Antibacterial and anti-inflammatory activity of plant species used in traditional poultry ethnomedicine in Zimbabwe: A first step to developing alternatives to antibiotic poultry feed additives.

ETHNOPHARMACOLOGICAL RELEVANCE: Global interest in phytogenic feed additives as alternatives to antibiotics in feed has been spurred by the banning of antibiotic growth promoters by several countries. Suitable plant extracts for development of phytogenic feed additives should have therapeutic value and should also be safe. AIM OF STUDY: The aim of this study was to evaluate the antibacterial, antioxidant and anti-lipoxygenase activities as well as cytotoxicity of selected plant species used in poultry ethnomedicine in Zimbabwe. METHODS: Antibacterial activity was determined against three ATCC strains (Staphylococcus aureus, Escherichia coli, Salmonella Enteritidis) and two clinical strains isolated from chickens (Escherichia coli and Salmonella Gallinarum) using a two-fold serial microdilution assay. Qualitative antibacterial bioautography was also carried out using the ATCC strains. Antioxidant activities of crude acetone and methanol extracts were determined using free radical scavenging assays whilst anti-lipoxygenase activity was evaluated using a ferrous oxidation-xylenol orange (FOX) assay. Cytotoxicity was evaluated using a tetrazolium-based colorimetric assay (MTT assay) on Vero monkey kidney cells. RESULTS: Erythrina abyssinica had the best antibacterial activity against both ATCC strains and clinical strains with minimum inhibitory concentration (MIC) values ranging from 0.02 to 0.156 mg/ml. Aloe greatheadii, Adenia gummifera (leaves), Senna singueana and Aloe chabaudii had moderate activity against the poultry pathogens. Bioautography showed that all ten plant species have antibacterial activity against the tested microorganisms with E. abyssinica and S. singueana having prominent bands of inhibition against both Gram-negative and Gram-positive bacteria. The acetone extract of S. singueana and the methanol extract of Euphorbia matabelensis had the most powerful antioxidant activities with mean IC50 values of 1.43 µg/ml and 1.31 µg/ml respectively in the ABTS assay which were comparable with those of the positive controls (ascorbic acid and trolox). Bobgunnia madagascariensis, A. chabaudii, E. abyssinica and Tridactyle bicaudata extracts had reasonable antioxidant activity. The S. singueana extract had the most potent anti-lipoxygenase activity with a mean IC50 value of 1.72 µg/ml. The cytotoxicity results showed that only the acetone extracts of A. greatheadii and S. singueana were relatively safe at concentrations that were active against the tested microorganisms (selective index >1). Regarding anti-lipoxygenase activity, extracts of B. madagascariensis, S. singueana, T. bicaudata and E. matabelensis were more active than toxic (selective index >5) indicating anti-inflammatory potential. CONCLUSIONS: This study showed that S. singueana had a cocktail of therapeutic activity and supports further investigation of this plant species for development of phytogenic poultry feed additives. Other plant species with noteworthy biological activities include B. madagascariensis, E. abyssinica, A. greatheadii, T. bicaudata and E. matabelensis.

A serological survey of anthrax in domestic dogs in Zimbabwe: a potential tool for anthrax surveillance.

Anthrax is an important disease caused by the bacterium Bacillus anthracis affecting both domestic and wild animals at the wildlife/livestock interface, defined here as a physical space in which wild and domestic species overlap in range and potentially interact. In endemic regions, sporadic anthrax outbreaks occur, causing significant deaths of both wildlife and livestock and sporadically, humans. However, it may also occur as isolated outbreaks with a few animals affected. Such isolated anthrax outbreaks maybe missed. High seroprevalence among carnivores suggests either regular non-fatal exposure to the pathogen circulating in a given environment, or contact with missed cases through consumption of anthrax carcases. To investigate the relevance of this potential indicator, a cross-sectional study was conducted to determine anthrax seroprevalence in domestic dogs (Canis lupus familiaris) from selected interface and non-interface areas of Zimbabwe with known history of anthrax outbreaks. Based on past anthrax outbreaks in the respective areas, the sites were further classified as high or low risk areas for anthrax outbreaks. Sera were collected from domestic dogs (n = 186) and tested for antibodies against B. anthracis protective antigens (PA) using an ELISA test. The overall seroprevalence was 51.6% (96/186; 95% CI 44.2-59.0). Sites from the non-interface areas recorded a significantly (P < 0.001) higher (72.1%) anthrax seroprevalence compared with those from the wildlife -livestock interface (41.5%). The results demonstrated a strong association (χ2 = 14.3; OR = 3.2, 1.6 < OR < 6.2, P < 0.001) between anthrax seropositivity and interface type. Low-risk sites (42.5%) had a significantly (P = 0.044) lower seroprevalence compared with high-risk sites (58.5%) but still demonstrated high seroprevalence for areas where anthrax was last reported more than 20 years back. Dogs from Tsholotsho South were more than 90-times (OR = 96.5, 13.5 < OR < 690.8) more likely to be seropositive compared with those from Hwange. The study demonstrated the potential to use domestic dogs as indicators of anthrax in the study areas to survey anthrax circulation in supposed low-risk areas and calls for a redefinition of both low and high risk areas for anthrax in Zimbabwe based on an improved surveillance.

