Goat Warble Fly Infestation (GWFI) is also known as subcutaneous myiasis caused by Przhevalskiana silenus (Diptera: Oestridae). It is widely distributed in tropical and sub-tropical areas of the world. In goats, WFI is usually detected through conventional procedure which underestimated the infestation. The current study was designed to determine the serodiagonsis of GWFI (through IDEXX Hypodermosis serum antibody test) and also aimed to investigate its seroepizootiological profile in Pothwar region, Pakistan from 2013-14. The results showed that average seropositivity (ELISA kit) of GWFI was 18.5% whereas, it was 11% by using conventional procedure (Palpation method) depicting a significant difference (p<0.05). Higher seropositivity (30.8%) was observed in Jhelum district as compared to e Attock district (6%). The L1 larvae were found in September, while nodules start appearing in October to December and last until the end of February. The month wise peaks of optical density (OD) was higher in December which gradually decrease along with the end of winter season. The prevalence of GWFI revealed no significant difference among three host breeds (Jattal, Beetal and Tedy). According to the results, high infestation rate (28%) was observed in young animals of age group < 1 year as compared to old animals (> 2 years). Topographically, hilly areas (33%) provide favourable climatic conditions for the propagating of larval stages. Sex difference showed no significant difference. The seroprevalence varied significantly with respect to age, month, districts and topographical location. The current study proved that serologic diagnosis (commercial ELISA kit) as more sensitive and accurate for timely diagnosis of GWFI than traditional method. The information on the epizootiology of P. silenus in goats of Pothwar region would help in devising effective control strategies.
In forensic entomology practice, it is more common to use raw animal tissue to breed dipteran larvae and it often brings unpleasant odour in the laboratory. Few studies suggested the use of synthetic diets, mainly agar-based media, as alternatives to animal tissue but it is rarely being practiced in forensic entomology laboratory. The present study observed the growth of a forensically important fly, Megaselia scalaris (Loew) on raw cow’s liver, nutrient agar, casein agar and cow’s liver agar. A total of 100 M. scalaris eggs were transferred each into the different media and placed in an incubator at 30°C in a continuous dark condition. Data on length and developmental period were collected by randomly sampling three of the largest larvae from each rearing media, twice a day at 0900 and 1500 hours until pupariation. M. scalaris larvae reared on raw cow’s liver recorded the highest mean length (4.23 ± 1.96 mm) followed by cow’s liver agar (3.79 ± 1.62 mm), casein agar (3.14 ± 1.16 mm) and nutrient agar (3.09 ± 1.11 mm). Larval length in raw liver and liver agar were significantly different from those in nutrient and casein agar (p < 0.05). Larvae bred in liver agar and raw liver recorded the shortest larval duration before entering the post-feeding stage (89 hours), followed by nutrient agar (119 hours) and casein agar (184 hours). Total developmental time from oviposition until adult emergence for M. scalaris in liver agar and raw liver was approximately 163 hours. All puparia in nutrient agar and casein agar failed to hatch. This research highlighted the potential use of cow’s liver agar as an alternative diet of raw liver to culture M. scalaris in laboratory.
Phorid flies play an important role in forensic cases and can cause myiasis in humans. Studies on phorid flies species diversity are still limited in Malaysia. This research was carried out to collect information about species and frequency distribution of phorid flies as to provide more information on their roles in forensic and medical entomology. Bait trap was used with 100 g beef liver as baits. The species of the flies were identified using identification keys from Disney as well as Brown and Oliver. There were 449 phorid flies found in Kepong, Kuala Lumpur including Megaselia scalaris, Megaselia spiracularis, Megaselia sp. and phorid flies of genus X. Female phorid flies (98.89%) were found more prone to be trapped compared to male phorid flies (1.11%). Most phorid flies trapped in Kepong, Kuala Lumpur were from genus Megaselia and consisted of female flies. A total of five species of phorid flies probably new to science were also discovered. This study showed that Megaselia flies were found indoors rather than in open spaces. This was corresponding to their discoveries among the decomposing corpse found inside premises.
A four-month cross-sectional study found five species of parasitoids parasitizing puparia of filth flies breeding at the Taman Beringin landfill in Kepong and a poultry farm in Sungai Pelek, Sepang, Selangor. Effect of monthly rainfalls towards density of flies and percentage of parasitoids emerging from collected puparia were also analyzed. Spalangia sp. was the most common, consisting of Spalangia endius Walker, S. cameroni Perkins and S. gemina Boucek. Other parasitoids collected were Pachycrepoideus vindemmiae Rondani and Exoristobia phillipinensis Ashmead. The parasitized fly hosts were Musca domestica Linn. and Chrysomya megacephala Fabricius. S. endius was the most common parasitoid attacking M. domestica at both locations. M. domestica was the most common fly found at the Sg. Pelek poultry farm whereas C. megacephala was the most numerous at the Taman Beringin landfill. During heavy rainfall month of November 2003, density of flies were high whereas the emerging parasitoids were low at both landfill and poultry farm. The present study revealed the endemic presence of parasitoids especially S. endius in both poultry farm and garbage landfill and the potential of the parasitoid species in fly control in Malaysia.
