salmonellosis is an economically important bacterial disease ailment inflicting
serious difficulty to the growth of poultry industry (Rajagopal & Mini,
2013). It is caused by Salmonella, a member of the family Enterobacteriaceae,
a Gram- negative facultative intracellular pathogen that is able to cause
different disease syndromes in a broad range of hosts. It constitutes two
species Salmonellabongori (S. bongori) and Salmonellaenterica (S. enterica) (Reeves, et al., 1989). Although there are greater
than 2,600 Salmonellaserovars (Rainier, et al.,, 2013), relatively few
serovar cause infection in most animal ( Saeki, et al., 2013; Zhu et al.,
2015). Hence, Salmonellaenterica
subsp. enterica serovar Enteritidis (S.
Enteritidis), serovar Gallinarum
biovars Gallinarum and pullorum and S. Typhimurium are generally isolated from poultry. Among these
serovar Gallinarum (S. Gallinarum) consists of two biovars,
Gallinarum and Pullorum that cause fowl typhoid and pullorum disease in adult
and young , respectively (Hossain, et al., 2006). Salmonellosis is one of the
major challenge of poultry farming that hinder its growth and development.
Therefore, it is necessary to screen virulence gene of Salmonellaat molecular
level for development of rapid and fast disease diagnosing technique.
Poultry industry is
uplifting as profitable sector in Nepal. Since, 1974 the remarkable improvement of
commercial poultry in Nepal. And had results in tremendous development of this
sector over the recent period of time. Since, 1974 the remarkable improvement of
commercial poultry in Nepal had been started (FAO, 2012). In our
country, 50 – 55 % of poultry birds are
commercially managed. And, poultry industry contributes about 3.5% in GDP.
Aditionally, provides an employment
opportunity creating an income along with improving the nutritional level of
the country. This also provides fulltime employment to about nine thousand and
partial employment to about ninety nine thousand people (CBS, 2015).
culture methods are used to identify Salmonellaand require at least 3-11
days. The standard protocol for isolating Salmonellaspecies include
non-selective pre-enrichment followed by selective enrichment and plating on
selective and differential media. These procedure are time consuming and labour
intensive (Menghistu, Rathore, Dhama, & Agarwal, 2011). In recent
years, particularly in developed countries, several methods such as Enzyme
Linked Immunosorbent Assay (ELISA), latex agglutination ,immunodiffusion, Polymerase
chain reaction (PCR), and real time PCR (RT PCR) had been introduced. In comparision to other method, polymerase chain reaction (PCR)
technology and real time PCR (RT PCR) have allowed the specific
amplification of particular target portion of DNA, which can be used for the diagnosis
of pathogen of veterinary importance. PCR tests have
demonstrated their utility as screening tools for Salmonellabroiler and
layer samples to reduce workloads and shorten time for Salmonellaevaluation
(Bautista et al., 2011).
S. enterica consist
of several virulence genes which encode products that help the organisms to
express its virulence in the host. Among the virulence genes, 16srna encode
for the confirmation of Salmonellaat genus level whereas invA for adhesion
and invasion of the pathogen in the host system, spv for systemic
disease state in the host cells, speffor ornithine decarboxylase of Salmonellagallinarum,
sdi and hilA encode for protein belonging to the transcriptional
regulators, fim H for fimbrin like protein, avrA to
modulate host cellular functions, agf
for diagnosis of Salmonellaarrayed on hydrophic grid membrane
filters, sivh gene for outer membrane protein and Stn for
enterotoxin genes of host pathogenic processes . While spvA, spvB, and Spvc virulence
gene codes for plasmid of pathogenic organism.
Thus, precise and systematic method should be adopted for screening
virulence genes from S. enterica isolates originated from the infected
samples (Murugkar et al., 2003).
Similarly, Polymerase chain reaction (PCR) had been discovered as a
high-throughput approach with a high degree of sensitivity and specificity for pathogen detection.
this research was undertaken in the clinical cases of broilers to screen the
virulence gene responsible for salmonellosis along with the antibiotic
resistance pattern of these sample. Salmonellaisolates will be screened
based on virulence gene profiling, focusing on virulence determinants
associated with SPIs, plasmids, toxins, fimbriae, and flagella that were
positive for the infection by Salmonellaas an intracellular pathogen