PHENOTYPIC AND BIOLOGICAL CHARACTERIZATION OF BRUCELLA STRAINS ISOLATED FROM NIGERIAN LIVESTOCK
PHENOTYPIC AND BIOLOGICAL CHARACTERIZATION OF BRUCELLA STRAINS ISOLATED FROM NIGERIAN LIVESTOCK
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Date
2005-05
Authors
REUBEN, ADAMA OCHOLI
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Abstract
In this study, bacteriological examination for the isolation of brucellae was
carried out in order to obtain the species and biovars. A total of 1,020 clinical samples
consisting of 283 milk, 10 hygroma, 581 vaginal swabs, 20 aborted fetuses, and 126
blood samples obtained from cattle, sheep, goats, pigs, dogs and horses drawn from
Plateau, Kaduna, Kano, Borno, Kogi, Nassarawa, Adamawa, Taraba, Bauchi and Enugu
States of Nigeria were investigated. The culturing of the samples, and identification and
biotyping of the isolates were done according to standard bacteriological techniques.
A total of 32 stored cultures of Brucella isolated from Nigerian cattle in previous
works between 1975 and 1982 whose identity were unknown were reactivated and rebiotyped.
To determine if the field strains of Brucella isolated in the present work were
virulent or attenuated in animals, the immune and pathological responses of BALB/c
mice inoculated with B. abortus strains MK5, SH3 and HS2 were compared with that of
B. abortus S19 (vaccine strain) and B. abortus strain w.544 (WHO pathogenic reference
strain).
Twenty five strains of Brucella were isolated, identified and biotyped. The
isolates were designated according to the laboratory identification number of the samples
from which the isolates were obtained. All the isolates belonged to one species and
biovar, Brucella abortus biovar 1. Most of the isolates did not require carbon dioxide
(CO2) for growth even on initial isolation. The isolates had identical reactions in the
various tests. All the isolates produced colonies with smooth shiny surface. Of the tests
used to differentiate species and biovars, all the isolates showed the following common
characteristics: produced hydrogen sulphide (H2S), grew in the presence of basic fuchsin
viii
(20ug ml-1), but not in the presence of thionin (20ug ml-1). In slide agglutination test, all
the isolates were agglutinated by the monospecific antisera A, and all the isolates were
lysed by Wb, Tb and Bk2 phages.
Successful isolation of Brucella was made only from clinical samples obtained
from cattle, sheep and horses. Out of the 25 isolates, 17 (68%) were from cattle, 5 (20%)
from sheep and 3 (12%) from horses. Of the isolation from clinical samples, 12 (48%)
were from milk samples, 6 (24%) from hygroma fluids, 5 (20%) from vaginal swabs and
2 (8%) from aborted fetuses. Isolations were made from samples from Plateau, Taraba,
Adamawa, Bauchi, Nassarawa and Sokoto States.
All the stored isolates belonged to one species, Brucella abortus. Out of the 32
strains, 31(96.9%) were of biovar 1 while only 1 (3.1%) was of biovar 2.
Following intraperitoneal injection of mice, B. abortus S19, and w.544, MK5, SH3 and
HS2 could be detected in the blood stream of infected mice as from 4 days post
inoculation (p.i.) to 21st, 24th, 70th, 42nd and 42nd days p.i. for the different strains
respectively. The splenic colony forming units (CFU) of B. abortus peaked at 7days p. i.
for S19, 14 days for w.544 and HS2 and 21days for MK5 and SH3. The B. abortus S19
was cleared from the spleen as at 12 weeks p.i. while the w.544, MK5, SH3 and HS2
persisted in the spleen for at least 20 weeks p.i. The splenic/body weight ratio peaked at
7 days p.i. for S19 and SH3, 14 days for w.544 and HS2, and 21 days for MK5. Sera
obtained from mice infected with all strains of B. abortus contained detectable antibodies
to Brucella as from 3 weeks p.i. Mice infected with the field strains of B. abortus
induced significant production of antibodies comparable to S19 and w.544-infected
mice. All the strains of B. abortus induced production and persistence of antibodies for at
least 20 weeks p.i. Small germinal centers and lymphoid depletion occurred in spleens of
ix
mice during the first week of infection with all strains of B. abortus. Large germinal
centers and disappearance of the macrophage accumulations in the spleen did not appear
until 10 weeks after infection of mice with S19 and until 20 weeks after infection of mice
with w.544, MK5, SH3 and HS2.
The epidemiological significance of these findings is discussed. Some
observations on the zoonotic and public health implications of Brucella infections in
Nigerian livestock are presented. A control programme involving improved
management, animal movement restrictions, public health education and mass
vaccination of animals is suggested.
These results indicate that the field strains of B. abortus are virulent and cannot
be used for testing as vaccine candidates.
More strains would have been obtained if molecular techniques for detection and
typing were used.
Description
A Dissertation Submitted to the
Ahmadu Bello University, Zaria
in Partial Fulfillment for the Degree of
Doctor of Philosophy (Ph. D)
Department of Veterinary Public Health and Preventive Medicine,
Faculty of Veterinary Medicine,
Ahmadu Bello University,
Zaria, Nigeria.
May, 2005
Keywords
PHENOTYPIC,, BIOLOGICAL CHARACTERIZATION,, BRUCELLA,, STRAINS ISOLATED,, NIGERIAN LIVESTOCK.