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  Table of Contents  
Year : 2019  |  Volume : 37  |  Issue : 3  |  Page : 426-432

Seroprevalence of brucellosis in Western Rajasthan: A study from a tertiary care centre

Department of Microbiology, All India Institute of Medical Sciences, Jodhpur, Rajasthan, India

Date of Submission25-Sep-2019
Date of Decision01-Nov-2019
Date of Acceptance07-Nov-2019
Date of Web Publication05-Dec-2019

Correspondence Address:
Dr. Ravisekhar Gadepalli
Department of Microbiology, All India Institute of Medical Sciences, Jodhpur, Rajasthan
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ijmm.IJMM_19_358

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 ~ Abstract 

Brucellosis is a significant bacterial zoonotic disease with a high seroprevalence in low-to-middle-income countries where there is a significant contact of humans with animals. This prospective study was conducted to observe the seroprevalence of brucellosis in 75 symptomatic patients with pyrexia of unknown origin (PUO) and 75 high-risk individuals (10 veterinarians, 15 milkmen and 50 healthy contacts of symptomatic patients) with possible exposure to brucellosis. Serum samples collected from these patients and individuals were subjected to rose Bengal test (RBT) and ELISA for the detection of IgM and IgG antibodies. RBT was positive in 50 samples (40 PUO and 10 high risk), of which 25 (33.3%) PUO cases had detectable IgM antibodies, whereas IgG antibodies were detected in 20 patients. None of samples from high-risk individuals was reactive for IgM/IgG antibodies by ELISA. The overall seroprevalence in this study was 16.7% (33.3% in PUO patients). Being a predominantly livestock rearing area, brucellosis is a significant health-care problem in this part of India with this entity being linked to 33% of PUO cases.

Keywords: Brucellosis, ELISA, pyrexia of unknown origin, zoonoses

How to cite this article:
Bansal Y, Aggarwal A, Gadepalli R, Nag VL. Seroprevalence of brucellosis in Western Rajasthan: A study from a tertiary care centre. Indian J Med Microbiol 2019;37:426-32

How to cite this URL:
Bansal Y, Aggarwal A, Gadepalli R, Nag VL. Seroprevalence of brucellosis in Western Rajasthan: A study from a tertiary care centre. Indian J Med Microbiol [serial online] 2019 [cited 2020 Oct 28];37:426-32. Available from:

 ~ Introduction Top

 Brucellosis More Details is a significant bacterial zoonotic disease with a high seroprevalence in low-to-middle-income countries where there is a significant contact of humans with animals. India ranks first in milk production and has the largest bovine population in the world[1] owing to which there is an inordinate exposure of workers in this industry to these animals and their products. The arid climate coupled with low economic investments in this part of western Rajasthan makes the majority of rural households rely on livestock for sustaining the household expenses.[2]

Early detection and treatment of brucellosis cases can be achieved by early recognition of the probable cases. This can be achieved by better understanding of the symptomatology and seroprevalence through epidemiological surveillance. Our study aims to determine the prevalence of human brucellosis in cases of pyrexia of unknown origin (PUO) and individuals at risk of exposure to  Brucella More Details in local population that included high-risk groups such as veterinarians, milkmen and contacts of symptomatic/confirmed cases.

 ~ Materials and Methods Top

The present study was a prospective study conducted from a period of January 2016 to June 2017 in a tertiary care centre of western Rajasthan. The study population comprised symptomatic individuals and a high-risk group. Patients with PUO were taken as symptomatic cases, whereas the high-risk group consisted of veterinarians, milkmen and other asymptomatic individuals having similar environmental exposure parameters as cases (such as patient's attendants) but without any evidence of the disease. A detailed history was collected from these individuals that included their name, age, occupation, nature of work, history of consumption of raw milk, history of fever (nature and duration) in the past and complaints of joint pains, if any and chronic illnesses.

Serum samples were collected from these individuals by obtaining 3–4 ml of blood sample in a Vacutainer without additives and separating the serum from these specimens. These sera were stored at −20°C until tested. The serum samples were analysed in two phases. In the first phase, serum samples were subjected to rose Bengal test (RBT). In the second phase, IgM and IgG antibodies against Brucella were detected by ELISA using Calbiotech Brucella IgG/IgM ELISA Kit (California, USA). This ELISA kit can detect IgG and IgM antibodies to Brucella in human serum or plasma.

