A study of diurnal and nocturnal distribution of flies was conducted in Putrajaya. Six different ecological habitats were selected, namely: botanical garden, lake-area, administration building, wetland, jungle fringes and housing areas. Two different type of traps, cylinderical and rectangular in shape were used in the study. Baits used in these traps were yeast, sugar, salted fish, shrimp paste and fresh liver. Traps were placed at the sites throughout the diurnal and nocturnal periods. The time for sunrise and sunset was determined using a Geographical Positioning System gadget (GARMIN) at the sites. Both type of traps were equally effective in trapping flies. There was no significant difference between both types of traps in their ability to trap flies (p > 0.05). A total of 1,534 flies were collected and identified from both types of trap using the multiple baits and habitats. The collection consisted of 23 species of flies classified under 6 families. The highest number of flies were caught from the lake-area followed by botanical garden, administration building, housing areas, wetland and jungle fringes. The most dominant species was Chrysomya megacephala, followed by species of Sarcophagidae and Musca domestica. Diurnal period had more numbers of flies (81.55%) compared to the nocturnal periods (18.45%). Some species of flies were strictly diurnal, some exibited both diurnal and nocturnal activities while only one species was strictly nocturnal.
Skeletal examination is an important aspect of forensic pathology practice, requiring effective bone cleaning with minimal artefact. This study was conducted to compare between chemical and entomology methods of bone cleaning. Ten subjects between 20 and 40 years old who underwent uncomplicated medico-legal autopsies at the Institute of Forensic Medicine Malaysia were randomly chosen for this descriptive cross sectional study. The sternum bone was divided into 4 parts, each part subjected to a different cleaning method, being two chemical approaches i.e. laundry detergent and a combination of 6% hydrogen peroxide and powder sodium bicarbonate and two entomology approaches using 2nd instar maggots of Chrysomyia rufifacies and Ophyra spinigera. A scoring system for grading the outcome of cleaning was used. The effectiveness of the methods was evaluated based on average weight reduction per day and median number of days to achieve the average score of less than 1.5 within 12 days of the bone cleaning process. Using maggots was the most time-effective and costeffective method, achieving an average weight reduction of 1.4 gm per day, a median of 11.3 days to achieve the desired score and an average cost of MYR 4.10 per case to reach the desired score within 12 days. This conclusion was supported by blind validation by forensic specialists achieving a 77.8% preference for maggots. Emission scanning electron microscopy evaluation also revealed that maggots especially Chrysomyia rufifacies preserved the original condition of the bones better allowing improved elucidation of bone injuries in future real cases.
Preservation of larvae retrieved from cadavers is important in ensuring the quality and integrity of entomological specimens used for the estimation of post-mortem interval (PMI). The process of killing and preserving larvae could distort the larvae leading to inaccurate estimation of PMI. In this study, the effects of killing Chrysomya megacephala larvae with hot water at different temperatures and subsequent maintenance in various preservatives were determined. Larvae not killed by hot water but preserved directly were used as control. The types of preservative used were 10% formalin, 70% ethanol and Kahle's solution. The morphological features examined were length, turgidity, curvature and coloration of larvae. Larvae killed in 80ºC hot water have shorter mean length (12.47 ± 2.86 mm) compared to those in 60ºC hot water (12.95 ± 2.69 mm). Increasing the duration of preservation in all types of preservative caused elongations of larvae treated or untreated with hot water. There were no significant changes in larval turgidity preserved in Kahle's solution compared to other two preservatives and were unaffected by the duration of storage. Larvae preserved in Kahle's solution experienced the least changes in coloration and shape compared to other preserved larvae in 70% ethanol or 10% formalin. Larvae directly immersed alive in 70% ethanol experienced the most changes in curvature, coloration and turgidity. This study suggested that killing larvae with hot water at 80ºC and preservation in Kahle's solution is the optimum method resulting in least changes in morphological features of Ch. megacephala larvae.
