Seroprevalence of small ruminant brucellosis in selected districts of Borana zone, southern Ethiopia

Godana Haro; Desie Sheferaw

doi:10.14405/kjvr.20240045

Korean J Vet Res > Volume 65(2); 2025 > Article

Haro and Sheferaw: Seroprevalence of small ruminant brucellosis in selected districts of Borana zone, southern Ethiopia

Review Article

Epidemiology

Korean Journal of Veterinary Research 2025;65(2):e15.

Published online: June 30, 2025

DOI: https://doi.org/10.14405/kjvr.20240045

Seroprevalence of small ruminant brucellosis in selected districts of Borana zone, southern Ethiopia

Godana Haro¹, Desie Sheferaw^2,^*

¹Dida Tuyura Borana Cattle Breeding and Improvement Center, Yabello 85, Ethiopia

²Faculty of Veterinary Medicine, Hawassa University, Hawassa 05, Ethiopia

^*Corresponding author: Desie Sheferaw Faculty of Veterinary Medicine, Hawassa University, Hawassa 05, Ethiopia
Tel: +251-916832419 E-mail: mereba480@gmail.com

Received July 16, 2024 Revised October 4, 2024 Accepted October 16, 2024

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Brucellosis is a zoonotic disease of considerable economic and public health significance worldwide. No information available about brucellosis status in the Gomole and Elewaye districts, Borana pastoral area, southern Ethiopia. Hence, aim this study was to estimate the seroprevalence of brucellosis in small ruminants and the associated risk factors in the study districts. A cross-sectional study design was conducted to estimate the seroprevalence of brucellosis in small ruminants and to assess the associated risk factors. The study animals were managed under extensive pastoral production system. Small ruminants above 6 months of age were included in the study. Approximately 10 mL of aseptic blood samples were collected from the jugular vein, and sera were extracted and stored at −20°C until the laboratory tests were conducted. All sera samples were screened using the rose Bengal plate test (RBPT). Those sera positive in RBPT were subjected to indirect enzyme-linked immunosorbent assay (iELISA) for confirmation. From 648 sera samples (i.e., 264 sheep and 384 goats), 40 (6.2%) were seropositive in iELISA, and the seroprevalence of brucellosis in sheep and goats was 3.4% and 8.1%, respectively. The seroprevalence of brucellosis was significantly higher in goats (odds ratio [OR], 2.15; p < 0.05) than in sheep, animals aged ≥ 5 years (OR, 11.23; p < 0.05), in good body condition (OR, 2.92; p < 0.05) and in larger flock sizes, ≥40 animals, (OR, 9.52; p < 0.05). The recorded seroprevalence was higher, and control measures, such as testing and culling of seropositive animals, could reduce the problem.

Keywords: Brucellosis, seroprevalence, iELISA, goats, sheep

Introduction

Sheep and goats play a crucial role in the livelihoods and the economies of developing countries. These animals are a means of income for producers and employment. Small ruminants are a crucial asset and safety net for people with low incomes, especially for women and pastoralist groups, and they provide an important source of nourishment for many rural and urban households [1,2].

Sheep and goats account for approximately 64.1% of domestic ruminants in Borana zone [3]. The recurring droughts in the area mean that most Borana pastoralists preferred to keep diversified and more drought-tolerant domestic animals as a coping strategy. First, they preferred to keep camels, followed by small ruminants [4,5].

Sheep and goats provide meat, milk, skin, and manure [6] and are the primary source of cash income [1,7]. Despite the enormous potential of small ruminants in the Borana (i.e., pastoral) area, their productivity is low for several reasons, such as genotype, drought, feed shortage, and various diseases [6,8,9]. Brucellosis is one of the diseases that hamper the productivity of small ruminants. Brucellosis is a zoonotic disease of considerable global economic and public health significance [10,11]. Brucellosis is characterized by reproductive disorders such as abortion, stillbirth, and the birth of weak offspring in females, and orchitis and epididymitis in male animals, causing transient or permanent infertility [12]. The disease is caused by intracellular bacteria, Brucella, and it is a contagious and zoonotic disease. Sheep and goats are affected mainly by Brucella melitensis and Brucella ovis, and sporadic infections by Brucella abortus and Brucella suis can occur [13]. According to Wakjira et al. [14], the seroprevalence of small ruminant brucellosis in Ethiopia ranges from 0.7% to 13.7%.

