbils. Findings from this study suggest that B. pahangi infection protect against severe anemia and hypoglycemia, which are manifestations of PbA infection.

Prevalence and risk factors of Strongyloides stercoralis infection among Orang Asli schoolchildren: new insights into the epidemiology, transmission and diagnosis of strongyloidiasis in Malaysia.

This cross-sectional study aimed to determine the prevalence and risk factors of S. stercoralis infection among 1142 Orang Asli primary schoolchildren in six different states of Peninsular Malaysia. Fecal samples were examined using direct smear, formalin-ether sedimentation (FES), agar plate culture (APC) and PCR techniques. Overall, 15.8% of the children were found to be infected with S. stercoralis. The prevalence was 0.2, 1.3, 15.2 and 13.7% by direct smear, FES, APC and PCR, respectively. Multivariate analysis showed that an age of >10 years, being male, belonging to a Proto-Malay tribe, belonging to the Senoi tribe, indiscriminate defecation, using an unimproved water source for drinking water and not wearing shoes when outside were the significant risk factors of infection among these children. In conclusion, we provide new evidence on the occurrence of S. stercoralis in Malaysia to show that there is a relatively high prevalence of infection among Orang Asli schoolchildren. Therefore, the use of specific methods for detecting S. stercoralis should be considered when screening these children for intestinal parasites. Moreover, prevention and control measures specific to S. stercoralis should be integrated into the intestinal parasitic infections control programme in Malaysia.

Molecular Diagnosis of Microsporidia among Immunocompromised Patients in Kuala Lumpur, Malaysia.

Microsporidia are obligate intracellular parasitic fungi causing chronic diarrhea, particularly among immunocompromised patients. The current method used for diagnosis is based on conventional microscopy, which does not differentiate parasites at the species level. The present study was carried out to identify microsporidian species in immunocompromised patients. From March 2016 to March 2017, a total of 289 archived stool samples were examined microscopically for microsporidian spores using Gram-chromotrope Kinyoun (GCK) stain. Positive stool samples by microscopy were subjected to polymerase chain reaction and sequencing for species identification. Based on microscopy examination, the overall prevalence of microsporidian infections was 32.2% (93/289; 95% CI = 27.1-37.8). Of these stool samples, 45 samples were successfully amplified and confirmed as Enterocytozoon bieneusi. No Encephalitozoon intestinalis was detected. Accurate identification of species might help clinicians to decide appropriate management strategies as dissemination risks and treatment response vary for different species, hence improving the management of microsporidian infections.

Laboratory Evaluation of a Rapid IgG4 Antibody Test (BLF Rapid™) for Bancroftian Filariasis.

At the end phase of the Global Programme to Eliminate Lymphatic Filariasis, antibody testing may have a role in decision-making for bancroftian filariasis-endemic areas. This study evaluated the diagnostic performance of BLF Rapid™, a prototype immunochromatographic IgG4-based test using BmSXP recombinant protein, for detection of bancroftian filariasis. The test was evaluated using 258 serum samples, comprising 96 samples tested at Universiti Sains Malaysia (in-house) and 162 samples tested independently at three international laboratories in the USA and India, and two laboratories in Malaysia. The independent testing involved 99 samples from Wuchereria bancrofti microfilaria or antigen positive individuals and 63 samples from people who were healthy or had other infections. The in-house evaluation showed 100% diagnostic sensitivity and specificity. The independent evaluations showed a diagnostic sensitivity of 84-100% and 100% specificity (excluding non-lymphatic filarial infections). BLF Rapid has potential as a surveillance diagnostic tool to make "Transmission Assessment Survey"-stopping decisions and conduct post-elimination surveillance.

Occurrence of Acanthamoeba genotypes in Central West Malaysian environments.

