Intestinal Parasites and the Occurrence of Zoonotic Giardia duodenalis Genotype in Captive Gibbons at Krabokkoo Wildlife Breeding Center, Thailand.

Intestinal parasitic infections can have an impact on health and growth of wildlife. The current study aims were to determine the prevalence of intestinal parasites and to molecular characterize Giardia duodenalis and Cryptosporidium spp. in captive gibbons at Krabokkoo Wildlife Breeding Center, Thailand. Fifty-five gibbons, 2 agile- (Hylobates agilis), 38 lar- (Hylobates lar) and 15 pileated gibbons (Hylobates pileatus) were included in this study. Fecal samples were collected individually at Krabokkoo Wildlife Breeding Center, Chachoengsao province, eastern Thailand, in November 2013. Intestinal parasitic infections were examined by zinc sulfate centrifugation flotation and by a commercially available immunofluorescent assay (IFA) for detection of G. duodenalis and Cryptosporidium spp.. Polymerase chain reaction targeting the Giardia glutamate dehydrogenase (gdh), beta- giardin (bg), triose phosphate isomerase (tpi) genes, and the Cryptosporidium small subunit-rRNA and heat-shock protein (hsp70) following by DNA sequencing were performed on the IFA positive samples. The overall prevalence of intestinal parasitic infection in gibbons at Krabokkoo Wildlife Breeding Center was 12.7% (95%CI: 5.3-24.5), Strongyloides spp. eggs or larvae were present in all positive samples. Co-infections with G. duodenalis were detected in 1.8% (95%CI: 0.1-9.7) of the samples. Based on the sequencing results of the three genes, the IFA Giardia positive isolate typed as the zoonotic genotype B. Since the data reveals the occurrence of zoonotic Giardia genotype, good hygiene management is suggested to prevent the transmission of this pathogen from gibbon to human, and vice versa.

Visceral pentastomiasis caused by Armillifer armillatus in a captive striped hyena (Hyaena hyaena) in Chiang Mai Night Safari, Thailand.

Visceral pentastomiasis (porocephalosis) caused by Armillifer armillatus larvae was incidentally diagnosed in a female striped hyena (Hyaena hyaena) of unknown age which died unexpectedly in 2013. The hyena had been imported from Tanzania 8years earlier and have been since then in a zoo in Chiang Mai, northern Thailand. Pathological examination revealed visceral nymph migrans of pentastomes throughout the intestine, liver, diaphragm, omentum and mesentery, spleen, kidneys, and urinary bladder. Polymerase chain reaction and sequencing that targeted the pentastomid-specific 18S rRNA gene determined 100% identity with reference sequence for A. armillatus, suggesting that its ova can infect the hyena to serve as an intermediate host for the parasite. Further studies to identify the source of infection, its risk factors, and host range for A. armillatus are important to determine its zoonotic potential and to better prevent and manage the disease to protect animal and human health.

Identifying fly puparia by clearing technique: application to forensic entomology.

In forensic investigations, immature stages of the fly (egg, larva, or puparia) can be used as entomological evidence at death scenes, not only to estimate the postmortem interval (PMI), analyze toxic substances, and to determine the manner of death but also to indicate the movement of a corpse in homicide cases. Of these immature stages, puparia represent the longest developmental time, which makes them of useful. However, in order for forensic entomologists to use puparia effectively, it is crucial that they are able to accurately identify the species of fly found in a corpse. Typically, these puparia are similar in general appearance, being coarctate and light brown to dark brown in color, which makes identification difficult. In this study, we report on the clearing technique used to pale the integument of fly puparia, thereby allowing observation of the anterior end (second to fourth segments) and the profile of the posterior spiracle, which are important clues for identification. We used puparia of the blowfly, Chrysomya megacephala (F.), as the model species in this experiment. With placement in a 20% potassium hydroxide solution daily and mounting on a clearing medium (Permount(R), New Jersey), the profile of the posterior spiracle could be clearly examined under a light microscope beginning on the fifth day after pupation, and the number of papillae in the anterior spiracle could be counted easily starting from the ninth day. Comparison of morphological features of C. megacephala puparia with those of other blowflies (Chrysomya nigripes [Aubertin], Chrysomya rufifacies [Macquart], Chrysomya villeneuvi [Patton], Lucilia cuprina [Wiedemann], and Hemipyrellia ligurriens [Wiedemann]) and a housefly (Musca domestica L.) revealed that the anterior ends and the profiles of the posterior spiracles had markedly distinguishing characteristics. Morphometric analysis of the length and width of puparia, along with the length of the gaps between the posterior spiracles of seven fly species, displayed differences among them. This provides a key to identifying the puparia of these seven fly species.