Chikungunya infection has become a public health threat in Malaysia since the 2008 nationwide outbreaks. Aedes albopictus Skuse has been identified as the chikungunya vector in Johor State during the outbreaks. In 2009, several outbreaks had been reported in the State of Kelantan. Entomological studies were conducted in Kelantan in four districts, namely Jeli, Tumpat, Pasir Mas and Tanah Merah to identify the vector responsible for the virus transmission.
A body of an unknown adult female was found within a shallow burial ground in Malaysia whereas the skull was exposed and visible on the ground. During autopsy examination, nine insect larvae were recovered from the interior of the human skull and subsequently preserved in 70% ethanol. The larvae were greyish in appearance, each with a posterior elongated breathing tube. A week after the autopsy, more larvae were collected at the burial site, and some of them were reared into adults. Adult specimens and larvae from the skull and from the burial site were sequenced to obtain DNA barcodes. Results showed all adult flies reared from the burial site, as well as the larvae collected from the skull were identified as Eristalinus arvorum (Fabricius, 1787) (Diptera: Syrphidae). Here, we report the colonization of E. arvorum larvae on a human corpse for the first time.
Dengue has enormous health impacts globally. A novel approach to decrease dengue incidence involves the introduction of Wolbachia endosymbionts that block dengue virus transmission into populations of the primary vector mosquito, Aedes aegypti. The wMel Wolbachia strain has previously been trialed in open releases of Ae. aegypti; however, the wAlbB strain has been shown to maintain higher density than wMel at high larval rearing temperatures. Releases of Ae. aegypti mosquitoes carrying wAlbB were carried out in 6 diverse sites in greater Kuala Lumpur, Malaysia, with high endemic dengue transmission. The strain was successfully established and maintained at very high population frequency at some sites or persisted with additional releases following fluctuations at other sites. Based on passive case monitoring, reduced human dengue incidence was observed in the release sites when compared to control sites. The wAlbB strain of Wolbachia provides a promising option as a tool for dengue control, particularly in very hot climates.
Piophila casei (Linnaeus) (Diptera: Piophilidae) is reported from human cadavers in two separate forensic cases for the first time in Malaysia. Both bodies were found indoors. The first case, was that of a male of unknown nationality and age and also contained maggots of the muscid Ophyra spinigera (Stein). The second case was a female Chinese whose body also contained other species of maggots but these were not identifiable.
A study on insect succession of monkey carcass in a forested area in Ulu Gombak, Selangor, Malaysia was conducted from 9 May to 18 June 2007. The third instar of the housefly, Musca domestica (Linnaeus) (Diptera: Muscidae) were only found on dry stage of a decomposed (Day-33) monkey carcass (Macaca fascicularis Raffles). This observation revealed that M. domestica maggots were found together with other muscid fly maggots, Hydrotaea (=Ophyra) spinigera (Stein) (Diptera: Muscidae) on dry stage of a carcass. However, the role of M. domestica on forensic entomological study remains unknown. This study recorded the first finding of M. domestica maggots on primate carcass in Malaysia.
The present study aims to investigate the susceptibility status of Aedes albopictus, Culex quinquefasciatus, and Cx. vishnui collected from a pig farm in Tanjung Sepat, Selangor, toward 11 insecticides representing the classes of organochlorines, carbamates, organophosphates, and pyrethroids. The results of a World Health Organization adult mosquito bioassay revealed that Ae. albopictus, Cx. quinquefasciatus, and Cx. vishnui exhibited different susceptibility toward various insecticides. Overall, pyrethroids were able to induce rapid knockdown for all test mosquito species. The pyrethroids lambdacyhalothrin and etofenprox were able to cause high mortality (> 80%) of all 3 species. The findings of the present study will benefit local authorities in selecting appropriate dosage of insecticides to be used in mosquito control in this area.
