Baha Latif

Cryptosporidiosis as threatening health problem: A review

The protozoa under the genus Cryptosporidium is a zoonotic apicomplexan obligate intracellular parasite. Cryptosporidiosis, the term used to designate infection caused by Cryptosporidium sp., is considered as one of the most common food and waterborne diseases with worldwide spread, acting as a common cause of diarrhoea in animals and man. In immunocompetent individuals, Cryptosporidium typically induces self-limiting diarrhoea, which may resolve on its own after 2-3 d. However, cryptosporidiosis may turn life-threatening and subsequently lead to death in small children, the elderly and immunocompromised person, especially in AIDS patient. The diagnosis for Cryptosporidium infection is usually carried out through examination of stool for the presence of oocysts which measured 4-6 μm with spherical appearance. Morphometric identification is often difficult because of the diminutive size and obscure internal structure of the protozoa. Often, the identification of Cryptosporidium is realised through the combination of methods incorporating data from morphometrics, molecular techniques, and host specificity. However, limitations to some of these techniques still exist whether because of cost, duration, expertise, or reliability. Drugs combination is implemented in treatment of cryptosporidiosis. The efficiency of paromomycin, an aminocyclitol antibiotic isolated from Streptomyces, can be effective when combined use with protease inhibitors or recombinant IL-12. Since there is no drug that achieves the complete removal of Cryptosporidium from the host, supportive therapy was preferred in both human and domestic animals.

Human Pentastomiasis Caused by Armillifer moniliformis in Malaysian Borneo

We report a case of visceral pentastomiasis caused by Armillifer moniliformis in a 70-year-old aboriginal farmer from rural Malaysian Borneo. The patient complained of upper abdominal pain, jaundice, and loss of weight. Radiological investigations and subsequent histopathological examination revealed an adenocarcinoma of the pancreas with an adjacent liver nodule containing a nymph of A. moniliformis. This report constitutes the first documented human pentastomid infection in the whole of Malaysia after nearly 40 years, and it is the third description from Malaysian Borneo. Cases of human and animal pentastomiasis in Malaysia are discussed.

The Importance of Habitat in the Ecology of Decomposition on Rabbit Carcasses in Malaysia: Implications in Forensic Entomology

ABSTRACT The stages of decomposition and the faunal succession on rabbit carcasses in three different habitats, namely jungle, rural, and highland areas, were studied. Three New Zealand White rabbit (Oryctolagus cuniculus) carcasses weighing ∼2kg were sampled daily until the decomposition process was completed. Representative specimens of adult flies, larvae, pupa, and mites were collected from the carcasses and processed in the laboratory. There were differences in decomposition rate and faunal succession between the carcasses. The fastest rate of decomposition was recorded in rural area, and the slowest rate of decomposition was recorded in highland area. The carcasses exhibited the same pattern of colonization by adult flies, but the dominant species of larvae and adult flies on each carcass in specific habitats were different. The primary species of flies recorded in jungle were Chrysomya megacephala F., Chrysomya rufifacies (Macquart), Chrysomya chani Kurahashi, Chrysomya villenuevi Patton, Chrysomya nigripes Aubertin, Chrysomya pinguis (Walker), Hemipyrellia ligurriens (Wiedemann), Hemipyrellia tagaliana (Bigot), Hypopyiopsis fumipennis (Walker), Hypopygiopsis violacea (Macquart), and Hydrotaea spinigera Stein represented by both adults and larvae. Musca domestica L., Atherigona sp., Lioproctia pattoni (Senior-White), Lioproctia saprianovae Pape & Bänziger, and Seniorwhitea princeps (Wiedemann) were represented by adults only. The biodiversity of flies in the rural area were C. megacephala, C. rufifacies, H. ligurriens, Fannia canicularis L., Hydrotaea chalcogaster (Wiedemann), and Hyd. spinigera represented by both adults and larvae, meanwhile M. domestica, Atherigona sp., Boettcherisca peregrina (Robineau-Desvoidy), Parasarcophaga taenionota Wiedemann, Parasarcophaga scopariiformis Senior-White, and S. princeps were represented by adults only. The species of flies collected in the highland area were Lucilia porphyrina (Walker), C. megacephala, C. rufifacies, C. villenuevi, C. pinguis, H. ligurriens, Hyd. spinigera, Hyd. chalcogaster, F. canicularis, and Boettcherisca highlandica Kurahashi & Tan represented by both adults and larvae, whereas C. nigripes, Chrysomya thanomthini Kurahashi & Tumrasvin, M. domestica, Atherigona sp., Parasarcophaga albiceps Meigen, P. taenionota, Sepsidae, Phoridae, and Millichidae were represented by adults only. Faunal succession followed the sequence of dominant flies, i.e., Calliphoridae, Sarcophagidae, Muscidae, Sepsidae, and lastly Stratiomyidae for jungle, or Sepsidae for rural and highland studies. Mites, from suborders Mesostigmata, Prostigmata, Astigmatina, and Oribatida, were also recovered throughout decomposition, which could be used for future implementation in forensic investigations. The data obtained from this study could provide more accurate indicators for local forensic scientists in solving criminal cases especially on the determination of time and primary location of death.

