Prevalence of small ruminant trypanosomosis in Assosa and Homosha districts , Benishangul Gumuz Regional State , North West of Ethiopia

A cross sectional study was conducted at Assosa zone of Benishangul Gumuz, North West Ethiopia, to determine the prevalence of trypanosomosis in local breeds of sheep and goats. Blood sample from 384 randomly selected sheep and goats (177 from Assosa and 207 from Homosha districts) of different species, sex, age groups were examined by dark phase contrast buffy coat and thin smear examination for species identification of trypanosome. Among the animals examined during the study period, 10 (2.6%) were infected with trypanosomes. From this survey, Trypanosoma vivax was found to be the major cause of trypanosomosis (1.82%), followed by Trypanosoma congolense (0.52%) and Trypanosoma brucei (0.26%). There was no statical difference (p > 0.05) in infection between sex, species, and among age groups. Mean packed cell volume (PCV) of the parastemic animals was significantly lower than (P < 0.05) that of aparastemic animals. In attempt to identify the vector involved in transmition, tsetse flies group (Glossina morsitans submorsitans) and mechanical vectors of trypanosomosis that belonging to Tabanidae (Tabanus, Stomoxys and Haematopota) were captured in both districts at an altitude range of 1270 to 1507 m above sea level. The results of the prevalence of the disease in small ruminants and its vectors indicate that an effective management and control measures for the disease and transmitting vectors should be designed and implemented.


INTRODUCTION
Trypanosomosis is a major constraint on ruminant livestock production in many areas of Africa including Ethiopia.Many animal species can be affected by the different trypanosomes, thus severely impairing the economic efficiency in endemic areas.From an economic point of view, the disease is particularly important in cattle, although other mammals can also be affected (Gupta et al., 2003;Singla et al., 2009;Sharma et al., 2012;Kumar et al., 2012).Although geographical differences have been observed, but the most affected animal species are buffalo, horses, camels, cattle and dog (Singh et al., 2003;Singla et al., 2012;Sharma et al., 2013;Sumbria et al., 2014).Other hosts, including small ruminants, can be affected (Corbera et al., 2006).
Trypanosomosis is a parasite disease caused by species of flagellated protozoa belonging to the genus trypanosome which inhibit the blood plasma, various blood tissues and liquid of vertebrate host (Rodostitis et al., 2000;Juyal et al., 2005).The infection caused by different species of trypanosomes is mainly transmitted cyclically by tsetse flies and mechanically by biting flies belonging to families Tabanidae and Hippoboscidae if they feed on more than one host within a short interval (Radostits et al., 2000;OIE, 2008).It is a serious constraint to agricultural production in extensive areas of the tsetse-infested Ethiopian lowlands (Slingenbergh, 1992).Out of the nine regions of Ethiopia, five (Amhara, beneshangul-Gumus, Gambella, Oromiya and SNNPR) are infested with more than one species of tsetse flies (NTTICC, 2004).
The epidemiology of typanosomosis depends on factors such as distribution of vectors, virulence of parasite and response of host.The disease follows the distribution and intensity of various species of tsetse fly (Bursell, 1960).The most important tsetse flies species that are distributed and infested parts of Ethiopia are Glossina morsitans submorsitans, Glossina pallidipes, Glossina fuscipes fuscipes, Glossina tachinoides and Glossina longipennis (Langridge, 1976;Getachew and Hunduma, 2005).Sheep and goats are naturally infected with Trypanosoma congolense, Trypanosoma vivax, Trypanosoma brucei and Trypanosoma evansi which produce acute, subacute, chronic, or subclinical forms of disease in these animals (Getachew, 2005;Corbera et al., 2006).Even though small ruminants are naturally infected and presenting clinical disease, it is commonly believed that they are highly resistant to infection, but it is only sporadic, and that the disease in these animals is of little economic consequence (Corbera et al., 2006).The role of small ruminant in epidemiology of the disease in nature is still not well known.But the current epidemiological information indicates that the sheep and goats could act as reservoirs for the spread to animal and human trypanosomsis (Dede et al., 2005).
In view of this, the present study was designed to estimate the prevalence rate of typanosomosis in small ruminants, identity species of trypanosomes, determine the packed cell volume (PCV) values on infected and non infected animals and the vector density tsetse and other biting flies of the study area.

