The current study investigated the occurrence of gastrointestinal tract (GIT) protozoa and associated risk factors in free range pigs in Busia District, Kenya. A total of 306 pigs from 135 farms in 6 Divisions were sampled for feces, which were analysed for parasites using direct smear and McMaster floatation methods. Associations between the occurrence of the parasites and explanatory variables (sex, age, division of origin and rainfall) were undertaken using ANOVA, chi-square and Pearson’s correlation statistics. The following gastrointestinal protozoan parasites were identified: Entamoeba spp. (87%), Balantidium coli (64%), Tritrichomonas suis (42%) and Coccidia spp (33%). The mean coccidial oocysts per gram (OPG) of all the sampled pigs was 1,276 (range = 0-28,000 OPG) and the proportions of the species included: Eimeria debliecki (40%), E. suis (26%), E. porci (16%), E. scabra (13%) and E. polita (5%). There was negative correlation between the amount of rainfall in the division of pig origin and prevalence of Eimeria spp, Tt. suis, and Entamoeba spp, but a positive correlation with prevalence of B. coli. The prevalences of Eimeria spp., Entamoeba spp. and Tt. suis were higher in males than females; but it was only the sex-differences for Tt. suis which were statistically significant (p <0.05) . The prevalences of Tt. suis in sows were significantly (p <0.05) lower than that of growers and piglets. It was concluded that GIT protozoan parasites of economic and zoonotic significance occur in pigs in the study area and effective control strategies should be implemented.
The vervet monkey (Chlorocebus aethiops) model of sleeping sickness was used to evaluate the effectiveness of TrypTectCIATT in assessing the success of trypanocidal therapy. A retrospective study was therefore conducted on sera collected from monkeys infected with Trypanosoma brucei rhodesiense and treated either curatively with melarsoprol or sub-curatively with diminazene aceturate. In the human survey, 440 sera collected from 96 human patients were tested. These patients were treated with either suramin or melarsoprol depending on the stage of the disease. An extra 56 parasitologically positive pre-treatment samples were also tested to aid in determination of the test sensitivity. Results indicated that between 21-28 days post-infection, the test detected trypanosomal antigens in 84.2% (16/19) of animal samples that were parasitologically positive by the hematocrit centrifugation technique（HCT). In curatively treated animals, 77.8% (7/9) exhibited positive reaction up to 9 months post-treatment. One animal was positive for trypanosomal antigens for the entire 12 months while one was a non-reactor. From the sub-curatively treated group, 80% (8/10) were detected positive for the entire 12 months while, 2 animals were non-reactors. In the human survey, 3 patterns of antigen profiles were observed. In some patients, there was fluctuation of antigen levels throughout the 12 months follow-up period. In others, antigens were detected for the entire 12 months but in decreasing levels. The last group was that of patients with antigens decreasing at different rates to undetectable levels at 12 months post-treatment. The presence of trypanosome positive but antigen negative samples during the study raises a few questions with regards to the sensitivity of the test. It is however evident that the test was able to detect trypanosomal antigens in over 80% of positive monkey and human serum samples. Consequently, TrypTectCIATT may be an important additional tool in reduction of the follow-up period and determination of success of chemotherapy in sleeping sickness.
Parasitemia and rectal temperatures were determined in Yankasa sheep experimentally infected with Trypanosoma congolense and immunomodulated with levamisole. Group A (6 sheep) and group B (6 sheep) were infected with 2 x 106T. congolense parasites. In addition Group B sheep were each administered with 2.5 mg/kg levamisole hydrochloride subcutaneously on the day of infection and later weekly throughout the experimental period. Group C (5 sheep) served as the uninfected controls. Parasitemia in the T. congolense infected immunomodulated group appeared 2 days earlier than the group with T. congolense infection only and the parasitemic level was also higher in the infected immunomodulated group at the first parasitemic peak. Rectal temperatures were significantly (p < 0.001) higher in the infected groups when compared the control but were not significantly different (p > 0.05) between the infected groups. It appears that levamisole immunomodulation of T. congolense sheep enhanced the early appearance of parasitemia but was not associated with mortality as all the infected animals survived the experimental period which lasted 6 weeks.
The efficacy of a bull cryopreservation medium Triladyl® in preservation of the various pathogenic trypanosomes was evaluated against the commonly used glycerol ethylenediamine tetraacetic acid (EDTA) saline glucose (ESG). Trypanosoma brucei rhodesiense, T. evansi, T. b. brucei and T. congolense were first propagated in Swiss White mice. At peak of parasitemia, blood was collected from the mice by cardiac puncture and 1 x 102 trypanosomes/ml dilution made. This dilution was then divided into 2 equal portions which were cryopreserved in either ESG 10% glycerol or Triladyl® 50%. After 1 month of cryopreservation, 2 irradiated mice were intraperitoneally administered with 0.2 ml of the cryopreserved sample and then monitored for parasitemia development. Triladyl-cryopreserved T .evansi and ESG-cryopreserved T. congolense had shorter although not significant prepatent period (PPP) compared to their counterparts in the alternate medium. The peak parasitemia was significantly higher for Triladyl®-cryopreserved T. evansi trypanosomes compared to that preserved in ESG. The parasitemia pattern for T. evansi, T. b. brucei and T. congolense were characterized by 1 wave while T. b. rhodesiense had 2 waves. For T. b. brucei, T. b rhodesiense and T. congolense the number of days taken to develop peak parasitemia were shortest for ESG cryopreserved trypanosomes and this was opposite for T. evansi. In conclusion, Triladyl® can be used as an alternative cryopreservative medium for trypanosomes and would be the cryopreservative medium of choice for T. evansi trypanosomes based on the PPP, peak parasitaemia and days to peak parasitaemia.
