Due to the growing public health and tourism awareness, Cimex hemipterus Fabricius (Hemiptera: Cimicidae) has gained a great interest in increasing reported infestation cases in tropical regions of the world, including Malaysia. Since the information on the molecular ecology and population biology of this species are tremendously lacking, the isolation and development of molecular markers can be used to determine its genetic structure. In this study, novel microsatellite primers isolated from enriched genomic libraries of C. hemipterus were developed using 454 Roche shotgun sequencing. Seven validated polymorphic microsatellite primers were consistently amplified and characterized from 70 tropical bed bugs collected from seven locations throughout Malaysia. The number of alleles per locus identified ranged from 6 to 14. Comparison of loci for overall and between population were done with mean observed and expected heterozygosity were determined at 0.320 and 0.814, 0.320 and 0.727, respectively. Polymorphic information criteria (PIC) valued the markers as highly informative as PIC >0.5. Overall population, they are possibly in Hardy-Weinberg equilibrium with loci Ch_09ttn, Ch_01dn, and Ch_13dn showing signs of a null allele. There were no scoring errors caused by stutter peaks, no large allele dropout was detected for all loci and showed no evidence of linkage disequilibrium. In conclusion, all seven molecular microsatellite markers identified can be beneficially used to gain more information on the population genetic structure and breeding patterns of C. hemipterus as well as the relationship of dispersal and infestation.
Behavioral responses of Aedes aegypti male populations developed for Release of Insects Carrying a Dominant Lethal (RIDL) technology and a Malaysian wild-type population of two age groups (4-5 and 8-10 d old) were tested under laboratory conditions against chemical irritants and repellents using the high-throughput screening system device. Results indicate that all male Ae. aegypti test populations showed significant (P < 0.01) behavioral escape responses when exposed to alphacypermethrin, DDT, and deltamethrin at the test dose of 25 nmol/cm2. In addition, all populations showed significant (P < 0.05) spatial repellent responses to DDT, whereas alphacypermethrin and deltamethrin elicited no directional movement in the assay. These data suggest that genetic modification has not suppressed expected irritancy and repellency behavior. Age effects were minimal in both contact irritant and spatial repellent assays. The magnitude of irritant response, based on percentage responding, was stronger in the RIDL test cohorts as compared with the wild-type Malaysian population, but the impact, if any, that this increased behavioral sensitivity might have on the success of a RIDL strategy has yet to be defined. Information of the type reported in the current study is vital in defining the effects of genetic modification on vector behavior and understanding how these behaviors may influence the success of RIDL technology as they relate to other vector control interventions implemented in the same disease-endemic locale.
We report a case of human intestinal myiasis in a 41-yr-old female patient presented at a clinic in Seri Kembangan, Selangor, Malaysia. Larvae passed out in the patient's feces were sent to the Department of Parasitology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia. DNA barcoding confirmed the second case of intestinal myiasis in Malaysia involving the larvae of Clogmia albipunctatus (Duckhouse) (Diptera: Psychodidae). We review reported cases of myiasis and discuss the present case of intestinal myiasis in an urban patient.
The potential of integrating the mycoinsecticide, Metarhizium anisopliae (Met.), into house fly control programs is tremendous. However, the interaction between the fungus and insecticide, when applied at poultry farms, remains poorly understood. This study investigated the interaction between M. anisopliae and two selected insecticides, cyromazine and ChCy (a mixture of chlorpyrifos and cypemethrin), with three objectives: to assess the compatibility of M. anisopliae and the insecticides by measuring fungal vegetative growth and conidia production in the presence of insecticides; to evaluate the effect of M. anisopliae on these insecticides by analyzing insecticidal residue using ultra performance liquid chromatography; and to study the synergistic effects of M. anisopliae and the insecticides by applying sublethal concentrations of insecticides with M. anisopliae to house fly larvae. Metarhizium anisopliae was more tolerant to ChCy than to cyromazine, as M. anisopliae showed significantly more growth when grown with this insecticide. The M. anisopliae + ChCy combination resulted in significantly less chlorpyrifos residues compared to the ChCy plate, and 62-72% house fly larva mortality occurred when M. anisopliae and sublethal concentrations of ChCy were combined, implicating synergistic effects of the fungus with low concentrations of ChCy. Integrating M. anisopliae with compatible chemical at right concentration is crucial for poultry farm house fly control programs.