Natural peptides in insect vectors played an important role in the control of
pathogens. Musca domestica Linnaeus and Chrysomya megacephala Fabricius were
two species of local fly chosen to detect presence of antimicrobial peptide substance.
The screening of the antimicrobial activity was carried using a spectrophotometric
method. Results were obtained much quicker and less laborious. The results showed
larva hemolymph of M. domestica lysed Bacillus subtilis and two Gram negatives,
Escherichia coli and Pseudomona. aeruginosa. The pupae hemolymph only lysed E.
coli. Whereas, the hemolymph of C. megacephala larva showed bactericidal effect
against both of the Gram positives tested, i.e. B. subtilis and Staph. aureus. and no
effect was against the Gram negatives. The pupa showed lytic activity against Staph.
aureus and P. aeruginosa. As a conclusion, the larva and pupa hemolymph of M.
domestica and C. megacephala demonstrated antibacterial activity. However, larva
hemolymph of M. domestica and C. megacephala has broader antibacterial activity
against both Gram positive and negative bacteria.
The plants of the enigmatic genus Rafflesia are well known for their gigantic flowers and their floral features such as pungent floral scent and vivid dark color, which mimics the food/brood sites of carrion. However, information on the pollination biology of this plant group remains limited and mostly anecdotal. In the present paper, we studied the floral volatiles of R. cantleyi Solms-Laubach and their role in pollinator attraction. To achieve these aims, the floral scent was collected in situ in the field using a dynamic headspace method followed by chemical analysis via GC-MS. The olfactory preferences of pollinators to the identified chemical compounds, were tested singly and in blends, in flight tunnel bioassays and compared with responses to headspace floral extracts. In addition, flower-visiting calliphorid flies and the local carrion fly community were sampled and identified. Five species of calliphorid flies (subfamilies of Chrysomyinae and Calliphorinae), all females, were found on the flowers, whereas nine species were found in the traps that were baited with tainted meat in the surrounding habitat. However, only flower visitors of one blow fly species, Chrysomya chani Kurahashi, were observed to carry R. cantleyi pollen after visiting male flowers. The floral volatiles emitted by male flowers in full bloom were dominated by two sulphur-containing compounds, dimethyl disulfide (DMDS) and dimethyl trisulfide (DMTS). These were accompanied by other minor compounds such as benzenoids (4), monoterpenoids (4), trace amounts of aliphatic compounds (1), and sesquiterpenes (1). In flight-tunnel bioassays, a female-specific positive response of C. chani flies to individual DMDS, DMTS, and a blend of DMDS and DMTS was evident. Our findings suggest that R. cantleyi biochemically mimics carrion and that relative ratio of oligosulfides in the floral scent play a key role in sex-biased pollinator specialization, attracting only female C. chani carrion flies to the flowers.
Chrysomya bezziana is an obligate, myiasis-causing fly in humans and warm-blooded animals throughout the tropical and subtropical Old World. We report a case of cutaneous myiasis due to C. bezziana in a dog from Guangxi province in China. A total of 35 maggots were removed from the lesions. Direct sequencing of the mitochondrial cytochrome b gene showed that the specimen belonged to haplotype CB_bezz02, which was previously reported in Malaysia and the Gulf region. This paper also reviews reported cases of screwworm myiasis from humans and animals in China. Geographical records indicate that the distribution of C. bezziana is expanding, suggesting that an integrated pest management control should be taken into consideration in China.
Myiasis occurs when living tissues of mammals are invaded by eggs or larvae of flies, mainly from the order of Diptera. Most of the previousty reported cases are in the tropics and they were usually associated with inadequate personal hygiene, sometimes with poor manual dexterity. This report describes two cases of oral myiasis in cerebral palsy patients in Seremban General Hospital, Malaysia. This article also discusses the therapeutic property of maggots and highlights the importance of oral health care in the special needs patients.