The emergence of multidrug-resistant bacterial strains has prompted the reintroduction of maggot therapy in the treatment of chronic, infected wounds. Many previous studies have demonstrated the potent antibacterial activity of larval excretions/secretions of the blowfly Lucilia sericata (Meigen) (Diptera:Calliphoridae) against bacteria. However, the antibacterial activity of its sibling species, Lucilia cuprina (Wiedemann) (Diptera:Calliphoridae) against a wide range of pathogenic bacteria has never been determined. The aim of this study was to develop a new procedure to produce whole body extract of larvae of L. cuprina via methanol extraction as well as to demonstrate the in vitro antibacterial activity of this extract against seven selected wound pathogens (Staphylococcus aureus, methicillin-resistant S. aureus, S. epidermidis, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa and Escherichia coli). The turbidimetric assay demonstrated that L. cuprina larval extract was significantly potent against all bacteria tested (P
Bioassay test against malathion had been carried out with larval and adult stages of Aedes aegypti. The mosquitoes were under selection pressure against malathion for forty-five consecutive generations. The rate of resistance development was measured by LC(50) and LT(50) values. The larvae and adult females, after subjection to malathion selection for 45 generations, developed high resistance level to malathion, with resistance ratio of 52.7 and 3.24 folds, respectively over control mosquitoes. Cross-resistance towards the same and different groups of insecticides was determined using the F44 and F45 malathion-selected adult females. Insecticides tested were DDT (4.0%), permethrin (0.75%), propoxur (0.1%), fenitrothion (1%), λ-cyhalothrin (0.05%) and cyfluthrin (0.15%). Results indicated that the mosquitoes were highly resistant to DDT and fenitrothion, moderately resistant to propoxur, tolerant to permethrin and λ-cyhalothrin, and very low resistant to cyfluthrin.
Aedes albopictus was bioassayed to determine resistance development to malathion (OP). Two methods were applied, including WHO larval bioassay to determine the susceptibility to lethal concentration (LC), and adult bioassay to determine lethal time (LT). Larvae from colonies that had undergone selection pressure with malathion to yield 50% mortality were further subjected to selection for subsequent 10 generations. Selection of Ae. albopictus with malathion could relatively induce a consistent resistance ratio of 1.0 throughout 10 generations. It was noted that Ae. albopictus larvae showed less susceptibility to malathion compared to adults. The susceptibility test of adult mosquitoes to diagnostic dosage of 5.0% malathion-impregnated paper showed a variety of susceptibility to malathion when compared to the susceptible strain. Bioassay results indicated that the LT50 values of malathion-selected Ae. albopictus ranged between 11.5 - 58.8 minutes for ten consecutive generations. Biochemical enzyme studies indicated that there was a significant difference (p < 0.05) in esterase level in malathion-selected mosquitoes compared to non-selected control. Electrophoretic patterns of non-specific esterases at different life stages in malathion-selected Ae. albopictus suggested that non-specific esterases do not play a role in resistance of malathion-selected Ae. albopictus.
Chikungunya virus (CHIKV) is maintained in the sylvatic cycle in West Africa and is transmitted by Aedes mosquito species to monkeys. In 2006, four verified CHIKV isolates were obtained during a survey of arboviruses in monkeys (Macaca fascicularis) in Pahang state, Peninsular Malaysia. RNA was extracted from the CHIKV isolates and used in reverse transcription polymerase chain reactions (RT-PCR) to amplify PCR fragments for sequencing. Nucleic acid primers were designed to generate overlapping PCR fragments that covered the whole viral sequence. A total of 11,238 base pairs (bp) corresponding to open reading frames (ORFs) from our isolates and 47 other registered isolates in the National Center for Biotechnology Information (NCBI) were used to elucidate sequences, amino acids, and phylogenetic relationships and to estimate divergence times by using MEGA 7.0 and the Bayesian Markov chain Monte Carlo method. Phylogenetic analysis revealed that all CHIKV isolates could be classified into the Asian genotype and clustered with Bagan Panchor clades, which are associated with the chikungunya outbreak reported in 2006, with sequence and amino acid similarities of 99.9% and 99.7%, respectively. Minor amino acid differences were found between human and non-human primate isolates. Amino acid analysis showed a unique amino acid at position 221 in the nsP1region, at which a glycine (G) was found only in monkey isolates, whereas arginine (R) was found at the same position only in human isolates. The time to the most recent common ancestor (MRCA) estimation indicated that CHIKV probably started to diverge from human to non-human primates in approximately 2004 in Malaysia. The results suggested that CHIKV in non-human primates probably resulted from the spillover of the virus from humans. The study will be helpful in understanding the movement and evolution of CHIKV in Malaysia and globally.
