Frequency of Extended Spectrum Beta
Lactamase Producing Escherichia coli in
Fresh and Frozen meat
Presented By
Amna Mamtaz
Roll No.231434
Mphil 1st Semester
INTRODUCTION:
• E.coli is gram Negative Rod.
• It is a Coliform.
• Responsible for gastrointestinal
issues including diarrhea, UTI and
Neonatal Meningitis.
• Poultry is main source of
E.coli dissemination.
(Gallardo et al.,2017)
What is extended spectrum Beta Lactamase??
• Enzymes produced by bacteria that break the beta-lactam
ring, inactivating the beta-lactam antibiotic.
E.coli Subtypes:
E.coli subtypes including
Entrotoxigenic Escherichia coli(ETEC),
Enterohemorrhagic Escherichia coli(EHEC)
Extraintestinal pathogenic Escherichia coli (ExPEC)
(Ranjan et al.,2017)
Mode Of Transmission
Meat can
Contaminatd Consumption
become
tools of
contaminated
contaminated
during Equipment food
slaughter
Antibiotics usage and it's Resistance in Food
animal production
• Antimicrobial resistance in E.coli( ESBL producing) is one
of most serious public health threat
• This resistance is shifted to humans by consumption of
meat and its product
AIM Of STUDY:
Purpose of this study is to determine Frequency of ESBL
producing E.coli in Fresh and Frozen Meat samples and it's
Microbial Resistant To detect Different ESBL-EC genes.
Materials & Methods
Sample
Collection
Isolation
Identification
Biochemical
tests
Antimicrobial
Susceptibility
Tests
Statistical tools
Samples collection
Total (n=100) fresh and frozen meat samples (poultry and
mutton)collected.
Packaged into polyethylene bags.
labeled properly.
Transferred in the icebox to the laboratory.
Isolation of Escherichia coli
25gm of each meat specimen was mixed aseptically in
the 225 ml of sterilize peptone water and subjected to
overnight enrichment.
Inoculation on MacConkey’s agar or Eosin Methylene
Blue (EMB) agar (Oxoid, UK) and incubated for 24-48
hours at 37°C.
(Nawaz et al.,2019)
Identification of E.coli
• Pinkish colonies on MacConkey agar
• Metallic green colonies on EMB agar
• Confirmed by Gram’s staining and biochemical tests.
• Preservation of isolates was done in brain heart infusion
with 30% glycerol
• Stored at -80°C
(Rahman et al., 2018).
RESULTS
Antimicrobial susceptibility testing:
Phenotypic detection of ESBL production
Modified double disc synergy test (MDDST)
Used for detection of ESBL producing E. coli using cefotaxime
(CTX, 30μg), ceftazidime (CAZ, 30 μg), ceftriaxone (CRO, 30 μg)
and amoxicillin-clavulanate (20/10 μg) discs.
Amoxicillinclavulanate (20/10 μg) disc was placed to center of the
agar plate and other antibiotics discs were placed at 15 to
20mm distance.
The ESBL production considered as positive as any increase in
zone in the direction of disc of amoxicillin-clavulanate.
Frequency of ESBL producing Escherichia coli:
The double disk synergy test showed that phenotypically 17
out of 36 (47.22%) isolates of E.Coli were ESBL producers.
Among these ESBL positive (n=17) isolates,bla CTX-M
gene was identified from 5 (29.41%) isolates followed by bla
OXA-48 from 4 (23.52%) and bla TEM gene was detected
1(5%)
Identification of ESBL encoding genes:
The isolates of Escherichia coli showing positive
phenotypic ESBL detection were processed for the
identification of extended spectrum beta lactamase
encoding genes using specific primers by PCR.
The targeted ESBL genes were bla CTX-M, bla TEM and
bla OXA-48 .
Molecular confirmation of Escherichia coli by
PCR
Discussion and Conclusion:
The results of current study concluded that
poultry and mutton meat samples were contaminated with
MDR-Escherichia coli and this contamination was more in
fresh meat samples as compared to Frozen meat.
Chicken meat is highly contaminated with E. coli than
mutton.
Mostly these isolates are MDR in nature, which increase
the challenge for their treatment and eradication.
FUTURE PROSPECTUS
This study revealed that vigilant control
procedures should be implemented all over the
food chain and effective surveillance should also
be performed at national level to minimize the
spread of MDR and ESBL producing Escherichia
coli from raw meat.
REFERENCCE:
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Prevalence of toxins and antimicrobial resistance among E.
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Isolation and molecular characterization of multidrug-
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