Bovine Tuberculosis:

Prevalence and Risk Factor Assessment in Cattle and Cattle Owners in Wuchale-Jida District, Central Ethiopia

G. Ameni, DVM*†

K. Amenu, DVM†

M. Tibbo, DVM^‡

*Institute of Pathobiology, †Faculty of Veterinary Medicine, Addis Ababa University

‡International Livestock Research Institute (ILRI)

Addis Ababa, Ethiopia

KEYWORDS: Mycobacterium bovis; prevalence; risk factors; zoonosis

Abstract

A cross-sectional study was conducted between December 2001 and April 2002 on 94 households and 763 (188 indigenous and 575 crossbred) cattle to determine the prevalence of bovine tuberculosis (BTB) and assess its public health implications in smallholder farms in Wuchale-Jida District, Central Ethiopia. Cluster sampling, single intradermal tuberculin (SIDT) and comparative intradermal tuberculin (CIDT) tests, a questionnaire, and mycobacteriology were used. Based on the CIDT test, herd and individual animal prevalences of BTB were 42.6% and 7.9%, respectively. The individual animal prevalence was significantly affected by herd size (P<0.01), age (P<0.0001) and body condition (P<0.05). Among the interviewed households, 24.5% (23 of 94) had experienced at least one human tuberculosis case in the family. Of these families, 43.5% (10 of 23) had reactor cattle. Nevertheless, no statistically significant association (P>0.05) was observed between reactor cattle and human tuberculosis cases in households. The habit of milk and meat consumption was affected by occupation (P<0.0001) and location of household residence (P<0.001). Although the level of education influenced the habit of milk consumption (P<0.05), it did not impact the habit of meat consumption (P>0.05). Less than half (38.3%; 36 of 94) of the respondents knew about BTB, and only 30.8% (29/94) of the respondents were conscious of its transmission from cattle to humans. Secondary data analysis from Muka-Turri clinic indicated that 85.6% of the human tuberculosis cases were from rural parts of the district. Although the BTB prevalence seems low, its potential risk to public health was important based on food consumption, poor sanitary measures, and the lack of understanding about its zoonosis.

Introduction

In the tropics, dairy production based on indigenous cattle alone would not be a fast or suitable option for meeting the increasing demand for milk and milk products. The most popular alternative thus far has been crossbreeding to incorporate the hardiness of Bos indicus cattle with the productive capacity of Bos taurus.1 This should be accompanied by minimizing major animal health constraints such as bovine tuberculosis (BTB), which affects a higher proportion of exotic dairy breeds than indigenous Zebu cattle.2 Some studies conducted in Ethiopia also indicate the economic and public health consequences of BTB in farms owning cross/exotic cattle.3–6

Bovine tuberculosis is an infectious disease of cattle mainly caused by Mycobacterium bovis and characterized by progressive development of tubercles in any tissue or organ of the body.7 In countries in which BTB is uncontrolled, most human tuberculosis cases due to M. bovis occur in young individuals and result from drinking or handling contaminated milk. As a result, cervical lymphadenopathy, intestinal lesions, chronic skin tuberculosis (lupus vulgaris), and other nonpulmonary forms are particularly common. Such cases may, however, also be caused by Mycobacterium tuberculosis.8 M. bovis infection is certainly an occupational hazard to agricultural workers who may acquire it by inhaling cough spray from infected cattle and develop typical pulmonary tuberculosis.9 The epidemic of HIV and AIDS in developing countries is making M. bovis infection a serious public health threat.10,11

The introduction of exotic and crossbred cattle into the central highlands has created a favorable condition for the spread of BTB, leaving cattle, cattle owners, and consumers of raw cattle products at risk for infection from M. bovis. Wuchale-Jida district is one of the 12 districts in the North Shewa Zone (Ethiopia) where dairying is commonly practiced using small herd size (smallholder farms). However, the extent of BTB in the district in both cattle and humans was not known. This study was formulated to determine the prevalence of BTB and assess associated risk factors in cattle and cattle owners in Wuchale-Jida District, central Ethiopia.

