Conventional and molecular Characterization of mycobacterial Strains Isolated from Cattle and Humans
Conventional and molecular Characterization of mycobacterial Strains Isolated from Cattle and Humans
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Date
2015-04-01
Authors
Hassan, Manal
Journal Title
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Publisher
university Of Khartoum
Abstract
In the present study, a total of 167 caseated, enlarged and
inflammed lymph nodes and tuberculous organs were collected
from cattle slaughtered at Slaughterhouses in Khartoum State.
Also twenty lyophilized mycobacterial strains previously isolated
from both human and animal cases were examined in this study
using PCR. The aim of this study was to estimate the prevalence
of bovine tuberculosis and to improve identification and to compare
between identification methods.
The purulent materials from the suspected specimens were
decontaminated with 2% NaOH and concentrated by
centrifugation. The smears made from decontaminated purulent
materials were stained by Ziehl-Neelsen method and culture onto
Lowenstein-Jensen medium was done. Results from culture and
microscopy were compared to a PCR method by amplifying an
insertion element IS6110 which is specific for Mycobacterium
tuberculosis complex.
Out of the 167 specimens examined, 35 (20.96%) were
found to have acid-fast bacteria when smears were examined
microscopically. Out of the 35 acid-fast bacteria, 22 (62.86%)
showed branching filaments and were identified tentatively as
Mycobacterium farcinogenes. The remaining 13 (37.14%) were
bacilli and were identified tentatively as Mycobacterium bovis. Of
the 167 specimens, 12 (7.19%) showed visible growth on
Lowenstein-Jensen medium when incubated aerobically at 37°C
between 4 and 8 weeks. None of the 22 specimens with branching
filaments grew using the above decontamination and growth
conditions. Ten isolates were nitrate-negative and urease-positive,
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and were thus identified as M. bovis, whereas the remaining tow
strains were nitrate-positive and urease-negative but one was
urease-positive.
Polymerase chain reaction (PCR) assay for the rapid
diagnosis of M. tuberculosis complex (M. tuberculosis, M. bovis, M.
microti and M. africanum) was done using oligonucleotide primers
TB41 and TB42 to amplify fragments of the IS6110, an insertion
sequence specific for M. tuberculosis complex. Of the 12 grown
culture, ten (83.3%) gave PCR product for IS6110 on agarose gel
electrophoresis, which confirmed their identity as M. tuberculosis
complex.
Ten isolates were already found nitrate negative and thus
were identified as M. bovis, none of the isolates were found to be
M. tuberculosis when using this test combination.
In conclusion, IS6110 amplification, urease and nitrate tests
provide a species identity to M. bovis. The results of the above
study is as follows: of the 167 caseated and purulent specimens
examined 10 (5.99%) were M. bovis, 2 (1.2%) were
mycobacterium other than tuberculosis (MOTT), 22 (13.17%) were
tentatively identified as M. farcinogenes, 23 (13.77%) were acidfast
bacteria (AFB) that failed to grow on L.J medium and 132
(79.04%) were caseous infections caused by non-acid-fast
bacteria.
Description
99 Pages
Keywords
Conventional, molecular,mycobacterial,Strains Isolated,Cattle,Humans;Bovine tuberculosis;Wildlife reservoirs;Pathogenesis;Pathogenesis;