To advance our limited knowledge of global mosquito biogeography, we analyzed country occurrence records from the Systematic Catalog of the Culicidae (http://www.mosquitocatalog. org/main.asp), and we present world maps of species richness and endemism. A latitudinal biodiversity gradient was observed, with species richness increasing toward the equator. A linear log-log species (y)-area (x) relationship (SAR) was found that we used to compare observed and expected species densities for each country. Brazil, Indonesia, Malaysia, and Thailand had the highest numbers of species, and Brazil also had the highest taxonomic output and number of type locations. Brazil, Australia, the Philippines, and Indonesia had the highest numbers of endemic species, but excluding small island countries, Panama, French Guiana, Malaysia, and Costa Rica had the highest densities of total species and endemic species. Globally, 50% of mosquito species are endemic. Island countries had higher total number of species and higher number of endemic species than mainland countries of similar size, but the slope of the SAR was similar for island and mainland countries. Islands also had higher numbers of publications and type locations, possibly due to greater sampling effort and/or species endemism on islands. The taxonomic output was lowest for some countries in Africa and the Middle East. A consideration of country estimates of past sampling effort and species richness and endemism is proposed to guide mosquito biodiversity surveys. For species groups, we show that the number of species of Anopheles subgenus Anopheles varies with those of subgenus Cellia in a consistent manner between countries depending on the region. This pattern is discussed in relation to hypotheses about the historical biogeography and ecology of this medically important genus. Spatial analysis of country species records offers new insight into global patterns of mosquito biodiversity and survey history.
The effects of four temperatures (15, 23.5, 28, and 35 degrees C) on the biological characteristics of the rove beetle Paederus fuscipes Curtis were studied, and its cuticular permeability also was measured. Specimens successfully developed to adulthood at each temperature tested, but development time of each preadult stage significantly decreased with increasing temperature. Both egg and L1 stages required at least 80 degree days above a threshold of approximately 10 degrees C to develop to the subsequent stage. The lengthy development time and high survival rate of preadults at 15 degrees C suggests that P. fuscipes can survive in a harsh environment during cold weather by hibernating, and this ability could allow preadults to succeed ecologically in temperate countries. However, adult longevity was short, and no fecundity was recorded at 15 degrees C. At 28 degrees C, P. fuscipes exhibited a high survival rate of adults, which had a longer life span and high fecundity; thus, the population had the highest intrinsic rate of increase (0.0788 +/- 0.0051 d(-1)) and the shortest mean generation time (48.57 +/- 1.43 d) at 28 degrees C. At this temperature, the population might reach a size that could facilitate invasion into residential areas. However, in the absence of a hygric environment, P. fuscipes was unable to survive despite favorable temperature. Unlike in adults and pupae, high cuticular permeability values were found in the larval stages. This indicates that larvae are highly susceptible to desiccation, and it explains why the distribution of P. fuscipes is restricted to moist habitats.
The susceptibility of three strains of Aedes aegypti (L.) to four strains of Ascogregarina culicis were studied under laboratory conditions. The parasite was found to be pathogenic to the mosquito; although pathogenicity varied among geographical strains of A. culicis and susceptibility varied among geographical strains of Ae. aegypti. In addition to affecting the survival of the mosquito larvae, infection with the parasite was found to shorten larval development time. Selected strains of A. culicis may be useful as a biological control agent against Ae. aegypti.