A study was undertaken to identify the immunogenic proteins in whole cell lysate (WCL) antigens (Ags) of Trypanosoma evansi isolated from 3 different Indian hosts; cattle, buffalo and horse, to characterize the antigenic epitopes of WCL Ags responsible for antigen-antibody reaction, using naturally T. evansi infected cattle sera (immune sera) by Western blot analysis. This study was conducted to identify the common immunoreactive Ags within these 3 isolates, and to observe the cross reaction of this sera with WCL Ags prepared from cattle, buffalo and horse isolates, so that T. evansi isolate of 1 species of animal could be used for sero-diagnosis of T. evansi infection in other species of animals. Naturally T. evansi infected cattle sera recognized a total of 13 numbers of immunogenic proteins of relative molecular weight (MW) range 159 to 14 kDa, as common immunogenic proteins in the WCL T. evansi Ags prepared from buffalo, cattle and horse isolates. Out of these 13 polypeptides, 9 polypeptides of MW ranges 73-74, 59-61, 46-48, 38-40, 30-33, 24-25, 18, 16 and 14 kDa were considered as major immunedominant polypeptides and 4 polypeptides of MW ranges 159, 92, 83-84 and 21 were considered as minor immunogenic polypeptides. Thus, it has been concluded from this study that naturally T. evansi infected cattle sera could react with WCL Ags prepared from T. evansi of cattle origin i.e. with homologous Ags as well as cross react with Ags prepared from T. evansi of buffalo and horse origins i.e. with heterologous Ags, which might be helpful for diagnosis in respect of detection of Abs in a particular species of host using the Ags prepared from T. evansi isolated from different species of host.
Two chronically infected cattle suspected for Trypanosoma evansi infection were studied for confirmative diagnosis and isolation of parasites. One zebu cattle with symptoms of intermittent rise of temperature (38.3℃ to 41.1℃), bottle jaw and chronic emaciation, found positive for T evansi in Giemsa stained blood smears, but found negative for T evansi after mouse inoculation test (MIT). Other crossbred cattle with symptoms of intermittent rise of temperature (38.3℃ to 41.7℃) and decreased milk yield, found negative for T evansi infection in Giemsa stained blood smears, but found positive after repetition of MIT. In such MIT only 25% inoculated mice of batch showed positivity in MIT. The presence of anti-T. evansi antibody was detected in serum samples of both of these cattle, as observed in indirect ELISA test. It has been concluded that cattle positive for T evansi infection in blood smears may not always positive in MIT and negative in blood smears for T. evansi, may be positive in MIT, which may need repeated inoculation of suspected blood to mice in different time intervals and in different batches to get positive result in MIT, in field condition. Repetition of MIT for successful result for isolation of T evansi in field condition is reported.
The anti-trypanosomal potential of aqueous stem extract of Jatropha gossypiifolia var gossypiifolia was investigated in vitro and the resultant effect on the infectivity of Trypanosoma congolense in Wistar rats was evaluated. Motility and mortality of the parasite was monitored over different incubation periods (0 min. - 1 hr 45 min at 37℃) with serial concentrations of the extract (6.25 mg/ml, 12.5 mg/ml, 25.0 mg/ml, 50.0 mg/ml and 100 mg/ml) prepared in phosphate buffered saline (PBS, pH 7.2). Activity was recorded at the higher concentrations when compared with the standard drug, isometamidium chloride (samorin MERIAL). This dose dependent activity delayed the onset/establishment of T. congolense infection in infectivity test in rats with resultant increase in prepatent period. This suggests possible anti-trypanosomal effect of this extract in vivo.