Forensic entomology cases in Thailand: a review of cases from 2000 to 2006.

This paper presents and discusses 30 cases of cadavers that had been transferred for forensic entomology investigations to the Department of Forensic Medicine, Faculty of Medicine, Chiang Mai University, northern Thailand, from 2000 to 2006. Variable death scenes were determined, including forested area and suburban and urban outdoor and indoor environments. The fly specimens found in the corpses obtained were the most commonly of the blow fly of family Calliphoridae, and consisted of Chrysomya megacephala (F.), Chrysomya rufifacies (Macquart) Chrysomya villeneuvi Patton, Chrysomya nigripes Aubertin, Chrysomya bezziana Villeneuve, Chrysomya chani Kurahashi, Lucilia cuprina (Wiedemann), Hemipyrellia ligurriens (Wiedemann), and two unknown species. Flies of the family Muscidae [Hydrotaea spinigera Stein, Synthesiomyia nudiseta (Wulp)], Piophilidae [Piophila casei (L.)], Phoridae [Megaselia scalaris (Loew)], Sarcophagidae [Parasarcophaga ruficornis (F.) and three unknown species], and Stratiomyiidae (Sargus sp.) were also collected from these human remains. Larvae and adults of the beetle, Dermestes maculatus DeGeer (Coleoptera: Dermestidae), were also found in some cases. Chrysomya megacephala and C. rufifacies were the most common species found in the ecologically varied death scene habitats associated with both urban and forested areas, while C. nigripes was commonly discovered in forested places. S. nudiseta was collected only from corpses found in an indoor death scene.

Field investigation of Bithynia funiculata, intermediate host of Opisthorchis viverrini in northern Thailand.

A survey of freshwater snail, Bithynia funiculata, was conducted in four locations, Doi Saket, Mueang, Saraphi and Mae Rim Districts of Chiang Mai Province, northern Thailand, between June and October, during the rainy season of 2004. A total of 2,240 snails was collected and classified into 7 families and 15 genera; of which 352 B. funiculata were obtained. B. funiculata was found most abundant in July and September. The infection rate of trematode cercariae in B. funiculata was 9.6% (19/352), while that of pleurolophocercous was 0.3% (1/352). Virgulate cercaria was the most common type, followed by lophocercous, monostome and pleurolophocercous. B. funiculata prefers habitats with clear water, which was less than 30 cm depth, temperatures between 24.48 and 31.78 degrees C, dissolved oxygen 2.03-7.66 mg/l, saturated dissolved oxygen 26.70-95.00%, conductivity 0.000-0.2642 mS/cm, turbidity 16.00-288.00 NTU and pH 6.58-7.56. Geographic Information System (GIS) analysis using loose soil mixed with clay revealed that the breeding grounds of this snail species was in the paddy fields and village environs of the Ping, Kuang and Fang river basins.

Morphological comparison between Chrysomya rufifacies (Macquart) and Chrysomya villeneuvi Patton (Diptera: Calliphoridae) puparia, forensically important blow flies.

In Thailand, the hairy maggots of the blow flies, Chrysomya rufifacies (Macquart) and Chrysomya villeneuvi Patton, are of forensic importance. Both flies are closely related species, not only in the morphological appearance of their larvae and puparia, but also on the aggressive feeding habit of the former. In our continuing studies of forensically important flies, identification of immature ones needs particular attention. In this study, we reported the morphological comparison between the puparia of these two blow fly species using scanning electron microscopy (SEM). Observation revealed that the cuticular sculpture of tubercles along the dorsal and lateral segments had markedly different features: with C. rufifacies having many sharp spines assembling only at the tip, while of C. villeneuvi bore stout spines throughout the tubercle. A larger number of globules at the bubble membrane on the dorsolateral border of the fifth segment was found in C. villeneuvi (average 225) than in C. rufifacies (average 35), and more papillae was observed on the anterior spiracle in C. villeneuvi (13-15) than in C. rufifacies (9-12). However, the morphology of distinct net-like patches of the integument and structure of the posterior spiracle of both species were almost identical. Morphological comparison in this study permitted identification of the puparia of both fly species, particularly in areas where they co-exist.

Morphological observation of puparia of Chrysomya nigripes (Diptera: Calliphoridae) from human corpse.

This article presents Chrysomya nigripes Aubertin as a blow fly species of forensic importance in Thailand, and morphological observation of fly puparia using scanning electron microscopy (SEM). Morphologically, we focused on the characteristics of puparia used to accurately identify fly species. Numerous puparia of C. nigripes were found aggregated, adhering side by side, on the tibia of a skeletonized corpse, which was recovered from a forested area of Chiang Mai, northern Thailand. In the triangular shape of the anterior end of the puparia, three thoracic segments and broad hairy patches beginning dorsolaterally at the sixth segments were distinguishing characteristics. This study, showing pupariation of the flies along the bone of a corpse as well as morphological features, provides important guidance in identifying C. nigripes puparia. A key to differentiate puparia of C. nigripes from the other flies of forensic importance in Thailand is given.

