This study was conducted to determine the seroprevalence of human
brucellosis and identify the potential risk factors in a rural area of
Western Anatolia, Turkey. A simple random-sampling method was used for
identifying 1,052 subjects for the study. Blood samples, collected from
all the subjects, were studied following the methods of Rose Bengal
slide agglutination and standard tube agglutination tests. One thousand
and one samples (95.2%) were seronegative, and 51 (4.8%) were
seropositive. There was a statistically significant correlation between
seropositivity and age, sex, consuming fresh cheese and cream made from
unboiled milk (p values 0.005, 0.019, <0.001, and <0.001
respectively). Seropositivity was not related to educational level
(0.270). It is concluded that pasteurization of milk and dairy products
and education regarding eating habits must be pursued for eradication
of human brucellosis from rural areas. The findings of the study
suggest that human brucellosis is still an important public-health
problem in the western Anatolia region of Turkey, especially in rural
areas

Ihsan H. Ciftci

Orhan C. Aktepe

Reha Demirel

Zafer Cetinkaya

University of Toronto Research Repository

Seroprevalence of Human Brucellosis in a Rural Area of Western Anatolia, Turkey

TSpace Repository

This study was conducted to determine the seroprevalence of human
brucellosis and identify the potential risk factors in a rural area of
Western Anatolia, Turkey. A simple random-sampling method was used for
identifying 1,052 subjects for the study. Blood samples, collected from
all the subjects, were studied following the methods of Rose Bengal
slide agglutination and standard tube agglutination tests. One thousand
and one samples (95.2%) were seronegative, and 51 (4.8%) were
seropositive. There was a statistically significant correlation between
seropositivity and age, sex, consuming fresh cheese and cream made from
unboiled milk (p values 0.005, 0.019, &lt;0.001, and &lt;0.001
respectively). Seropositivity was not related to educational level
(0.270). It is concluded that pasteurization of milk and dairy products
and education regarding eating habits must be pursued for eradication
of human brucellosis from rural areas. The findings of the study
suggest that human brucellosis is still an important public-health
problem in the western Anatolia region of Turkey, especially in rural
areas

Cetinkaya, Zafer

Aktepe, Orhan C.

Ciftci, Ihsan H.