Acanthamoeba species are ubiquitous free-living protozoa that can be found worldwide. Occasionally, it can become parasitic and the causative agent of acanthamoebic keratitis (AK) and Granulomatous Amoebic Encephalitis (GAE) in man. A total of 160 environmental samples and 225 naturally-infected animal corneal swabs were collected for Acanthamoeba cultivation. Acanthamoeba was found to be high in samples collected from environments (85%, 136/160) compared to infected animal corneas (24.89%, 56/225) by microscopic examination. Analysis of nucleotide sequence of 18S rRNA gene of all the 192 cultivable Acanthamoeba isolates revealed 4 genotypes (T3, T4. T5 and T15) with T4 as the most prevalent (69.27%, 133/192) followed by T5 (20.31%), T15 (9.90%) and T3 (0.52%). Genotype T4 was from the strain of A. castellanii U07401 (44.27%), A. castellanii U07409 (20.83%) and A. polyphagaAY026243 (4.17%), but interestingly, only A. castellanii U07401 was detected in naturally infected corneal samples. In environmental samples, T4 was commonly detected in all samples including dry soil, dust, wet debris, wet soil and water. Among the T4, A. castellanii (U07409) strains were detected high occurrence in dry (45%) followed by aquatic (32.50%) and moist (22.50%) samples but however A. castellanii (U07401) strains were dominant in dry samples of soil and dust (93.10%). Subsequently, genotype T5 of A. lenticulata (U94741) strains were dominant in samples collected from aquatic environments (58.97%). In summary, A. castellanii (U07401) strains were found dominant in both environmental and corneal swab samples. Therefore, these strains are possibly the most virulent and dry soil or dusts are the most possible source of Acanthamoeba infection in cats and dogs corneas.

Pathogenesis of Plasmodium berghei ANKA infection in the gerbil (Meriones unguiculatus) as an experimental model for severe malaria.

BACKGROUND: As the quest to eradicate malaria continues, there remains a need to gain further understanding of the disease, particularly with regard to pathogenesis. This is facilitated, apart from in vitro and clinical studies, mainly via in vivo mouse model studies. However, there are few studies that have used gerbils (Meriones unguiculatus) as animal models. Thus, this study is aimed at characterizing the effects of Plasmodium berghei ANKA (PbA) infection in gerbils, as well as the underlying pathogenesis. METHODS: Gerbils, 5-7 weeks old were infected by PbA via intraperitoneal injection of 1 × 106 (0.2 mL) infected red blood cells. Parasitemia, weight gain/loss, hemoglobin concentration, red blood cell count and body temperature changes in both control and infected groups were monitored over a duration of 13 days. RNA was extracted from the brain, spleen and whole blood to assess the immune response to PbA infection. Organs including the brain, spleen, heart, liver, kidneys and lungs were removed aseptically for histopathology. RESULTS: Gerbils were susceptible to PbA infection, showing significant decreases in the hemoglobin concentration, RBC counts, body weights and body temperature, over the course of the infection. There were no neurological signs observed. Both pro-inflammatory (IFNγ and TNF) and anti-inflammatory (IL-10) cytokines were significantly elevated. Splenomegaly and hepatomegaly were also observed. PbA parasitized RBCs were observed in the organs, using routine light microscopy and in situ hybridization. CONCLUSION: Gerbils may serve as a good model for severe malaria to further understand its pathogenesis.

Case Report: A Symptomatic Case of Hymenolepis diminuta Infection in an Urban-Dwelling Adult in Malaysia.

A case of Hymenolepis diminuta infection in a 43-year-old Malaysian male with persistent abdominal colicky pain is reported. Endoscopy revealed whitish worms in the lumen of the small intestine, which were identified as H. diminuta after microscopy. Patient was successfully treated with a single dose of praziquantel (25 mg/kg).

Needs of exploring the burden of recent onset seizures due to neurocysticercosis and challenges in southeast Asia focusing on scenario in Malaysia.

Seizures due to neurocysticercosis (NCC) is a neglected human-to-human transmitted disorder and an emerging problem worldwide. A substantial portion of recent onset seizures is known to be attributed to NCC in Taenia solium (T. solium) endemic areas where populations which neither raise pigs nor eat pig meat are also at risk. High prevalence of NCC causing epilepsy has been reported in the underdeveloped areas of Southeast Asia (SEA) however, only fragmentary information on its incidence is available in countries like Malaysia. In Malaysia T. solium infection was previously thought to be infrequent due to Muslim population majority and the religious prohibition of consuming pork, but it is not totally absent. There is an evident lack of knowledge and awareness of the actual burden, routes of transmission, and the impact of NCC in this region. The problem is assumed to be more prevalent particularly in cities because of the frequent inflow of possibly T. solium infected individuals or carriers among those who migrate from neighboring endemic countries to Malaysia. The issue of imported cases that are likely to be emerging in Malaysia is highlighted here. An accurate quantification of regional burdens of epilepsy due to NCC in Malaysia is warranted considering the disease emergence in its neighboring countries. It is suggested that the importance of NCC be recognized through quantification of its burden, and also to collect epidemiological data for its subsequent elimination in line of World Health Organization's mission for control of cysticercosis as a neglected tropical disease. In this review the need as well as a strategy for neuro-care center screening of epilepsy cases, and various issues with possible explanations are discussed. It is also proposed that NCC be declared as a reportable disease which is one of the eradicable public health problems in SEA.