Rickettsia felis (Rickettsiales: Rickettsiaceae) is an emergent human pathogen that causes febrile illnesses in various parts of the world. This study describes the identification and growth characteristics of a R. felis-like organism (designated as Rickettsia sp. TH2014) cultured from Ctenocephalides orientis fleas in rural Malaysia. In this study, culturing of rickettsiae from filtered triturated flea lysates was performed in Aedes albopictus C6/36 cells. Cytopathic effects were observed from one of the samples 4 d post-inoculation. Electron microscopy revealed actively replicating intracytosolic coccobacillary organisms in the rickettsia-infected cells. Sequence analysis of amplified citrate synthase (gltA) gene fragment shows complete match of the rickettsia with Rickettsia sp. Rf31 in Southeast Asia, and 'Candidatus Rickettsia senegalensis' strain PU01-02 in Africa. The whole-genome sequence of Rickettsia sp. TH2014 was determined and assembled. The estimated genome size and guanine + cytosine content of the rickettsia are 1.37 Mb and 32.9%, respectively. The high values of average nucleotide identity and tetra-nucleotide signature correlation index obtained from pairwise genome comparison study suggest the identification of the rickettsia as R. felis. The whole-genome single-nucleotide polymorphism analysis demonstrates close genetic relatedness of the rickettsia with R. felis and Rickettsia asemboensis. However, based on sequence analyses of rickettsial genes (16S rDNA, gltA, ompB, and sca4), Rickettsia sp. TH2014 is found to be distinct from R. felis and R. asemboensis. The sequence analyses reveal that Rickettsia sp. TH2014 is highly similar to 'Ca. Rickettsia senegalensis' detected in fleas from Africa, Asia, and North America. Further investigation to provide insights on pathogenic potential and transmission dynamics of the rickettsia is warranted.
Pediculosis capitis caused by Pediculus humanus capitis (De Geer) is endemic all over the world, and children are mostly affected, particularly those living in overcrowded institutions. Several studies have shown that P. h. capitis carried human pathogenic bacteria, suggesting the potential role of head lice in the transmission of pathogens to humans. In this study, we determined the genetic diversity of head lice collected from welfare homes sheltering underprivileged children by using DNA barcoding and demonstrated the presence of Acinetobacter spp., Serratia marcescens, and Staphylococcus aureus in head lice, which have never been investigated before in Malaysia. Cox1 DNA barcoding identified the head lice, P. h. capitis collected from welfare homes across two geographical areas of Peninsular Malaysia as belonging to clades A, B, and D. Acinetobacter bacteria: Acinetobacter guillouiae, Acinetobacter junii, Acinetobacter baumannii, and Acinetobacter nosocomialis were detected in head lice belonging to clades A and also D. In addition, DNA from S. marcescens and S. aureus were also detected in both clades A and D. To our knowledge, this is the first report on the genetic diversity of head lice in Malaysia through DNA barcoding, as well as the first to provide molecular evidence on the type of bacteria occurring in head lice in Malaysia. It is anticipated that the DNA barcoding technique used in this study will be able to provide rapid and accurate identification of arthropods, in particular, medically important ectoparasites.
The tropical bed bug is scientifically recognized as a significant public health problem. While there is an increased awareness about their resurgence by medical and life science committees, efficient bed bug management still remains unresolved. The solution may soon arise, as information about bed bugs' infestation dynamics and systematics are becoming more distinguishable. Recent developments in studies about bed bugs are based on molecular intervention by determining their genetic variation and phylogeography. The aim of this study is to assess the phylogenetic relationships and genetic diversity among the populations of tropical bed bugs inhabiting Malaysia. A molecular genotyping study was conducted with 22 tropical bed bug populations composed of three individuals per population. The mitochondrial (COI) gene was used as a marker. The data obtained were analyzed using the T-Coffee, ClustalX, MEGA 6.0, and PAUP software. The results showed one main monophyletic clade that consisted of two groups: Ch01 and Ch02. Ch02 consists of samples from the Bandar Hilir population, differing from the other populations studied by one singleton base. However, as there were no changes in the amino acid, this singleton genetic variation was considered to have no effect on genetic differentiation. Ch01 shows similarity with some sequence of Cimex hemipterus (F.) from Thailand, suggesting an international diversity connection. The disparity index apparently suggests that all isolates are homogeneous populations and are supported by the low value of the mean pairwise distance between isolates. This study will increase the knowledge about phylogeographic diversity of tropical bed bug in Malaysia.
