Ivermectin at single doses of 0.2-1.0 mg/kg body weight reduced the microfilarial counts of subperiodic Brugia malayi in Presbytis cristata by 59.9%-89.6% of initial counts, 4 weeks after treatment. Adult filaricidal activity was poor, live adult worms being recovered from all animals at autopsy. There was no serious side effect at these doses.
Gonadectomized male mice aged 5 weeks were given 5 mg testosterone propionate daily for 14 days. The treatment significantly decreased the number of blood leukocytes. The number of all individual types of leukocytes except basophils in vehicle-treated gonadectomized mice was increased. Testosterone-treated mice consistently had a lower number of leukocytes after being infected with Plasmodium berghei than did vehicle-treated mice. The results suggest that testosterone suppresses the production of leukocytes and that testosterone-treated mice become more susceptible to parasite infection.
Presbytis cristata monkeys infected through the inoculation of between 200 and 400 subperiodic Brugia malayi infective larvae (L3) in the right thigh, in both thighs or in the dorsum of the right foot were followed up for varying periods of up to about 8 months after infection. All 148 inoculated animals became patent, with mean prepatent periods being between 66 and 76 days. In animals injected in the thigh, the patterns of microfilaraemia were similar, there being a rapid rise in the geometric mean counts (GMCs) of microfilariae during the first 10-12 weeks of patency, which then plateaued at levels of greater than 1000/ml. Adult worm recovery, expressed as the percentage of the infective dose, was significantly higher in animals injected with 100 L3 in each thigh, being 9.4% as compared with 2.8%-4.8% in other groups. It is therefore recommended that animals should be injected with 100 L3 in each thigh and that the testing of potential filaricides in this model be carried out during the phase of rapid increase in microfilaraemia to ensure that any microfilaricidal effect can easily be detected.
Gonadectomized male albino rats aged 7 weeks were given 1.5 mg/kg testosterone propionate daily and inoculated with 50 third-stage larvae of Angiostrongylus malaysiensis. The treatment significantly increased the number of larvae and adult worms recovered from the brain and pulmonary arteries, respectively, and the rats exhibited smaller thymus glands. The total numbers of leukocytes, monocytes, neutrophils, and especially eosinophils increased significantly post-infection, but the counts were higher in the untreated infected controls. Presumably, immunosuppressive effects of testosterone may at least partly be responsible for the higher loads of A. malaysiensis worms found in male rats as compared with females in the field.
Established criteria for morphological typing of sarcocysts was applied to a large series of cases of human skeletal muscle sarcocystosis in Malaysia to determine the type of sarcocyst present. We also wanted to test the general usefulness of this classification and to determine if there are any new cyst types. Three-dimensional (3-D) reconstruction was done to see if the sarcocyst has a distinct 3-D morphology. A total of 66 sarcocysts from 21 cases of human muscle sarcocystosis obtained from a previous prevalence study were examined. Tissue sections (5 microns thick) were stained with haematoxylin and eosin and studied under the light microscope. For 3-D reconstruction, an image analyser was used to align and reconstruct the sarcocyst after microscopic images had been captured with a charge-coupled device (CCD) camera. All the cysts best fit into the type 4 category. This classification is generally useful, although cyst wall characteristics and zoite size appear to be the most reliable criteria for classification. The cyst width averaged 77 microns (range, 30-137.5 microns). Cyst walls were smooth, had no cytophaneres and were less than 1 micron thick. No secondary cyst wall or surrounding inflammation was evident. Numerous cyst merozoites with diameters averaging 1 micron filled the cyst lumen. Although septa were not apparent, in many cysts, zoites were arranged in a unique, curvilinear fashion that suggested their presence. 3-D reconstruction showed the sarcocyst to be a long, tortuous "cylinder" with no branching or other distinguishing feature.
Gonadectomized male laboratory rats were given 0.06 mg/kg estradiol benzoate daily for 14 days before being inoculated with 50 third-stage larvae of Parastrongylus malaysiensis. Hormone treatment was continued until the rats were killed. The numbers of larvae in the brain and of adult worms in the pulmonary area of the rats were determined every 7 days after the inoculation. It was found that the rats treated daily with estradiol benzoate had significantly and consistently higher numbers of larvae and adult worms as compared with the controls. The number of total leukocytes increased significantly after the rats were infected. The results show that estradiol-treated rats become susceptible to P. malaysiensis infection, which may indicate that the immunosuppressive effects of testosterone observed in earlier studies may partly be caused by estradiol that was peripherally aromatized from testosterone.