A community-based survey of Toxoplasma gondii infection among pregnant women in rural areas of Taiz governorate, Yemen: the risk of waterborne transmission.

BACKGROUND: Toxoplasma gondii is a zoonotic coccidian parasite causing morbidity and mortality. In Yemen, T. gondii infection has been reported among pregnant women seeking healthcare in the main cities. However, no data are available on the prevalence of T. gondii infection and its associated risk factors among pregnant women in the rural communities of the country. Thus, the present study aimed to determine the seroprevalence of T. gondii and identify its risk factors among pregnant women in the rural communities of Taiz governorate, Yemen. METHODS: A total of 359 pregnant women living in the rural communities of Taiz governorate were enrolled in this study by house-to-house visits. Data were collected using a pre-designed questionnaire, and blood samples were collected and tested for the detection of anti- T. gondii IgM and IgG antibodies by enzyme-linked immunosorbent assay. RESULTS: The prevalence of T. gondii infection among pregnant women in this study was 46.2% (166/359). Bivariate analysis identified the age of ≥ 30 years (odds ratio [OR] = 1.7; 95% confidence interval [CI] = 1.09-2.65, P = 0.019) and unimproved water sources (OR = 2.2; 95% CI = 1.10-4.55, P = 0.023) as factors associated with T. gondii infection among pregnant women. The multivariable analysis, however, identified unimproved water sources as an independent risk factor (adjusted OR = 2.4; 95% CI = 1.16-5.0, P = 0.018) associated with T. gondii infection among pregnant women. CONCLUSIONS: Pregnant women in the rural communities of Taiz, Yemen are at high risk of contracting T. gondii infection. Unimproved water sources (wells, water streams and water tanks) are significantly associated with T. gondii infection and should be considered in prevention and control strategies, especially among pregnant women.

Epidemiological Characteristics of Strongyloidiasis in Inhabitants of Indigenous Communities in Borneo Island, Malaysia.

Epidemiological study on strongyloidiasis in humans is currently lacking in Malaysia. Thus, a cross-sectional study was carried out to determine the prevalence of Strongyloides stercoralis infection among the inhabitants of longhouse indigenous communities in Sarawak. A single stool and blood sample were collected from each participant and subjected to microscopy, serological and molecular techniques. Five species of intestinal parasites were identified by stool microscopy. None of the stool samples were positive for S. stercoralis. However, 11% of 236 serum samples were seropositive for strongyloidiasis. Further confirmation using molecular technique on stool samples of the seropositive individuals successfully amplified 5 samples, suggesting current active infections. The prevalence was significantly higher in adult males and tended to increase with age. S. stercoralis should no longer be neglected in any intestinal parasitic survey. Combination of more than 1 diagnostic technique is necessary to increase the likelihood of estimating the 'true' prevalence of S. stercoralis.

Seroprevalence of Sparganosis in Rural Communities of Northern Tanzania.

In this study, the seroprevalence of sparganosis and its relationship with sociodemographic factors in northern Tanzania have been assessed. A total of 216 serum samples from two rural districts, Monduli and Babati, were tested for sparganosis using an enzyme-linked immunosorbent assay. The seroprevalence of anti-sparganum IgG antibodies was 62.5% (95% confidence interval [CI] = 56.1-68.9) in all age groups. There were significant associations between district (relative risk [RR] = 1.95, 95% CI = 1.42-2.69), education (RR = 1.40, 95% CI = 1.15-1.70), and pet ownership with seropositivity (RR = 1.48, 95% CI = 1.02-2.16) based on univariate analysis. However, only the district was significantly associated with seropositivity (odds ratio = 4.20, 95% CI = 1.89-9.32) in binary logistic regression analysis. Providing health education to people residing in sparganosis-endemic areas is likely to improve the efficacy of preventative measures and reduce human disease burden.

Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia.

Infections of humans with the zoonotic simian malaria parasite Plasmodium knowlesi occur throughout Southeast Asia, although most cases have occurred in Malaysia, where P. knowlesi is now the dominant malaria species. This apparently skewed distribution prompted an investigation of the phylogeography of this parasite in 2 geographically separated regions of Malaysia, Peninsular Malaysia and Malaysian Borneo. We investigated samples collected from humans and macaques in these regions. Haplotype network analyses of sequences from 2 P. knowlesi genes, type A small subunit ribosomal 18S RNA and cytochrome c oxidase subunit I, showed 2 genetically distinct divergent clusters, 1 from each of the 2 regions of Malaysia. We propose that these parasites represent 2 distinct P. knowlesi types that independently became zoonotic. These types would have evolved after the sea-level rise at the end of the last ice age, which separated Malaysian Borneo from Peninsular Malaysia.

