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1 Seroprevelance of Brucellosis in Different Animals Species in Northern State (Sudan) 1 Zein A.M, 2 Adris M.A 1 Department of Microbiology and Parasitology, Faculty of Medicine, University of Dongola, Sudan 2 Department of Biochemistry, Faculty of Medicine and Health Sciences, University of Dongola, (SUDAN) 1 Zain1971@gmail.com ABSTRACT Brucellosis is considered to be one of the most important zoonotic infectious diseases because of its impact on public health and the economy. In Northern State, Sudan, brucellosis is prevalent in Dongola, Daba and Marawe localities and new cases are reported annually (Anon, 2001), but the exact situation of the disease is not known in human and animals in the Northern State. Objectives of the study were to: Investigate the prevalence of the disease in different animal species in the Northern State. Prospective study was conducted in Northern State, Sudan. Blood Samples collected over 6 Months in the period January ( 201 to June 2011) form (7607) domesticated animals (2009 cattle, 2170 Sheep, 2303 goats and 1225 camels) were tested using. RBPT and CELISA. Amongst live stocks 23.8% of cattle, 11.2% of sheep 16.3% of goats and 23.6%. Of camels were seropostive. The overall prevalence of brucellosis in the 7607 domesticated animals was found to be 17.9% in Northern State. In conclusion, there is an urgent need to perform a control strategy at national level on animal brucellosis. Keywords: Brucellosis, domesticated animals 1. INTCTIONRODU Brucellosis is a contagious disease which affects different animal species including cattle, camels, small ruminants, swine, dogs and man (Nicoletti, 1989). The disease is caused by Brucella abortus, Br. melitensis, Br. ovis, Br. suis, Br. canis and Br. neotomae. The disease cause different clinical manifestation, abortion and to less extent orchitis and infertility in males (Solera, 1997). It is essentially a disease of animals caused by bacteria of the genus Brucella, with humans as accidental host (Taylor et al. 1989). The disease is usually transmitted to human by consumption of unpasteurized milk and dairy products or by direct contact with infected animals or both through surveys and countries reports, it was confirmed that animal brucellosis is wide spread in Africa (Pandey, et al. 1999). Several published reports indicate that, animal brucellosis is a common disease in Sudan. Control measures were adopted, but still the disease is widely spread and new cases were reported annually (Anon 2001). This Study was carried out to investigate the prevalence of the disease in the Northern State, Sudan, in different animal species. study area are delivered through district health net-work, consisting of a local hospital and a few health centers in the cities. 2.2 Localities of the Northern State The state was divided into seven localities for the purpose of the study, and the localities were divided into three geographical areas: a. Northern area: Consisted of Wadi Halfa and Dangola localities which were divided into five units (Wadi Halfa, Abre, Dalgo, Berka, and Farage). b. Middle area: Consisted of Alburgage, Dongola, and Goled Localities which were divided into seven units (Hafeer, Karma, Argo, Dongola city, Goled, Kaddar and shark Elneel). c. Southern area: Consisted of Dabba and Marawe localities which were divided into seven units (Gaba, Dabba City, Tadamon, Karema City, Marawe city, Gorear and Alshohada) 2. MATERIALS AND METHODS 2.1 Area of the study The study was carried in Northern State which is one of the biggest States in the Sudan. It is about 348,796 Km 2. It is bordered by Egypt in the North and Libya and North Darfur in the West, Nahr Elneel in the East, Khartoum and Northern Kordofan State in the South. The population of the Northern State is about 699,065while the population of animals is about 3,150,000 distributed in seven localities, Wadi Halfa, Dalgo, Al burgage, Dongla, Goled, Dabba, and Marawe. The Northern State is mainly an agricultural State (dates, wheat and beans). The veterinary services facilities depend on veterinary hospitals in all parts of the State as well as dispensaries and primary veterinary care units. Health services in the 210

2 2.3 Mapif the Northern State and localitcs N Egypt Northern Darfur State Naher Alneel State Libya Khartoum State 211