Effect of microwave irradiation on the blow fly Chrysomya megacephala (F.) (Diptera: Calliphoridae).

Chrysomya megacephala (F) is a blow fly species of medical importance, being pestiferous and a mechanical carrier of various microorganisms to human food. Various strategies to reduce the fly population in endemic areas are needed. We investigated the effects of microwaves at 2,450 MHz, with third-instar exposure times of 7, 10, 15, 30 and 60 seconds on the larvae. Survival of the larvae was monitored by their emergence. About 60% of the larvae lived after 7 seconds of exposure, most died by 15 seconds, and all were dead at 30 and 60 seconds. The number of survivals decreased with increasing exposure times. This indicates the effect of microwave irradiation on fly larval survival.

Morphology of second and third instars of Chrysomya villeneuvi Patton (Diptera: Calliphoridae), a fly species of forensic importance.

The morphology of the second and third instars of Chrysomya villeneuvi Patton, a fly species of forensic importance, was presented by use of light microscopy. Both instars were of hairy appearance, bearing elongated tubercles along the abdominal and caudal segments. The anterior spiracle had 13-15 papillae. Minute dark spots were observed to thoroughly cover the tubercle's surface, with 4-6 strong dark tips. Regarding the third instar, the intersegmental spines between the prothorax and mesothorax were heavily pigmented. The posterior spiracle had a thick and heavily pigmented incomplete peritreme. The surface and tip of the tubercles was covered with heavily pigmented sharp spines. The integument of the body was covered with numerous distinct net-like patches. A comparison with another well-known hairy maggot, Chrysomya rufifacies (Macquart), was discussed.

First report of human myiasis caused by Chrysomya megacephala and Chrysomya rufifacies (Diptera: Calliphoridae) in Thailand, and its implication in forensic entomology.

We report a forensic entomology case associated with human myiasis in Chiang Mai Province, northern Thailand. The remains of a 53-yr-old-male were concurrently infested with third instars of the two blow fly species, Chrysomya megacephala (F.) and Chrysomya rufifacies (Macquart), near a severe tumor lesion presented on the lower right leg. The presence of third instars, approximately 5 d old, on the day following postmortem indicated that myiasis occurred before death. This is the first report of both fly species acting as a myiasis-producing agent in Thailand. Unsynchronized data between the age of fly larvae due to myiasis premortem and verified age/ condition of the corpse suggest a potential complication and error in the estimation of postmortem interval if other predisposition fly infestations are not considered.

Larval morphology of Chrysomya nigripes (Diptera: Calliphoridae), a fly species of forensic importance.

The morphology of all instars of Chrysomya nigripes Aubertin, a blow fly species of forensic importance, is presented with the aid of both light microscopy and scanning electron microscopy (SEM). Morphological features of the cephalopharyngeal skeleton, anterior spiracle, posterior spiracle, and dorsal spines between the prothorax and mesothorax are highlighted. No consistent features were found, even using SEM, for distinguishing the first instar of C. nigripes from that of Chrysomya megacephala (F.) or Chrysomya rufifacies (Macquart), two other commonly associated blow fly species in corpses in Thailand. Several features observed in second and third instars proved to be valuable characteristics for separating these species.

Antennal sensilla of some forensically important flies in families Calliphoridae, Sarcophagidae and Muscidae.

Antennal sensilla of some forensically important fly species in the families Calliphoridae (Chrysomya megacephala, Chrysomya rufifacies, Chrysomya nigripes and Lucilia cuprina), Sarcophagidae (Parasarcophaga dux) and Muscidae (Musca domestica) were studied using scanning electron microscopy. Five types of sensilla were observed: trichoid, basiconic, coeloconic, styloconic and sensory pit. Only trichoid sensilla are found on the scape of the antenna, while both trichoid and styloconic sensilla are located on the antennal pedicels of all species studied. Basiconic sensilla are the most numerous of the sensilla found on the antennae of both sexes of all fly species studied and are comprised of two subtypes: large and small basiconic sensilla. Coeloconic sensilla are characterized by short pegs, with either grooved or smooth surfaces, that are sunken into deep depressions. No marked difference was observed in the number, morphological structure or distributional pattern of any of the sensilla among the species studied, with the exception of there being more numerous sensory pits detected in female P. dux compared to the other species. The suggested function of each antennal sensillum was based on comparison with results of other investigations on similar sensilla.