45 research outputs found
Molecular epidemiology of antimicrobial resistance (AMR) and Shiga toxin producing E. coli (STEC) in dairy herds of central Zambia
No full textAntimicrobial resistance (AMR) is a worldwide public health concern. While it is
evident that the use of antibiotics creates selection pressure for the evolution of
antibiotic resistance genes, there are still considerable knowledge gaps relating to the
status quo of antibiotic use, emergence of resistant pathogens in different livestock
production systems and spread within human and animal communities.
This thesis includes a survey of antibiotic use in the dairy sector within a specific area
of Zambia and analysis of AMR and virulence factors in E. coli isolated from dairy
cattle and diarrhoea human patients with the following objectives.
1. To investigate the usage of antibiotics in the dairy sector and the drivers for
use.
2. To determine the prevalence and patterns of antimicrobial resistance in E. coli
isolated from faecal samples of dairy cattle.
3. To use whole genome sequencing (WGS) to investigate the molecular
epidemiology of resistance determinants in E. coli strains isolated from both
dairy cattle and humans.
4. To assess the zoonotic potential of isolated E. coli focusing on Shiga toxin-producing
E. coli (STEC) and relationship to STEC associated with clinical
disease in the UK.
In view of these objectives, the first part of the work was carried out in Zambia and
involved a questionnaire, a field survey, isolation of E. coli from dairy cattle faecal
samples and phenotypic testing for AMR. In addition, E. coli isolates were obtained
from another study that was focused on human patients presenting with diarrhoea at
the University Teaching Hospital in Lusaka. The second part involved whole genome
sequencing and molecular analyses of E. coli for resistance and virulence genotypes at
the Roslin Institute (UK). For the field study, a stratified random sample of 104 farms
was studied, representing approximately 20% of all dairy farms in the region. On each
farm, faecal samples were collected from a random sample of animals and a
standardised questionnaire on the usage of antibiotics was completed. An E. coli
isolate was obtained from 98.67% (371/376) of the sampled animals and tested for
resistance against the six types of antibiotics (tetracycline, ampicillin,
sulfamethoxazole/trimethoprim, cefpodoxime, gentamicin and ciprofloxacin). These
E. coli were then analysed together with those from humans for genotypes in the
laboratory and from Illumina short read whole genome sequences using bioinformatics
tools.
Tetracylines and penicillin were the commonly used antibiotics in dairy herds. This
finding was in line with the resistance phenotypes detected in E. coli isolated from the
dairy cattle. The most prevalent AMR was to tetracycline (10.61; 95%CI: 7.40-13.82),
followed by ampicillin (6.02; 95%CI: 3.31-8.73), sulfamethoxazole/ trimethoprim
(4.49; 95%CI: 2.42-6.56), cefpodoxime (1.91; 95%CI: 0.46-3.36), gentamicin (0.89;
95%CI: 0.06-1.84) and ciprofloxacin (0%). The risk analysis indicated that AMR was
associated with livestock diseases (lumpy skin disease and foot rot), exotic breeds
(Jersey and Friesian), location, farm size and certain management practices.
Analysis of whole genome sequences showed that isolates from humans had both
higher levels and a greater diversity of resistance alleles than the cattle isolates.
Common genotypes in both populations were: tetA (16%), tetB (10%), tetC (2%) for
cattle isolates with tetA (32%), tetB (22%) and tetD (1%) in human isolates. Other
common genotypes were blaTEM (56%), sul1 (29%), sul2 (66%), strA4 (57%) and
strB1 (64%) in isolates of human origin while blaTEM (15%), sul1 (3%), sul2 (17%),
strA4 (13%) and strB1 (19%) were in the cattle isolates.
Whilst the E. coli isolates from cattle encoded resistance to common antibiotics of
limited significance to human clinical medicine, isolates from humans had additional
extended spectrum beta-lactamases (blaOXA, blaCMY, blaNDM, and blaDHA, blaOKP and
blaCTX-M) that encode for resistance to essential antibiotics such as third generation
cephalosporins and carbapenems. This was an evidence that AMR is an ongoing public
health subject in Zambia but the exclusivity of certain resistances in the human
population points to limited or no exchange of genotypes between E. coli of human
origin and those from cattle. AMR in humans was probably independently selected by
the use of antibiotics of clinical importance such as cephalosporin and
fluoroquinolones.