Dengue is the most important mosquito-borne viral disease. No specific treatment or vaccine is currently available; traditional vector control methods can rarely achieve adequate control. Recently, the RIDL (Release of Insect carrying Dominant Lethality) approach has been developed, based on the sterile insect technique, in which genetically engineered 'sterile' homozygous RIDL male insects are released to mate wild females; the offspring inherit a copy of the RIDL construct and die. A RIDL strain of the dengue mosquito, Aedes aegypti, OX513A, expresses a fluorescent marker gene for identification (DsRed2) and a protein (tTAV) that causes the offspring to die. We examined whether these proteins could adversely affect predators that may feed on the insect. Aedes aegypti is a peri-domestic mosquito that typically breeds in small, rain-water-filled containers and has no specific predators. Toxorhynchites larvae feed on small aquatic organisms and are easily reared in the laboratory where they can be fed exclusively on mosquito larvae. To evaluate the effect of a predator feeding on a diet of RIDL insects, OX513A Ae. aegypti larvae were fed to two different species of Toxorhynchites (Tx. splendens and Tx. amboinensis) and effects on life table parameters of all life stages were compared to being fed on wild type larvae. No significant negative effect was observed on any life table parameter studied; this outcome and the benign nature of the expressed proteins (tTAV and DsRed2) indicate that Ae. aegypti OX513A RIDL strain is unlikely to have any adverse effects on predators in the environment.
In forensic entomology, breeding of fly larvae in a controlled laboratory environment using animal tissue is a common technique to obtain insect developmental time for the estimation of postmortem interval. Previous studies on growth media are mostly on the effect of different diets on fly development. However, the interaction effects between temperature and food type used have not been explored. The objective of this study was to compare the use of cow's liver agar and raw liver on the development of a forensically important fly, Megaselia scalaris (Loew). This study also determined the interaction between different temperatures and different food types on the growth of this species. A total of 100 M. scalaris eggs were transferred into each of the two media mentioned above. Liver agar was prepared by adding dried ground liver into nutrient agar, whilst raw liver was naturally prepared from the same animal source. This experiment was conducted at 27, 30 and 33 °C in an incubator in a continuously dark condition. Length and weight of larvae, puparia and adult samples were determined. Total developmental times for larvae feeding on liver agar at each temperature were approximately 7-15 h slower than those feeding on raw liver. Survival rates were almost equal in both diets but were lower at 33 °C. Mean larva length in both diets did not differ significantly at all temperatures, but larvae feeding on liver agar had lower mean weight values than those in raw liver at 30 and 33 °C. The effect of temperature was significant in female puparia weight and male adult weight whereas the effect of diet types was significant in both male and female puparia size and weight. Interaction effects of temperature and food type on M. scalaris puparium size and adult weight were significant, indicating that puparium size and adult weight depended on both food type and temperature. This experiment highlighted the use of cow's liver agar as an alternative diet to breed M. scalaris in the laboratory and the importance of considering the interaction effect between temperatures and food types when deciding the most suitable medium in fly larva rearing.
A 73-year-old Chinese man was admitted to the Accident and Emergency Premorbid Ward of a local hospital in Malaysia. The patient complained of shortness of breath with cough and was in a semi-conscious state. He was later admitted to an intensive care unit (ICU) of the hospital. Six days after admission 5-6 maggots were recoverd from the nasal cavity. The maggots were identified as the third-instar larvae of Lucilia cuprina Wiedmann (Diptera: Calliphoridae) based on the morphological characteristics. This patient was classified as having nosocomial myiasis. The presence of the third instar larvae indicated that the infestation was not more than three to four days. An adult sarcophagid identified as Parasarcophaga ruficornis (Fabricius) caught in the ICU where the patient was warded provided further evidence of the potential for the nosocomial infestation.
Fourth instars larvae of freshwater midge Chironomus javanus (Diptera, Chironomidae) were exposed for a 4-day period in laboratory conditions to a range of copper (Cu), cadmium (Cd), zinc (Zn), lead (Pb), nickel (Ni), iron (Fe), aluminium (Al) and manganese (Mn) concentrations. Mortality was assessed and median lethal concentrations (LC(50)) were calculated. LC(50) increased with the decrease in mean exposure times, for all metals. LC(50)s for 96 hours for Cu, Cd, Zn, Pb, Ni, Fe, Al and Mn were 0.17, 0.06, 5.57, 0.72, 5.32, 0.62, 1.43 and 5.27 mg/L, respectively. Metals bioconcentration in C. javanus increases with exposure to increasing concentrations and Cd was the most toxic to C. javanus, followed by Cu, Fe, Pb, Al, Mn, Zn and Ni (Cd > Cu > Fe > Pb > Al > Mn > Zn > Ni). Comparison of LC(50) values for metals for this species with those for other freshwater midges reveals that C. javanus is equally or more sensitive to metals than most other tested dipteran.
