Molecular typing of Brucella species isolates from Human and livestock bloods in Isfahan province

Authors

1 Department of Microbiology and Virology, Isfahan University of Medical Sciences, Isfahan, Iran

2 Department of Genetics and Molecular Biology, Medical School, Isfahan University of Medical Sciences, Isfahan, Iran

3 Center of Veterinary and Clinic Network, Isfahan University of Medical Sciences, Isfahan, Iran

4 Department of Public Health, Isfahan University of Medical Sciences, Isfahan, Iran

Abstract

Background: Human brucellosis is caused by infection with certain species of the genus Brucella and is characterized by bacterial persistence and inflammation of many host tissues. Handling all live Brucella involves risk of laboratory infection and very strict biosafety rules must be observed. In order to avoid these disadvantages, method based on the PCR-RFLP shows excellent typeability, reproducibility, stability, and epidemiological concordance. The omp2 locus contains two gene copies (named omp2a and omp2b) coding for porin proteins and has been found particularly useful for molecular typing and identification of Brucella at the species, biovar, or strain level. This study is designed to evaluate the molecular epidemiology of Brucella spp from human and livestock in Isfahan province, central region of Iran in order to use the findings in efficient disease prevention programs.
Materials and Methods: One hundred ninety blood samples were collected from human and cattle with active brucellosis and 40 aborted ewes fetuses were collected and genotyped using PCR-RFLP technique, DNA polymorphisms such as the restriction patterns of the PCR-amplified omp2a and omp2b genes.
Results: The molecular characterization performed to assess the species and the biovar of the Brucella strains. Analysis of the 230 isolates examined in this study generated three unique RFLP profiles. One of the profiles was the most common being present in 134/180.
Conclusion: Our findings confirm abundance of B. melitensis, particularly biovar 1 in human and sheep are identical but B. abortus
 biovar 3 as the etiological agent of cattle brucellosis most frequently isolated in the Isfahan area.

Keywords

1.
Pappas G, Papadimitriou P, Akritidis N, Christou L, Tsianos EV. The new global map of human brucellosis. Lancet Infect Dis 2006;6:91-9.  Back to cited text no. 1
    
2.
Godfroid J, Scholz HC, Barbier T, Nicolas C, Wattiau P, Fretin D, et al. Brucellosis at the animal/ecosystem/human interface at the beginning of the 21 st century. Prev Vet Med 2011;102:118-31.  Back to cited text no. 2
    
3.
Vizcaino N, Kittelberger R, Cloeckaert A, Marin CM, Fernandez-Lago L. Minor nucleotide substitutions in the omp31 gene of Brucella ovis result in antigenic differences in the major outer membrane protein that it encodes compared to those of the other Brucella species. Infect Immun 2001;69:7020-8.  Back to cited text no. 3
    
4.
Cloeckaert A, Vizcaino N, Paquet JY, Bowden RA, Elzer PH. Major outer membrane proteins of Brucella spp.: Past, present and future. Vet Microbiol 2002;90:229-47.  Back to cited text no. 4
    
5.
Carvalho Neta AV, Mol JP, Xavier MN, Paixão TA, Lage AP, Santos RL. Pathogenesis of bovine brucellosis. Vet J 2010;184:146-55.  Back to cited text no. 5
    
6.
Salehi M, Pishva E, Salehi R, Rahmani R. Isolation of Brucella abortus using PCR-RFLP analysis. Iran J Pub Health 2006;35:22-27.  Back to cited text no. 6
    
7.
De Massis F, Di Girolamo A, Petrini A, Pizzigallo E, Giovannini A. Correlation between animal and human brucellosis in Italy during the period 1997-2002. Clin Microbiol Infect 2005;11:632-6.  Back to cited text no. 7
    
8.
Corbel MJ. Identification of dye-sensitive strains of Brucellamelitensis. J Clin Microbiol 1991;29:1066-8.  Back to cited text no. 8
    