Daily intramuscular injection with thyroxine (T4) at a dose of 2.5 micrograms/100 g body weight decreased the larvae and adult worm burden of Parastrongylus malaysiensis in the brain and pulmonary arteries of male Sprague-Dawley albino rats. In contrast, rats treated with propyl thiouracil (PTU), an antithyroid drug, at a dose of 3.75 mg/100 g body weight retained greater numbers of larvae and adult worms. The results may reflect the contrasting immunomodulatory effects of T4 and PTU that influence the susceptibility of the host.
The intraspecific variation of four laboratory-reared isolates of Taenia taeniaformis the SRN and KRN isolates from Norway rats, Rattus norvegicus, captured in Japan and Malaysia, respectively; the BMM isolated from a house mouse, Mus musculus, captured in Belgium; and the ACR isolate from a gray red-backed vole, Clethrionomys rufocanus bedfordiae, captured in Japan was examined by various criteria. Eggs of each of the four isolates were orally inoculated into several species of intermediate host. They were most infective to the rodent species from which the original metacestode of each isolate had been isolated in the field, and only the ACR isolate was infective to the gray red-backed vole. Although little difference was found between the SRN, KRN, and BMM isolates by the other criteria, including the morphology of rostellar hooks, the protein composition of the metacestode, and restriction endonuclease analysis of DNA, the ACR isolate was clearly different from the others. It was considered that the ACR isolate was independent as a strain distinct from the other three isolates.
A new sanguinicolid blood fluke, Parasanguinicola vastispina, is described from sea bass Lates calcarifer cultured in Malaysia. It is distinguished by its massive armature and widely spaced genital pores, the female pore being pre-ovarian. P. vastispina inhabits the branchial arteries, dorsal aorta, mesenteric venules and renal artery of its host. No pathological effect was observed in infected fish.
For elucidation of the taxonomic status of the Japanese Fasciola species, whole mitochondrial DNA of Fasciola hepatica from Australia, F. gigantica from Malaysia, and Fasciola sp. from Japan was digested with three four-base-cutting endonucleases: HinfI, MspI, and RsaI. The resulting digestion patterns showed that for each enzyme there were some bands specific for each geographical isolate and that the Japanese Fasciola sp. shared more bands with F. gigantica than with F. hepatica. Nucleotide sequences of two regions, the second internal transcribed spacer (ITS2) of the nuclear ribosomal RNA cluster and mitochondrial cytochrome c oxidase subunit I (COI), were also compared among them. The ITS2 sequence was highly conserved among the three isolates. F. gigantica and the Japanese Fasciola sp. were identical, but they differed from the Australian F. hepatica at six sites, one of which was a deletion. The COI sequence was less conserved but implied a similar relationship between the isolates. There seems no reason to regard the Japanese Fasciola sp. as anything other than a strain of F. gigantica.
The shedding pattern of the protozoan parasite, Blastocystis hominis, is investigated in man and in experimental animal infections. The shedding pattern of the vacuolar and cystic forms of Blastocystis hominis in infected individuals have been shown in the present study to be irregular. The study shows that there is marked fluctuation in the shedding of the parasite from day to day, varying from as high as 17 to 0 per x40 microscopic field. The cystic stages when estimated in 8 Blastocystis-infected individuals ranged from as high as 7.4x10(5) cysts per gram of stool to 0. The shedding of cystic and vacuolar forms observed over a period of 20 days in experimentally-infected Wistar rats were not only shown to be irregular but the amount varied from host to host. The study has important diagnostic implications in that the stool samples must be collected more than once from patients showing clinical signs and symptoms to eliminate the cause of it to Blastocystis. The study also shows that there are asymptomatic individuals who pass a large amount of cysts as such individuals should be treated to prevent transmission to others.
A total of 20 isolates of Blastocystis were characterized using a single set of polymerase chain reaction (PCR) primers. The amplification product revealed five types of pattern. All four isolates from Singapore yielded PCR products quite different from those of the local isolates. However, most of the local isolates showed a major product at either 280 or 500 bp, or both. We also suspected that the amplification product detected at 280 bp might be an indicator of the pathogenicity of this parasite. One isolate (M12) obtained from a monkey showed patterns similar to those of human isolates (10203 and KP1) and probably belongs to the same strain. The results indicate that the intraspecific or interstrain variations in these 20 Blastocystis isolates belong to 5 different patterns. The differences among isolates of the same strain revealed by the presence or absence of certain amplification products showed further intrastrain variations in this parasite.
