Towards disentangling the classification of freshwater fish trypanosomes

Currently,new species of freshwater fish trypanosomes,which are economically important parasites,are being described based on subjectively selected features,i.e.,their cell morphology and the host species.We have performed detailed phylogenetic and haplotype diversity analyses of all 18S rRNA genes available for freshwater fish trypanosomes,including the newly obtained sequences of Trypanosoma carassii and Trypanosoma danilewskyi.Based on a sequence similarity of 99.5%,we divide these trypanosomes into 15 operational taxonomic units,and propose three nominal scenarios for distinguishing T.carassii and other aquatic trypanosomes.We find evidences for the existence of a low number of freshwater fish trypanosomes,with T.carassii having the widest geographic and host ranges.Our analyses support the existence of an umbrella complex composed of T.carassii and two sister species.

The diurnal activity, movement and trypanosome infection rates of Glossina fuscipes fuscipes （Diptera： Glossinidae） in Buvuma Island, Lake Victoria, Uganda

The diurnal activity patterns, trypanosome infection rates and movement of Glossinafuscipesfuscipes （Diptera： Glossinidae） were investigated in Buvuma Island, Lake Victoria, Uganda. Hourly trapping of tsetse flies was undertaken to determine their activity rhythm while a capture-mark-release-recapture method was conducted to assess the movement and dispersal of tsetse flies between lakeshore, hinterland and further inland sites along a transected area. Dissection of tsetse flies was also undertaken to determine the trypanosome infection rates in salivary glands, proboscis and mid-gut. Results indicated a bimodal diurnal activity profile for G. f fuscipes on the Island, both on the lakeshore and in the hinterland. Movement and dispersal of G. f fuscipes tsetse flies occurred between lakeshore, hinterland and further inland sites with a greater tendency of flies to move to the lakeshore. Trypanosome infection rates of 4.32% for Trypasoma vivax and 1.15% for 7. congolense were found in G. f. fuscipes.

Patterns of tsetse abundance and trypanosome infection rates among habitats of surveyed villages in Maasai steppe of northern Tanzania

Background:Changes of land cover modify the characteristics of habitat,host-vector interaction and consequently infection rates of disease causing agents.In this paper,we report variations in tsetse distribution patterns,abundance and infection rates in relation to habitat types and age in the Maasai Steppe of northern Tanzania.In Africa,Tsetse-transmitted trypanosomiasis negatively impacted human life where about 40 million people are at risk of contracting the disease with dramatic socio-economical consequences,for instance,loss of livestock,animal productivity,and manpower.Methods:We trapped tsetse flies in dry and wet seasons between October 2014 and May 2015 in selected habitats across four villages:Emboreet,Loiborsireet,Kimotorok and Oltukai adjacent to protected areas.Data collected include number and species of tsetse flies caught in baited traps,PCR identification of trypanosome species and extraction of monitored Normalized Difference Vegetation Index(NDVI)data from Moderate Resolution Imaging Spectrometer(MODIS).Results:Our findings demonstrate the variation of tsetse fly species abundance and infection rates among habitats in surveyed villages in relation to NDVI and host abundance.Results have shown higher tsetse fly abundance in Acacia-swampy ecotone and riverine habitats for Emboreet and other villages,respectively.Tsetse abundance was inconsistent among habitats in different villages.Emboreet was highly infested with Glossina swynnertoni(68%)in ecotone and swampy habitats followed by G.morsitans(28%)and G.pallidipes(4%)in riverine habitat.In the remaining villages,the dominant tsetse fly species by 95%was G.pallidipes in all habitats.Trypanosoma vivax was the most prevalent species in all infected flies(95%)with few observations of co-infections(with T.congolense or T.brucei).Conclusions:The findings of this study provide a framework to mapping hotspots of tsetse infestation and trypanosomiasis infection and enhance the communities to plan for effective control of trypanosomiasis.