Materials and Methods

Study Area

The study was performed in Wuchale-Jida District, central Ethiopia, from December 2001 to April 2002. Muka-Turri is the town and seat of administrative bodies of the district and is located 80 km north of Addis Ababa. The district, with a total area of 1,111 km2 (sharing 9.5% of the total area of the Zone) is divided into 3 agroclimatic zones: temperate (badda), subtropical (badda-dare), and tropical (gamoji), each sharing 87%, 11%, and 2% of the total area of the district, respectively. The district receives a mean annual rainfall of 1,026 mm. The mean minimum and maximum temperatures were 11.23˚C and 20.86˚C, respectively. The altitude of the district ranges from 1,000 to 3,000 meters above sea level.12 The climate of the area appears to be favorable for crop and livestock production. The district is the main milk shed for Addis Ababa.

The human population of the district is estimated at 107,138, with a crude population density of 96 persons per km2. The total number of households in the district was 20,830, and each household included an average of 5 individuals. The cattle population of the district was estimated at 132,023 cattle, of which 17,000 (12.9%) of the cattle were crossbred (mainly Friesian and Jersey with indigenous Zebu). The district was ranked second of the total 12 districts of the North Shoa Zone in its cattle population.13 The production system is mainly mixed crop-livestock production. Smallholder farmers rear the majority of farm animals at a subsistence level. Farmers owning crossbred cows sell milk to the milk collection units established by a group of interested farmers across the main road. Other groups of farmers sell their milk to private milk processing enterprises, which move to the area, collect milk from individual farmers, and then transport it to Addis Ababa.

Study Subjects and Epidemiological Design

The study included 94 households owning at least one-crossbred cattle in their herd. A total of 763 cattle (188 indigenous and 575 crossbred) belonging to these households were included in this study. The households were sampled from Muka-Turri town and the surrounding 5 villages. Cattle in the sampled households were of both sexes and above 6 months of age. Secondary data on the human health records in Muka-Turri clinic were used to assess the status of human tuberculosis in the district between April 2001 and March 2002.

The selection of the study area, Muka-Turri and the surrounding villages, was based on the concentration of crossbred cattle at the sites. The list of heads of the households, obtained from the district’s Agricultural Department, was used as sampling frame. The household was the primary sampling unit. A herd was defined, in this study, as cattle owned by a household.

Calculating the number of households required for this study using the formula described by Martin et al14 to estimate sample size would be appropriate. However, this was not done, becaue between-herd (household) variation in the prevalence of BTB was not determined earlier. As a result, simple random sampling was assumed for calculation of sample size, but the actual method used was cluster sampling, with a household as a cluster unit. In cluster sampling, to be on the safe side, 4 times as many animals were proposed to be included as for a simple random sampling method.14

Briefly, because the average herd size was 8 animals per herd (Desalegn, Personal communication, 2002), the expected prevalence of BTB was 14.2% based on the prevalence in Wolaita Soddo (Ethiopia).6 The statistical confidence level was decided to be 95%, and desired absolute precision was 5%. Accordingly, the sample size was determined to be 188 for simple random sampling, which corresponds to 23.5 households. Conversion to cluster sampling gave 94 households (23.5 x 4).

Questionnaire

The head of the household was interviewed using a predesigned questionnaire. The questions were focused on determining the respondent’s awareness about transmission of tuberculosis from cattle to humans and vice versa, habits of milk and meat consumption, recent history of tuberculosis cases in the family, and, if present, the type of tuberculosis. Local names were used for all scientific terms during the interview. Questions related to general livestock husbandry in the area were also included.