In an effort to develop a more effective technique in dispersing a microbial control agent, Bacillus thuringiensis (Bt), a truck-mounted ultra low volume (ULV) generator (Scorpion) was used to disperse B. thuringiensis israelensis (Bti) and Bti with malathion. Complete larval and adult mortalities for all tested mosquito species within the first 70-80 feet from the ULV generator were achieved. Beyond that distance less than 50% mortality was achieved as insufficient sprayed particles reached the area. A minimum of 10(3) Bti colony forming units per ml is required to cause 100% larval mortality. The sprayed Bti larvicidal toxins were persistent in the test water 7 days post ULV. The effectiveness of B. thuringiensis jegathesan (Btj), a new mosquitocidal Bt serotype was also evaluated. Similar mortality results as Bti were achieved except that the Btj toxins underwent degradation in the test water, since less than 50% less in larval mortality was observed in 7 days post ULV samples. This ULV method has the potential to disperse Bt and malathion effectively for a simultaneous control of mosquito adults and larvae.
Ovitrap surveillance was conducted in four dengue endemic areas in Kuala Lumpur and Selangor, Malaysia to determine the distribution and percentage of mixed breeding of both Aedes aegypti and Aedes albopictus. The percentage of mixed breeding in all study sites both indoors and outdoors accounted for 10 to 32 % from the total ovitraps collected. Ae. aegypti was found at a higher frequency than Ae. albopictus in these ovitraps. This study again indicates that ovitrap is a sensitive tool to attract gravid females of more than one mosquito species to oviposit in the container.
In this paper, we will start off by introducing the classical Ross-Macdonald model for vector-borne diseases which we use to describe the transmission of dengue between humans and Aedes mosquitoes in Shah Alam, which is a city and the state capital of Selangor, Malaysia. We will focus on analysing the effect of using the Mosquito Home System (MHS), which is an example of an autodissemination trap, in reducing the number of dengue cases by changing the Ross-Macdonald model. By using the national dengue data from Malaysia, we are able to estimate λ, which represents the initial growth rate of the dengue epidemic, and this allows us to estimate the number of mosquitoes in Malaysia. A mathematical expression is also constructed which allows us to estimate the potential number of breeding sites of Aedes mosquitoes. By using the data available from the MHS trial carried out in Section 15 of Shah Alam, we included the potential effect of the MHS into the dengue model and thus modelled the impact MHS has on the spread of dengue within the trial area. We then extended our results to analyse the effect of the MHSs on reducing the number of dengue cases in the whole of Malaysia. A new model was constructed with a basic reproduction number, R0,MalaMHS, which allows us to identify the required MHSs coverage needed to achieve extinction in Malaysia. Numerical simulations and tables of results were also produced to illustrate our results.
DNA identification of blow fly species can be a very useful tool in forensic entomology. One of the potential benefits that mitochondrial DNA (mtDNA) has offered in the field of forensic entomology is species determination. Conventional identification methods have limitations for sibling and closely related species of blow fly and stage and quality of the specimen used. This could be overcome by DNA-based identification methods using mitochondrial DNA which does not demand intact or undamaged specimens. Mitochondrial DNA is usually isolated from whole blow fly and legs. Alternate sources for mitochondrial DNA isolation namely, egg, larva, puparium and empty puparium were explored in this study. The sequence of DNA obtained for each sample for every life cycle stage was 100% identical for a particular species, indicating that the egg, 1st instar, 2nd instar, 3rd instar, pupa, empty puparium and adult from the same species and obtained from same generation will exhibit similar DNA sequences. The present study also highlighted the usefulness of collecting all life cycle stages of blow fly during crime scene investigation with proper preservation and subsequent molecular analysis. Molecular identification provides a strong basis for species identification and will prove an invaluable contribution to forensic entomology as an investigative tool in Malaysia.