Assessment of community awareness and risk perceptions of zoonotic causes of abortion in cattle at three selected livestock-wildlife interface areas of Zimbabwe.

A study was conducted to assess the awareness of cattle abortions due to brucellosis, Rift Valley fever (RVF) and leptospirosis, and to compare frequencies of reported abortions in communities living at the periphery of the Great Limpopo Transfrontier Conservation Area in southeastern Zimbabwe. Three study sites were selected based on the type of livestock-wildlife interface: porous livestock-wildlife interface (unrestricted); non-porous livestock-wildlife interface (restricted by fencing); and livestock-wildlife non-interface (totally absent or control). Respondents randomly selected from a list of potential cattle farmers (N = 379) distributed at porous (40·1%), non-interface (35·5%) and non-porous (26·4%), were interviewed using a combined close- and open-ended questionnaire. Focus group discussions were conducted with 10-12 members of each community. More abortions in the last 5 years were reported from the porous interface (52%) and a significantly higher per cent of respondents from the porous interface (P < 0·05) perceived wildlife as playing a role in livestock abortions compared with the other interface types. The odds of reporting abortions in cattle were higher in large herd sizes (odds ratio (OR) = 2·6; 95% confidence interval (CI) 1·5-4·3), porous (OR = 1·9; 95% CI 1·0-3·5) and non-porous interface (OR = 2·2; 95% CI 1·1-4·3) compared with livestock-wildlife non-interface areas. About 21·6% of the respondents knew brucellosis as a cause of abortion, compared with RVF (9·8%) and leptospirosis (3·7%). These results explain to some extent, the existence of human/wildlife conflict in the studied livestock-wildlife interface areas of Zimbabwe, which militates against biodiversity conservation efforts. The low awareness of zoonoses means the public is at risk of contracting some of these infections. Thus, further studies should focus on livestock-wildlife interface areas to assess if the increased rates of abortions reported in cattle may be due to exposure to wildlife or other factors. The government of Zimbabwe needs to launch educational programmes on public health awareness in these remote areas at the periphery of transfrontier conservation areas where livestock-wildlife interface exists to help mitigate the morbidity and mortality of people from some of the known zoonotic diseases.

Brucellosis in Sub-Saharan Africa: Current challenges for management, diagnosis and control.

Brucellosis is a highly contagious zoonosis caused by bacteria of the genus Brucella and affecting domestic and wild mammals. In this paper, the bacteriological and serological evidence of brucellosis in Sub-Saharan Africa (SSA) and its epidemiological characteristics are discussed. The tools available for the diagnosis and treatment of human brucellosis and for the diagnosis and control of animal brucellosis and their applicability in the context of SSA are presented and gaps identified. These gaps concern mostly the need for simpler and more affordable antimicrobial treatments against human brucellosis, the development of a B. melitensis vaccine that could circumvent the drawbacks of the currently available Rev 1 vaccine, and the investigation of serological diagnostic tests for camel brucellosis and wildlife. Strategies for the implementation of animal vaccination are also discussed.

Living at the edge of an interface area in Zimbabwe: cattle owners, commodity chain and health workers' awareness, perceptions and practices on zoonoses.

