Despite the occurrence of S. Typhimurium infections, little is known on the genetic
diversity, virulence characteristics and antimicrobial resistance profiles of poultry S. Typhimurium
in South Africa. Therefore, S. Typhimurium (n=141) isolated from organs (n=115) and
environments (n=26) of diseased poultry between 1995 and 2002 were screened by PCR for
bacteriophages, plasmids and Salmonella pathogenicity islands (SPIs) - encoded virulence
genes (virulotyping) which are essential for invasion (invA, sopB, gtgB, sspH1, sopE, spvC, and
pefA), survival (sifA, gipA, sodC1, gtgE, mig5, and sspH2) and serum killing (rck, and srgA) of the
pathogen in the host. Isolates were also characterized by: pulsed field gel electrophoresis (PFGE)
for genetic relatedness, and plasmid profiling (n=43). Furthermore, isolates (n=141) were tested
for susceptibility to 16 antimicrobials by disk diffusion and further screened by PCR for the
carriage of 27 resistance genes, and integrons. Multi-resistant S. Typhimurium definitive phage
type (DT) 104 were determined by disk diffusion and confirmed by PCR.
All isolates carried SPIs-encoded genes: invA, sopB, and sifA. Bacteriophages-encoded
genes (sspH2, sspH1, sodC1, gtgB, and gtgE) occurred in more than 74.5% of the isolates expect
for gipA (57.6%), and sopE (19%). The occurrence of plasmid-encoded genes (pefA, mig5, rck, spvC, and srgA) ranged from 48.2% to 74.5%. Two sample t - test showed that virulence genes:
gtgB, spvC, gipA, gtgE, mig5, rck and srgA were more frequent (p ? 0.05) in S. Typhimurium
isolates from environments. Virulotyping clustered 141 isolates into 59 virulotypes with 97 isolates
clustering in 5 predominant virulotypes while 44 were single isolate virulotypes. PFGE grouped
140 isolates into 55 pulsotypes with 66 isolates clustering in 5 major pulsotypes, 51 isolates
clustering in small pulsotypes (containing less than 5 isolates) while 33 were single isolate
pulsotypes. Ten plasmid profiles ranging from 2kb to 90kb were observed. The most common
plasmid profile contained the 90kb plasmid and was observed in 12/43 isolates. Major virulotypes
and plasmid profiles corresponded approximately to pulsotypes and clustered isolates recovered
from the same farms or during the same period. Virulotyping and PFGE showed identical
discriminatory index (D=0.93). Multidrug resistance (resistance to ? 2 antimicrobials) was
observed in 97.2% of isolates. High levels of resistance phenotypes and their respective
resistance genes were observed for: streptomycin (94.3%) conferred by ant3'Ia (60.3%) and str
(50.4%), sulphonamides (87.2%) conferred by sul1 (66%) and sul3 (31.9%), ciprofloxacin (79.4%)
conferred by qnrA (79.4%), tetracycline (61%) conferred by tetB (35.5%) and tetG (28.4%), and
cefotaxime (55.3%) conferred by blaSHV (57.4%). Two sample t - test revealed that isolates from
poultry organs were more resistant (p?0.05) to ampicillin, amoxicillin clavulanic acid,
chloramphenicol, tetracycline and sulfamethoxazole - trimethoprim while isolates collected from
poultry environments were more resistant to cephalothin, cefotaxime, ceftazidime, colistin
sulphate and nalidixic acid. Using the Kappa statistics, there were agreements ranging from good
to perfect between phenotype and genotype. In addition, for every phenotypic resistance
recorded, at least one corresponding resistance gene was detected. DT104 strains and class1
integrons were observed in 34.7% and 83% of the isolates respectively. Multi-resistant S.
Typhimurium (97.2%) also carried SPIs - encoded virulence genes involved in invasion and
survival in the host. In addition, more than 50% of resistant isolates to each of the antimicrobials
also carried at least 12 virulence genes: invA, sopB, sifA, sspH2, sspH1, sodC1, gtgB, gtgE, pefA, mig5, spvC, and srgA. A significant number (44.9%) of the DT104 strains that were clustered in
the same pulsotype X25 also belonged to virulotype V3a which contained 13 virulence genes:
invA, sopB, sifA, sspH2, sspH1, sodC1, gtgB, gtgE, pefA, rck, mig5, spvC, and srgA. Most of
isolates that belonged to the same antimicrobial resistance profile (phenotype and genotype)
carried at least 8 common virulence genes.
In conclusion, these data indicate that S. Typhimurium isolated from diseased poultry carry
virulence genes that are usually incriminated in Salmonella human outbreaks. Virulotyping and
PFGE showed the same discriminatory index (D=0.93) indicating that virulotyping can be an
alternative subtyping method in laboratories where PFGE is not available. Salmonella
Typhimurium are also genetically diverse since they were recovered from multiple farms and
during a period spanning 8 years. Furthermore, isolates were resistant to multiple antimicrobials
used in poultry operations (streptomycin, sulphonamides, and tetracycline) and those used to
treat human salmonellosis: ciprofloxacin, and cefotaxime. Multidrug resistant isolates carried
most of virulence genes. This relationship between virulence and antimicrobial resistance
suggests that the adaptation of isolates against antimicrobial effects may induce expression of
virulence factors. The increasing incidence of DT104 threatens the public health since DT104
strains are associated with hospitalizations and deaths in humans. Salmonella Typhimurium
carried mobile genetic elements (bacteriophages, integrons and plasmids) which pose a public
hazard as they propagate virulence and resistance genes with emerging new pathogenic bacteria
as a result. Therefore, monitoring and surveillance of salmonellosis and prudent antimicrobials
use need more efforts to ensure animal health and food safety for consumers in South Africa. / Dissertation (MSc)--University of Pretoria, 2016. / Paraclinical Sciences / MSc / Unrestricted
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:up/oai:repository.up.ac.za:2263/60275 |
| Date | January 2016 |
| Creators | Ntivuguruzwa, Jean Bosco |
| Contributors | Van Heerden, Henriette, boscus2@gmail.com |
| Publisher | University of Pretoria |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Rights | © 2017 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. |
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