3 2.4 Population of the study and sampling The study was focused on: adult sexually mature non vaccinated cattle, sheep, goats, and camels in the study area. 2.5 Rose Bengal Plate Test A standardized RBPT was provided by the Central Veterinary Research laboratory, Soba, Sudan, was used to screen the samples as by Alton et al. (1975): a. The serum sample and antigen were brought to room temperature (22± C). b. A 30µl of each serum sample was placed on a white enamel or plastic plate. c. After shaking gently the antigen bottle, an equal volume of the antigen was placed near each serum spot. d. Immediately after the last drop of antigen has been added to the plate, the serum and antigen were mixed thoroughly, using a clean glass or plastic rod for each test, to produce a circular or oval zone approximately 2 cm in diameter. e. The mixture was agitated gently for 4 minutes at ambient temperature on a rocker. f. The result of agglutination was read at the end of the 4 minutes period and reported as positive or negative according to the OIE (2008). 2.6 Competitive Enzyme-linked Immuno-Sorbent Assay (c-elisa) The RBPT positive serum samples were confirmed by the c-elisa (OIE, 2008). The test was performed using commercial kits (Sabnova- Sweden). Material and reagents were provided by the University of Dongola, Faculty of Medicine. The test was performed according to the method described by the producing company as follows: 2.7 Test Procedure a. The PBS-Tween Solution 20x concentrate 1/20 was diluted in distilled water. 500 ml were prepared by adding 25ml PBS-Tween Solution to 475 ml distilled water and mixed thoroughly. b. The freeze dried mab was reconstituted immediately before use with 6ml sample dilution buffer by adding the buffer carefully into the bottle. c. All reagents were equilibrated to the room temperature before use. d. The samples and controls were diluted by adding them to the wells of the plates prefilled with buffer. e. A50µl of each serum controls (positive, weak positive and negative) were added into each of the appropriate wells, respectively. Each control was run in duplicate. f. A5µl of each sample dilution buffer were added into two appropriate wells (designated as Conjugate control, Cc). g. A5µl of each test sample was added to each of appropriate well. For confirmation each samples was run in duplicates. h. A50µl of mab-solution were added to all wells of the samples and controls used for controls within 10 minutes. i. The test plate was sealed and the reagents were mixed thoroughly by tipping the sides of the plate. j. The plate was incubated at room temperature for 30 minutes. k. The plate was rinsed 4 times with PBS-Tween using Sanofi Pasteur washer. l. A100 µl substrate solutions were added to each well and incubated for 10 minutes at room temperature timing began after the first well was filled. m. The reaction was stopped by adding 50µl of stop solution in the same order as the substrate solution was added to each well and mixed thoroughly. n. The optical densities (OD) of the controls and samples were measured using a micro plate photometer at 450 nm after 15 minutes from the addition of the stop solution. 2.8 Calculations a. The mean OD-values for each of the controls and samples were calculated. b. The percentage inhibition (PI) values for the controls and samples were calculated using the following formula: PI = 100 (mean OD samples / control 100) Mean OD conjugate control Cc 2.9 Test Validity To ensure validity of the test, the values of the controls must fall within the following limits: OD Cc = PI Positive Control = PI Weak Positive Control =35 65 PI Negative Control = (-10) Interpretation of the test The status of the test was determined as follows: PI Status < 30% Negative 30% Positive 3. RESULTS AND DISCUSSION 3.1 Field Observations A surveillance was carried out during (2011) in Northern State in Northern Sudan. Indigenous domesticated animals includes sheep, goats, cattle, camels and equine and foreign breeds were kept in the area of the study to supply milk and meat for private consumption and also to support the farmer s income. The animals were kept in backyards of the houses and villagers raised 211