Sarcocystosis among Wild Captive and Zoo Animals in Malaysia

Sarcocystis sp. infection was investigated in 20 necropsied captive wild mammals and 20 birds in 2 petting zoos in Malaysia. The gross post-mortem lesions in mammals showed marbling of the liver with uniform congestion of the intestine, and for birds, there was atrophy of the sternal muscles with hemorrhage and edema of the lungs in 2 birds. Naked eye examination was used for detection of macroscopic sarcocysts, and muscle squash for microscopic type. Only microscopically visible cysts were detected in 8 animals and species identification was not possible. Histological examination of the sections of infected skeletal muscles showed more than 5 sarcocysts in each specimen. No leukocytic infiltration was seen in affected organs. The shape of the cysts was elongated or circular, and the mean size reached 254 x 24.5 µm and the thickness of the wall up to 2.5 µm. Two stages were recognized in the cysts, the peripheral metrocytes and large numbers of crescent shaped merozoites. Out of 40 animals examined, 3 mammals and 5 birds were positive (20%). The infection rate was 15% and 25% in mammals and birds, respectively. Regarding the organs, the infection rate was 50% in the skeletal muscles followed by tongue and heart (37.5%), diaphragm (25%), and esophagus (12.5%). Further ultrastructural studies are required to identify the species of Sarcocystis that infect captive wild animals and their possible role in zoonosis.

Pentaplex PCR Assay for Detection of Hemorrhagic Bacteria from Stool Samples

ABSTRACT Diarrheal diseases cause illness and death among children younger than 10 years in developing countries. Conventional testing for the detection of hemorrhagic bacteria takes 2 to 5 days to yield complete information on the organism and its antibiotic sensitivity pattern. Hence, in the present study, we developed a molecular-based diagnostic assay that identifies common hemorrhagic bacteria in stool samples. A set of specific primers were designed for the detection of Salmonella spp., Shigella spp., enterohemorrhagic Escherichia coli (EHEC), and Campylobacter spp., suitable for use in a one-tube PCR assay. The assay in the present study simultaneously detected five genes, namely, ompC for the Salmonella genus, virA for the Shigella genus, eaeA for EHEC, 16S rRNA for the Campylobacter genus, and hemA for an internal control. Specific primer pairs were successfully designed and simultaneously amplified the targeted genes. Validation with 20 Gram-negative and 17 Gram-positive strains yielded 100% specificity. The limit of detection of the multiplex PCR assay was 1 × 103 CFU at the bacterial cell level and 100 pg at the genomic DNA level. Further evaluation of the multiplex PCR with 223 bacterium-spiked stool specimens revealed 100% sensitivity and specificity. We conclude that the developed multiplex PCR assay was rapid, giving results within 4 h, which is essential for the identification of hemorrhagic bacteria, and it might be useful as an additional diagnostic tool whenever time is important in the diagnosis of hemorrhagic bacteria that cause diarrhea. In addition, the presence of an internal control in the multiplex PCR assay is important for excluding false-negative cases.