BACKGROUND: In the great Limpopo transfrontier conservation area (GLTFCA), there is an increased interface between wildlife and domestic animals, because rural households move their cattle into the game park in search of grazing and watering resources. This creates opportunities for inter-species transmission of infectious diseases, including zoonoses like brucellosis and tuberculosis, which may also pose a health risk to the local rural communities. This study investigated the awareness, perceptions and practices on zoonoses amongst rural cattle owners, commodity chain- and health-workers in three different localities around Gonarezhou National Park (GNP), Zimbabwe, where the interface between wild and domestic animals varies. METHODS: A cross-sectional study was conducted in Malipati, Chikombedzi and Chiredzi that are considered to be high-, medium- and low-domestic animal-wildlife interface areas, respectively. Data was collected from cattle owners, commodity chain and health-workers using a semi-structured questionnaire. To determine the public health risk of food-borne zoonoses, their practices with regard to meat and milk consumptions, and measures they take to prevent exposure to infections were assessed. Data were analyzed using descriptive statistics and principal component analysis. RESULTS: Most respondents (52.8 %, 102/193) were cattle owners, followed by health (30.1 %, 58/193) and lastly commodity chain workers (17.1 %, 33/193). Overall 67.4 % (130/193) of the respondents were aware of zoonoses with respective 48, 81.8, and 93.1 % of cattle owners, commodity chain, and health workers, being aware. Significantly more cattle owners (P < 0.05) from medium and low interface areas were aware of zoonoses compared to those from high interface areas. All categories of respondents cited anthrax (69.2 %), rabies (57.7 %), tuberculosis (41.5 %) and brucellosis (23.9 %) as important zoonoses. About half (46.1 %; 89/193) of the respondents perceive wildlife as important reservoirs of zoonoses. High proportions 98.4 % (190/193) and 96.4 % (186/193) of the respondents indicated that they consume meat and milk, respectively. Access to game meat and milk from informal markets was closely associated with consumption of raw meat and milk. CONCLUSIONS: Fewer cattle owners from a high interface area of Malipati are aware of zoonoses compared to other areas due to combined effects of limited education and other factors disadvantaging these marginalised areas. This may increase their risk of exposure to zoonoses, considering that consumption of raw meat and milk is common. Thus, awareness campaigns may reduce the public health impact of zoonoses at the interface.

Spatial modelling of Bacillus anthracis ecological niche in Zimbabwe.

Anthrax continues to cause significant mortalities in livestock, wildlife and humans worldwide. In Zimbabwe, anthrax outbreaks have been reported almost annually over the past four decades. In this study we tested whether anthrax outbreak data and a set of environmental variables can be used to predict the ecological niche for Bacillus anthracis using maximum entropy modelling for species geographical distribution (Maxent). Confirmed geo-referenced anthrax outbreaks data for the period 1995-2010 were used as presence locations and a set of environmental parameters; precipitation, temperature, vegetation biomass, soil type and terrain as predictor variables. Results showed that the environmental variables can adequately predict the ecological niche of B. anthracis (AUC for test data=0.717, p<0.001), with soil type as the most important predictor followed by variance of vegetation biomass and maximum temperature. These results imply that the model we tested may be used by animal health authorities in devising better control strategies for anthrax.

Awareness and attitude toward zoonoses with particular reference to anthrax among cattle owners in selected rural communities of Zimbabwe.

We conducted a cross-sectional study to assess cattle owners' awareness, perceptions, and attitudes toward zoonoses, with particular emphasis regarding anthrax. Data on awareness of zoonoses, clinical signs of anthrax in animals and human, its routes of transmission and methods of prevention, the families' consumption habits of anthrax-infected carcasses, and other family activities that increase exposure to anthrax were collected using an interviewer-administered questionnaire. A total of 41.4% (135/326) of the farmers were from high-anthrax-risk districts, whereas 28.5% and 30.1% were from medium- and low-risk districts, respectively. Overall, the level of awareness amongst the farmers for the named zoonoses were rabies (88.7%), anthrax (71.5%), and brucellosis (20.9%). Except for anthrax, awareness of other zoonoses did not differ significantly (p>0.05) among the district categories. Farmers from anthrax high-risk districts were significantly more aware of anthrax compared to those from moderate- (p=0.000) and low- (p=0.000) risk districts. All of the farmers were aware that anthrax occurs in cattle, and 73% indicated the presence of unclotting blood oozing from natural orifices as a consistent finding in cattle that died of anthrax, whereas 86.7% of them indicated the presence of skin lesions as the most common sign of the disease in humans. The good efficacy of human anthrax treatment (58.3%), slaughter of moribund cattle and selling of meat from cattle found dead to unsuspecting consumers (59.8%), reluctance to lose animals (47.9%), and forgetting about anthrax (41.1%) were cited as the major reasons for consuming anthrax-infected carcasses. Given that 75.2% of cattle owners indicated that they would not consume meat from cattle found dead, because they were discouraged by veterinary authorities, introducing meat inspection services is likely to have a positive impact in preventing human anthrax outbreaks in Zimbabwe.