4 small flocks ranging from 2-12 animals which are usually managed by women or children. No housing was available except night shelters (small holdings). Nomadic and semi-nomadic systems of breeding were adopted in southern area of the State. Local green fodders were given in the morning and evening to the animals kept in small holdings and concentrates were not fed except to animals in governmental farms and those kept in confined system. Sources of water for the livestock are wells and the River Nile. Natural breeding is used in the area of the study and the farmers exchange breeding bulls. Animal diseases were reported in the area included mainly pneumonia, external and internal parasites and nutritional deficiency diseases. There are several small abattoirs, which were poorly equipped or even lacked facilities. Sometimes they are next to meat shops in the vicinity of houses. Home slaughter and consumption of sheep, goats, cattle and camel meat was wide spread and their offal s are usually given to dogs. Dead animals are often abandoned and or are usually thrown into the River Nile. Fresh milk and dairy products, meat and other animal products are often consumed raw or insufficiently cooked and constituted a special risk. In some areas, dried animal feces are kept near human dwellings and used for cooking.there are houses which lack running water and toilets. Small markets are scattered all over the State and large- permanent markets were only in big towns. 3.2 Prevalence of brucellosis in different animal s species The (7607) animals examined, 1368 (17.9%) were positive for brucellosis (Table 1). Accordingly, sero-prevalence of brucellosis in sheep, goats, cattle and camels in the study area was found to be 11.2%, 16.3, 23.8% and 23.6%, respectively. Table 1: The prevalence of brucellosis in the different animal according to species Animal species Total of samples examined Number positive (%) positive 1-Sheep % 2-Goat % 3-Cattle % 4-Camel % Total % 3.3 Prevalence of brucellosis in different areas The serological survey showed that 13.6% of the animals examined in the Northern area were positive for brucellosis. The prevalence rate of Brucella antibodies in Wadi Halfa locality was 13.2% while in Dongola locality was 14%. The overall prevalence of brucellosis in 386 sheep, 360 goats, 311 cattle and 158 camels in the Northern area was found to be 9%, 9.7%, 13.8% and 34.1, respectively. The serological survey showed that 13.3% of the animals examined in the middle area were positive for brucellosis. The prevalence rate of Brucella antibodies in Dongola locality was 10.6%, Alburgage locality was 11.7% and Al Goled locality was 17.1%.The overall prevalence of brucellosis in the 954 sheep, 949 goats, 995 and 573 camels examined was found to be 7.7%, 17.4%, 17.1%, 16.7 respectively. The serological survey showed that 15% of the animals examined in the southern area were positive for brucellosis. The prevalence rate of brucella antibodies in Aldaba locality was 10.4% while in Marawe locality was 19.9%. The overall prevalence of brucellosis in the 830 sheep, 894 goats, 703 cattle and 494 camels equine examined in this area was found to be 14.3%, 14.9%, 23.4% and 22% respectively. In the present study the overall prevalence of brucellosis in the7607 domesticated animals was found to be 17.9%. This was higher than those reported by Suliman (1987) in El Gazera and Khartoum States 11.6% and 15.8%, respectively and lower, than that reported by Musa (1995) in Darfur States 20%. The results indicated that, there was a marked increase in brucellosis in animals in the Northern State due to the absence of brucellosis control programmer. The prevalence reported here was also higher than those reported in some Arabian and African countries; 12% in Moroco, 4.8% in Egypt, 5.2% in Kuwait, 8% in Jordan and 3.8% in Libya (Refai, 2002) and 3.8% in Chad, 6% in Ivory Coast, 7.5% in Cameron, 8% in Upper Volta and 10% in South Africa (Kubuafor et al. 2000) Several reports indicated that, brucellosis in animals is common in the Sudan. El Nasri (1960) reported 14% to 18% sero-prevalence of brucellosis among cattle, 6.6% in goats and 35% in sheep in Southern Sudan. Abdalla (1964) reported prevalence of 3% in cattle and 1.7% and 1.5% in sheep and goats in Northern Sudan, respectively. In Darfur States Western Sudan, Musa (1995) found 13.9%, 3.5%, 5.9% and 7.7% prevalence in cattle, sheep, goats and camels, respectively. In Gezera State Central Sudan, Dafalla (1962) reported 