Anisakiasis Causing Acute Dysentery in Malaysia.

Human anisakiasis is a zoonosis acquired by eating raw or undercooked infected seafood. Herein, we report a case of acute dysentery caused by anisakiasis in a 64-year-old man in Malaysia. A colonoscopy was performed and a nematode larva was found penetrating the mucosa of the ascending colon. Bleeding was observed at the site of penetration. Y-shaped lateral epidermal cords were seen from the cross section of the worm, which is a prominent feature of Anisakis larva. Molecular analysis using polymerase chain reaction of cytochrome oxidase 2 (cox2) gene confirmed the specimen to be larva of Anisakis simplex.

Molecular markers associated with resistance to commonly used antimalarial drugs among Plasmodium falciparum isolates from a malaria-endemic area in Taiz governorate-Yemen during the transmission season.

Since 2005, artesunate (AS) plus sulfadoxine/pyrimethamine (SP) combination has been adopted as the first-line treatment for uncomplicated malaria in Yemen in response to the high level of Plasmodium falciparum resistance to chloroquine (CQ). Therefore, the aim of the present study was to determine the frequency distribution of molecular markers associated with resistance to CQ and AS plus SP combination among P. falciparum isolates from a malaria-endemic area in Taiz governorate, Yemen. Fifty P. falciparum isolates were collected during a cross-sectional study in Mawza district, Taiz, in the period from October 2013 to April 2014. The isolates were investigated for drug resistance-associated molecular markers in five genes, including P. falciparum CQ resistance transporter (pfcrt) 76T and P. falciparum multidrug resistance 1 (pfmdr1) 86Y as markers of resistance to CQ, mutations in the Kelch 13 (K13) propeller domain for resistance to AS, and P. falciparum dihydrofolate reductase (pfdhfr) and P. falciparum dihydropteroate synthase (pfdhps) genes for resistance to SP. Nested polymerase chain reaction was used to amplify target genes in DNA extracts of the isolates followed by restriction fragment length polymorphism for detecting 76T and 86Y mutations in pfcrt and pfmdr1, respectively, and by DNA sequencing for detecting mutations in K13, pfdhfr and pfdhps. All the investigated isolates from Mawza district were harboring the pfcrt 76T mutant and the pfmdr1 N86 wild-type alleles. The pfdhfr 51I/108N double mutant allele was found in 2.2% (1/45) of the isolates; however, no mutations were detected at codons 436, 437, 540, 581 and 613 of pfdhps. All P. falciparum isolates that were successfully sequenced (n=47) showed the K13 Y493, R539, I543 and C580 wild-type alleles. In conclusion, the pfcrt 76T mutant allele is fixed in the study area about six years after the official withdrawal of CQ, possibly indicating its over-the-counter availability and continued use as a self-medication in the study area. However, the almost predominant wild-type alleles of the genes associated with resistance to AS and SP among P. falciparum isolates in the present study indicates the sustained efficacy of the currently adopted first-line treatment of AS plus SP in the study area.

First Case Report of Canthariasis in an Infant Caused by the Larvae of Lasioderma serricorne (Coleoptera: Anobiidae).

We report an unusual cause of gastrointestinal infection occurring in a 1-year-old infant patient who was brought to a public hospital in Kuala Lumpur, Malaysia. Larvae passed out in the patient's feces were confirmed by DNA barcoding as belonging to the species, Lasioderma serricorne (F.), known as the cigarette beetle. We postulate that the larvae were acquired from contaminated food and were responsible for gastrointestinal symptoms in the patient. To our knowledge, this the first report of human canthariasis caused by larvae of L. serricorne.

First Molecular Characterization of Leishmania Species Causing Visceral Leishmaniasis among Children in Yemen.

Visceral leishmaniasis (VL) is a debilitating, often fatal disease caused by Leishmania donovani complex; however, it is a neglected tropical disease. L. donovani complex comprises two closely related species, L. donovani that is mostly anthroponotic and L. infantum that is zoonotic. Differentiation between these two species is critical due to the differences in their epidemiology and pathology. However, they cannot be differentiated morphologically, and their speciation using isoenzyme-based methods poses a difficult task and may be unreliable. Molecular characterization is now the most reliable method to differentiate between them and to determine their phylogenetic relationships. The present study aims to characterize Leishmania species isolated from bone marrows of Yemeni pediatric patients using sequence analysis of the ribosomal internal transcribed spacer-1 (ITS1) gene. Out of 41 isolates from Giemsa-stained bone marrow smears, 25 isolates were successfully amplified by nested polymerase chain reaction and sequenced in both directions. Phylogenetic analysis using neighbor joining method placed all study isolates in one cluster with L. donovani complex (99% bootstrap). The analysis of ITS1 for microsatellite repeat numbers identified L. infantum in 11 isolates and L. donovani in 14 isolates. These data suggest the possibility of both anthroponotic and zoonotic transmission of VL-causing Leishmania species in Yemen. Exploring the possible animal reservoir hosts is therefore needed for effective control to be achieved.