Tuberculin Test

Single intradermal tuberculin (SIDT) tests were performed on each of the study cattle, following the manufacturer’s instructions. The skin of the mid neck was shaved and examined for presence of any gross lesion. Thickness of the skin fold was measured with callipers, after which 0.2 mL (1,400 TU/mL) of bovine PPD (Bovitubal, strain AN5, Bioveta, Czech Republic) was injected intradermally using a semiautomatic syringe. The thickness of the skin fold at the site was measured again 72 hours after tuberculin inoculation. Animals were classified as negative, doubtful, or positive when the change in skin thickness was less than 2 mm, 2 mm or greater but less than 4 mm, or 4 mm of more, respectively. After the result of the SIDT test was read, positive and doubtful reactors were subjected to comparative intradermal tuberculin (CIDT) tests two months after the first test. Two sites at the middle of the neck 12.5 cm apart were shaved on the same side of neck, and skin fold was measured with a calliper. Then 0.2 mL (1,400 TU/mL) of bovine PPD (Bovitubal, strain AN5, Bioveta, Czech Republic) and 0.1 mL (28,000 TU/mL) of avian PPD (Avitubal, strain D4ER, Bioveta, Czech Republic) were injected intradermally. The sites were examined and skin thickness was measured 72 hours later. Interpretation of the result was made according to OIE.15

Milk Collection and Processing

About 20 mL of milk was drawn from 4 quarters of 24 tuberculin-positive cows under aseptic conditions and transported to the Institute of Pathobiology at 4˚C and stored at -20˚C until processing. The specimens were processed according to procedures given by Kazwala et al.16 Briefly, the milk was added to screw-capped sterile test tubes and centrifuged at 3,000 rpm for 15 minutes at room temperature. The cream was removed with sterile spatula, the supernatant was decanted, and the sediment was decontaminated with 3% NaOH. It was centrifuged again in the same condition and neutralized with concentrated HCl.

Bromocresol was used to monitor neutralization, which was achieved when the suspension changed from deep purple to yellow. The sediment was inoculated onto 2 Lowenstein-Jensen media (one with pyruvate and the other with glycerol). The culture was incubated at 37˚C for a week in a slant position and for 10 weeks in an upright position.

Data Analysis

Individual animal level prevalence was defined as the number of positive reactors per 100 animals tested. Herd level prevalence was computed as the number of herds with at least one-reactor cattle divided by the total number of herds tested. The variations between different factors were analyzed using Chi-square (c2) test. Odds ratio (OR) was calculated to assess strength of association of different factors to the occurrence of BTB in cattle and its potential risks in humans. For the analysis of the effect of different risk factors on tuberculosis status of the animals, doubtful results were considered negative. A P-value of <0.05 was considered statistically significant.

Results

Prevalence

Herd level characteristics: A herd prevalence of 42.6% (40/94) was recorded by CIDT test. The difference in prevalence among the different herd size classes was statistically significant (c2 = 13.3; df = 2; P<0.01). Cattle in herds of 16 or more heads of cattle had an about 11 times (OR=10.8; 95% Confidence interval [CI] = 2.5; 46.7) higher chance of infection with BTB than those in herds of 5 heads or less (Table 1).

The result of interviews indicated that 24.5% (23 of 94) of the households had experienced cases of human tuberculosis in the family, of which 43.48% (10 of 23) owned a herd with reactor cattle. Overall, 25 tuberculosis cases were reported from the households, of which 19 (76%) were pulmonary and the rest 6 (24%) were extrapulmonary type. However, the association between reactor cattle in the herd and human tuberculosis patients in the household was not statistically significant (c2=0.01; df =1; P>0.05).

Individual animal level characteristics: Individual animal prevalences of 11.3% (86 of 763) and 7.9% (60 of 763) were recorded by SIDT and CIDT tests, respectively. As body condition score increased from poor to medium and then to good, the risk of a positive result was increased (c2=11.09; df=2; P<0.001). The animals in good body condition had 5 times the risk (OR=5.09; CI=1.85;13.94) of reacting to tuberculin when compared with animals in poor condition. However, the odds of reactivity for animals in medium condition compared with those in poor condition was not significant. As indicated in Table 2, reactivity of the individual animals to tuberculin increased with age until 96 months and then decreased. The difference in tuberculin reactivity among the different age categories was statistically significant (c2=32.17; df=3; P<0.0001). There was no significant difference (c2=2.46; df=5; P>0.05) in the prevalence of BTB among the villages. Lactation and pregnancy did not significantly affect the prevalence rate.