Demirel, Reha

Bioline International

J HEALTH POPUL NUTR    2005 Jun;23(2):137-141 © 2005 ICDDR,B: Centre for Health and Population ResearchISSN 1606-0997        $ 5.00+0.20Seroprevalence of Human Brucellosis in a RuralArea of Western Anatolia, TurkeyZafer Cetinkaya1, Orhan C. Aktepe1, Ihsan H. Ciftci1, and Reha Demirel21Department of Microbiology and 2Department of Public Health, Faculty of Medicine,Afyon Kocatepe University, Gazligöl Yolu, 03200 Afyon, TurkeyABSTRACTThis study was conducted to determine the seroprevalence of human brucellosis and identify the poten-tial risk factors in a rural area of Western Anatolia, Turkey. A simple random-sampling method wasused for identifying 1,052 subjects for the study. Blood samples, collected from all the subjects, werestudied following the methods of Rose Bengal slide agglutination and standard tube agglutinationtests. One thousand and one samples (95.2%) were seronegative, and 51 (4.8%) were seropositive.There was a statistically significant correlation between seropositivity and age, sex, consuming freshcheese and cream made from unboiled milk (p values 0.005, 0.019, <0.001, and <0.001 respectively).Seropositivity was not related to educational level (0.270). It is concluded that pasteurization of milkand dairy products and education regarding eating habits must be pursued for eradication of humanbrucellosis from rural areas. The findings of the study suggest that human brucellosis is still an impor-tant public-health problem in the western Anatolia region of Turkey, especially in rural areas.Key words: Brucella; Brucellosis; Seroepidemiologic studies; TurkeyCorrespondence and reprint requests should be addressed to:Dr. Zafer CetinkayaMaresal Cakmak MahallesiAdnan Menderes Bulvari No. 8/11   TR-03000 AfyonTurkeyEmail: zfcetinkaya@hotmail.comFax: 0090 (272) 214 2065INTRODUCTIONBrucellosis is a public-health problem in many devel-oping countries, including Turkey. The disease is zoo-notic, and virtually all infections are derived directly orindirectly from animal exposure (1,2). Brucella strains cause diseases mainly in domesticanimals, such as sheep, goats, cows, and pigs, resultingin important economic losses. Human brucellosis isnotoriously a multisystem disease with varied manifes-tations, and the onset may be either acute or insidious.The diverse and sometimes deceptive manifestationsof localized, sub-acute or chronic infection may lead tomissing or delaying the diagnosis if the attending clini-cian has a low index of suspicion (1,3). Brucellosis is usually transmitted to humans bydirect contact through abraded skin or mucosal sur-faces, by consumption of contaminated milk andmilk products, or by inhalation. Agricultural wor-kers, veterinarians, and others involved in hand-ling of animals or animal carcasses are at higherrisk of direct inoculation. Individuals who ingestunpasteurized dairy products, especially from areasof endemic infection, are at significant risk of food-borne brucellosis (1,2,4,5). Brucellosis can, there-fore, be termed as an occupational disease. Person-to-person transmission of Brucella is extremelyrare (5).Although the prevalence of Brucella is notexactly known in Turkey, the seropositivity hasbeen reported to be about 2-6% (6). The sero-prevalence of brucellosis in sheep and cows inthis country has been reported to be 1.97% and1.43% respectively (7). Afyon city is located inCentral Western Anatolia. Raising of domestic ani-mals and processing of animal products is wide-spread. The majority of the people live in ruralareas and work in animal farming and agriculture.Cetinkaya Z et al.J Health Popul Nutr Jun 2005138The important characteristics of the studied region arethe production and consumption of cream from unboiledmilk and of fresh cheese from unpasteurized milk.In this study, we aimed at determining the seropre-valence of Brucella by Rose Bengal slide agglutinationand standard tube agglutination methods in a regionwhere domestic animal farming and agriculture is themain occupation.MATERIALS AND METHODSThe study was carried out in the rural areas of sevendistricts of Afyon during April-May 2003. Nearly half(453,405 of 846,420) of the population of the regionlive in rural areas. The sample size that should be takento ensure a 95% confidence