Study area
This study was conducted in two districts (Assosa and Homosha) of Assosa zone of Benishangul Gumuz Regional State.The study was limited to only two peasant associations from each district due to their scatteredness and limited resources.These are Ashura and Bamadon from Homosha district; Ura and Tsetse Adurnon from Assosa district.Assosa is found at 675 km far from capital city, of Addis Ababa, North Western part of Ethiopia at 34° 02' 20E to 36°30'E and 9°30'N to 11°39'N.The maximum and minimum temperature of the area ranges from 35 to 25°C, respectively with the altitude ranging from 600 to 2731 m above sea level and the mean annual rain fall of 1000 mm.The area was gaining the rain for six month duration which starts on April and ends on October.The main occupation of the population is mixed farming practice of crop and livestock.The major livestock reared in the area are bovine, sheep, goats, donkeys and poultry (RSA, 2007).According to the publication bureau of Assosa Zone Agricultural office, the number of animal population in the area were 39133 Cattle, 6977 Sheep, 17675 Goat and Equines 2558 in Assosa district and 3665 cattle, 1350 Sheep, 3752 goat and 317 Equines in Homosha district.
The vegetations type of the area is savanna grass with scattered trees of different species dominated by bamboo trees that covers large area of the region with fat low land.Among the wild game few of them are antelope, bush pig, baboons and warthog (NTTICC, 2004).

Sample size
The sample size was determined using Thrusfield (2005) formula.Totally, 384 samples were collected from four sites of the two districts, namely, Ashura, Bamadon, Ura and Tsetse Adurnon by simple random sampling method.

Study animals
The study was conducted on 384 local breeds of the sheep and goats.From each selected peasant association, a proportional number to the total population of the study animals were sampled.The sample size were 86, 121, 58 and 119 in Ashura, Bamadon, Ura and Tsetse Adurnon, respectively.Examination and evaluation of body condition were accomplished during sample collection.They were classified as very thin (1), thin (2) moderate(3), good(4) and very good (5) by observing the body condition of animals in the field (Cooper and Thomas, 1985).The age of animal were also estimated by examining dentations (Kripali et al., 2010) and information obtained from owner.

Study design
The study was based on entomological and parasitological survey.This cross-sectional study were conducted in two districts.

Parasitological and hematological examination
Blood sample were collected randomly from small ruminants of four sites during the study period.Blood was collected from ear vein using sterile blood lancet and capillary tube.Pair of heparinazed capillary tube were filled with blood from small ruminants to 3/4 of its height and sealed at one end with crystal seal.The capillary tube were loaded on the microhaematocrit centrifuge symmetrically and centrifuged at 12000 rpm for 5 min (Murray et al., 1997).Packed cell volume (PCV) was determined using Haematocrit reader (Woo, 1970).After the PCV has been read, capillary tubes were broken in 1 mm below the Buffy coat which includes red blood cell layer and the content were expressed on microscopic slide and covered with a 22 × 22 mm cover slip.The content was examined under ×40 objective using dark ground Buffy coat technique (Murray et al., 1997).From positive sample, thin smear were made, fixed with methanol for 5 min and stain with geimsa solution for 30 min and examined using oil immersion under × 100 objective to detect the species of trypanosome.