9.
Al Dahouk S, Scholz HC, Tomaso H, Bahn P, Göllner C, Karges W, et al. Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system. BMC Microbiol 2010;10:269.  Back to cited text no. 9
    
10.
Zerva L, Bourantas K, Mitka S, Kansouzidou A, Legakis NJ. Serum is the preferred clinical specimen for diagnosis of human brucellosis by PCR. J Clin Microbiol 2001;39:1661-4.  Back to cited text no. 10
    
11.
Lucero NE, Ayala SM, Escobar GI, Grayon M, Jacques I. A new variant of Brucellamelitensis. Clin Microbiol Infect 2006;(12):593-6.  Back to cited text no. 11
    
12.
Cloeckaert A, Vizcaino N, Paquet JY, Bowden RA, Elzer PH. Major outer membrane proteins of Brucella spp.: Past, present and future. Vet Microbiol 2002;90:229-47.  Back to cited text no. 12
    
13.
Cloeckaert A, Verger JM, Grayon M, Paquet JY, Garin-Bastuji B, Foster G, et al. Classification of Brucella spp. isolated from marine mammals by DNA polymorphism at the omp2 locus. Microbes Infect 2001;3:729-38.  Back to cited text no. 13
    
14.
Al Dahouk S, Tomaso H, Prenger-Berninghoff E, Splettstoesser WD, Scholz HC, Neubauer H. Identification of Brucella species and biotypes using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Crit Rev Microbiol 2005;31:191-6.  Back to cited text no. 14
    
15.
Nagalingam M, Shome R, Balamurugan V, Shome BR, NarayanaRao K, Vivekananda, et al. Molecular typing of Brucella species isolates from livestock and human. Trop Anim Health Prod 2012;44:5-9.  Back to cited text no. 15
    
16.
Leal-Klevezas DS, Martinez-Vazquez IO, Lopez-Merino A, Martinez- Soriano JP. Single-step PCR for detection of Brucella spp. From blood and milk of infected animals.J Clin Microbiol 1995;33:3087-95.  Back to cited text no. 16
    
17.
Marianelli C, Ciuchini F, Tarantino M, Pasquali P, Adone R. Molecular characterization of the rpoB gene in Brucella species: New potential molecular markers for genotyping. Microbes Infect 2006;8:860-5.  Back to cited text no. 17
    
18.
Cloeckaert A, Grayon M, Grepinet O. Identification of Brucellamelitensis vaccine strain Rev.1 by PCR-RFLP based on a mutation in the rpsL gene. Vaccine 2002;20:2546-50.  Back to cited text no. 18
    
19.
Baddour MM, Alkhalifa DH. Evaluation of three polymerase chain reaction techniques for detection of Brucella DNA in peripheral human blood. Can J Microbiol 2008;54:352-7.  Back to cited text no. 19
    
20.
Pishva E, Salehi M, Gharavi M. Relationship between Brucella immunocomplex and glomerulopathies. J Clin Infect Dis 2008;5:127-32.  Back to cited text no. 20
    
21.
Whatmore AM, Murphy TJ, Shankster S, Young E, Cutler SJ, MacMillan AP. Use of amplified fragment length polymorphism to identify and type Brucellaisolates of medical and veterinary interest. J Clin Microbiol 2005;43:761-9.  Back to cited text no. 21
    
22.
Ancora M, De Santis P, Di Giannatale E, Luciani M, Alessiani A. Molecular typing of Brucella field strains isolated in Italy. Vet Ital 2005;41:51-5.  Back to cited text no. 22
    
23.
Kutlu G, Ertem GT, Coskun O, Ergun U, Gomceli YB, Tulek N, et alBrucella: A cause of peripheral neuropathy. Eur Neurol 2009;61:33-8.  Back to cited text no. 23
    
24.
Cloeckaert A, Verger JM, Grayon M, Grepinet O. Restriction site polymorphism of the genes encoding the major 25 kDa and 36 kDa outer membrane proteins of Brucella. Microbiology 1995;141:2111-21.  Back to cited text no. 24