interval was considered tobe 1,117 subjects, assuming the prevalence of humanbrucellosis at 3% and a 1% error rate. Seven (Bolvadin,Sandikli, Cobanlar, Sultandagi, Emirdag, Dinar, andSuhut) of 17 towns in the region of Afyon were chosenat random. Of 1,330 subjects invited to participate inthe study, 213 did not agree to participate. Blood sam-ples were collected from 1,117 subjects living in sevenrural towns (Bolvadin-143, Sandikli-221, Cobanlar-22,Sultandagi-78, Emirdag-134, Dinar-276, and Suhut-243).The subjects were chosen by a simple random-samp-ling method from the area. Sixty-five samples wereexcluded from the study because of haemolysis, and1,052 samples were finally included in the study.   The subjects were informed about the nature of thestudy, and written consents were obtained. The partici-pants were questioned as to whether they had ever eatenfresh cheese or raw cream until prior to three weeksbefore the date of questioning. Data were collectedusing a questionnaire comprising three groups of ques-tions: (a) personal data (age, gender, education), (b)consumption of cream and fresh cheese from unboiledmilk, and (c) direct contact with animals (milking, clean-ing, shearing, handling, and feeding). The same physi-cian (Z.C.) completed the questionnaire by personalinterview. The correlation between the habits of con-sumption of cream and fresh cheese from unboiled milkand brucellosis was examined.   Serum was harvested from blood collected from theperipheral venous vessels. Whole blood was centrifuged,and serum was separated. The sera were stored at -400C. All the serum samples were studied by Rose Bengalslide agglutination test (RB) (Biotech, Barcelona, Spain)and standard tube agglutination test (SAT) (WellcomeDiagnostics, London, UK). All sera were routinely dilutedfrom 1/20 to 1/1280. Each batch of the test included apositive control and a negative saline control. A defi-nite agglutination of the suspension was read as a posi-tive reaction. Agglutination was not seen in negativesamples. For positive samples, the lowest positive titrewas determined. Titration of 1/80 was accepted as expo-sure to Brucella, and 1/160 was accepted as brucellosisinfection. Therefore, the titrations of 1/80 and overwere accepted as positive (6). The positive sampleswere screened with control serums four weeks later.We treated all the positive serum samples simultane-ously with the 2-mercaptoethanol test for the detectionof antibody classes either IgM or IgG. Although IgMtitres are the first to rise after infection with brucel-losis, IgM paradoxically remains elevated longer thanIgG; thus, measures of IgG are traditionally used formonitoring the effectiveness of therapy. The control samples that had increased or stabilizedserum-specific IgG levels were referred to the localhealth institutions for follow-up and treatment. Othercases were informed about the results of laboratory in-vestigations.   Statistical analysisData were processed using the SPSS software version10.0 (SPSS Inc, Chicago, IL, USA). The mean age wasgiven as M±SD. Statistical significance was tested withthe chi-square test and Fisher's Exact test.RESULTSThe subjects (n=1,052) enrolled in the study were aged7-80 (median 37) years with a mean of 38.3±16.9 years.Five hundred seventy-five were female, and 477 weremale. The seroprevalence of human brucellosis was 4.8%.Four percent was established as false negative accord-ing to SAT by the RB test. False positivity was notdetected with the RB test. When compared with SAT,the sensitivity of RB was 96%, and its specificity wasthe same. Serum levels of specific IgG increased in twoof the positive cases after four weeks. The seropreva-lence of human brucellosis was lower in cases aged lessthan 50 years compared to other age groups (p=0.005).The seroprevalence of human brucellosis was higheramong women (6.26%) than among men (3.14%) (oddsratio [OR]: 2.1, 95% confidence interval [CI] 1.1-3.8,p=0.019). The educational status of the subjects was Seroprevalence of Brucella in rural Anatolia 139not statistically significant between the seropositive andthe seronegative group (p=0.270). Thirty percent of the subjects had consumed creamfrom unboiled milk, and 9.5% of them were seropositive,while 70% had consumed cream from