To better understand the population dynamics and dispersal ability of insect species, it is often helpful to derive a life table containing fundamental demographic data. The aim of this study was to determine a life table for the predatory necrophagous species Synthesiomyia nudiseta (van der Wulp, 1883) on a pig liver diet and under controlled laboratory conditions (29.5 ± 2. 5°C, RH 50 ± 15%, and a photoperiod of 12:12). This species has medical and veterinary importance and its distribution extends in tropical and subtropical areas and now it has been established in the southwestern of Europe. The mean adult longevity was 36. 18 ± 2. 06 d and the net reproduction rate, R, was 27.65 offspring/female, the mean generation time, T, was 22. 09 d, the finite rate of increase, λ, was 1. 16 d-1, and the intrinsic rate of increase, r, was 0. 15 d-1. These results indicate that S. nudiseta cannot be considered an r-strategist as the most common synanthropic necrophagous blowflies due to its predatory behavior; however, its invasive and colonist abilities are discussed. This is the first life table study of this species from Palearctic region to analyze the effect of its dispersal ability.
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.
The relationship among body size (as indicated by wing length), age (as indicated by parity dissections), and malaria infection were observed in host-seeking Anopheles maculatus Theobald females collected in aboriginal villages of peninsular Malaysia. Both ELISA and salivary gland dissections were used to determine malaria infection. The wings of parous females were significantly longer than those of nulliparous females, suggesting that larger females live longer than smaller ones, and thus have a higher vectorial capacity. Body size differences were not detected between infected parous and uninfected parous females. Females infected with only oocysts were significantly larger than females infected with sporozoites. No correlation was found between the number of oocysts or sporozoites and body size in this small sample.
Susceptibility status of Aedes aegypti (L.) and Aedes albopictus Skuse larvae obtained from 12 states in Malaysia were evaluated against five insect growth regulators (IGRs), namely, pyriproxyfen, methoprene, diflubenzuron, cyromazine, and novaluron under laboratory conditions. Field populations of Ae. aegypti exhibited moderate resistance toward methoprene and low resistance toward pyriproxyfen, with resistance ratios of 12.7 and 1.4, respectively, but susceptibility to diflubenzuron, cyromazine, and novaluron. On the other hand, field populations of Ae. albopictus exhibited low resistance against diflubenzuron and novaluron, with resistance ratio of 2.1 and 1.0, respectively, but susceptibility to other tested IGRs. Our study concluded that the tested IGRs provide promising results and can be used to control field population of Ae. aegypti and Ae. albopictus, especially cyromazine. The use of IGR should be considered as an alternative when larvae develop resistance to conventional insecticides.
A basic tenet of forensic entomology is development data of an insect can be used to predict the time of colonization (TOC) by insect specimens collected from remains, and this prediction is related to the time of death and/or time of placement (TOP). However, few datasets have been evaluated to determine their accuracy or precision. The black soldier fly, Hermetia illucens (L.) (Diptera: Stratiomyidae) is recognized as an insect of forensic importance. This study examined the accuracy and precision of several development datasets for the black soldier fly by estimating the TOP of five sets of human and three sets of swine remains in San Marcos and College Station, TX, respectively. Data generated from this study indicate only one of these datasets consistently (time-to-prepupae 52%; time-to-eclosion 75%) produced TOP estimations that occurred within a day of the actual TOP of the remains. It is unknown if the precolonization interval (PreCI) of this species is long, but it has been observed that the species can colonize within 6 d after death. This assumption remains untested by validation studies. Accounting for this PreCI improved accuracy for the time-to-prepupae group, but reduced accuracy in the time-to-eclosion group. The findings presented here highlight a need for detailed, forensic-based development data for the black soldier fly that can reliably and accurately be used in casework. Finally, this study outlines the need for a basic understanding of the timing of resource utilization (i.e., duration of the PreCI) for forensically relevant taxa so that reasonable corrections may be made to TOC as related to minimum postmortem interval (mPMI) estimates.