Antitrypanosomal potentials of the extract and fractions of Abrus precatorius seeds

Objective:To evaluate the in vivo trypanocidal activity of the methanol extract and fractions of Abrus precatorius seeds in mice.Methods:Parasiteamia was induced unto mice by intraperitoneal injection of 1.25×105 Trypanosoma in normal saline.Five days when a high level of parasiteamia was established treatment commenced until ten days.The mice were treated with 10,20 and 40 mg/kg bt.of the extract and 5 and 10 mg/kg bt.of the fraction（F2）,respectively for 5 days.Diminazene acelurate at the dose of 3.5 mg/kg bt.for two days was used as the reference drug.The level of parasitaemia and packed cell volume（PCV） of the animals estimated. Results:At doses of 10,20 and 40 mg/kg the crude extract showed a sharp reduction in the level of parasitaemia in mice compared with the untreated group.The mice treated with F,at doses of 5 and 10 mg/kg showed a sharp reduction in the level of parasitamia to zero in day 9,and a gradual recovery from the 12th day of treatment.This effect is comparable to that of the mice treated with 7 mg/kg of standard drug diminazene aceturate.The PCV of the treated showed a gradual decrease with time,but not as much as the untreated group.Phytochemical screening revealed the presence of glycosides,alkaloids,carbohydrates,tannins and proteins in the Abrus precatorius powder while F2 was rich in alkaloids.Conclusions:This study shows that both the extract and the fractions of Abrus precatorius seeds exhibited a promising trypanocidal property.Alkaloids may be responsible for the observed activity.

The effects of combination of methanolic leaf extract of Azadirachta indica and diminazene diaceturate in the treatment of experimental Trypanosoma brucei brucei infection in rats

Objective:To investigate the effects of combination therapy of methanolic leaf extract of Azadirachta indica(A.indica) and diminazene diaceturate(DDA) in the treatment of experimental Trypanosoma brucei brucei(T.brucei brucei) infection in rats.Methods:Acute toxicity study of the drug and extract combinations were done.Selection of the best drug and extract combinations was carried out using fifty four rats of both sexes separated into 9 groups.Three dose combinations were derived from selection of the best drug and extract combinations used for the final study viz:7 mg/kg bw DDA plus 125 mg/kg bw extract(group B),3.5 mg/kg bw DDA plus 250 mg/kg bw extract(group C),and 1.8 mg/kg bw DDA plus 500 mg/kg bw extract(group D).The final study had in addition to the three groups derived from the dose response study,four other groups viz:uninfected untreated negative control(group F),infected and treated with 3 000 mg/kg bw extract alone(group E),infected and treated with 7 mg/kg bw DDA alone(group A),and infected untreated positive control(group G).The parameters assessed were onset of parasitaemia(OP), level of parasitaemia(LOP),clearance of parasites post treatment(COPPT),relapse infection period(RIP),post infection survival period(PIST).Results:There was no significant difference in OP between the groups(P【0.05).One day post treatment,the mean LOP of groups A,B,and C were found to be significantly lower than that of group D which in turn was lower than that of group E and G respectively.The mean LOP of group E was significantly lower than group G two days post treatment and this trend continued throughout the experimental period.Mean COPPT of group D was significantly longer than that of groups A,C and B.There was no significant difference in the mean COPPT among groups B,C and A.The mean RIP of group D was significantly shorter than group C,and that of group C was significantly shorter than that of group A.There was no relapse of infection in group B.The PIST of group E did not differ significantly from group G.Conclusions:This experiment stands to conclude that combination of 125 mg/kg bw extract and 7 mg/kg bw DDA is very effective in the treatment of trypanosomosis,caused by T. brucei.This combination therapy proved to be better than single therapy of DDA.

Improved Detection of Sleeping Sickness Cases by LED Fluorescence Microscopy: Evidence from a Prospective Multi-Centric Study in the Democratic Republic of the Congo