8.7% to 10.7% sero prevalence of the disease in cattle, 4.2% to 50% in sheep and 2.5% to 30% in goats. In Khartoum State, Suliman (1987) found a prevalence of 15.8% among cattle and more recently Ali (2007) reported a prevalence of 27.4% in cattle. The results confirm the finding of Omer, et al. (2007) who reported that the prevalence of brucellosis increased during the last years among sheep, goats, cattle and camels in Kassala area, Eastern Sudan. Therefore, an urgent organized program for monitoring and control of animal brucellosis should be adopted current study revealed a prevalence of 23% in cattle, which is high, according to the criteria of Thimm and Wundit (1976) who considered that a percent range between16-25 was high. The prevalence was higher in cattle compared with that in other animals in the study area 11.2% in sheep, 15.6% in goats. Only camels showed a similar prevalence rate of 23.6%. This might be due to the system of breeding adopted in the study area that large ruminants were kept in small holdings. The results are in agreement with these, obtained in Togo (22.5%) (Akakpo et al. 1981), Rwanda (25.7%) (Kabagmbe et al. 1988), India (Punjab) (22.6%) (Aulakh et al., 2008) and Iran (22.8%) (Moshelani et al. 2011). However, were lower than, those found in Kenya (77.5%) ( waghela, et al. 1978), Cameroon (40%) (Kubuafor et al. 2000) and Turkey 212

5 (68.1%) (Gen et al. 2005). The prevalence of bovine brucellosis is increasing in the Northern State when it is compared with the previous study by Abdalla (1966) who found 3% prevalence in cattle. The overall prevalence of brucellosis in camels, in Northern, middle and southern areas of the Northern State was found to be 34.1%, 16.7% and 22%, respectively. The prevalence was higher in the Northern area which might be due to the expansion of large-scale trading of camels transported from Western Sudan through the area to Egypt without proper attention being payed to the possibility of disease transmission. Some of these camels are kept in feed-lots before being exported across the Nubian lake to Egypt. The 23.6% prevalence of the disease in camels in the area is in agreement with that reported by Musa (1995) who found 23.8% prevalence in Darfur, Western Sudan, but is considerably higher than those recorded by several investigators from different parts of the country. Yagoub et al and Bitter, 1986) and lower than those found by Majid et al., 1999; Fayza et al and Omer, The results were also higher than Kenya (8%) (Paling et al. 1998), than that from Iran (10.5%) (Ahmed et al, 1995), from Libya (4.1%) (Gameel et al. 1993), from Nigeria (11.42%) (Junaidu et al., 2006) and from Somalia (3.9%) (Gen, et al. 2005). In countries where Br. melitensis is prevalent in sheep and goats, other farm animals such as pigs, cattle and camels, may become infected from them (Altons, 1985). In the present study the prevalence of brucellosis in sheep and goats in Northern, middle and southern areas of the Northern State was found to be 9%, 7.7%, and 14.3% in sheep and 9.7%, 17.4% and 15% in goats, respectively. The prevalence of the disease was higher in the middle area as large numbers of small ruminants were distributed in the farmers in this area in the years 2007 and 2008, without testing for brucellosis. Furthermore, some of these animals when showed signs of the disease were not excluded and attempts were made to treat them with a local therapeutic (Om-regala). Treated animals might have constituted foci of infection to the herds in the area. Results showed that the prevalence of ovi-caprine brucellosis was increasing when compared with the 1.7% in sheep and 1.5% in goats which reported by Abdalla (1966). This increase might be due to the spread of the disease due to absence of control measures and to improvement of monitoring and surveillance procedures applied. Although the prevalence recorded in this study was 15.6%, it is lower than those reported in Turkey (31%) (Gen et al. 2005), in Iran (24.6%) (Yahya et al. 2009) in Jordan (27%) (Al-majali, 2004) and in Egypt (31%) (Mayada et al.2010). From the results it appears that brucellosis is more prevalent in cattle and camels than in sheep and goats and this is in agreement with the finding of Omer et al. (2007). Sheep and goat flocks in the study area were mobile. Free movement of infected sheep and goats and mixing them with other animals can spread brucellosis. Control of movement of sheep and goats and control or elimination of infection in them can reduce spread of the disease to other animals. ACKNOWLEDGEMENTS I would like to thank to the staff of Veterinary Research Institute, Dept. of Brucella, Dr. Enaam Elsanosi and Mr.Salsh who provided the Rose Bengul antigen and assets of the Department on hand to accomplish this work. My thanks extend to Ministry of Health, Northern State, Sudan for supporting this work. REFERENCES [1] Abdalla,A.