Various reports from different parts of Ethiopia have been published [15-21]. On the other hand, no study has been conducted in the Gomole and Elewaye districts, Borana zone, southern Ethiopia. Therefore, the objectives of this study were to estimate the seroprevalence of brucellosis in sheep and goats and to assess the associated risk factors in these districts.

Materials and Methods

Ethical statement

Ethical clearance was obtained from the College of Natural and Computational Sciences (CNCS) Ethical Committee, Hawassa University (reference number: CNCS-REC016/24).

Study areas

This study was carried out in Gomole and Elewaye districts in the Borana zone. The sheep and goat populations of the Borana zone were 420,512 and 657,479, respectively [3]. Generally, the Borena plateau represents a lowland area where the altitude gently slopes from the northern region, 1,650 meter above sea 1evel, to the southern region, 1,000 meter above sea 1evel [22]. The area is characterized by erratic rainfall, and the average annual rainfall is less than 450 mm. The main rainy time extends from March to May (locally known as “Ganna”) and from the last week of September to early November (locally termed as “Hagaya”). In Borana, livestock shares common grazing and browsing areas and watering points. Five or more village herds/flocks are often maintained together to reduce labor demand.

Study design

A cross-sectional study design was used to estimate the prevalence of small ruminant brucellosis and assess the potential risk factors associated with the disease. The study was conducted from January 2021 to August 2021. The list of districts in the Borana zone was collected, and 2 out of 13 districts in the zone were then selected, Gomole and Elwaye, based on the absence of information and lack of study in the areas (Fig. 1). From the 2 districts, the target Kebeles was selected based on small ruminant flock size.

Study animals and risk factors

The study animals were sheep and goats managed under an extensive pastoral production system. Sheep and goats above 6 months of age were included in the study. The study areas have no vaccination history for brucellosis. The study flock and animals were selected using systematic random sampling methods. The risk factors considered during this study included age, sex, flock size, parity number, and history of reproductive problems (i.e., mainly abortion and retained fetal membrane). Based on the local community flock size grouping system (i.e., mixed species of sheep and goats), the flock size was categorized into 3 levels: large (≥ 40), medium (16-40), and small (≤ 15).

Sample size determination

The sample size required for the study was calculated by considering the prevalence of 6.1% and 9.2% of sheep and goats, respectively [23]. The formula described by Thrusfield [24], 95% confidence interval (CI), and 5% desired absolute precision were used for the sample size determination. Accordingly, the computed sample size was 216 (i.e., 88 for sheep and 128 for goat). The sample size was increased threefold to increase the accuracy, and 264 sheep and 384 goats (i.e., a total of 648 animals) were studied.

Study methodology

Blood sample collection

The selected animals were properly restrained, and a 10 mL aseptic blood sample was collected from the jugular vein using a sterile vacutainer needle and plain vacutainer tubes. The collected blood samples were labelled with all necessary information using a permanent marker. The samples were transported to Yabello Regional Veterinary Laboratory in an ice box. Upon arrival at the laboratory, blood samples were left to stand overnight by placing the tubes in slant positions to allow blood clotting. The sera were extracted, filled into sterile cryo vials, and stored at −20°C until the laboratory tests were conducted.

Laboratory tests

All sera samples were screened by rose Bengal plate test (RBPT) as described by Office International des Epizooties (OIE) [13] for Brucella agglutinin. The test antigen was obtained from the Animal and Plant Health Agency, New Haw, Addle Stone, Surrey, KT15 3NB, UK. During the RBPT procedure, 25 μL of the antigen was placed on a glass slide, and 75 μL of sheep or goat serum was added. The antigen and the test serum were mixed thoroughly with a plastic applicator, shaken for 4 minutes, and the result was read immediately. The sample was checked for the presence or absence of agglutination by the naked eye. The result was recorded as 0 (no agglutination), + (barely visible agglutination), ++ (fine agglutination) and +++ (coarse clumping). Those samples with no agglutination were recorded as negative, while the others were recorded as positive.