Risk Factors for the Transmission of
M. Bovis to Humans
Habit of milk consumption: The result of the questionnaire survey showed that 52.1% (49 of 94) household heads had the habit of raw milk consumption. Conversely, 44.7% (42 of 94) boil fresh milk before consumption although they consume soured milk products without heat treatment. Only 3% (3 of 94) of household heads responded that they did not consume raw milk and milk products. Fortunately, 90% (85 of 94) of the respondents had the habit of boiling milk for babies. The habits of raw milk consumption were significantly (P<0.0001) different among the diverse occupational groups, residence groups, and levels of formal education. It was found that farmers, rural dwellers, and less educated respondents showed the habit of raw milk consumption more than their respective counterparts (Table 3).

Habit of meat consumption: Regarding meat consumption, 90% (85 of 94), 1% (1 of 94), and 9% (8 of 94) of the respondents consume mixed (raw and cooked meat), cooked, and raw meat, respectively. Unlike milk consumption, the habit of meat consumption was not affected by the educational background of the respondents (c2=5.78; df=6; P>0.05). However, there was significant difference between rural and urban dwellers (c2=11.91; df= 2; P<0.05), and among the different occupational groups (c2=17.79; df= 6; P<0.05) in their habit of raw meat consumption (Table 4).

Knowledge of Respondents About BTB Assessment of the knowledge of cattle owners about BTB showed that 38.3% (36 of 94) of the respondents knew that cattle can have tuberculosis, and 30.8% (29 of 94) recognized that BTB is zoonotic (Table 5). The results also indicated that the respondents’ knowledge of BTB transmission from cattle to humans and vice versa was influenced by educational background (c2=22.36; df=3; P<0.0001).

Of the 94 households interviewed, 13 used milk and milk products for home consumption. Regarding the marketing of milk and milk products, 36% (29 of 81) of the households sold milk and milk products directly to local people, and 56.5% (46 of 81) formed an association through which they pool fresh milk to nearby milk collection centers, process, and sell milk products. Only 7.5% (6 of 81) of the households sold milk to the commercial dairy processing industries in Addis Ababa.

Isolation of Mycobacterium From Milk

No growth of mycobacteria was observed on Lowenstein-Jensen medium from 24 milk samples collected from the tuberculin-positive cows.

Human Tuberculosis

Of a total of 97 tuberculosis cases registered at Muka-Turri clinic, 83.5% (81 of 97) and 16.5% (16 of 97) were diagnosed as pulmonary and extrapulmonary tuberculosis, respectively. Classification of the cases based on age indicated that 74.2% of the patients were between the ages of 16 and 45 years. Furthermore, 85.6% of them were rural dwellers. The frequency of extrapulmonary tuberculosis cases was high (87.5%) in rural areas of the district.

Discussion

The fact that 42.6% of the study herds contained at least one reactor head of cattle showed that BTB is widespread in Wuchale-Jida District. As stated by O’Reilly and Daborn,17 if all tuberculin-positive cattle are regarded as “open” cases of tuberculosis potentially capable of transmitting infection to other animals and humans, the animals and the community in the area are at risk of infection with M. bovis.

The increasing risk of BTB in cattle as the herd size increased seen in this study is consistent with previous reports by Cook et al18 and Asseged et al.19 This might be due to the fact that risk of an individual animal introducing tuberculosis infection into a negative herd may increase with herd size, and the lateral spread of infection within the herd may also be favored.