Apart from the positive and negative controls supplied by the manufacturer along with the ELISA kits, additional controls were also run along with the samples. The serum collected from confirmed cases of brucellosis (isolated on repeating sampling and blood culturing) was used as a strong positive serum control. The serum samples taken from the healthy individuals with no history of contact with animals or other such high-risk behaviour/exposure were used as the negative control serum. A moderate positive control was prepared by diluting the strong positive serum in negative serum (1:20 fold dilution). The sera to be tested were diluted 1:21 before the ELISA procedure.

For the detection of IgM antibodies, the patient's serum was diluted with a serum diluent provided with the ELISA kit that contained a sorbent to remove rheumatoid factor and human IgG interference. This diluted serum was added to the wells coated with purified antigen. The rest of the procedure/steps were standard ELISA procedural steps as mentioned in the supplied kit literature. The study was conducted after obtaining due ethical clearance from the institute's ethical clearance committee.

 ~ Results Top

A total of 150 samples were collected during the study period that included 75 symptomatic cases (PUO) and 75 high-risk individuals that included 10 veterinarians, 15 milkmen and 50 healthy contacts of symptomatic cases such as patient attendants. The mean age of the participants was 31.4 years (range, 2–78 years). One hundred and six (70.7%) were male, whereas the rest 44 (29.3%) were female.

All symptomatic cases were investigated as cases of PUO in the institute and the symptoms in these patients are given in [Table 1]. A total of 50 (33.3%) samples were positive by RBT, which was followed by the ELISA testing of all 150 samples to detect IgM and IgG antibodies. Of 150 samples, IgM antibodies were detected in 25 (16.7%) samples, whereas IgG antibodies were detected in 20 (13.3%) samples. All samples with detectable IgG antibodies were reactive for IgM antibodies as well. Hence, 13.3% (20/150) of samples were positive for IgM antibodies as well as IgG antibodies, whereas 3.3% (5/150) were reactive for IgM antibodies only. None of the samples tested were reactive only for IgG antibody.
Table 1: Symptomatology, serological tests and their performance characteristics

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All ELISA positive samples (reactive for IgM ± IgG antibodies) belonged to the symptomatic group. None of the patients from high-risk group had a detectable antibody level though 10/75 (13.3%) were positive by RBT. The group-wise distribution of serology results along with their sensitivity, specificity, positive predictive value and negative predictive values is shown in [Table 1].

 ~ Discussion Top

The present study was done to assess the burden of brucellosis in the region as the true incidence of this disease in a developing country like India is still unknown despite agriculture, livestock and cattle rearing being a significant source of income for majority of the population in low- and middle-income groups in rural and semiurban areas.[3],[4] The area to which our institute caters to is known to be an agriculture and livestock rearing area where humans maintain a close contact with animals such as cows, goats, sheep, buffaloes and camels for the past few centuries.[2]

There are two main sources of dairy products in this region. The commercial supply is through industries that supply pasteurised products with maintenance of the cold chain, whereas cattle dwellers that supply raw, unpasteurised products also contribute significantly to the dairy product consumption. Kachhawaha et al. have documented the problem of brucellosis in animals in this region.[5] The prevalence of brucellosis in these animals ranges from 17.7% to 75% and is said to be increasing as a result of the ban on cow slaughter.[5] Hence, a high prevalence of brucellosis is expected in this region.

In the present study, the overall seroprevalence of brucellosis was 16.7%, whereas the seroprevalence in PUO patients and high-risk group was 33.3% and 0%, respectively. The surprising lack of seropositivity in high-risk group could be due to low numbers of milkmen and veterinarian in our study. The seroprevalence from other parts of India has been reported in a few studies. A study from Chandigarh in PUO patients showed a Brucella positivity of 9.94% by serology over a period of 10 years.[6] Another study from Karnataka over a period of 10 years (2006–2015) showed 5.1% seropositivity in suspected patients (PUO).[7] Various studies from India have documented brucellosis seroprevalence ranging from 2% to 18%.[8]

In an earlier study from our region, the seroprevalence in PUO patients was found to be 13% in which the diagnosis was made only using IgG ELISA.[9] We used both IgM and IgG ELISA for establishing the diagnosis, and a positive patient was defined as a patient with a positive IgM serology with or without IgG antibodies. This could be one of the reasons for significantly higher association of brucellosis and PUO in our study. Furthermore, the fact that our institute is a referral hospital in this region could be another reason as there is a delay between onset of symptoms and the patient's presentation to our institute. This results in time for IgM and/or IgG antibody formation in the patient.

