Publikationsdatenbank

Molecular epidemiology, genotyping and antimicrobial susceptibility studies on brucella spp. isolated from livestock (2020)

Art

Hochschulschrift

Autor

Khan, Aman Ullah (WE 10)

Quelle

Berlin: Mensch und Buch Verlag Berlin, 2020 — v, 120 Seiten

ISBN: 978-3-96729-072-1

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/28837

Kontakt

Institut für Tier- und Umwelthygiene

Robert-von-Ostertag-Str. 7-13
14169 Berlin
+49 30 838 51845
tierhygiene@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Brucellosis is a notifiable zoonotic disease of public health importance affecting animals and humans globally. Although the disease is controlled and eradicated from the developed world, it is still an endemic and often neglected zoonosis in developing countries including Egypt. The disease is emerging and remains endemic at high prevalences in ruminants particularly at established intensive breeding farms. The disease is also continuously introduced by the import of animals especially camels from neighboring brucellosis endemic countries like Sudan. Serological surveillance programs in the country have given indirect proof of brucellosis in cattle, buffaloes, sheep, goats, camels, equines, dogs, cats, and pigs. Published scientific literature has highlighted the dominance of B. melitensis bv3 and B. abortus bv1 infecting animals and humans in the country. The disease can be transmitted to other animals via direct or indirect contact. Clinically healthy animals may be a carrier of infection and a source of disease spread. Humans get infected via the consumption of unpasteurized milk and milk products and contact with infected animals. Movement of infected animals can also spread the infection to atypical hosts that become carriers. Although brucellosis is endemic in Egypt, reports were scarce regarding its epidemiology in pigs. Thus, seroepidemiological surveillance was performed in pigs using serological and molecular assays. To achieve this objective, 331 serum samples were collected from two governorates (Cairo and Giza) having pig production. Samples were analyzed using competitive and indirect Brucella ELISAs. Anti-Brucella antibodies were detected in 4.83% and 10.8% pig sera by i-ELISA and c-ELISA, respectively. Molecular assays identified Brucella DNA in 3.02% of tested serum samples and identified as B. melitensis (7/10) and B. suis (3/10). The identification of both, B. melitensis and B. suis DNA, in serum samples of pigs was never reported till now. The results of this study provide help to develop effective control strategies for animal and human brucellosis in Egypt (Chapter 2). The transmission of host-specific Brucella spp. to non-preferred hosts may occur due to the mixed rearing of farm animals. Data regarding Brucella spp. in the camel population in Egypt are scarce. Thus, this thesis focused on the seroepidemiology of camel brucellosis in three governorates (Giza, Aswan, and Al-Bahr Al-Ahmar (The Red Sea)) with the highest camel population and the largest camel markets in Egypt to determine the seroprevalence and to identify the Brucella spp. of local camel holdings. Serologic assays conducted on 381 serum samples of camels showed that anti-Brucella antibodies were detected in 59 (15.5%), 87 (22.8%), 77 (20.2%) and 118 (31.0%) of sera by RBPT, i-ELISA, c-ELISA and CFT, respectively. Using real-time PCR, Brucella DNA was amplified from 32 (8.4%) seropositive samples and DNA of all species supposingly prevalent in Egypt was found: Brucella abortus (25/32), Brucella suis (5/32) and Brucella melitensis (2/32). Identification of B. suis DNA in camel sera reflects the complex epidemiological situation of brucellosis in Egypt. A high prevalence of brucellosis was observed in male animals. No significant association of brucellosis was found with age, sex, breed, and regions. The role of open markets in crossspecies transmission and dissemination of brucellae nationwide and regionally is highlighted (Chapter 3). The complex epidemiological situation of brucellosis in livestock poses risk to public health also. In this study, 29 B. abortus and B. melitensis strains were isolated from lymph nodes, milk, and fetal abomasal contents of infected cattle, buffaloes, sheep, and goats from nine districts. The phenotypic and genotypic patterns of antimicrobial resistance in local Brucella isolates were investigated. Notably, massive phenotypic resistance was observed in B. melitensis strains against ciprofloxacin, erythromycin, imipenem, rifampicin, and streptomycin, 76.2%, 19.0%, 76.2%, 66.7%, and 4.8%, respectively. Whereas, 25.0%, 87.5%, 25.0%, and 37.5% of B. abortus strains were resistant to ciprofloxacin, erythromycin, imipenem, and rifampicin, respectively. Molecular analysis identified mutations in the rpoB gene associated with rifampicin resistance. These mutations were identified in all phenotypically resistant isolates. Mutations in gyrA and gyrB genes associated with ciprofloxacin resistance were also identified in B. melitensis. Repeated mutations were detected at positions 676, 677 (TAC to CTC/tyrosine to leucine) and 1435 (AAG to CAG/lysine to glutamine) in the rpoB gene of phenotypic resistant B. melitensis isolates while the same was recorded at position 2890 (CGT to GGT/arginine to glycine) in the rpoB gene for B. abortus isolates. Similarly, mutations in the gyrA gene were detected at positions 167 (ATG to AGG/methionine to arginine), 197 (CCC to CGC/proline to arginine), 202 (CGC to AGC/arginine to serine), 235 (GGT to CGT/glycine to arginine), 941 (GCC to GAC/alanine to aspartic acid), 944 (GTG to GAG/valine to glutamic acid), 944-945 (GTG to GGA/valine to glycine), 946 (GCC to TCC/alanine to serine) and 962 (AAC to ACC/asparagine to threonine) in B. melitensis strains. Mutations detected in genes associated with antimicrobial resistance unravel the molecular mechanisms of resistance in Brucella isolates from Egypt. The mutations in the rpoB gene in phenotypically resistant B. abortus isolates in this study were reported for the first time for Egypt. The results of this study advocate for more research on the epidemiology and spread of antimicrobial resistance in Brucella strains (Chapter 4). Several studies from Egypt highlight the endemicity of brucellosis in the country. However, there are limited data available on the genetic diversity of brucellae circulating in Egypt. In this study, WGS was used to discriminate the genotypes along with virulence-associated factors and genetic markers responsible for antimicrobial resistance in 29 Brucella isolates (21 B. melitensis and 8 B. abortus) recovered from livestock in Egypt. In silico classical MLST identified sequence type-11 and sequence type-1 of B. melitensis and B. abortus strains, respectively in animal species in Egypt. These sequence types (ST-11 and ST-1) share a close relationship with Mediterranean and Asian, American, or European lineages respectively. In silico MLVA-16 analysis divided B. abortus and B. melitensis strains into 2 and 10 distinct genotypes, respectively. These strains types circulate in different governorates. However, cgSNPs analysis provided higher resolution and discriminated B. abortus and B. melitensis strains into 3 and 14 genotypes. Additionally, SNP variants were identified in rpoB and gyrA genes responsible for antimicrobial resistance in brucellae against rifampin and ciprofloxacin, respectively. These mutations are present at position 297-GAT to GAA/Asp-Glu in gyrA and two-point mutations were found in the rpoB gene at position 2784 (CGC to CGT/Arg-Arg) and 2394 (ACG to ACT/Thr-Thr) in 3 and 4 strains of B. melitensis, respectively (Chapter 5).