Those positive reactor sera for RBPT were further subjected to indirect enzyme-linked immunosorbent assay (iELISA) for confirmation. This test was conducted at Yabello Regional Veterinary Laboratory on a microplate coated with inactivated antigens according to the manufacturer’s instructions. One hundred microliters of pre-diluted sera and controls (1:400) were added to a microplate and incubated for one hour at 37°C. The plate was then washed 3 times. Subsequently, 100 μL of conjugate was added to each well. The palates were then covered and incubated for one hour at 37 °C. The plates were then washed 3 times. Finally, 100 μL of substrate was added to each well and incubated at 26°C for 15 minutes. In the next step, 100 μL of stop solution was added to each well, and the result was read at a wavelength of 450 nm. The results are expressed as the percentage of the ratio between the corrected sample optic density (OD) and positive control OD. The test was conducted according to the manufacturer guidelines (ID.VET Innovative Diagnostic ID Screen@ Brucellosis Serum Indirect Multispecies, BRU-MS-ver. 1,014 GB. ID-vet, 310, rue Louis Pasteur-Grabels-FRANCE). According to Praud et al. [25], the sensitivity and specificity of iELISA were 91.7% (88.2%-99.2%) and 95.2% (47%-100%), respectively. The diagnostic kit, iELISA, used in this study, was diagnosed to detect antibodies directed against B. abortus, B. melitensis, and B. suis.

Case definition

The animal was considered positive if it was seropositive for RBPT and iELISA tests. Flock was considered seropositive when at least one animal in the flock was positive for both tests. All seropositive results were considered natural infections because there was no vaccination history against brucellosis in the Borana area.

Data analysis

All data collected during the study were entered into a Microsoft Excel spread sheet, edited, and coded. It was summarized using descriptive statistics, such as frequency, mean, and percentages. The association between the risk factors considered for this study and the seroprevalence of brucellosis was analyzed at the individual animal and flock level using univariable logistic regression. Those variables with p ≤ 0.25 in univariable logistic regression analysis were tested for collinearity, and the non-collinear variables were subjected to multivariable logistic regression analysis. A multivariable model was applied using non-collinear variables. Finally, the model fitness was assessed using the Hosmer-Lemeshow goodness-of-fit test [26]. Further selection of the variables in the final model was based on a stepwise backward elimination procedure. The data were analyzed using STATA ver. 14.0 for Windows (Stata 14 Corp. College Station, USA). The study considered the 95% CI and the 5% desired level of precision.

Results

Sera were collected from 648 small ruminants (i.e., 264 sheep and 384 goats) selected from 76 flocks using a systematic random technique. The overall seroprevalence of brucellosis was 6.8% and 6.2% with RBPT and iELISA, respectively (Table 1). Among 76 flocks selected, 25 (32.9%) had at least one animal positive for Brucella antibodies.

Risk factors analysis

Flock-level univariable and multivariable logistic regression analyses showed no significant variation among the districts, animal species, and flock size (Table 2). Table 3 lists the animal-level univariable and multivariable logistic regression analyses of the potential risk factors. The seroprevalence of brucellosis was higher in goats than sheep, small ruminants aged ≥ 5 years, and in flocks of greater or equal to 40. Those risk factors with p ≤ 0.25 (animal species, age, sex, parity, body condition score, reproductive problems, and flock size) in univariable logistic regression were checked for collinearity. None of them were collinear. Hence, they were subjected to multivariable logistic regression analysis (Table 3). Multivariable logistic regression analysis showed that the age of the animal, flock size, and body condition score significantly (p < 0.05) affected the seroprevalence of brucellosis.

Discussion

This study assessed the serological prevalence of small ruminant (sheep and goats) brucellosis and associated risk factors in 2 selected districts of the Borana zone, southern Ethiopia. Information on the status of brucellosis is essential for planning to control and eradicate brucellosis. From the 648 small ruminant sera collected, 44 (6.8%) and 40 (6.2%) were positive, according to RBPT and iELISA, respectively. This diagnostic kit, iELISA, mainly detects antibodies against B. abortus, B. melitensis, and B. suis. The seroprevalence of brucellosis in sheep and goats was 3.4% (n = 264) and 8.1% (n = 384), respectively. This finding generally agrees with Wubishet et al. [23] from the Yabello district, Borana zone. This prevalence falls in the range of small ruminant seroprevalence brucellosis reported in Ethiopia, 0.07% to 13.7% [16,17]. Borana has no history of vaccination for brucellosis (personal communication); hence, all seropositive animals could be considered natural infections.