The gold standard for the diagnosis of brucellosis is blood culture, but its aerosolisation during culture procedures coupled with a very low infective dose of 10–100 bacilli makes it a very potent laboratory-acquired pathogen.[10] There are very few articles on culture-confirmed brucellosis from India such as Mani et al., where they reported 22 culture-confirmed cases of brucellosis.[11] Because the facility (BSL-3 laboratory) for the same is lacking in our institute, the isolation of Brucella from the blood cultures was not attempted as part of the study.

Our study shows the relative utility of each test for serological diagnosis. Many previous serology-based studies used ELISA tests that either was not able to distinguish between IgM and IgG or only detected IgG antibodies, thereby introducing a possibility of missing acute cases. Among serological tests, we designated the combined use of IgM and IgG antibody detection as the gold standard for serological diagnosis and compared the performance characteristics of other tests with it. RBT suffered specificity issues, whereas IgG ELISA suffered sensitivity issues as it missed five acute cases.

 ~ Conclusions Top

Brucellosis is a significant health-care problem in this part of India, with almost one-third of PUO cases being attributable to this entity. Being predominantly an agriculture and livestock rearing area, the on-going transmission between man and cattle is hard to break and a good effort from the public health authorities needs to be made to spread awareness regarding high-risk behaviour such as the consumption of unboiled/unpasteurised milk.

Financial support and sponsorship

Supported by intramural grant from AIIMS, Jodhpur.

Conflicts of interest

There are no conflicts of interest.

 ~ References Top

Deka RP, Magnusson U, Grace D, Lindahl J. Bovine brucellosis: Prevalence, risk factors, economic cost and control options with particular reference to India – A review. Infect Ecol Epidemiol 2018;8:1556548.  Back to cited text no. 1
Patidar M, Patel AK, Misra AK, Sirohi AS, Kumawat RN, Meghwal PR, et al. Improving Livelihood of Farmers through Livestock Interventions in Nagaur District of Rajasthan. Jodhpur: Central Arid Zone Research Institute (CAZRI); 2014. 56. Available from: [Last accessed on 2019 Sep 14].  Back to cited text no. 2
Shome R, Kalleshamurthy T, Shankaranarayana PB, Giribattanvar P, Chandrashekar N, Mohandoss N, et al. Prevalence and risk factors of brucellosis among veterinary health care professionals. Pathog Glob Health 2017;111:234-9.  Back to cited text no. 3
Rubach MP, Halliday JE, Cleaveland S, Crump JA. Brucellosis in low-income and middle-income countries. Curr Opin Infect Dis 2013;26:404-12.  Back to cited text no. 4
Kachhawaha S, Singh K, Tanwar RK. Serological survey of Brucellosis in cattle and buffaloes of Jodhpur Region. Vet Pract 2005;6:43-4.  Back to cited text no. 5
Appannanavar SB, Sharma K, Verma S, Sharma M. Seroprevalence of brucellosis: A 10-year experience at a tertiary care center in North India. Indian J Pathol Microbiol 2012;55:271-2.  Back to cited text no. 6
  [Full text]  
Patil DP, Ajantha GS, Shubhada C, Jain PA, Kalabhavi A, Shetty PC, et al. Trend of human brucellosis over a decade at tertiary care centre in North Karnataka. Indian J Med Microbiol 2016;34:427-32.  Back to cited text no. 7
[PUBMED]  [Full text]  
Mantur BG, Amarnath SK, Shinde RS. Review of clinical and laboratory features of human brucellosis. Indian J Med Microbiol 2007;25:188-202.  Back to cited text no. 8
[PUBMED]  [Full text]  
Ali S, Bohra GK, Kothari D, Kumar D, Vyas T. Seroprevalence of brucellosis in Western Rajasthan. J Med Sci Clin Res 2014;2:332-8.  Back to cited text no. 9
Noviello S, Gallo R, Kelly M, Limberger RJ, DeAngelis K, Cain L, et al. Laboratory-acquired brucellosis. Emerg Infect Dis 2004;10:1848-50.  Back to cited text no. 10
Mani SS, Gunasekaran K, Iyyadurai R, Prakash JA, Veeraraghavan B, Mishra AK, et al. Clinical spectrum, susceptibility profile, treatment and outcome of culture-confirmed brucellosis from South India. Indian J Med Microbiol 2018;36:289-92.  Back to cited text no. 11
[PUBMED]  [Full text]  


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