The virulence analysis focused on STEC, 11% (41/371) of E. coli isolates from cattle
contained Shiga toxin genes (stx) while none (0/73) of the human isolates were
positive. Phylogenetic analysis showed a random distribution of bovine STEC, with
no indication of clonal spread. Although 89% (16/18) of the STEC tested had a
cytotoxic effect on Vero cells, indicative of Shiga toxin production, only three (O45,
O111, O157) belonged to one of the seven serogroups (O26, O157, O111, O103,
O121, O145 and O45) associated with life-threatening enterohaemorrhagic E. coli
(EHEC) infections in humans. In line with this, only the O157 serotype encoded a type
3 secretion system. This shows that, while Stx-encoding strains are common in these
dairy herds of Zambia, they are not strain backgrounds known to pose an immediate
threat to human health as they lack colonisation factors that are found in typical human
EHEC. However, we must remain vigilant as emergence of EHEC strains in these
animals remains an ever-present threat
Quick PCR to detect M. tuberculosis and M. bovis in swine blood samples
No full textThe disease control strategies in livestock are the key areas to enhance improvement of livestock production and safety of livestock product consumers. Therefore, reliable techniques for quick detection and specific identification of disease causing agents need to be developed. In this study a nested PCR was developed using primer primier 5.0 software for primer design targeting IS1081 gene conserved regions in both M. tuberculosis and M. bovis. Two sets of primers TB-Q1/TB-Q2 and TB-B1/TB-B2, were designed to detect M. tuberculosis and M. bovis DNA extracted from blood samples. The findings were, the assay is sensitive enough to detect up to 1.35fg of the antigen DNA with 100% specificity.</p
Degumming of Vegetable Oil by a New Microbial Lipase
No full textLecitase® Ultra lipaza je kiselinska lipaza s maksimalnom aktivnosti pri pH=5,0. Enzim omogućava hidrolizu fosfolipida i triglicerida. Pri temperaturi višoj od 40 °C prevladava fosfolipazna aktivnost, a lipazna je aktivnost djelomično potisnuta. Enzim se može koristiti za degumiranje biljnog ulja, a fosfatidi u ulju smanjeni su enzimskom kataliziranom hidrolizom na manje od 10 mg/kg tijekom 5 sati pri 50 °C.Lecitase® Ultra lipase is an acidic lipase which exhibits maximal activity at pH=5.0. It takes part in the activity towards both phospholipid and triglyceride structures. When the temperature is over 40 °C, the phospholipase activity predominates, and the lipase activity is partly suppressed. It could be applied for the degumming of vegetable oil, and the phosphatides in the oil were easily converted by enzymatic catalyzed hydrolysis to less than 10 mg/kg within 5 h at 50 °C
Erratum:Prevalence and patterns of antimicrobial resistance among Escherichia coli isolated from Zambian dairy cattle across different production systems (Scientific Reports (2015) 5 (12439) DOI: 10.1038/srep12439)
No full textThe original version of this Article contained an error in the spelling of the author Paul R. Bessell, which was incorrectly given as Paul B. Bessell. This has now been corrected in both the PDF and HTML versions of the Article
Identification of MHC Alleles Associated with Disease Resistance/Susceptibility in Smallholder Cattle in Zambia
No full textThe occurrence of Major Histocompatibility Complex (MHC) alleles associated with resistance to Mastitis, Bovine Leukaemia Virus (BLV), Theileriosis, Foot and Mouth Disease (FMD) and susceptibility Dermatophilosis in African cattle is ill defined.
The researchers used manual annotation to screen for five MHC alleles previously known to be associated with resistance/susceptibility to these diseases from a database of alleles sequenced from 846 cattle in Zambia.
Overall, the researchers found twenty-eight (3.3%), twenty-one (2.6%), fifty-five (6.5%), and fifteen (1.8%) animals with resistance alleles to Mastitis, BLV, Theileriosis, FMD; and thirty-nine (4.6%) animals with susceptibility alleles to Dermatophilosis, respectively.
This study provides the first evidence of resistance/susceptibility alleles in smallholder cattle in Zambia and the data could aid strategies for breeding cattle with enhanced resistance to disease in endemic countries
Degumming of Vegetable Oil by a New Microbial Lipase
No full textLecitase® Ultra lipase is an acidic lipase which exhibits maximal activity at pH=5.0. It takes part in the activity towards both phospholipid and triglyceride structures. When the temperature is over 40 °C, the phospholipase activity predominates, and the lipase activity is partly suppressed. It could be applied for the degumming of vegetable oil, and the phosphatides in the oil were easily converted by enzymatic catalyzed hydrolysis to less than 10 mg/kg within 5 h at 50 °C
Resistance to change: AMR gene dynamics on a commercial pig farm with high antimicrobial usage
No full textGroup antimicrobial administration is used to control disease in livestock, but we have little insight into how this impacts antimicrobial resistance (AMR) gene dynamics. Here, a longitudinal study was carried out during a single production cycle on a commercial pig unit with high historic and current antimicrobial usage. Quantitative PCR, 16S rRNA gene metabarcoding and shotgun metagenomic sequencing were used to track faecal AMR gene abundance and diversity and microbiome alpha diversity. Shotgun metagenomic sequencing identified 144 AMR genes in total, with higher AMR gene diversity present in young pigs compared to dry sows. Irrespective of in-feed antibiotic treatment or changes in microbiome diversity, mean AMR gene copy number was consistently high, with some AMR genes present at copy numbers comparable to the bacterial 16S rRNA gene. In conclusion, AMR gene prevalence and abundance were not influenced by antibiotic use, either during the production cycle or following whole-herd medication. The high levels of certain genes indicate they are widely disseminated throughout the microbial population, potentially aiding stability. Despite the high and relatively stable levels of resistance genes against the main antimicrobials used, these compounds continue to control production limiting diseases on this unit