Isolation of Bacillus anthracis from soil in selected high-risk areas of Zimbabwe.

AIMS: To isolate Bacillus anthracis from cattle carcass burial sites from high-risk districts in Zimbabwe. METHODS AND RESULTS: Soil samples were collected from carcass burial sites from seven areas, including two national game parks. Samples were collected from top 5-10 cm, and for spore extraction, 25 g of soil was suspended in sterile distilled water overnight. Supernatants were filtered through 0.45-µm pore cellulose nitrate, deposits suspended in 5 ml phosphate-buffered saline, aliquoted and heated at temperature regimen of 65, 70, 75 and 80 °C for 15 min. Samples were plated onto PLET agar. B. anthracis isolates were identified using growth morphology and PCR detecting pXO1 and pXO2 virulence plasmids. From samples heated at 75 °C for 15 min, B. anthracis were isolated from 9 of 81 (11.1%) soil samples representing five of the seven sampled areas. CONCLUSIONS: We isolated B. anthracis from soil collected from carcass burial sites. PCR targeting virulence plasmids provided a rapid confirmation of B. anthracis. SIGNIFICANCE AND IMPACT OF THE STUDY: The positive isolation indicated that some carcass burial sites may retain viable spores for at least 12 months after the previous outbreak, which suggests that they may be important sources of B. anthracis and new disease outbreaks.

Distribution of Salmonella serovars and antimicrobial susceptibility of Salmonella Enteritidis from poultry in Zimbabwe.

A study was carried out to determine the distribution and antimicrobial susceptibility profiles of Salmonella serovars from chickens from large-scale commercial (LSC), small-scale commercial (SSC), and rural free-range (RFR) farms of Zimbabwe. Pooled cloacal swabs were collected for culture and isolation of Salmonella spp. A chi-square test was used to assess distribution differences of salmonellas among the farming sectors. Approximately 10% (283/2833) of the swabs were positive for Salmonella enterica, with only subspecies enterica (98.6%) and arizonae (1.4%) being detected. The prevalence of S. enterica varied significantly (P<0.05) among areas, with Harare (27.8%) and Buhera (1.3%) recording the highest and the least prevalence, respectively. S. enterica was only isolated from LSC and SSC farms, with the former having a significantly (P<0.001; χ(2)=155.3) higher prevalence than the latter. S. arizonae was only isolated from the SSC farms while none were obtained from the RFR farms. The serovars isolated were Salmonella Enteritidis (72.8%), Group C (20.1%), Group B (4.2%), Salmonella Typhimurium (1.1%) and Salmonella Gallinarum (0.4%). S. Enteritidis predominated in the urban/periurban areas. Approximately 26% (53/206) of S. Enteritidis isolates were resistant to one or more antimicrobial agents. Resistance to tetracycline was the most common, while no resistance was detected for furazolidone, neomycin and trimethoprim-sulfamethoxazole. There were 12.1% multi-drug-resistant S. Enteritidis isolates, and the resistance to ampicillin/kanamycin was predominant. The identification of multi-drug-resistant S. Enteritidis is of public health concern. Thus, stringent control of S. Enteritidis will reduce the public health risk of human salmonellosis.

Brucella seroprevalence of the Kafue lechwe (Kobus leche kafuensis) and Black lechwe (Kobus leche smithemani): exposure associated to contact with cattle.

We investigated Brucella seroprevalence in Kafue (Kobus leche kafuensis) and Black (Kobus leche smithemani) lechwe antelopes to assess Brucella infections in relation to presence/absence of cattle interaction on the wetlands. Accordingly, two study populations based on cattle interaction were assesed: Kafue lechwe from Kafue flats which interact with cattle; and the Black lechwe with no known interaction with cattle from the Bangweulu swamps. Fourteen Kafue lechwe and thirty Black lechwe were slaughtered between October and December 2009 using special research licenses obtained from the Zambia wildlife authority to investigate diseases in lechwe antelope. For the purpose of this study, blood was collected and sera separated for Rose Bengal and indirect ELISA tests. Seroprevalence of Brucella in the Kafue lechwe was estimated at 42.9% [95% CI: 15.2-70.5] while that in Black lechwe was 0% [95% CI:0.0-11.6]. On the Kafue flats, cattle were spotted grazing in the same areas as lechwe while there was no evidence of cattle presence on the Bangweulu swamps. These differences in seroprevalence between Kafue lechwe and Black lechwe were assumed to be associated with interaction between Kafue lechwe and Brucella infected cattle, and no such contact existed between cattle and the Black lechwe. Our study suggests that brucellosis in the Kafue lechwe may have originated from cattle but has now established a reservoir in wild animals. It is also important to keep in mind that the Black lechwe can easily become infected with Brucella spp. once cattle are introduced in the surrounding areas.