Acanthamoeba sp. is a free-living amoeba known to cause chronic central nervous system infection or eye infection in humans. Many cases remain undetected for want of a good detection system. We report for the first time a rapid staining method to facilitate the identification of Acanthamoeba sp. using the modified Field's staining technique. A. castellanii, which was used in the present experiment, is maintained in our laboratory in mycological peptone medium (Gibco). The cultures were pooled together and smears were made on glass slides for staining purposes. Different types of stains such as Field's stain, modified Field's stain, Wright's stain, Giemsa stain, Ziehl-Neelsen stain, and trichrome stain were used to determine the best stain for the identification of this amoeba. The concentration of various stains and the duration of staining were varied to provide the best color and contrast for each stain. Acanthamoeba was also obtained from the brain of experimentally infected mice and was stained with various stains as mentioned above to determine the best stain for use in identifying the presence of this parasite in experimentally infected animals. The modified Field's stain gives a very good color contrast as compared with other stains. Furthermore, it takes only 20 s to be carried out using the least number of reagents, making it suitable for both laboratory and field use.
The present study investigated whether people working closely with animals were at higher risk of getting infected with Blastocystis hominis. The prevalence of the parasite was determined in two population groups, i.e., animal handlers and normal healthy individuals who did not work with animals. In all, 105 stool samples were collected from animal handlers from 2 local research institutions, a local zoo, and a local abattoir and 163 stool samples were collected from normal healthy individuals residing in high-rise flats in the city. The in vitro culture method used in the study detected that 41% of 105 animal handlers and 17% of 163 flat-dwellers in the city were positive for Blastocystis. This statistically significant finding (P = 0.0000313) shows that people who work closely with animals do stand at risk of acquiring Blastocystis infection.
In this study, the genome of the Plasmodium falciparum Gombak A strain was examined for the presence of a gene encoding falcipain-2, a cysteine protease, using homology-based polymerase chain reaction cloning. The nucleotide sequence obtained from the gene cloned (designated pFG1) is approximately 99% homologous to other falcipain-2 genes from different strains. Comparatively, it is 69% homologous to falcipain-3 genes. Direct cloning of the falcipain-2 gene and its resemblance to the reported corresponding mRNA transcript suggests the absence of introns in this gene. Sequence alignment and comparison revealed four amino acid differences at positions 15, 51, 59 and 414 in the falcipain-2 from P. falciparum Gombak A as compared to other falcipain-2 proteins from different strains.
A gene encoding the larval excretory-secretory antigen TES-120 of the dog ascarid worm Toxocara canis was cloned into the methylotrophic yeast Pichia pastoris. Specificity of the recombinant TES-120 antigen produced by the yeast was investigated. Forty-five human serum samples from patients infected with different()parasitic organisms, including 8 cases of toxocariasis, were tested against the recombinant antigen in immunoblot assays. Results from the assays showed that the recombinant TES-120 antigen reacted with sera from toxocariasis patients only. This highly specific recombinant TES-120 antigen can potentially be used for the development of an inexpensive serodiagnostic assay for human toxocariasis.
Blastocystis hominis is one of the most common human parasites that inhabit the intestinal tract. Conflicting reports continue to exist regarding the existence and the functional role of the amoeboid forms in the life cycle of the parasite. The present study investigates the presence of these forms in 20 isolates obtained from ten symptomatic and asymptomatic patients respectively. A total of 10,000 parasite cells per ml from each isolate were inoculated into three culture tubes each containing 3 ml of Jones' medium supplemented with 10% horse serum, incubated at 37 degrees C. The contents were examined daily for 10 days. Irregular and polymorphic amoeboid forms with multiple extended pseudopodia were observed in all isolates from symptomatic patients, while none of the isolates from asymptomatic patients showed the presence of the amoeboid forms. The amoeboid forms were initially noted on day 2 and the percentages increased from 2% to 28%, with peak percentages from day 3 to day 6. Transmission electron microscopy revealed two types of amoeboid forms; one containing a large central vacuole completely filled with tiny electron-dense granules, and the other which revealed multiple small vacuoles within the central body. The cytoplasm contained strands of electron-dense granules resembling rough endoplasmatic reticulum, which is suggestive of active protein synthesis. The surface coat of the amoeboid form surrounding the parasite showed uneven thickness. Acridine orange stained the central body yellow and the periphery orange, indicating activity at the level of nucleic acids. The amoeboid form could either be an indicator of pathogenicity of B. hominis, or the form likely to contribute to pathogenicity and be responsible for the symptoms seen in patients.