well-boiled milk,and 2.9% of them were seropositive (OR: 3.6, 95% CI2.0-6.4, p<0.001). Human brucellosis was 15 times more prevalentamong subjects consuming fresh cheese made from un-pasteurized milk than among others (OR: 14.9, 95% CI6.3-35.3, p<0.001). Asian country where brucellosis is endemic and con-stitutes a serious public-health and economic problemin certain rural areas (1,2,6). Many people are exposedto the disease each year and encounter physical inva-lidity, causing loss of work force (8). Results of studies conducted in Southern Italy, Jordan,and Southern Saudi Arabia showed that the overall pre-valence of brucellosis was 3.1%, 16.7%, and 19.2% res-pectively (4,9,10). The seropositivity of Brucella wasassessed in various studies conducted in Turkey. Theseropositivity of Brucella was reported to be 0.8-11.9%in various occupational groups of different countries (6).Table. Distribution of seroprevalence of human brucellosisCharacteristics of study subjects Seroprevalence    Number (base)   Odds ratio          95 % CI           p value Age (years) groups 0.005<50 3.6 797 1.050-64 8.5 142 2.4 1.2-4.9 0.01*>65 8.8 113 2.6 1.2-5.4 0.021†Sex 0.019Male 3.1 477 1.0Female 6.3 575 2.1 1.1-3.8Education 0.270Illiterate 7.2 139 1.0Primary school                   4.2 778 0.6 0.3 -1.2 0.129‡Secondary school and graduate-level college 5.9 135 0.8 0.3-2.2 0.672¶Consumption of raw creamNo 2.9 737 1.0 <0.001Yes          9.5 315 3.6 2.0-6.4Consumption of fresh cheese           <0.001No 0.9 672 1.0Yes          11.8 380 14.9 6.3-35.3Have contact with animals 0.897No 5.0 301 1.0Yes          4.8 751 0.9 0.5-1.8* <50 with 50-64 years; † <50 with >65 years; ‡ Illiterate with primary school; ¶ Illiterate with secondaryschool and graduate-level college; CI=Confidence intervalThere was no statistically significant difference in sero-positivity in means of contact with animals (p=0.897).Contact with animals was defined as dealing with cattle,such as milking with automatic machines, handling,and feeding. Not all of the persons who came in con-tact with animals were involved in the jobs of handmilking, shearing, cleaning, and parturition.DISCUSSIONBrucellosis is highly endemic in the Mediterraneanbasin, Western Asia, the Middle East, Africa, and LatinAmerica. Turkey is both a Mediterranean and western The seropositivity of brucellosis was found to be3.2% in the elderly population in mid-Anatolia ofTurkey (6). Seropositivity among farmers was 6.2%(11). Seropositivity in rural areas and dairy farms wasreported to be 7.2% and 5.7% respectively (12,13).The seroprevalence of human brucellosis found in thecurrent study (4.8%) is similar to that found in otherstudies conducted in Turkey. We are unaware of anystudy about the prevalence of human brucellosis andanimal brucellosis in our region. We have previouslyinvestigated 100 milk samples collected from brucel-losis-suspected animals that had a history of abortion andfound the prevalence of brucellosis to be 35% with Cetinkaya Z et al.J Health Popul Nutr Jun 2005140by machinery at definite periods. In this region, moststables are fenced with barbed wire, and the domesticanimals live in a natural area which is called uncoveredstable system.Based on the findings of our study, we conclude thatthe main risk factor for brucellosis is consumption offresh, unpasteurized dairy produce. This study also con-firms that brucellosis continues to be a major public-health concern in the rural area of Western Anatolia.Increased risk has been attributed to certain dietarypreferences, particularly fresh cheese, unpasteurized andunboiled milk, and cream. Production and consumptionof cream made from boiled milk and consumption ofcheese made from pasteurized milk or mature cheese(more than three months) may be the main protectivefactors for brucellosis. REFERENCES1. Young EJ. Brucella species. In: Mandell GL, DolinR, Bennett JE, editors. Principles and practice ofinfectious diseases. 