Paraquat poisoning is commonly associated with suicide or homicide in Malaysia. In a case involving advanced body decomposition, pathological analysis regarding the cause of death may become difficult or almost impossible. Insects serve as common alternative matrix for poison detection in forensic analysis. Paraquat detection via secondary bioaccumulation in fly larvae tissue has never been reported. In this study, tissues from blowfly larvae collected from a rabbit carcass with paraquat poisoning were analyzed for secondary bioaccumulation. Larvae samples were collected and analyzed using liquid-liquid extraction. The detection was performed via reduction of quaternary ammonium presence in paraquat and analyzed using gas chromatography-mass spectrometry (GC-MS) with selected ion monitoring mode (SIM mode). GC-MS showed the elution of reduced paraquat was at retention time 12.8 min. Blowfly larvae tissue has proven useful as a secondary detector in paraquat-related deaths.
Most tropical mammal species are threatened or data-deficient. Data collection is impeded by the traditional monitoring approaches which can be laborious, expensive and struggle to detect cryptic diversity. Monitoring approaches using mammal DNA derived from invertebrates are emerging as cost- and time-effective alternatives. As a step towards development of blowfly-derived DNA as an effective method for mammal monitoring in the biodiversity hotspot of Peninsular Malaysia, our objectives were (i) to determine the persistence period of amplifiable mammal mtDNA in blowfly guts through a laboratory feeding experiment (ii) to design and test primers that can selectively amplify mammal COI DNA mini-barcodes in the presence of high concentrations of blowfly DNA. The persistence period of amplifiable mammal mtDNA in blowfly guts was 24 h to 96 h post-feeding indicating the need for collecting flies within 24 h of capture to detect mammal mtDNA of sufficient quantity and quality. We designed a new primer combination for a COI DNA mini-barcode that did not amplify blowfly DNA and showed 89% amplification success for a dataset of mammals from Peninsular Malaysia. The short (205 bp) DNA mini-barcode could distinguish most mammal species (including separating dark taxa) and is of suitable length for high-throughput sequencing. Our new DNA mini-barcode target and a standardized trapping protocol with retrieval of blowflies every 24 h could point the way forward in the development of blowfly-derived DNA as an effective method for mammal monitoring.
The majority of true parasitoids manipulate their host's physiology for their own benefit. In this study, we documented the physiological changes that occurred in major soldiers of the subterranean termite Macrotermes gilvus (Hagen) (Isoptera: Termitidae) parasitized by the koinobiont larval endoparasitoid Misotermes mindeni Disney and Neoh (Diptera: Phoridae). We compared the metabolic rate, body water content, body water loss rate, cuticular permeability, and desiccation tolerance between parasitized and unparasitized major soldiers. The metabolic rate of parasitized hosts was significantly higher than that of unparasitized termites. Mean total body water content of parasitized major soldiers (64.73±3.26%) was significantly lower than that of unparasitized termites (71.99±2.23%). Parasitized hosts also had significantly lower total body water loss rates (5.72±0.06%/h) and higher cuticular permeability (49.37±11.26 μg/cm/h/mmHg) than unparasitized major soldiers (6.75±0.16%/h and 60.76±24.98 μg/cm/h/mmHg, respectively). Parasitized major soldiers survived almost twice as long as unparasitized termites (LT(50)=6.66 h and LT(50)=3.40 h, respectively) and they had significantly higher tolerance to water loss compared to unparasitized termites (45.28±6.79% and 32.84±7.69%, respectively). Body lipid content in parasitized hosts (19.84±6.27%) was significantly higher than that of unparasitized termites (6.17±7.87%). Finally, parasitized hosts had a significantly lower percentage of cuticular water content than unparasitized major soldiers (10.97±1.84% and 13.17±2.21%, respectively). Based on these data, we conclude that the parasitism-induced physiological changes in the host are beneficial to the parasitoids as the alterations can clearly increase the parasite's chances of survival when exposed to extreme environmental conditions and ensure that the parasitoids are able to complete their larval development successfully before the host dies.
This is prospective case-control study of more than 18 months performed to assess the effectiveness of maggot debridement therapy (MDT) with the sterile larvae of Lucilia cuprina (a tropical blowfly maggot) for the treatment of diabetic foot ulcers. Literature thus far has only reported results with the temperate maggot, Lucilia sericata. This study documents outcome in diabetic foot wounds treated with maggot debridement versus those treated by conventional debridement alone. In this series of 29 patients treated with MDT, 14 wounds were healed, 11 were unhealed and 4 were classified under others. The control group treated by conventional debridement had 30 patients of which 18 wounds were healed, 11 unhealed and 1 classified under others. There was no significant difference in outcome between the two groups. The conclusion that can be made from this study is that MDT with L. cuprina is as effective as conventional debridement in the treatment of diabetic foot ulcers. It would be a feasible alternative to those at high risk for surgery or for those who refuse surgery.