Evaluation of sensitivity and specificity of RBT, c-ELISA and fluorescence polarisation assay for diagnosis of brucellosis in cattle using latent class analysis.

The sensitivity (Se) and specificity (Sp) of the Rose Bengal test (RBT), competitive ELISA (c-ELISA), serum (sFPA) and blood (bFPA) fluorescence polarisation assay for brucellosis were evaluated using latent class analysis using sera and whole blood collected from infected cattle reared in smallholder dairy farms of Zimbabwe. The latent class model allowed estimation of Se and Sp in the absence of a gold standard test. The c-ELISA had the highest Se (99.0%; 95% credible posterior interval (CPI): 94.8; 100%), while the RBT and sFPA had the highest Sp (99.0%; 95% CPI: 98.0; 99.6%). The bFPA had the lowest Se (71.3%; 95% CPI: 56.2, 83.5%), while its Sp (96.3%; CPI: 93.9; 98.0%) was marginally higher than that of the c-ELISA (95.4% CPI: 93.7; 96.8%). Therefore based on these data, test regimen using the RBT and c-ELISA could be suitable for diagnosis of brucellosis in smallholder dairies in Zimbabwe. Based on cost and ease of performance, the sFPA may be adopted as a confirmatory test, but its performance may be optimised by altering cut-off points to suit the Zimbabwean conditions. Thus, latent class models provide an alternative method for evaluating Se and Sp of diagnostic tests, which could be used to optimise test performance in different cattle populations.

Risk factors for Brucella spp. infection in smallholder household herds.

Risk factors for Brucella infection, the association and impact of Brucella seropositivity on abortions were investigated in cattle (n=1291) reared in smallholder household herds (n=203) from six geographical areas of Zimbabwe between September 2004 and 2005. Data on management, abortion and herd structure were collected. Sera were tested for Brucella antibodies using the Rose Bengal test and a competitive enzyme-linked immunosorbent assay. Data were analysed by generalized estimating equation and logistic regression models. Brucella antibodies were estimated at 5·5% and 22·9% for individual cattle and herds, respectively. Abortions were reported in 3·2% of cows and 22·0% herds. The age of cows and Brucella seropositivity predicted abortion. For herds, Brucella seropositivity, geographical area, purchase of cattle and large herd size were independently associated with increased odds of abortion. Exposure to Brucella had a significant impact on abortion. These results highlight the important risk factors for Brucella spp. infection in smallholder herds. Thus, brucellosis control programmes which take these factors into consideration will be beneficial.

Herd-level factors for Brucella seropositivity in cattle reared in smallholder dairy farms of Zimbabwe.

A cross-sectional study was conducted to investigate factors for Brucella seropositivity in smallholder dairy cattle herds from Gokwe, Marirangwe, Mushagashe, Nharira, Rusitu and Wedza areas located in different agro-ecological regions of Zimbabwe between September 2004 and November 2005. Sera were collected from cattle aged > or = 2 years from 203 herds. Data on herd-level and management variables were collected using a structured questionnaire. Sera were screened for anti-Brucella antibodies using the Rose Bengal test (RBT) and confirmed by competitive ELISA (c-ELISA). A herd was classified as Brucella seropositive if at least one animal tested seropositive on both tests. The herd-level factors for Brucella seropositivity were tested using multivariable logistic model with herd infection status as dependent variable while the levels of exposure in individual animals within-herds were analysed by negative binomial regression using the number of positive animals as the outcome. Of the 203 herds tested, 52 (25; 95% CI: 18.1, 31.9%) tested seropositive for brucellosis. Using the logistic regression model we identified area, with both Rusitu (OR=0.26; 95% CI: 0.07, 1.03) and Wedza (OR=0.07; 95% CI: 0.01, 0.49) having lower Brucella seropositivity compared to Gokwe. Keeping mixed cattle breeds (OR=8.33; 95% CI: 2.70, 25.72) compared to single breed herds, was associated with increased herd seropositivity. The farmer's knowledge of brucellosis was associated with lower odds (OR=0.17; 95% CI: 0.05, 0.55) of farms testing seropositive. The odds of Brucella seropositivity were progressively higher with increasing stocking density and herd size. Using the negative binomial regression model we identified area, keeping mixed breed herds, stocking density and herd size as independently associated with increased counts of seropositive cattle in a herd.