The association between reports of human cases of tuberculosis in the households and reactor cattle in the household’s herd was not significant. This result differed from previous reports by Cook et al18 and Ameni et al,6 who reported statistically significant associations between human tuberculosis cases and reactor herds.

Based on the sensitivity (90.9%) and the specificity (100%) of CIDT tests reported by Ameni et al20 in Ethiopia and the formula described by Putt et al,21 the true individual animal prevalence was estimated to be 8.6%. The presently recorded apparent prevalence of BTB (7.9%) is lower than the previous reports in intensive dairy farms in Ethiopia5,22 and in Eritrea.23 It is also lower than the 14.2% prevalence reported by Ameni et al6 in smallholder farms in southern Ethiopia. However, the 11.3% prevalence obtained by SIDT test in this study was in agreement with the 10.3% prevalence reported by Asseged et al19 using the same test. Cosivi et al8 indicated that breed of cattle, housing, and gathering of animals at watering and grazing sites have influenced the prevalence of BTB. This study was conducted in smallholder farms with smaller herds, in which animals are kept in the open air. This might have minimized the transmission and prevalence of BTB.

Breed-based analysis indicated no significant superiority of the indigenous cattle in resisting infection with M. bovis. This finding contradicts previous studies.5,6,16 This difference might be associated with the conservative way of classifying study animals into crossbred and indigenous without considering blood levels because of poor records on the animals.

Lower reactor levels were recorded in young animals, and reactivity increased with age to 96 months, after which it diminished. This finding was in agreement with previous studies.6,18,19 Infection might not have been established in young animals, but as the animals get older, the chance of being infected is higher because of sufficient exposure time. It has been reported that the disease is progressive in cattle,2,7 and progressive lesions provide sufficient repeated antigenic stimulation to cause temporary depression of skin reactivity.24 This depression of skin reactivity in progressive lesions could be due to T-helper 2 cell response predominating over the T helper 1 cell response.25,26

M. bovis can be transmitted to humans through inhalation of the cough spray from infected animals and ingestion of infected animal products. In addition, there are other routes of transmission of minor importance. Kleeberg9 indicated that one cow with tuberculous mastitis can excrete enough viable tubercle bacilli to contaminate the milk of up to 100 cows when milk pooling and bulk transportation is used. The same author noted tubercle bacilli has been found in milk products such as yogurt and cheese made from non-pasteurized milk 14 days after processing and in butter as long as 100 days after processing. Because most of the farmers either sell their milk to local people or pool milk in units for selling milk products without treating it with heat, risk of milk contamination with M. bovis is a potential major health hazard to consumers. Furthermore, the cattle owners’ poor understanding of BTB exacerbates the situation.

M. bovis was not isolated from the 24 milk specimens. Grange and Yates26 reported that tuberculosis in cattle was principally a pulmonary disease; only 1% of the tuberculous cows excrete tubercle bacilli in their milk, which shows that cows transmit the disease by erogenous route. In a study performed in Tanzania, of 805 milk samples, M. bovis was isolated and confirmed in only 2 milk specimens.16

The fact that the information obtained from Muka-Turri clinic indicated that a large proportion of the human tuberculosis cases originated in rural parts of the district might be associated with poor awareness of the transmission of BTB from cattle to humans, human-cattle interaction (eg, for housing), and the consumption behavior of people (eg, raw milk) in rural areas compared with people in urban areas.

In the near future, the number of private dairy farms in the study area is expected to increase in response to the demand for dairy products and the prevalence of BTB. This is mainly because of the absence of control measures in cattle and poor understanding of BTB and the open tuberculosis cases in human beings becoming a potential risk for both humans and cattle because of HIV/AIDS (data not presented). Therefore, awareness should be created through education of the public on the potential risk of BTB, proper food handling, and personal hygiene. Furthermore, more detailed studies are needed to assess and evaluate the scale of the problem and to design feasible and cost-effective control methods through close collaboration between the medical and veterinary professions.

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