A higher seroprevalence was recorded in goats (p < 0.05) than in sheep, approximately 2.15 more likely to occur in goats. This finding aligns with Sintayehu et al. [16] and Tadeg et al. [17]. Generally, if the nutritional need of a buck is fulfilled and the health status is good, it can mate with 10 to 40 does in a day. On the other hand, a healthy ram can usually mate with 3 to 4 3 to 4 ewes per day. Therefore, the chance for the spread of the disease in the goat flock is comparatively higher in goats.

The seroprevalence of brucellosis increases significantly with the age of the study animals. The risk of acquiring Brucella infection was 11.23 times higher in animals 5 or more years old (χ² = 3.67, p < 0.05). This finding generally agrees with reports from various parts of the country [15,18-21]. Higher seroprevalence of brucellosis was observed in adults because of the increasing susceptibility following sexual maturity [27]. Generally, the chance of animal contact and free mating should increase when the age of the animals and flock size increases, especially in extensive livestock management systems; hence, this might increase the opportunity for the disease to spread in the flock. Moreover, following infection, antibodies like immunoglobulin (Ig) M and IgG can be monitored, but these antibodies fade after the animal improves, often over 18.5 to 24 months [28]. This extended period could increase the probability of Brucella seropositivity.

The flock size was significant (χ² = 2.17, p < 0.05) related to the increasing seropositivity of Brucella in small ruminants. In a Borana livestock management system, a flock comprises a mixture of sheep and goats; hence, the flock-level analysis was not done separately for each species. Multivariable logistic regression analysis showed that the odds of seropositivity in larger flock size, ≥ 40 animals, were 9.53 times higher than the small flock, ≤ 15 animals. This finding aligns with the reports from various parts of the country [15,29-35]. According to Al-Majali [35], larger flocks are likelier to have at least one positive animal than smaller flocks, possibly because of the higher close contact between the animals and ease of transmission. In general, higher seroprevalence is expected in areas where small ruminant density is higher [32].

The seroprevalence of brucellosis was statistically higher (odds ratio [OR], 2.92; p < 0.05) in good body condition animals than in medium to poor body conditions, which was also reported by Ullah et al. [36]. It was also significantly higher (OR, 2.15; p < 0.05) in goats (8.1%; 95% CI, 5.7%-11.3%) than in sheep (3.4%; 95% CI, 1.8%-6.4%). These observations might be because of the difference in the mating behavior of the 2 species. The mating behavior of sexually matured animals that are in good nutritional and body conditions could be influenced positively. Hence, their mating desire and frequency could increase. Sheep and goats in such conditions might serve up to 40 females (i.e., does and ewes) per day. Therefore, if the male animals (i.e., rams and bucks) are infected, then the transmission in the flock is faster, and a large number of female animals will be infected. Herding mixed livestock (i.e., cattle, sheep, goats, and camel) is a standard practice in the Borana pastoral area [37]. This could increase the animal density, leading to higher contact and the spread of Brucella infections [20,37-39]. Moreover, Olsen and Palmer [40] reported that goats are more susceptible to infection than sheep.

The seroprevalence of small ruminant brucellosis was higher and spread in both districts of Borana zone. The seroprevalence of brucellosis was significantly higher in goats, adult animals (≥ 5 years), and large flock sizes (≥ 40 animals). Implementing control measures (for example, testing and culling reactor animals) and raising public awareness are recommended. Hence, further study will be needed to identify Brucella species circulating in the areas.

Notes

The authors declare no conflict of interest.

Author’s Contributions

Conceptualization: all authors; Data curation: Haro G; Formal analysis: Sheferaw D; Investigation: all authors; Methodology: all authors; Supervision: Sheferaw D; Writing-original draft: Haro G; Writing-review & editing: Sheferaw D.

Acknowledgments

The authors would like to extend their gratitude to the staff members of Yabello Regional Veterinary Laboratory for their technical and material support.

This paper was published with special support from the Korean Society of Veterinary Science.

Data Availability Statement

The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.

Fig. 1.

Map showing the study area (Borana Zone of Oromia Region).

Table 1.