Whole Genome Sequence Analysis Reveals Lower Diversity and Frequency of Acquired Antimicrobial Resistance (AMR) Genes in E. coli From Dairy Herds Compared With Human Isolates From the Same Region of Central Zambia
No full textAntibiotic treatment of sick dairy cattle is critical for the sustainability of this production system which is vital for food security and societal prosperity in many low and middle-income countries. Given the increasingly high levels of antibiotic resistance worldwide and the challenge this presents for the treatment of bacterial infections, the rational use of antibiotics in humans and animals has been emphatically recommended in the spirit of a “One Health” approach. The aim of this study was to characterize antimicrobial resistance (AMR) genes and their frequencies from whole genome sequences of Escherichia coli isolated from both dairy cattle and human patients in central Zambia. Whole genome sequences of E. coli isolates from dairy cattle (n = 224) and from patients at a local hospital (n = 73) were compared for the presence of acquired AMR genes. In addition we analyzed the publicly available genomes of 317 human E. coli isolates from over the wider African continent. Both acquired antibiotic resistance genes and phylogroups were identified from de novo assemblies and SNP based phylogenetic analyses were used to visualize the distribution of resistance genes in E. coli isolates from the two hosts. Greater acquired AMR gene diversity was detected in human compared to bovine E. coli isolates across multiple classes of antibiotics with particular resistance genes for extended-spectrum beta lactamases (ESBL), quinolones, macrolides and fosfomycin only detected in E. coli genomes of human origin. The striking difference was that the Zambian or wider African human isolates were significantly more likely to possess multiple acquired AMR genes compared to the Zambian dairy cattle isolates. The median number of resistance genes in the Zambian cattle cohort was 0 (0–1 interquartile range), while in the Zambian human and wider African cohorts the medians and interquartile ranges were 6 (4–9) and 6 (0–8), respectively. The lower frequency and reduced diversity of acquired AMR genes in the dairy cattle isolates is concordant with relatively limited antibiotic use that we have documented in this region, especially among smallholder farmers. The relatively distinct resistant profiles in the two host populations also indicates limited sharing of strains or genes
Alternate thermoregulation and functional binding of Escherichia coli Type 1 fimbriae in environmental and animal isolates
No full textType 1 fimbriae (T1F) are well characterised cell surface organelles expressed by Escherichia coli and required for adherence to mannosylated host tissue. They satisfy molecular Koch's postulates as a virulence determinant and a host-adapted role has been reinforced by reports that T1F expression is repressed at sub-mammalian temperatures. Analysis of a group of 136 environmental and animal E. coli isolates that express T1F at 37 °C showed that 28% are also capable of expression at 20 °C, in a phase variable manner. The heterogeneous proportions varied widely, and although growth temperature impacted the total proportion expressing T1F, there was no direct correlation between growth at 37 and 20 °C, indicative of differences in thermoregulation of the genetic switch (fimS) that controls phase variation. Specificities of the adhesin (FimH) also varied between the isolates: most bound to α-(1-3) mannan and yeast extracts as expected, but some recognised β-(1-4)-mannans and N-linked glycoproteins from plants, and T1F from two of the isolates mediated binding to plant roots. The results expand our view of a well described adherence factor to show alternative expression profiles and adhesin specificities, which in turn may confer an advantage for certain isolates in alternative hosts and habitats
Factors Associated with Occurrence of Antibiotic Resistance in Broiler Chickens in Selected Districts, Zambia
No full textA cross-sectional study was conducted in five districts of Zambia. The study aimed to investigate the risk factors associated with antimicrobial resistance (AMR) in broiler chickens in Zambia. A total of 109 broiler farms were included in the study, of which 104 were small-scale farms, two were medium-scale, and three were commercial-scale. A semi-structured questionnaire was administered alongside the collection of cloacal swabs and litter samples to determine the risk factors associated with the occurrence of AMR of food pathogen microorganisms on the selected broiler farms. Data collected was analysed using STATA version 12. The broiler farm-level AMR prevalence was 84.4% (n= 109). Gender, marital status, administration of drugs by veterinary personnel or the farmer and the poultry house drainage destination were among the predictors of AMR on broiler chicken farms. The high prevalence of AMR in broiler chicken farms suggests a high dependence on antimicrobials in poultry production in Zambia. In addition, socio-demographics such as gender and age, as well as farm characteristics and management practices, may primarily contribute to the high prevalence of antibiotic resistance. This study has demonstrated the association between the AMR of foodborne pathogens and the risk factors for AMR during poultry production. It is, therefore, necessary for risk management measures such as biosecurity to be put in place to target the identified AMR predictor variables. There is also a need to strengthen AMR surveillance to ensure intervention strategies are appropriate to the existing risk