Little information is available on human anaplasmosis and ehrlichiosis in Southeast Asia despite increasing reports of the detection of Anaplasma spp. and Ehrlichia spp. in the ticks. We report herein the serological findings against the tick-borne pathogens in a group of animal farm workers (n = 87) and indigenous people (n = 102) in Peninsular Malaysia. IgG antibodies against Ehrlichia chaffeensis were detected from 29.9% and 34.3% of farm workers and indigenous people, respectively, using commercial indirect immunofluorescence assays. Comparatively, only 6.9% of the indigenous people but none of the animal farm workers were seropositive to Anaplasma phagocytophilum. A polymerase chain reaction (PCR) assay targeting the 16S rRNA gene of Anaplasmataceae was used to identify Anaplastamataceae in ticks collected from various locations adjacent to the areas where the serological survey was conducted. In this study, a total of 61.5% of ticks infesting farm animals, 37.5% of ticks infesting peri-domestic animals in rural villages, 27.3% of ticks collected from wildlife animals, and 29.1% of questing ticks collected from forest vegetation were positive for Anaplasmataceae DNA. Sequence analyses of 16S rRNA gene region (238 bp) provide the identification for Anaplasma marginale, Anaplasma bovis, Anaplasma platys, A. phagocytophilum, and Anaplasma spp. closely related to Candidatus Cryptoplasma californiense in ticks. E. chaffeensis DNA was not detected from any ticks, instead, Ehrlichia sp. strain EBm52, Ehrlichia mineirensis and Candidatus Ehrlichia shimanensis are the only Ehrlichia sp. identified from cattle ticks in this study. Further investigation is required to ascertain the occurrence of zoonotic transmission of Ehrlichia and Anaplasma infections in Peninsular Malaysia.
Toxicities of three pyrethroids, d-phenothrin, decamethrin, and permethrin, were evaluated in the laboratory against Leptotrombidium fletcheri (Womersley & Heaslip). The susceptibilities between populations of the species infected and noninfected with scrub typhus were investigated. The three pesticides exhibited different toxicities to the chiggers. D- phenothrin was the most toxic, followed by decamethrin, then permethrin. There were no significant differences between susceptibilities of the infected and noninfected populations. Log-probit regression lines indicated that the species was most sensitive to increasing concentrations of d-phenothrin and least sensitive to permethrin. The results show that the three pesticides are potential candidates for chemical control of L. fletcheri. It may be possible in the future to conduct similar bioassays only with the noninfected population, thus reducing risk of infection to workers conducting the bioassays. Similarly, there may not be a need to separate field-collected chiggers into the two populations before performing the bioassays.
The effect of temperature and humidity on the survival and water loss of the tropical bed bug, Cimex hemipterus (F.), was studied using two field-collected strains. Insects were exposed to temperatures ranging from 20 to 45 degrees C and relative humidities (RHs) of 33, 75, and 100%. C. hemipterus survived longest under the interaction of low temperature (20 degrees C) and high RH (75-100%). Survival and water loss were significantly affected (P < 0.01) by temperature and RH (either singly, or in interaction). Strain and sex significantly (P < 0.01) influenced bed bug survival, but not on water loss. Eggs, first instars, and adults reached their upper thermal lethal limit within 1 h at 39 degrees C, 44 degrees C, and 46 degrees C, respectively. The survival and water loss profiles showed that starved C. hemipterus started to die after losing 35-45% of their body weights.
This study examined the effects of different life stages (first, second, third, fourth, and five instars; adult females and adult males) and feeding regimes (starved and blood fed) on the active movement activity of the tropical bed bug, Cimex hemipterus (F.), under mixed-stage conditions. We used an extended arena made from Tygon tube coils and observed the movement frequency and movement distance at selected time intervals up to 120 h. The fifth instars and adult males and females showed significantly (P < 0.01) greater movement frequency compared with the other stages. The first and second instars showed limited movement (< 8 m) over the experimental period. Starved bed bugs showed greater movement frequency compared with blood-fed bed bugs, with the exception of adult females. Blood-fed adult females exhibited significantly (P < 0.01) greater movement frequency and distance compared with starved females. Blood-fed females moved up to 42.3 m over 120 h. Regression analysis between movement distance of the fifth instars and adults and the time intervals revealed a positive relationship (r2 = 0.6583; P < 0.01), suggesting that delays in bed bug control efforts will increase the risk of the greater infestation. During bed bug inspection, the presence of only late instars and adults in premises would indicate a new infestation, whereas an established infestation likely would consist of mixed stages.