Autochthonous Human Schistosomiasis, Malaysia

To the Editor: In Malaysia, the only histologically diagnosed autochthonous cases of human schistosomiasis were reported in the 1970s, all in rural aborigine (Orang Asli) populations (1–3) (Technical Appendix Figure 1). The fact that the infection had been found only among aborigines had led to the proposal of a distinct unknown schistosome with an animal reservoir causing sylvatic infections (2,3). Consequently, during the 1980s, Schistosoma malayensis n. sp. was described from intermediate snail (Robertsiella sp.) and final mammalian hosts (Rattus muelleri and R. tiomanicus [4]). S. malayensis is closely related to S. mekongi and differs genetically from the latter by ≈10%. Both species differ from S. japonicum by 25% (5), and adult and ova morphologies are similar (4). Few transmission sites for this new S. japonicum–complex schistosome species were identified in rural areas (4). We report after 30 years the histologic finding of S. malayensis–like eggs in the liver of a Malay man and discuss public health implications. A 29-year-old male nonaboriginal Malay from Subang Jaya in Selangor State, Peninsular Malaysia, had died suddenly of an intoxication in 2011. According to his mother, he had reported hematuria and dysuria during adolescence. Similar symptoms had reoccurred 10 years later, accompanied by constipation. The patient had never been outside of Malaysia, and he had gone bomb fishing for many years in Sungai Lepar Utara, a river near his village (Felda Tekam Utara, Jerantut, Pahang; 3°52’30”N, 102°49’2”E). No tests on blood or feces were performed before his death. An autopsy was conducted in Sungai Buloh Hospital, and gross pathology showed a normal heart, kidneys, and brain. The lungs were edematous and congested. The liver also was congested, but no macroscopic lesions were seen. Toxicology investigations showed methadone and a derivative in his blood and urine. During a routine histologic examination, several granulomas with intensive lymphocyte, monocyte, and eosinophil infiltration surrounding clusters of ovoidal eggs were found in the liver (Figure; Technical Appendix Figure 2). Serial sectioning showed that the eggs contained miracidia and had the overall appearance of S. malayensis–like ova 50 μm long × 28 μm wide. The ova were not operculated and had no bipolar plugs; the thin yellowish shell was not striated, but a knob-like structure was seen laterally. Morphologic differential diagnoses included eggs of Capillaria hepatica (bipolar striated ova in liver), Dicrocoelium (slightly smaller operculated ova typically found in feces or bile), and the similar Eurytrema (thick-walled operculated ova in feces). Figure Close-up of liver granuloma with section through 3 Schistosoma malayensis-–like ova embedded in dense fibrous tissue. The thin-walled, nonstriated helminth ova are not operculated and contain nonvital miracidial cells. Hematoxylin and eosin stain; ... Schistosomiasis is endemic in many developing countries and infects >207 million persons living in rural agricultural areas (6). In Asia, S. japonicum, S. mekongi, and S. malayensis cause human infection (7), with S. japonicum being the most dangerous. In Malaysia, S. malayensis, in addition to S. spindale, S. nasale, S. incognitum, Trichobilhazia brevis, and Pseudobilharziella lonchurae, is known to occur in wildlife (8). The first known case of human schistosomiasis in Malaysia was discovered in 1973 during an autopsy of an aborigine. Schistosoma eggs resembling those of S. japonicum were found in liver tissue (1). A subsequent retrospective autopsy study revealed additional cases with these Schistosoma japonicum–like ova in the rural aboriginal population, resulting in an overall prevalence of 3.9% (2). Several attempts to recover eggs from feces from the Orang Asli population in peninsular Malaysia (3), a biopsy-positive Orang Asli (3), and serologically positive persons (9, and others) were unsuccessful, however, which was attributed to the zoonotic nature of S. malayensis and thus missing adaptation to the human host. Whether hematuria, a typical sign of S. haematobium infection, as seen in the patient reported here also was caused by S. malayensis disease remains unclear because symptoms of the latter have not been reported. Serologic surveys for schistosomiasis in peninsular Malaysia showed prevalences of 4%–25% in selected rural populations (9). Because infected Robertsiella snails had been found almost exclusively in small rivers (4,9)—habitats like the Sungai Lepar Utara River in our current report—we suspect that the patient most likely became infected while fishing. The travel history may not be accurate because it was obtained from a relative, and possible unreported drug-related travel by the patient to neighboring countries cannot be fully excluded. R. muelleri, the jungle rat and definitive host for S. malayensis, is often seen at river banks (4), and rodent feces could have contaminated the water with schistosome eggs. Future field studies are needed to identify focal hot spots of sylvatic transmission by snail examination and seroprevalence studies of persons living in rural areas, especially the Orang Asli population. Moreover, in light of growing ecotourism, which also encompasses stays at remote Orang Asli villages and canoeing on small streams (10), appropriate public health measures, such as rodent and snail control near tourist sites, should be implemented. Technical Appendix: Places of residence in Peninsular Malaysia of the schistosomiasis-infected Orang Asli from the 1970s and section of liver showing the helminth egg granuloma. 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Preliminary observations of necrophagous dipteran activities and a new host record of parasitoids on a high-rise building

Abstract A preliminary study on the ecological activities of necrophagous Diptera on a highrise building in Malaysia was conducted from April to August 2010. Fresh chicken livers (approximately 500 g) serving as bait were placed on the rooftop (101.6 m from the ground). Three replicates were carried out, each replicate lasting 30 days with daily observations and fly sampling. Maggots found in the bait were collected at random: some were preserved in 70% ethanol while the others were reared to the adult stage. Results showed that the flies arrived on the bait between 8 hours and 3 days after placement. The oriental latrine blow fly Chrysomya megacephala (Diptera: Calliphoridae) was the first to reach this altitude, followed by the scuttle fly Megaselia scalaris (Diptera: Phoridae) and the flesh fly Liopygia ruficornis (Diptera: Sarcophagidae). The larvae of Chrysomya megacephala were first recovered from the bait. However, they were succeeded by sarcophagid larvae. The predominant sarcophagid larvae were then raised to the adult stage and the resulting adults were identified as Parasarcophaga dux. The other sarcophagid pupae were either parasitized by hymenopteran parasitoids or dead. We recorded two species of parasitoids emerging from Liopygia ruficornis pupae: Exoristobia philippinensis (Hymenoptera: Encyrtidae) and Dirhinus himalayanus (Hymenoptera: Chalcididae). Liopygia ruficornis is recorded as a new host for these parasitoids for the first time.