A survey of Tsetse and animal Trypanosomiasis was carried out in 2 Local Government Areas (LGA) of Bauchi state. These are Alkaleri LGA and Toro LGA. Five settlements in Duguri district comprising; Yankari game reserve, Lim, Birim, Yuli and Kungibar in Alkaleri LGA was surveyed, while 8 villages in 3 districts of Toro LGA were surveyed. These are Toro (Tilded-Fulani, Bujel, funde and Ribina), Jama’a (Fanshanu) and Lame (Gumau, Sanga and Laru). A total of 93 biting files were caught in the Toro LGAs. Seventy seven (82.8%） from Duguri districts comprising of 60 (78.0%) Glossina tachinoides, 10 (13.0%) G. palpalis, 3 (4.0%) Tabanus, 3 (4.0%) Stomoxys and 1 (1.0%) Chrysops. In Toro district 8 (8.6%) biting flies were caught; 4 (50.0%) G. tachinoides, 1 (12.5%) G. palpalis, 1 (12.5%) Tabanus, 1 (12.5%) Stomoxys and 1 (12.5%) Chrysops. In Lame districts 4 (4.3%) biting flies were caught 1 (25.0%) Tabanids and 3 (75.0%) Stomoxys, catches of 4 (4.3%) of biting flies were made in Jama’a district, 1 (25 .0%) G. tachinoides, 2 (50.0%) Tabanus and 1 (25.0%) Stomoxys. A total of 770 animals blood samples from ruminants (cattle, sheep, goat and dog) were analysed. Four hundred and forty eight blood samples were collected; from cattle (277), sheep (90) and Goats (81) in Duguri districts. While 322 (41.8%) blood samples from ruminants (cattle, sheep, Goat and dog) from 3 districts were analysed for Trypanosoma, 23 (47.9%) samples in Duguri district was positive; for Trypanosoma brucei 1 (4.3%), T. vivax 9 (39.1%), T. congolense 1 (4.3%) and Babesia spp. 12 (52 .2%). In Toro district 10 (6.7%) were positive; for T. brucei 2 (20.0%), T. vivax 2 (20.0%), T. congolense 1 (10.0%), Babesia spp. 4 (40.0%) and microfilaria 1 (10.0%). In Jama'a district 2 (3.8%) were positive, for T. brucei 1 (50.0%) and Babesia spp 1 (50.0%). In Lame district 11 (9.2%) blood samples were positive; for T. brucei 1 (9.1%), T. vivax 4 (36.3%), T. congolense 1 (9.1%) and Babesia spp. 5 (45.5%). The parasites have no sex or animal species preference. The public health significance of these findings to human health is discussed.
A novel gene encoding a 47-kDa surface protein was identified by bioinformatic analyses of Expressed Sequence Tags (EST) prepared from mRNA of Babesia gibsoni. The Bgp47 cDNA encodes a 47-kDa polypeptide having an-N terminal signal peptide, a central hydrophilic core and a C-terminal transmembrane domain. This protein shares homology with 50-kDa (BgP50) and 32-kDa (BgP32) proteins of B. gibsoni. The expressed recombinant protein without the signal peptide and the transmembrane domain (BgP47t) was approximately 46-kDa after glutathione S-transferase （GST) cleavage, similar to an approximately 47-kDa mature native protein identified in the parasite merozoite after probing with mouse anti-BgP47t serum. A 32-kDa band was also identified using the same antiserum suggesting that BgP47 and BgP32 might share common B cell epitopes. The BgP47 was localized predominantly on the parasite cell surface. An enzyme-linked immunosorbent assay (ELISA) revealed that the rBgP47t did not cross-react with B. canis subspecies and other genetically related apicomplexans indicating that the antigen is specific. Out of 106 sera from dogs with B. gibsoni-like symptoms, 79.2% were seropositive by rBgP47t-ELISA. Therefore, the BgP47 is prospective serodiagnostic antigen.
It is not known whether precursors of methotrexate, such as 2, 4-diamino-6-hydroxymethy1-pteridine (DAP) and 2, 4-diamino-N10-methy1-pteroic acid (DAMPA), could target the dihydrofolate reductase-thymidylate synthase (DHFR-TS) enzyme of Babesia and inhibit the parasite growth. Therefore, we have determined whether DAP and DAMPA as well as other chemically related compounds like pteroic acid (PA) and N10-Triflouropteroic acid (N10TFPA) could target the DHFR-TS enzyme of B. gibsoni and inhibit its growth. DAMPA was a more-potent inhibitor of the B. gibsoni growth in vitro (50% inhibition concentration [IC50] = 2.4 ± 0.20 µM) [mean ± standard error of the mean] than DAP (IC50 = 78 ± 15 µM). Moreover, DAMPA potently inhibited enzymatic activity of recombinant DHFR-TS of B. gibsoni (IC50 = 2.6 ± 0.15 µM) than DAP (IC50 > 100 µM). In contrast, PA and N10-TFPA did not inhibit the activity of the recombinant enzyme and growth of B. gibsoni. The inhibition of the recombinant enzyme activity by DAMPA mirrored with
inhibition of the parasite growth indicating that the purified recombinant enzyme could be used for preliminary screening of some antifolate precursors. Therefore, both DAP and DAMPA inhibit growth of B. gibsoni by targeting the DHFR-TS enzyme of the parasite.
Apicomplexan parasites have been shown to elicit a potent, cell-mediated immune response that can not only efficiently control the growth of the invading parasite, but also lead to nonspecific resistance to unrelated pathogens. In this study, we identified a cross-protective antigen of Plasmodium falciparum able to control Toxoplasma gondii infection.