Abstract
Brucellosis is a worldwide zoonotic infection of economic and public health importance
caused by bacteria belonged to the genus Brucella. These are non-motile, facultative,
anaerobic, intracellular, Gram-negative coccobacilli. In cattle brucellosis is caused by B.
abortus which results in abortion, infertility, still birth and reduced milk production.
Isolation, identification and molecular characterization of Brucella spp. circulating in
animals are essential to undertake appropriate control and preventive measures in a
particular geographical area. This study was conducted for isolation, identification and
molecular characterization of Brucella spp. in dairy cattle. Twenty two dairy farms housed
1285 dairy cattle located at Mymensingh, Savar, Gazipur, Jamalpur and Dinajpur were
selected. Sera (n=496), uterine discharge (n=45), milk (n=115), vaginal swab (n=71), placenta (n=7) and aborted fetus (n=2) were collected from dairy cattle with the history of abortion,
retained placenta, infertility and still birth. Sera of cattle were tested for the presence of
Brucella abortus specific antibody response by the Rose Bengal Plate Test (RBPT). Samples
were streaked duplicate onto Brucella selective agar and inocubated at 37-C for 7-14 days in
an incubator supplied with 5% CO2 for isolation of Brucella spp. Brucella spp. recovered from
the samples were identified at species and biovar levels by routine bacteriological and
classical biotyping (CO2 requirement for growth, H2S production and growth in presence
of thionine and basic fuchsin) methods, Molecular confirmation of Brucella both at genus
and species level were also done by genus and species specific PCR assays. Molecular
characterization of Brucella spp. was performed by multi-locus variable- number tandemrepeat assay-16 (MLVA-16). Thirty nine sera (7.86%) were tested positive by RBPT. Ten
Brucella isolates were recovered from uterine discharge (n=7), milk (n=2) and vaginal swabs
(n=1), which were confirmed as B. abortus by enhanced AMOS-ERY PCR assay. The classical
biotyping method confirmed all 10 B. abortus isolates belonged to the biovar 3. The MLVA-16
assay indicates that all B. abortus isolates were identical and shared the same genotype 40,
based on panel 1 MLVA-8. Dendrogram and minimum spanning tree (MST) analysis
when compared with 1633 global isolates of B. abortus revealed that B. abortus isolates of
this study were genetically similar with three isolates of Brazil and one isolate of France.
The MST analysis also revealed that 10 Bangladeshi B. abortus isolates belonged to clade
C1 and sub-group 3b of biovar 3. In Bangladesh, this is the first report of isolation,
identification, biotyping and genetic characterization of B. abortus from dairy cattle.