Prevalence and risk analysis of bovine brucellosis in Asella organized dairy farm, Oromia Regional State, South East Ethiopia

A cross-sectional study was conducted on organized Dairy Farm at Asella, Oromia Regional State of Ethiopia to determine seroprevalence and risk analysis of bovine brucellosis in May, 2016. A total of 304 samples were collected; all were tested and confirmed serologically using Rose Bengal plate test (RBPT) and complement fixation test (CFT). Out of 304 samples tested, overall seroprevalence of RBPT and CFT results was12.48% (38) and 9.87% (30) respectively, which was higher in animals above two years age than younger one. History of abortion and retained fetal membrane were found to be significantly (p<0.05) associated with occurrence of bovine brucellosis. A statistically not significant difference (p >0.05) was observed between cross and local dairy cattle. The result showed the high prevalence of bovine brucellosis in the farm. Hence, culling of the positive dairy cattle and practicing good management should result in a control and prevent of the brucellosis.


INTRODUCTION
Brucellosis is a highly contagious, zoonotic, and economically important bacterial disease of animals worldwide (OIE, 2009). It is endemic in many developing countries and caused by Brucella species that affect man, domestic and some wild animals, and marine mammals (Seleem et al., 2010). It is primary reproductive disease clinically characterized by abortion in the last trimester and retained placenta in the female whereas orchitis and epididymitis with frequent and sterility occur in male (Radostits et al., 2007). Sources of infection for isolation of bacterial include aborted fetuses, fetal membranes, vaginal discharges and milk from infected cows. The most common route of transmission in cattle is through direct contact with an aborting cow and the aborted foetus or by indirect contact with contaminated fomites. Ingestion of contaminated pasture, feed, fodder and water may also play a secondary role (Godfroid et al., 2010). Susceptibility of animals to brucellosis depends on their natural resistance, level of immunity and environmental stress (Radostits et al., 2007). Mature animals are much more susceptible to infection, regardless of sex. In female animals, pregnancy has *Corresponding author. E-mail: eyoba20000@gmail.com.
Author(s) agree that this article remain permanently open access under the terms of the Creative Commons Attribution License 4.0 International License positive contribution to the degree of susceptibility than their age. Bulls are relatively resistant than sexually mature heifers and less resistant than sexually immature heifers (Godfroid et al., 2010). A precise diagnosis of Brucella spp. infection is important for the control of the disease in animals and consequently in man. Clinical diagnosis is based usually on the history of reproductive failures in livestock, but it is a presumptive diagnosis that must be confirmed by laboratory methods (Poester et al., 2010). Laboratory methods also help to differentiate from other infectious causes of abortions (Juyal et al., 2011). No single test is appropriate in all epidemiological situations; all have their own limitations. The first serological test for brucellosis was used by Wright and Smith (OIE, 2009). Compliment fixation test is a standard method for the epidemiological surveillance of brucellosis (Köppel et al., 2007). Antibodies anti-Brucella have been demonstrated by the Rose Bengal plate test (RBPT), standard tube agglutination test (STAT), coombs test, complement fixation test (CFT), 2-mercaptoethanol test and enzyme-linked immunosorbentassay (ELISA) (OIE, 2009). In Ethiopia, the prevalence of bovine brucellosis has been intensively investigated in state owned dairy farms (Bekele et al., 2000). In smallholder farms in some parts of the country (Berhe et al., 2007) and in the central highlands of Ethiopia (Kebede et al., 2008). Thus, this study was carried out to determine the seroprevalence of bovine brucellosis and its associated risk factors in Asella organized dairy farm.

MATERIALS AND METHODS
A cross sectional study was conducted in May, 2016 at Asella organized dairy farm managed under intensive system which is located at 175 km South East of Addis Ababa. In this study a bout 5-10 ml of blood was collected from the jugular vein of 304 cattle which are more than six month of age using plain vacationer tube to collect a serum samples. Information on individual animal such as age, sex, breed and history of abortion was recorded in separate sheet. The collected sera samples were screened for the presence of antibody against Brucella using the Rose Bengal Plate Test (RBPT) and Complement Fixation Test (CFT) as a confirmatory test were used in detecting antibody against Brucella antigen. RBPT undertaken at Asella regional veterinary laboratory and CFT was undertaken at the National Animal Health Diagnostic and Investigation Center, serology laboratory, sebata, Ethiopia. The procedure and interpretation of results described by OIE (2008) were followed. Finally, the collected data and the results of laboratory tests were analyzed by statistical package for social science (SPSS), to determine those variable that were significantly associated with seropositivity to Brucella.