Fly survey
The survey was under taken in two districts of Assosa zone in four peasant associations.A mono-pyramidal trap was used, to trap different flies, total of 48 h were used and 59 traps were deployed.The collected flies' were counted and identified using hand lenses.
Mono-pyramidal traps baited with acetone, octenol, and cow urine (Brightwell et al., 1987) were used for assessing the fly density.The site selection was to include suitable tsetse habitats like savanna area, river valleys, livestock grazing area and watering points and vicinity to assume wild game reserve area.In all the study sites, a total of 59 mono-pyramidal traps were deployed early in the morning and maintained in position for 48 h.Savanna tsetse (Glossina morsitans sub morsitans) can detect odour from about 200 m.So, the traps were spaced at about 200 m interval.
During trapping, acetone and octenol were dispensed from open vials through an approximately O-size hole.While cow urine from open bottle on a piece of tissue paper was included to facilitate odor diffusion.All odors were placed on the ground, above 20 cm up the wind of the trap.The traps poles were greased to exclude insect predators like ants.The different catches in the traps were counted, identified (Langride et al., 1976) and analyzed.The species of tsetse fly was identified based on characteristic morphology (Leak et al., 1993).Other biting flies are separated according to their morphological characteristic, such as size, color, wing venation structure and proboscis at genus level (Wall and Shearer, 1997).
Sexing was done just by observing the posterior end of the ventral aspect of the abdomen by hand lenses.As a result, male fly is easily identified by enlarged hypopygium in posterior ventral part of the abdomen (Challier, 1965).The fly apparent density is the mean catched in traps deployed, expressed as the number of fly catched per trap per day (Leak, 1999).

Data analysis
Statical analysis was employed by Chi-square ( 2 ) for data management and analysis using Stata Version 7.0 (2000).The tested hypothesis were prevalence of trypanosomes, PCV value, the relation between PCV value and prevalence of trypanosomes, the relation between age value and prevalence of trypanosome.

RESULTS
During this study, a total of 384 small ruminants of local breeds were examined in both study areas, out of these 207 were from Homosha and 117 from Assosa district.Generally, the overall prevalence rate of the two districts was 2.6% (Table 2.) The trypapanosome species encountered are T. vivax,T.congolense and T. brucei.T. vivax and T. congolense were found in two districts, but T. brucei was in Homosha only.The relative proportion of trypanosome species were 1.82, 0.52 and 0.26% for T. vivax, T. congolense and T. brucei, respectively (Tables 1 and 3).
There was no statistically significant difference (P>0.05) between trypanosome infection rates in the village of two Woredas; with prevalence rates of 3.83, 1.64, 3.45 and 2.52% in Ashura, Bamadon, Ura and Tsetse Adurnon, respectively (Table 4).
Rate of infection was 3.82 and 1.78% in ovine and caprine, respectively with an overall infection rate of 2.60%.There is no significant difference (P>0.05)above different species of small ruminant (Table 5).
The body condition of all sampled small ruminant was also evaluated by scoring method indicated by Cooper and Thomas (1985).Out of the infected small ruminant, 3.85% were with thin body condition, 2.68% with moderate body condition, and 2.06% with good body condition (Table 6).
Infection rate was 0 and 2.98% in less than two years and greater than two and/or equal to two, respectively (Table 7).There is no statistically significant difference (P>0.05) in different age groups.
Infection rate between different sexes were 2.25 and 2.9% for male and female, respectively (Table 8).There is no statistically significant difference (p>0.05) in different sex groups.
There was statistically significant (P<0.05)difference in means PCV of parastemic and aparastemic small ruminants.The mean PCV value of parastemic and aparastemic were 20.9 and 27.95%, respectively (Table 9).