Field evaluation of a PfHRP-2/pLDH rapid diagnostic test and light microscopy for diagnosis and screening of falciparum malaria during the peak seasonal transmission in an endemic area in Yemen.

BACKGROUND: Malaria is a public health threat in Yemen, with 149,451 cases being reported in 2013. Of these, Plasmodium falciparum represents 99%. Prompt diagnosis by light microscopy (LM) and rapid diagnostic tests (RTDs) is a key element in the national strategy of malaria control. The heterogeneous epidemiology of malaria in the country necessitates the field evaluation of the current diagnostic strategies, especially RDTs. Thus, the present study aimed to evaluate LM and an RDT, combining both P. falciparum histidine-rich protein-2 (PfHRP-2) and Plasmodium lactate dehydrogenase (pLDH), for falciparum malaria diagnosis and survey in a malaria-endemic area during the transmission season against nested polymerase chain reaction (PCR) as the reference method. METHODS: A household-based, cross-sectional malaria survey was conducted in Mawza District, a malaria-endemic area in Taiz governorate. A total of 488 participants were screened using LM and PfHRP-2/pLDH RDT. Positive samples (160) and randomly selected negative samples (52) by both RDT and LM were further analysed using 18S rRNA-based nested PCR. RESULTS: The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the RDT were 96.0% (95% confidence interval (CI): 90.9-98.3), 56.0% (95% CI: 44.7-66.8), 76.3% (95% CI: 69.0-82.3), and 90.4% (95% CI: 78.8-96.8), respectively. On the other hand, LM showed sensitivity of 37.6% (95% CI: 29.6-46.3), specificity of 97.6% (95% CI: 91.7-99.7), PPV of 95.9% (95% CI: 86.3-98.9), and NPV of 51.3% (95% CI: 43.2-59.2). The sensitivity of LM dropped to 8.5% for detecting asymptomatic malaria. Malaria prevalence was 32.8% (32.1 and 37.5% for ≥10 and <10 years, respectively) with the RDT compared with 10.7% (10.8 and 9.4% for age groups of ≥10 and <10 years, respectively) with LM. Among asymptomatic malaria individuals, LM and RDT-based prevalence rates were 1.6 and 25.6%, respectively. However, rates of 88.2 and 94.1% of infection with P. falciparum were found among patients who reported fever in the 48 h prior to the survey by LM and PfHRP-2/pLDH RDT, respectively. CONCLUSIONS: The PfHRP-2/pLDH RDT shows high sensitivity for the survey of falciparum malaria even for asymptomatic malaria cases. Although the RDT had high sensitivity, its high false-positivity rate limits its utility as a single diagnostic tool for clinical diagnosis of malaria. On the other hand, low sensitivity of LM indicates that a high proportion of malaria cases is missed, underestimating the true prevalence of malaria in the community. Higher NPV of PfHRP-2/pLDH RDT than LM can give a straightforward exclusion of malaria among febrile patients, helping to avoid unnecessary presumptive treatments.

Diagnosis of an imported Plasmodium ovale wallikeri infection in Malaysia.

Plasmodium ovale is rare and not exactly known to be autochthonous in Malaysia. There are two distinct forms of the parasite, namely P. ovale curtisi (classic form) and P. ovale wallikeri (variant form). Here, the first sequence confirmed case of an imported P. ovale wallikeri infection in Malaysia is presented. Microscopy found Plasmodium parasites with morphology similar to P. ovale or Plasmodium vivax in the blood films. Further confirmation using polymerase chain reaction (PCR) targeting the small-subunit rRNA gene of the parasite was unsuccessful. Genus-specific PCR was then performed and the product was sequenced and analysed. Sequence analyses confirmed the aetiological agent as P. ovale wallikeri. New species-specific primers (rOVA1v and rOVA2v) were employed and P. ovale wallikeri was finally confirmed. The findings highlight the need to look out for imported malaria infections in Malaysia and the importance of a constantly updated and validated diagnostic technique.