5th ed. New York: Livingstone,2000:2386-91.2. Chu MC, Weyant RS. Francisella and Brucella. In:Murray PR, Baron EJ, Pfaller MA, Jorgensen JH,Yolken RH, editors. Manual of clinical microbiol-ogy. 8th ed. Washington, DC: American Societyof Microbiology, 2003:797-804.3. Al-Eissa YA. Brucellosis in Saudi Arabia: past,present and future. Ann Saudi Med 1999;19:403-5.4. Abu Shaqra QM. Epidemiological aspects of bru-cellosis in Jordan. Eur J Epidemiol 2000;16:581-4.5. Fosgate GT, Carpenter TE, Chomel BB, Case JT,DeBess EE, Reilly KF. Time-space clustering ofhuman brucellosis, California, 1973-1992. EmergInfect Dis 2002;8:877.6. Sumer H, Sumer Z, Alim A, Nur N, Ozdemir L.Seroprevalence of Brucella in an elderly popula-tion in mid-Anatolia, Turkey. J Health Popul Nutr2003;21:158-61.7. Iyisan AS, Akmaz O, Duzgun SG, Ersoy Y,Eskiizmirliler S, Guler L et al. Sero-epidemiologyof brucellosis on cattle and sheep in Turkey. JPendik Vet Microbiol 2000;31:21-75.8. Bikas C, Jelastopulu E, Leotsinidis M, KondakisX. Epidemiology of human brucellosis in a ruralarea of north-western Pelopunnes. Eur J Epidemiol2003;18:267-74.Rink test and 25% with the culture method (unpub-lished data). Kaleli et al. reported the seropositivity ofbrucellosis as 8.9% in the 17-49-year age group and as15.5% in the 50-69-year age group (12). Al Sekait repor-ted the seropositivity of brucellosis to be 9.7%, 16.0%,18.7%, and 19.3% in the age groups of 0-14 years, 15-29 years, 30-44 years, and >45 years respectively (14).In our study, seropositivity was higher in the older agegroup in agreement with other studies.In the seroepidemiologic study carried out in a ruralarea of northwestern Greece, it was also reported thatthe incidence of brucellosis was higher in males (60.7%)than in females (39.3%) (8). Seropositivity in males andfemales has been reported to be similar in some stud-ies (4,6,14). In our study, the prevalence of brucellosiswas higher in females (6.3%) than in males (3.1%).In this study, the higher rate of seroprevalence ofbrucellosis in females compared to males was probablydue to an increased involvement of women in farmingdomestic animals and handling their products. Malesare generally employed in agricultural jobs. Contrary toour findings, seropositivity was higher (males 60.7%,females 39.3%) in males in a study in Greece (8). Theauthors claimed that women who are occupied withhousehold chores and raising of children spend lesstime with animals. There was no correlation between the educationallevel and the seropositivity of brucellosis in our study.We think that the habit of consuming fresh cheese andraw cream is an acquired taste in all social-cultural-educational groups. This may be responsible for thisfinding. In our study, the seroprevalence of brucellosis washigher in persons consuming fresh cheese and creamproduced from unpasteurized milk (9.5% and 11.8% res-pectively). Consumption of fresh cheese and milk creamproduced from unpasteurized milk has been reportedto be a significant risk factor for brucellosis (4-6,8,14).Contact with animals has been reported as one of theimportant risk factors for brucellosis (4,6,8,14). How-ever, contact with animals was defined as milking, clean-ing, shearing, and animal parturition. On the other hand,in our study, definition of contact with animals differedsomewhat from the previous studies. Contact with ani-mals was not so intense in our population. For example,milking was done by automatic machines, and shearingand animal parturition are performed by professionally,experienced persons. In addition, stable cleaning is doneSeroprevalence of Brucella in rural Anatolia 1419. Torre I, Ribera G, Pavia M, Angelillo IF. A sero-epidemiologic survey on brucellosis antibodies insouthern Italy. Infection 1997;25:150-3.10. Alballa SR. Epidemiology of human brucellosisin southern Saudi Arabia. J Trop Med Hyg 1995;98:185-9.11. Sonmez E, Durmaz B, Aladag M. Brucellosis pre-valence of Malatya (Turkey) province. TürkiyeMed J 1997;4:102-5.12. Kaleli I, Kocoglu T, Ozen N, Aksit F. The preva-lence of brucellosis in the Denizli  region. TurkishJ Infect 1999;13:231-3.13. Ozbakkaloglu B, Tunger O, Dinc G, Borand H,Orhon H, Degerli K et al. Brucellosis prevalenceof risk groups in Manisa (Turkey). Turkish J Infect1998;12:453-7.14. Al Sekait MA. Seroepidemiological survey ofbrucellosis antibodies in Saudi Arabia. Ann SaudiMed 1999;19:219-22.

Seroprevalence of Human Brucellosis in a Rural Area of Western
Anatolia, Turkey

Seroprevalence of Human Brucellosis in a Rural Area of Western Anatolia, Turkey

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