Forensic entomology applies knowledge about insects associated with decedent in crime scene investigation. It is possible to calculate a minimum postmortem interval (PMI) by determining the age and species of the oldest blow fly larvae feeding on decedent. This study was conducted in Malaysia to identify maggot specimens collected during crime scene investigations. The usefulness of the molecular and morphological approach in species identifications was evaluated in 10 morphologically identified blow fly larvae sampled from 10 different crime scenes in Malaysia. The molecular identification method involved the sequencing of a total length of 2.2 kilo base pairs encompassing the 'barcode' fragments of the mitochondrial cytochrome oxidase I (COI), cytochrome oxidase II (COII) and t-RNA leucine genes. Phylogenetic analyses confirmed the presence of Chrysomya megacephala, Chrysomya rufifacies and Chrysomya nigripes. In addition, one unidentified blow fly species was found based on phylogenetic tree analysis.
The study on biodiversity of forensically important Diptera in the tropical rain forest in Malaysia is scarce. Thus, a preliminary survey was conducted at a jungle fringe near Kampung Bahagia Bukit Lagong, Sungai Buloh, Selangor. A rat carcass was offered to attract carrion flies and we collected an adult female calliphorid, Hypopygiopsis fumipennis (Walker, 1856) during the fresh stage of carcass decomposition. The female fly was allowed to oviposit on chicken liver in a container and the resulting larvae were reared to the adult stage. Along the developmental process, several individuals from each instar were collected and preserved in 70% ethanol and then processed on the slides. We recorded the duration of development for each instar and described its larval features for the first time. The third instar larvae of H. fumipennis showed accessory oral sclerite present, anterior spiracle with 13-15 papillae, intersegmental spines mostly unicuspid with pointed end, and posterior spiracles heavily sclerotized with inter-slit projections. Some larval differences between H. fumipennis and Hypopygiopsis violacea were noted.
The natural and artificial mating of laboratory bred Aedes albopictus and transgenic Aedes aegypti RIDL-513A-Malaysian strain was conducted. The experiment consisted of crossmating of homologous Ae. aegypti RIDL female symbol X Ae. aegypti RIDL male symbol and reciprocal Ae. aegypti RIDL female symbol X Ae. albopictus WT male symbol. The other set comprised homologous Ae. albopictus WT female symbol X Ae. albopictus WT male symbol and reciprocal Ae. albopictus WT female symbol X Ae. aegypti RIDL male symbol. This study demonstrated that reproductive barriers exist between these two species. Cross insemination occurred between A. albopictus male and Ae. aegypti female and their reciprocals. There was 26.67% and 33.33% insemination rate in Ae. aegypti RIDL female cross-mating with A. albopictus WT male and Ae. albopictus female cross-mating with Ae. aegypti RIDL male, respectively. There was 0% hatchability in both directions of the reciprocals. There was also no embryonation of these eggs which were bleached. Although none of the female Ae. albopictus WT was inseminated in the cross-mating with Ae. albopictus WT female symbol X Ae. aegypti RIDL male symbol, a total of 573 eggs were obtained. The homologous mating was very productive resulting in both high insemination rate and hatchability rates. Generally there was a significantly higher insemination rate with artificial mating insemination of homologous than with artificial mating of reciprocal crosses. Interspecific mating between Ae. aegypti RIDL and Ae. albopictus wild type was not productive and no hybrid was obtained, indicating absence of horizontal transfer of introduced RIDL gene in Ae. aegypti to Ae. albopictus.