No association between the Plasmodium vivax crt-o MS334 or In9pvcrt polymorphisms and chloroquine failure in a pre-elimination clinical cohort from Malaysia with a large clonal expansion

Increasing reports of resistance to a frontline malaria blood-stage treatment, chloroquine (CQ), raises concerns for the elimination of Plasmodium vivax. The absence of an effective molecular marker of CQ resistance in P. vivax greatly constrains surveillance of this emerging threat. A recent genetic cross between CQ sensitive (CQS) and CQ resistant (CQR) NIH-1993 strains of P. vivax linked a moderate CQR phenotype with two candidate markers in P. vivax CQ resistance transporter gene (pvcrt-o): MS334 and In9pvcrt. Longer TGAAGH motif lengths at MS334 were associated with CQ resistance, as were shorter motifs at the In9pvcrt locus. In this study, high-grade CQR clinical isolates of P. vivax from a low endemic setting in Malaysia were used to investigate the association between the MS334 and In9pvcrt variants and treatment efficacy. Among a total of 49 independent monoclonal P. vivax isolates assessed, high-quality MS334 and In9pvcrt sequences could be derived from 30 (61%) and 23 (47%), respectively. Five MS334 and six In9pvcrt alleles were observed, with allele frequencies ranging from 2 to 76% and 3 to 71%, respectively. None of the clinical isolates had the same variant as the NIH-1993 CQR strain, and none of the variants were associated with CQ treatment failure (all P > 0.05). Multi-locus genotypes (MLGs) at 9 neutral microsatellites revealed a predominant P. vivax strain (MLG6) accounting for 52% of Day 0 infections. The MLG6 strain comprised equal proportions of CQS and CQR infections. Our study reveals complexity in the genetic basis of CQ resistance in the Malaysian P. vivax pre-elimination setting and suggests that the proposed pvcrt-o MS334 and In9pvcrt markers are not reliable markers of CQ treatment efficacy in this setting. Further studies are needed in other endemic settings, applying hypothesis-free genome-wide approaches, and functional approaches to understand the biological impact of the TGAAGH repeats linked to CQ response in a cross are warranted to comprehend and track CQR P. vivax

New leaf- and litter-dwelling species of the genus Pholcus from Southeast Asia (Araneae, Pholcidae)

We describe eight new species of the genus Pholcus, and document their microhabitats. Four species are assigned to the previously described Pholcus ethagala group: P. tanahrata Huber sp. nov., P. uludong Huber sp. nov., and P. bukittimah Huber sp. nov. from the Malay Peninsula, and P. barisan Huber sp. nov. from Sumatra. These species are all litter-dwellers that build domed sheet webs on the undersides of large dead leaves on the ground. The other four species are assigned to newly created species groups: the P. tambunan group with two species from northern Borneo: P. tambunan Huber sp. nov. and P. bario Huber sp. nov.; and the P. domingo group with two species from the Philippines, Mindanao: P. domingo Huber sp. nov. and P. matutum Huber sp. nov. These latter four species are leafdwellers that build barely visible silk platforms tightly attached to the undersides of live leaves. The main rationale for this paper is to provide part of the taxonomic and natural history background for upcoming phylogenetic and evolutionary (microhabitat shifts) analyses

Figs 48–53. Pholcus ledang Huber, 2011 in New leaf- and litter-dwelling species of the genus Pholcus from Southeast Asia (Araneae, Pholcidae)

Figs 48–53. Pholcus ledang Huber, 2011, SEM micrographs (ZFMK, Ar 15704–05). 48–49. Male and female prosomata, frontal views. 50. Comb hairs on male tarsus 4. 51–52. Right appendix and embolus, prolateral and distal views. 53. Female ALS. Scale bars: 48 = 300 µm; 49 = 200 µm; 50 = 10 µm; 51–52 = 100 µm; 53 = 20 µm

Figs 32–38. Live specimens. 32–35. Pholcus gombak Huber, 2011 in New leaf- and litter-dwelling species of the genus Pholcus from Southeast Asia (Araneae, Pholcidae)

Figs 32–38. Live specimens. 32–35. Pholcus gombak Huber, 2011, Kemensah (32) and Gunung Liang (33–35), ♂, ♀ with parasitized egg-sac seven days before eclosion of wasps (33), one day before eclosion (34), and at eclosion (35). 36–38. P. ledang Huber, 2011, Gunung Ledang, ♂ and ♀ with egg-sac