Seroprevalence levels of small ruminants (sheep and goats) brucellosis in Elwaya and Gomole districts

Species	No. tested	Animal level
		RBPT		iELISA
		Positive (n, %)	95% CI	Positive (n, %)	95% CI
Sheep	264	10 (3.8)	2.0-6.9	9 (3.4)	1.8-6.4
Goats	384	34 (8.9)	6.4-12.1	31 (8.1)	5.7-11.3
Total	648	44 (6.8)	5.1-9.0	40 (6.2)	4.6-8.3

RBPT, rose Bengal plate test; iELISA, indirect enzyme-linked immunosorbent assay; CI, confidence interval.

Table 2.

Flock-level logistic regression analyses of seroprevalence of small ruminant (sheep and goats) brucellosis

Variable	Category	No. examined	RBPT				iELISA
Variable	Category	No. examined	Positive (n, %)	95% CI	OR	p-value	Positive (%)	95% CI	OR	p-value
District	Elwaya	38	15 (39.5)	25.0-56.0	0.12	0.807	13 (34.2)	20.7-50.9	0.89	0.807
	Gomole	38	12 (31.6)	18.6-48.3	Ref.		12 (31.6)	18.6-48.3
Species	Sheep	25	6 (24.0)	10.9-45.0	Ref.		5 (20.0)	8.3-40.9
	Goat	51	21 (47.2)	28.3-55.4	0.95	0.100	20 (39.2)	26.6-53.5	1.04	0.086
Flock size	≤ 15	10	1 (10.0)	1.2-50.4	Ref.		1 (10.0)	1.2-50.4
	15-40	11	5 (45.5)	19.0-74.7	1.22	0.332	3 (27.3)	8.4-60.7	0.27	0.267
	≥ 40	55	21 (38.3)	26.1-51.9	1.72	0.116	21 (38.2)	26.1-51.9	1.87	0.096
Total		76	27 (35.5)	25.4-47.1			25 (32 9)	23.1-44.4

RBPT, rose Bengal plate test; iELISA, indirect enzyme-linked immunosorbent assay; CI, confidence interval; OR, odds ratio; ref., reference.

Table 3.

Logistic regression analysis of the seroprevalence of small ruminant brucellosis in selected districts of Borana, Ethiopia

Variable	Category	No. examined	Positive (n, %)	95% CI	Univariable		Multivariable
Variable	Category	No. examined	Positive (n, %)	95% CI	OR	p-value	OR	p-value
District	Gomole	337	20 (5.9)	3.9-9.0	Ref.
District	Elwaye	311	20 (6.4)	4.2-9.8	0.92	0.793
Species	Sheep	264	9 (3.4)	1.8-6.4	Ref.		Ref.
Species	Goats	384	31 (8.1)	5.7-11.3	2.49	0.019	2.15	0.039
Age (y)	0.5-2	179	3 (1.7)	0.5-5.1	Ref.		Ref.
	3-4	349	20 (5.7)	3.7-8.7	3.57	0.042	1.11	0.051
	≥ 5	120	17 (14.2)	8.9-21.7	9.68	0.000	11.23	0.000
Sex	Female	588	34 (5.8)	4.2-7.9	Ref.
Sex	Male	60	6 (10.0)	4.5-21.7	1.81	0.202	0.84	0.117
BCS	Poor	244	13 (5.3)	3.1-9.0	Ref.		Ref.
	Medium	254	14 (5.5)	3.3-9.1	1.04	0.928	1.11	0.807
	Good	150	13 (8.7)	5.1-14.4	1.69	0.199	2. 92	0.017
Parity	None	49	1 (2.0)	0.3-13.4	Ref.
	1-2	217	5 (2.3)	0.9-5.4	1.13	0.911	1.06	0.958
	≥ 3	322	28 (8.7)	6.1-12.3	4.57	0.140	4.39	0.154
Reproductive problem	No	512	27 (5.3)	3.6-7.6	Ref.
Reproductive problem	Yes	76	7 (9.2)	4.4-18.2	1.35	0.176
Flock size	≤ 15	90	1 (1.1)	0.2-7.6	Ref.		Ref.
	16-39	133	4 (3.0)	1.1-7.8	2.76	0.368	2.41	0.444
	≥ 40	425	35 (8.2)	5.9-11.3	7.99	0.042	9.53	0.030
Total		648	40 (6.2)	4.6-8.3

CI, confidence interval; OR, odds ratio; ref., reference; BCS, body condition score.

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