Effectiveness of Rose Bengal test and fluorescence polarization assay in the diagnosis of Brucella spp. infections in free range cattle reared in endemic areas in Zambia.

The effectiveness of Rose Bengal test (RBT) and fluorescence polarization assay (FPA) in diagnosing cattle brucellosis in endemic areas was assessed and RBT and FPA test agreement was compared (n = 319). The sensitivity of RBT and FPA in detecting low Brucella titres were evaluated in paired sera (n = 34). A logistic regression model was constructed to predict cattle test result in FPA using RBT as the main predictor and incorporating bio-data and animal history. There was 79.3% agreement between the RBT and FPA (Kappa = 0.59; Std error = 0.05; p = 0.000) and a high correspondence between high RBT scores and positive FPA results suggesting that sera with high RBT score may not require confirmation with tests such as competitive-ELISA or CFT. High FPA cut-off points were more likely to miss animals with low antibody titres. The RBT had a reduced ability in detecting low antibody titres compared to the FPA. FPA test interpretation was improved if a priori information, such as sex and age was used. Under the challenging disease surveillance conditions prevailing in rural Africa, field-testing methods that are sensitive and specific; allow single animal contact, low technical skills in data interpretation are suitable.

Responses to tuberculin among Zebu cattle in the transhumance regions of Karamoja and Nakasongola district of Uganda.

Responses to tuberculin in Zebu cattle of the transhumant pastoral farming system in Karamoja region and Nakasongola district in the north-eastern and mid-central regions in Uganda, respectively, were investigated using a comparative intradermal tuberculin skin test. Of the 1864 cattle tested from 30 large units (superherds) in Karamoja and 7 herds in Nakasongola, a total of 28 animals from 19 herds (51.4%) tested positive. Inter-district tuberculin reactor prevalence variations seemed to be influenced by climate, with impact on both the management patterns and transmissibility of agent. High herd tuberculin reactor prevalence (51.4%) was attributed to widespread contacts and mixing of animals between herds. Low individual animal tuberculin test positivity (mean = 1.4%) was attributed to low transmissibility of the agent under the Karamoja climate, which is semi-arid, and to increased resistance due to non-specific response to environmental mycobacteria and natural selection, since there was no active control against bovine tuberculosis. Owing to similarities in management practices in Karamoja and widespread risk factors, it was difficult to identify which were more important, but variations in sources of drinking water pointed to provision of lake and borehole water during dry season as reducing the risk. Positive bovine tuberculin reactor prevalence and skin reactor status were related to age.

Brucellosis surveillance and control in Zimbabwe: bacteriological and serological investigation in dairy herds.

Brucellosis in dairy cattle is endemic in Zimbabwe. The prevalence continues to be monitored intensively. Only milk and serum samples are routinely screened. Attempts to culture Brucella spp. from clinical specimens are seldom made. Consequently, incidence of various Brucella spp. within Zimbabwe is virtually unknown, despite the high serepositivity reported. This information is paramount in understanding the transmission cycle and is also significant to public health; particularly as B melitensis infects humans more often than do the other brucellae. This paper describes the results of bacteriological and serological investigations of brucellosis in a dairy from near Bulawayo. The said farm was selected for the present pilot study because of the high incidence of reported abortion. The milk ring test was employed to test the bulk pooled milk samples once a month for 14 months. The test was recorded highly positive on all 14 occasions. To locate reactors, milk samples from 36 individual cows were similarly tested. Of these, 21 (almost 59%) were found to be reacting positively. One hundred and seventy-seven animals were marked for serotesting. Of these 40 (approximately 25%) showed quite high serum titres (> 1:360) in both the STT and the Rosebengal test. The farmer was advised to havet all abortions full investigated. However, all the clinical material from cases of abortion, except one, were received in an advanced state of putrefaction. From this, Brucella was isolated on culture from stomach contents and cotyledons. The isolates from both the sites were characterized in detail, employing dye inhibition, phagetyping; the oxidative metabolic test and agglutination with monospecific sera. Both the isolates belonged to B. abortus biovar I, which was confirmed by the Central Veterinary Research Laboratory, Weybridge. The significance of isolation and the need to intensify similar studies have been discussed.