Background: Confirmatory diagnosis of Trypanosoma brucei gambiense human African trypanosomiasis (HAT) is based on demonstration of parasites by microscopy. However, the sensitivity of routine microscopy methods is very low, and many cases are missed and left untreated. A clinical study was conducted in the Democratic Republic of the Congo to evaluate the accuracy of improved microscopy methods in diagnosis of HAT. These included examination by fluorescence microscopy (FM) of acridine orange (AO) stained smears of whole blood and smears made following a new procedure for concentrating trypanosomes by selective lysis of red blood cells (RBC). Methodology/Principal Findings: Venous blood was collected from 213 HAT cases, 101 HAT suspects and 95 controls and used to determine the accuracy of four microscopy methods: bright field microscopy of Giemsa-stained thick blood smears, FM of AO-stained thick blood smears, FM of AO-stained thick blood smears prepared after RBC lysis and concentration, and FM of AO-stained thin blood smears prepared after RBC lysis and concentration. The sensitivity of FM using thick blood smears stained with AO was 3 times higher than bright field microscopy using Giemsa-stained thick blood smears [19.7% (95% CI: 14.9% - 25.6%) versus 6.1% (95% CI: 3.6% - 10.2%)]. When the RBC lysis and concentration procedure was included, sensitivity of the test was further enhanced to 23.0% (95% CI: 17.9% - 29.1%) with thick blood smears and 34.3% (95% CI: 28.2% - 40.9%) with thin blood smears. Specificity of all four microscopy methods was 100% (95% CI: 96.1% - 100.0%). However, the miniature anion exchange chromatography technique (mAECT) and capillary tube centrifugation (CTC) method remained more sensitive. Conclusions: These new methods have practical advantages, including shorter staining time, ease of demonstration of parasites, and the possibility of archiving slides. They could, therefore, be alternative methods to improve case detection where concentration procedures such as mAECT or CTC are not performed.

In vitro and in vivo trypanocidal action of aescin and aescin liposomes against Trypanosoma evansi in experimental mice

Objective:To verify the trypanocidal effectiveness of aescin and aescin liposomes against Trypanosoma evansi in vitro and in vivo.Methods:Aescin and aescin liposomes were used in vitro on trypomastigotes at different concentrations(0.5%,1.0%and 2.0%) and exposure times(0,1,3,6 and 9 h).In vivo tests were performed using mice as the experimental model.Trypanosome evansi infected mice were treated with aescin and aescin liposomes with doses of 60 and 100 mg/kg during 4 d.Results:The three concentrations tested in free form and nanoencapsulated showed trypanocidal activity in vitro,completely eliminating the parasites in small concentration after6 h of assay.Animals treated with aescin(100 mg/kg) and aescin liposomes(100 mg/kg)showed increase in longevity,however without curative effect.Conclusions:Active compounds present in natural products,such as aescin,may potentiate the treatment of trypanosomosis when used in association with other trypanocidal drugs.

Effect of experimental single Ancylostoma caninum and mixed infections of Trypanosoma brucei and Trypanosoma congolense on the humoural immune response to anti-rabies vaccination in dogs

Objective:To determine the effect of Ancylostoma caninum(A.caninum)and trypanosome parasites on the immune response to vaccination in dogs in endemic environments.Methods:Sixteen dogs for the experiment were grouped into 4 of 4 members each.Group I was the uninfected control one,and GPII was infected with A.caninum;GPIII was infected with A.caninum/Trypanosoma congolense(T.congolense),and GPIV was infected with Trypanosoma brucei(T.brucei)/A.caninum.The dogs were first vaccinated with antirabies vaccine before infecting GPII,GPIII and GPIV with A.caninum which were done 4 weeks after vaccination.By 2-week post-vaccination,trypanosome parasites were superimposed on both GPIII and GPIV.A secondary vaccination was given to GPI,GPII,GPIII,and GPIV by Week 12 of the experiment(4 weeks post treatment).Results:The prepatent period was(3.00±1.40)days,in the conjunct infection of T.brucei/A.caninum.It was(9.00±1.10)days,in conjunct T.congolense/A.caninum.The prepatent period of A.caninum was(14.0±2.0)days in the single A.caninum group and(13.0±1.0)days in the conjunct trypanosome/A.caninum.At the 1st week after vaccination,the antibody titer in all the vaccinated groups(GPI,GPII,GPIII,and GPIV)significantly increased(P<0.05)and peaked at the 3rd week after vaccination.Following infections,there were marked significant decreases(P<0.05)in the antibody production against rabies in GPII,GPIII and GPIV.The significant decrease(P<0.05)in antibody titer was highest in the conjunct groups(GPIII and GPIV)compared to the single infection(GPII).Treatment with diminazene aceturate and mebendazole did not significantly improve antibody response in the dogs.A secondary vaccination administered at the 12th week after the primary vaccination significantly increased(P<0.05)the antibody titer with a peak at the 3rd week after the secondary vaccination.Conclusions:It was therefore concluded that A.caninum,T.brucei and T.congolense induced immunosuppression in antirabies vaccination in dogs.