Control of
brucellosis in livestock depends on the reliability and sensitivity of the
methods used for detection and identification of bacteria which would create a
consequential control of the disease in wildlife and also in humans (Poester et
al., 2010). However, putting into consideration that the incubation period of
brucellosis varies considerably and is affected by several factors which
include; gestation, exposure dose, age, vaccination, and other unknown
host-resistance influence (Thomsen A., 1950) diagnosis would be cumbersome.
Most protection against infection by B.abortus
is largely a result of cell-mediated immunity (Cheers et al., 1983) but most
diagnostic tests developed rely on detecting a response in humoral immune
system (Stemshorn., 1983). Th1 cells in the spleen plays an important role in
conferring protective immunity to B.melitensis
infection (Vitry et al.,2014).

In this present
study, Brucella abortus specific histopathological changes were observed. The
recruitment of lymphocytes in the liver was evident forming micro-granulomas
which corresponded to a study done by Mariana et al., 2013; who showed in
IL-1Oflox/CD4cre mice exhibiting an inflammatory response in the liver as well
as the spleen. In addition, there was distinct evidence in architectural
changes of the spleen histology. However, Mariana et al marked an influx of
neutrophils and histiocytes in the spleen at an acute stage of infection, later
depicting that in chronic infections particularly in B. abortus infection, and
also granuloma formation comprised of mononuclear cells not depicted in this study. The Brucella organism’s predilection for
organs rich in reticuloendothelial cells (spleen, liver, bone marrow, lymph
nodes) and its intracellular location are responsible for the chronicity of the
disease, which can last for months or even years (Bothwell, 1963; Spink, 1964;
Robbins, 1968). The spleen and liver are organs that contain many bacterial
cells after Brucella invasion. (Hort
et al., 2003).

Although bacteriological diagnosis
being a gold standard diagnostic method for brucellosis, culture of the species
is challenging as it is a fastidious bacterium requiring rich media for primary
cultures (Hadush., 2013; Bricker., 2002). In the present study, results did
show that there was a high prevalence of Brucella
abortus strain 544 isolated in the vector control group than in the
immunized groups with a slight effectivity of the ST system given at a higher
dosage 5 X 1010 CFU/ml per animal over the animals immunized
using ST system 5 X 109 CFU/ml per animal. In addition, results suggest that the
bacterium may be readily isolated in lymph tissue rather than in the
reticuloendothelial tissue samples.

IHC allows in situ localization of the organism
within Brucella induced lesion (Poester
et al.,2006). Immunostaining does not require viable bacteria and thus a better
option to bacterial culture in those regards. In this study, it was noticed
that immunohistochemistry could not detect bacterial antigens exactly
corresponding to bacterial culture results also suggesting that the sectioning
of the tissue reduces the chances of locating the positive cells.

The present
study also explored the use of real-time
PCR approach using the SYBR Green I (double-stranded
DNA intercalating dye (Newby, 2003), the advantages being that it was faster as
they was no electrophoretic analysis and does not require post amplication handling of PCR products. However,
it was thus noticed that almost all DNA templates examined showed a fluorescent
signal. Even though SYBR GREEN I assays may not require probe design, the lack
of additional discrimination that can be gained by incorporating of one or more
probes may result in false positives. To overcome
such errors a melt analysis/ or gel analysis can be done but then what would be
the need of using it as a diagnostic tool (Newby et al). Real-time PCR can be used as a confirmatory
diagnostic alternative for problematic culturing of Brucella sp. The high detection sensitivity of real-time PCR can be
explained by the fact that it detects DNA from bacteria that are damaged and
non-inviable and therefore impossible to isolate by conventional cultures. In
the present study IS711 real-time PCR was
able to detect positive animals which were negative by bacterial isolation and
detect additional animals that were immunohistochemically negative. Therefore,
the importance of using more than one type of diagnosis for detection of
brucellosis in animals (Elfaki et al., 2005: Ilhan et al., 2008)