This paper discusses the colonization of the stratiomyid species Ptecticus melanurus (Walker) (Diptera: Stratiomyidae) in monkey carrion and its potential for the determination of the minimum time since death (PMI). A study was conducted in a tropical forest at Bangi, Malaysia from 13 November 2009 to 8 June 2011. Twelve monkey carcasses (Macaca fascicularis Raffles) were used and divided in equal number into three different field trials. Adults of P. melanurus were first observed on monkey carrions on the second day the carcasses were placed in the field while their penultimate instar larvae were found in the wet soil under and beside carcass from day 8 to 31 days postmortem.
Two performance (efficacy and attractiveness) comparisons of neonicotinoid baits QuickBayt® (imidacloprid) and Agita® (thiamethoxam) against filth flies were conducted under field conditions to determine suitability for use outdoors. The first experiment compared bait performance and the second compared effects of different applications on QuickBayt® performance. Applications compared were: (i) scattered in petri dish (SPD); (ii) wet-down in petri dish (WPD); (iii) scattered on cardboard (SCB) and (iv) painted on cardboards (PCB). Efficacy and attractiveness were assessed based on knockdown percentage (KD%) and number of flies feeding on baits, respectively. The KD% of QuickBayt® (34% ± 3.0%) was not significantly higher than Agita® (29% ± 1.3%) (t-test, P>0.05). Agita® (101 ± 5.7 flies) was significantly more attractive to flies than QuickBayt® (76 ± 4.8 flies) and the sugar solution (49 ± 7.2) (one-way ANOVA, P<0.05). The PCB and SCB applications were significantly more attractive to filth flies than WPD and SPD (One-way ANOVA, P<0.05), however differences in KD% were not significantly different (One-way ANOVA, P>0.05). The two baits provided the same level of efficacy in a wide-open area against filth flies of various species. QuickBayt® was more versatile; efficacy was not significantly affected by different applications. Surface area and moisture affects attractiveness of the bait.
A forensic entomological study was conducted using monkey carcasses (Macaca fascicularis Raffles) that were placed in either an outdoor or indoor environment at a coastal area in Tanjung Sepat, Selangor, Malaysia during May until August 2008. We collected pupae of Chrysomya rufifacies (Marquart) from the carcasses and kept them individually. The emergence of 13 parasitic microhymenopteran, from one of the pupae occurring within a week were identified as Exoristobia philippinensis Ashmead (Hymenoptera: Encyrtidae). Another observation was made whereby a pupa of C. rufifacies was predated by a muscid larva, Ophyra spinigera (Stein). The larva squeezed into the pupa and consumed the contents. This paper report C. rufifacies as a new host record for E. philippinensis in Malaysia and highlighted the predatory behavior of O. spinigera larva in natural environment.
Insects found associated with corpse can be used as one of the indicators in estimating postmortem interval (PMI). The objective of this study was to compare the stages of decomposition and faunal succession between a partially burnt pig (Sus scrofa Linnaeus) and natural pig (as control). The burning simulated a real crime whereby the victim was burnt by murderer. Two young pigs weighed approximately 10 kg were used in this study. Both pigs died from pneumonia and immediately placed in an oil palm plantation near a pig farm in Tanjung Sepat, Selangor, Malaysia. One pig was partially burnt by 1-liter petrol while the other served as control. Both carcasses were visited twice per day for the first week and once thereafter. Adult flies and larvae on the carcasses were collected and later processed in a forensic entomology laboratory. Results showed that there was no significant difference between the rate of decomposition and sequence of faunal succession on both pig carcasses. Both carcasses were completely decomposed to remain stage after nine days. The species of flies visiting the pig carcasses consisted of blow flies (Chrysomya megacephala, Chrysomya rufifacies, Hemipyrellia ligurriens), flesh fly (Sarcophagidae.), muscid fly (Ophyra spinigera), soldier fly (Hermetia illucens), coffin fly (Phoridae) and scavenger fly (Sepsidae). The only difference noted was in the number of adult flies, whereby more flies were seen in the control carcass. Faunal succession on both pig carcasses was in the following sequence: Calliphoridae, Sarcophagidae, Muscidae, Phoridae and lastly Stratiomyidae. However, there was overlap in the appearance of members of these families. Blowflies continued to oviposit on both carcasses. Hence postmortem interval (PMI) can still be estimated from the partially burnt pig carcass.