Ectoparasites of murids in peninsular Malaysia and their associated diseases

Abstract A considerable number of rat-borne ectoparasite studies have been conducted since the early 1930s in the Malayan Peninsula (now known as peninsular Malaysia). The majority of studies were field surveys and collections of specimens across the region, and were conducted primarily to catalogue the ectoparasite host distribution and discover novel species. This has generated a signification amount of information, particularly on the diversity and host distribution; other aspects such as morphology, host distribution and medical significance have also been investigated. Amongst the four main groups (mites, fleas, ticks, lice), rat-borne mites have received the most attention with a particular emphasis on chiggers, due to their medical importance. More recent studies have examined the distribution of ectoparasites in rats from different habitat type simplicating a high prevalence of zoonotic species infesting rat populations. Despite being capable of transmitting dangerous pathogens to human, the health risks of rat-borne ectoparasites appear to be small with no serious outbreaks of diseases recorded. Although an extensive number of works have been published, there remain gaps in knowledge that need to be addressed, such as, the distribution of under studied ectoparasite groups (listrophorids and myobiids), determining factors influencing infestation, and understanding changes to the population distribution over time

Figs 39–47. Pholcus gombak Huber, 2011 in New leaf- and litter-dwelling species of the genus Pholcus from Southeast Asia (Araneae, Pholcidae)

Figs 39–47. Pholcus gombak Huber, 2011, SEM micrographs (ZFMK, Ar 15701–02). 39–40. Male and female prosomata, frontal views. 41. Left procursus; asterisk marks large prolatero-dorsal process. 42– 43. Right appendix and embolus, prolateral and prolatero-distal views. 44. Female ALS. 45. Cuticular processes on whitish retrolateral area of left procursus. 46. Male gonopore. 47. Epigynum, ventral view. Scale bars: 39–40 = 300 µm; 41, 47 = 200 µm; 42, 43 = 100 µm; 44 = 20 µm; 45 = 10 µm; 46 = 30 µm

Is There a Risk of Suburban Transmission of Malaria in Selangor, Malaysia?

Background: The suburban transmission of malaria in Selangor, Malaysia's most developed and populous state still remains a concern for public health in this region. Despite much successful control efforts directed at its reduction, sporadic cases, mostly brought in by foreigners have continued to occur. In addition, cases of simian malaria caused by Plasmodium knowlesi, some with fatal outcome have caused grave concern to health workers. The aim of this study was to investigate the possibility of local malaria transmission in suburban regions of Selangor, which are adjacent to secondary rainforests.Findings: A malaria survey spanning 7 years (2006 - 2012) was conducted in Selangor. A total of 1623 laboratory confirmed malaria cases were reported from Selangor's nine districts. While 72.6% of these cases (1178/1623) were attributed to imported malaria (cases originating from other countries), 25.5% (414/1623) were local cases and 1.9% (31/1623) were considered as relapse and unclassified cases combined. In this study, the most prevalent infection was P. vivax (1239 cases, prevalence 76.3%) followed by P. falciparum (211, 13.0%), P. knowlesi (75, 4.6%), P. malariae (71, 4.4%) and P. ovale (1, 0.06%). Mixed infections comprising of P. vivax and P. falciparum were confirmed (26, 1.6%). Entomological surveys targeting the residences of malaria patients' showed that the most commonly trapped Anopheles species was An. maculatus. No oocysts or sporozoites were found in the An. maculatus collected. Nevertheless, the possibility of An. maculatus being the malaria vector in the investigated locations was high due to its persistent occurrence in these areas.Conclusions: Malaria cases reported in this study were mostly imported cases. However the co-existence of local cases and potential Plasmodium spp. vectors should be cause for concern. The results of this survey reflect the need of maintaining closely monitored malaria control programs and continuous extensive malaria surveillance in Peninsula Malaysia

Malaria case classification in Selangor (2006-2012).

<p>Malaria cases were classified as either local, imported, relapsed cases or unclassified/unknown. Classification was based on patient history. Cases of patients residing in Selangor for 2 weeks or less would be classified as imported. As shown above, there is an increase of local cases from 2006 to 2012 (except 2010), with the highest number of local cases (116) being reported in 2011. Imported cases, though decreasing from 2007, still remain higher than local cases with the highest number of imported cases (300) being reported in 2007. By 2012, there is an almost equal percentage of imported and local cases. Relapsed cases were also reported in all years except 2007 and 2012, with 17 cases being the highest in 2010. In all relapsed cases, <i>Plasmodium </i><i>vivax</i> was the infective agent. Some cases were denoted as unclassified due to lack of data. </p

Map of the study area showing the distribution of <i>Anopheles</i> species mosquitoes, macaque monkeys, and <i>P.knowlesi</i> malaria cases.

<p>Most of Selangor districts are exposed to <i>Anopheles </i><i>species</i> malaria vectors and <i>Macaca </i><i>fascicularis</i>. Only Hulu Selangor has reported <i>P. knowlesi</i> cases according to vector lab records.</p

Maps of confirmed reported malaria cases by district, 2006 to 2012.

<p>Malaria endemicity showing the distribution of malaria cases, as indicated by district-level estimates based on available survey data from the Klang Vector Laboratory, Selangor, Department of Health. The districts were generated into shaded maps, in which the values for the cases are represented in shades from light (low cases) to dark (high cases). (<b>A</b>) Map showing cases that originated from outside the state of Selangor (Imported cases) (<b>B</b>). Map showing malaria cases that originated locally within the State of Selangor (Local cases).</p