RESULTS
In dairy animals investigated during the study were above six month of age and 76(25%) and 228 (75%) were local Borena and cross-breeds of indigenous zebu and Holstein Friesian, respectively. in addition 95 (31.25%) of the animals were lactating cows, 30(9.87%) were pregnant, 42(13.82%) were bulls and the remaining 137(45.10%) were heifers. From the studied animals there was 45(14.8%) history of retained fetal membrane and 28(9.2%) abortion. Generally, the frequency distribution of breed, age group, and sex were summarized in Table 1. Out of 304 serum samples, 38 (12.5%) were positive for brucellosis using RBPT. The present study attempted to look into the existence of any association between seropositivity and breeds, age and sex of the animals. Thus, the prevalence of local Borena, and to cross breed animals was compared in Table 2. The sera prevalence of local Borena, and cross breed cattle was calculated as 1.32 and 8.55% having not a significant variation with P-value of 0.265, the sera prevalence of age for animals 6 month-3 year, 3-6 year and above 6 year which assess in Table 3 was intended as 2.3, 2.63 and 4.93% respectively which have significant variation with p-value 0.011 the prevalence of male and female which assess in Table 4 was intended as 0.99 and 8.88% have not a significant variation with pvalue of 0.523. The association of brucellosis with abortion and retained fetal membrane was tested using Chi-square. It was found that brucellosis was significantly associated with abortion and retained fetal membrane with p-value of 0.000 and 0.002, respectively (Table 5).

DISCUSSION
The present study revealed that the seroprevalence of anti-Brucella antibodies determined with CFT and RBPT was 9.87 and 12.48%, respectively. The overall seroprevalence of bovine brucellosis in the study area was 9.87%. This high seroprevalence is an agreement with previous finding of ( Kebede et al., 2008) with 11% in central highland, (Hunduma and Regassa, 2009) with 11.2% in east show and (Megersa et al., 2012) with 8% in pastoral region.
On the other hand, there were reports with a relatively higher sero-prevalence of bovine brucellosis in other parts of the country, (Sintaru, 1994) with 22% in a dairy farm in northeastern Ethiopia and (Bekele et al., 2000) with 11-15% in dairy farms and ranches in southwestern Ethiopia. Other investigator 0.14% in selected area of north Gondar (Tadese, 2003), 0.77% in selected site of Jima Zone (Tolosa et al., 2008), 0.45% in central highlands of Ethiopia (Lidia, 2008) and 0.05%, in Arsi Zone (Degefa et al., 2011) indicates lower overall prevalence when compared to our present study. The level of brucellosis infection tends to be relatively high in intensive farm than in extensive farm (Matope et al., 2011).
There is still disagreement between different authors among breed susceptibility to brucellosis. In this study breed has supposed one of the risk factors, consequently seroprevalence was found to be higher in cross breed animals (8.55%) than local (1.32%). Nevertheless, this difference was statistically not significant which is similar to reported in GutoGidadistrict (Moti et al., 2012) and in central highland of Ethiopia (Lidia, 2008). On the other hand Minda et al. (2016) and Jergefa et al. (2009) reported significant variation on serological prevalence of brucellosis with higher prevalence in cross-bred than in local ones. Age have association with occurrence of brucella. This could be explained by sexual maturity and pregnancy due to the influence of sex hormones and placenta erythritol on the pathogenesis of brucellosis (Radostits et al., 2007). This result in agreement with report of Lidia (2008) central highland of Ethiopia and Nuraddis et al. (2010) in selected site of Jima Zone. The presences of statistically significant contradict with the previous finding of Minda et al., (2016) and Magona et al. (2009). Even if there is high prevalence in adult animals there was seropositive reactor in less than 3 years of age this is an indication of variations in the management practices (level of intensification and hygienic practices). Even though sex is not significantly associated with Brucella seroposetivity (p> 0.05), high seroprevalence was found among female animals which is 8.88% in female and 0.99% in male animals. This finding was in agreement with the report done by Asfaw et al. (1998) in and around Addis Ababa, Tolosa et al. (2008) in Jima Zone and Desalegn et al. (2011) in Asella dairy farm. The lower prevalence of male reactors in this report could be as a result of smaller number of males tested as compared to female and it was also reported that the serological response of male animal to Brucella infection is limited (Mohammed et al., 2009). Female animals are more susceptible to Brucella organism in gravid uterus of pregnant animals than in testis due to the presence of erythritol in female reproductive tract which stimulates the growth of the organism (Godfroid et al., 2010).
In our study, individual animal sero-prevalence was positively associated with the occurrence of abortion and retained fetal membranes. This indicated that history of abortion or still birth and retained fetal membrane were significantly associated with brucellosis seropositivity. This could be explained by the fact that abortion or still birth and retained fetal membrane are typical outcome of brucellosis (Radostits et al., 2007). This result was in agreement with other investigators Desalegn et al. (2011) in Asella dairy farm and Berhe et al. (2007) in Tigray Region.

Conclusion
The study reflected higher prevalence of bovine brucellosis about 9.87% in the target dairy farm. The current findings indicated that the age, history of abortion and retained fetal membrane were the risk factors statistically significant associated with Brucella seropositivity for this study. Therefore, considering the economic and public health importance of brucellosis, regular screening of brucellosis for newly introduced and the whole farm animals, and culling of those positive one and practicing good farm management were recommended to reduce the risk incidence of bovine brucellosis in dairy farm and surrounding population.