(1966).Incidence of brucellosis in Wadi half a District.Sudan.J.Vet.Sci.Anim.Husb.,7:28-31 [2] Ahmed, R, M.A, Munir (1995) Epidemiological investigations of brucellosis in Pakistan. Pakistan.Vet.J.,15: [3] Akakapa, A.J,Bornarel, P, Sarradin, P. (1981). Bovine brucellosis in Togo. Bull. Anim. Hlth.Prod.,132: [5] Ali, A,A,I. (2007).Prevalence of brucellosis in Kuku dairy, Khartoum State and the susceptibility of the isolates to some chemotherapeutic agents. MSc. Thesis, Faculty of Pharmacy, University of Khartoum, Sudan. [6] Almajali, A.M,Talafha, A.Q, Ababenh, M.M. ( 2004). Sero-prevalence and risk factors for bovine brucellosis in Jordan.J. Vet. Sci.,10 : 6-15 [7] Alton GG, Jones LM, Pietz DE. (1975). laboratory techniques in brucellosis, World Health Organization, Geneva [8] Alton GG, Jones LM, Pietz DE. (1975). laboratory techniques in brucellosis, World Health Organization, Geneva [9] Anczkowski F (1972). Further Studies on bovine brucellosis.vet. Bull., 42: [10] Anon (2001) Annual Report of Sudan Veterinary Surices. [11] Aulakh, H.K,Patil, P.K, Sharma, S. ( 2008).Study on the epidemiology of bovine brucellosis in Punjab(India) using milk ELISA. Acta. Vet. Brono., 77: [13] Dafalla,E. N and Khan, A.(1962) The occurrence, epidemiology and control of animal Brucellosis in Sudan. 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6 [16] Gen, O,Otlu, S, Pahun, J.P. (2005). Seroprevalence of brucellosis and Leptospirosis in aborted dairy cows. Turk. J. Vet. Anim. Sci.,29: [17] Junaidu,A.U, Oboegbulem, S.I. Salihu, M.D (2006) Serological survey of Brucella antibodies in breeding herds. J. Microbiol. Biotech. Res., 1: [18] Kabagmbe, J.P,Elzer, P.H, Opuda, J, Smith, S.H. (1988). Risk factors for Brucella seropositivity in goats herds in eastern and western Uganda.Prev.Vet.Med.,52: [23] Omer, M.M, Abdelaziz, A.A, Abusalab, S.M.A and Ahmed, A.M (2007). Survey of brucellosis among sheep, goats, Camels and Caltle in Kassala Estern Sudan. J. Anim. Vet. Ad., 6(3): [24] Pandey, G.S, Kobayashi, K, Nomura, Y, Namota, A, Mwima, H.K, Suzuki, A.K. (1999). Studies on sero- prevalence of brucellosis Kafue lechwe in Zambia. Ind. Vet. J., 76: [25] Refai, M. (2002). Incidence and control of brucellosis in Near East region.vet.microbiol, 90: [19] Kubuafor, D.K, Awumbia, B, Akanmoi, B.D. (2000). Seroprevelance of brucellosis in cattle and human in the Akawapin-South district of Ghana. Acta.Trop.,76:45-47 [20] Moshelani, S, Jaraheri-Koupae, M, Rabiee, S. (2011). Detection of Brucella spp.and Liptospiras spp. By multiplex polymerase chain reaction from aborted bovine,ovine and caprine fetuses in Iran. Afr. J. Microbiol. Res.,5: [21] Musa, M.T. (1995). The magnitude and the problem of brucellosis in Darfur States.Ph.D.Thesis,University of Khartoum [22] Nicoletti, P. (1986). Diagnosis and Control of Brucellosis in the Near East. FAO, Rome. [26] Solera, J, Martinez- Alfaro, E, Espinosa, A. (1997). Recognition and optimum treatment of brucellosis. Drugs, 53: [27] Suliman, M.A. (1987). The prevalence of bovine brucellosis in Khartoum and Gezira regions.msc. Thesis, Fculty of Veterinary Science,University of Khartoum. [28] Waghela, S,Fazil, M.A, Gathuma, J.M. (1978). A serological survey of brucellosis in camels in northeastern province in Kenya.Trop.Anim.Hlth.Prod., 10: [29] Taylor JP, Perdue JN (1989). The changing epidemiology of human brucellosis in Texas, Am. J. Epidemiol., 130:

Detection of Brucellosis in sheep intended for export and local slaughter in Khartoum State, Sudan

Detection of Brucellosis in sheep intended for export and local slaughter in Khartoum State, Sudan African Journal of Microbiology Research Vol. 6(39), pp. 6805-6810, 11 October, 2012 Available online at http://www.academicjournals.org/ajmr DOI: 10.5897/AJMR12.1423 ISSN 1996-0808 2012 Academic Journals