DISCUSSION
The results indicated that trypanosomosis to be the important livestock disease in Assosa zone of North West Ethiopia as also reported by Tewolde et al. (2001) in Metekel district, North West of Ethiopia.Even though various conventional diseases induce livestock mortality and results in economic losses in Ethiopia, tsetse transmitted trypanosomosis has a crucial effect which is becoming unchallengable to treat.Vector control action as a strategy option is not widely implemented in Ethiopia (NTTICC, 2004).
The finding of tsetse survey revealed one type of tsetse species at Homosha and Assosa districts Woreda.The main vector were G. m. submorsitans which was similar with the previous result in Metekel district (Getachew and Hunduma, 2005).Typical habitat was found in the study area for savanna species, that is, G. m. submorsitans prefer savanna grass, riverine and forest ecology.G. m. submorsitans was found to be concentrated in low land areas as climatic condition were more favourable.Some flies however, were found as high as 170 m.Earlier works had established the tsetse geographical limit at 1600 m.Later, Slingenbergh (1992) found the inceased limit up to 2000 m.
During the tsetse survey in two Woredas of the Assosa zone 0.0086 and 0.018 flies/trap/day were captured in Homosha and Assosa districts, respectively.Small fly density was obtained because of dry season.There were uncontrolled bush fires few weeks prior to the survey period.Such circumstance might have suppressed the fly density and forced the fly to move the moisture area and river banks of extreme low altitude.The fly density was found to be relatively increased in late rainy season than dry season.This fact is in agreement with result of Leak et al. (1993).According to Leak et al. (1999),  There was no significant difference (p > 0.05) in infection rate among thin, medium and good body condition.This result is similar to the work of Goossens et al. (1998) and Snow et al. (1996).These researchers have indicated that small ruminant are not often selected by tsetse flies.Though these animals do not show signs of trypanosomosis.The same result is revealed by Getachew and Hunduma (2005) at South West of Ethiopia and report that trypanosomosis in sheep and goats is an important disease and small ruminant serve as potential reservoir of infection for other animals and do not show clinical sign and were in good body condition.
The present study revealed that when the age increased, the prevalence rate also increased.This may be due to the immune compromization in very old individuals.The findings from Muturi (1999) and Terzu (2004) support the present finding.Out of the three detected trypanosomes, T. vivax stands first and this is due to the fact that its ability is being transmitted by mechanically as well as cyclically and there is also little number of Glossina species when compared with other biting flies.This result is similar to the work of Kalu and Uzoigwe (1996), who reported that the area encountered low density of tsetse flies and T. vivax was a predominant species.This suggests that the biting flies would mediate T. vivax infection when tsetse fly density is low or absent.
There was no significant difference (p>0.05) in the infection rate among species of small ruminant, because in this study, area management, grazing area and nutritional status of the two species were the same.The same result is revealed by Coulibaly et al. (1995) and Defly et al. (1988) indicates that livestock species had a major effect on trypanosome prevalence.
It is known that the development of anemia is the most reliable indicator of the progress of trypanosome infection (ILRAD, 1998).But it can also be assumed that numerous concurrent diseases and nutritional factors interferes with anemic development (O.A.U/S.T.R.C., 1979) and PCV value are reliable indicator of anemia.During PCV determination, a value of 24 to 46 (Radostitis et al., 2000) was considered to be normal.
The mean PCV values of parastemic (20.9%) was found to be statically lower than (p>0.05) that of aparastemic (27.95%) small ruminant.Similar result were obtained at South West Ethiopia by (Getachew and Hunduma, 2005).Taylor (1998) indicated that the anemia persists during chronic infection of when parastemia is quite low, probably because of different mechanisms that are involved in its genesis during acute and chronic stage of infection (Singla et al., 1997).Thus suggest that control of parastemia is unrelated in chronic phase when immune system is depressed and anemia is sustained through dyserthropoisis.

Conclusion
Trypanosomosis was prevalent in small ruminant in the study area at low rate of 2.6% with different trypanosome species.Prevalence of mechanical transmitters or biting flies such as Stomoxys, Tabanus and Haematopota along with cyclically tsetse G. m. submorsitans species indicated the vectors for trypanosomes in the study area.Regarding the trypanosomosis in small ruminant, a severe clinical signs does not overt by infection, but they can be considered as an important reservoir for the majority of trypanosomes for other animals and humans.Therefore, in areas where the presence of small ruminants are important and trypanosomosis is prevalent, these animals should be taken into consideration in all programs to control the disease.

Table 1 .
Species and number of vectors identified at two districts of Assosa zone.

Table 2 .
Prevalence small ruminant trypanosomosis on district basis.

Table 3 .
Relative proportion of different trypanosome species on basis of district.

Table 4 .
Prevalence of small ruminant trypanosomosis on the basis of the study peasant associations.

Table 5 .
Prevalence of small ruminant trypanosomosis based on species.

Table 6 .
Prevalence of small ruminant trypanosomosis on the basis of body condition

Table 7 .
Prevalence of small ruminant trypanosomosis on the basis of age.

Table 8 .
Prevalence of small ruminant trypanosomosis on the basis of sex.

Table 9 .
Mean PCV value of parastemic and aparastemic small ruminant.