1) The study found Extended Spectrum Beta-Lactamase (ESBL) antibiotic resistant bacteria on packaged fruit samples, indicating that unwashed fruit can be a vector for these bacteria to enter the human food chain. 2) Of the 58 fruit samples tested, 10.3% of isolated bacteria were found to be ESBL producers carrying genes such as TEM, SHV, and CTXM that code for beta-lactamase enzymes which break down beta-lactam antibiotics. 3) Specifically, samples of strawberries, apples, and red grapes were found to contain ESBL bacteria carrying TEM, SHV, and CTXM genes, showing that fresh produce can be a source

1. Extended Spectrum Beta-Lactamase (ESBL) antibiotic
resistant bacteria found on food
outlet packaged fruit.
It is apparent that the overuse of antibiotics is leading to the increase of antibacterial
resistant bacteria. One method for these bacteria to enter the human food chain may
be the intake of fruit eaten without being washed by the consumer. (Ruimy et al. 2010).
The aim of this study is to find examples of these bacteria in the form of ESBLs.
58 samples of fruit were taken and sub-cultured (fig. 2A) to find any bacteria present.
All isolated bacteria were harvested (fig. 5) and E.coli and Coliforms were tested for
Cefpodoxime (CPD) susceptibility (fig. 2B), any resistant strains were then tested with a
comparison of ‘CPD’ and ‘CPD with Clavulanic acid (CV) to detect ESBL producers (fig.
2C).
William Hughes
Bangor University
School of Medical Sciences, Bangor LL57 2UW, UK
bsu2ff@bangor.ac.uk
Supervisor: Mr. Merfyn Williams
figure 2: (A) plates showing sub-culture of sample organisms, purple—E.coli, blue—
Coliform, turquoise—Enterococci (B) Cefpodoxime (CPD) Screening Test, (C) CPD and CPD
with Clavulanic Acid (CPDCV) sensitivity comparison. All samples with a difference >5mm
are confirmed ESBL producers.
This particular sequence shows the progression of confirmed ESBL sample 7AE, an E.coli
isolated from Strawberries grown in the United Kingdom.
figure 3: graph showing difference in Cefpodoxime (CPD) and
Cefpodoxime with Clavulnic acid (CPDCV) sensitivity,
indicating ESBL producers.
Beta-Lactams are a group of antibioticsthat include penicillins,
cephalosporins (G1, G2 & G3), monobactams and carbapenems that
contain a Beta-Lactam ring (fig. 1) that aid in the breakdown of cell
walls in bacteria.
ESBL producing bacteria are capable of producing the enzyme, Beta-
Lactamase that is capable of breaking down this Beta-Lactam ring,
rendering the antibiotic unviable.
figure 1: drawing of
a Beta-Lactam ring
References:
Ruimy, R., Brisabois, A., Bernede, C., Skurnik, D., Barnat, S.,
Arlet, G., Momcilovic, S., Elbaz, S., Moury, F. & Vibet, M.
2010, "Organic and conventional fruits and vegetables
contain equivalent counts of Gram‐negative bacteria
expressing resistance to antibacterial
agents", Environmental microbiology, vol. 12, no. 3, pp.
608-615.
Shaikh, S., Fatima, J., Shakil, S., Rizvi, S.M.D. & Kamal, M.A.
2015, "Antibiotic resistance and extended spectrum beta-
lactamases: Types, epidemiology and treatment", Saudi
Journal of Biological Sciences, vol. 22, no. 1, pp. 90-101.
Lynch, M., Tauxe, R. & Hedberg, C. 2009, "The growing
burden of foodborne outbreaks due to contaminated
fresh produce: risks and opportunities", Epidemiology
and infection, vol. 137, no. 03, pp. 307-315.
figure 5: table to show all 50 harvested samples from 58 fruits and the test results for
each, including results from PCR analysis.
figure 6: (A) Analysis of blaTEM amplicon on 2% agarose gel ; Lane 1: the 100-bp ladder, lanes 2–7: the 6 samples, lane 8: positive control . Two
samples 1BO [lane 3] and 3BO [lane 5] displayed high-intensity bands at 800 bp which corresponded to the blaTEM gene. Samples 1BE, 3AE & 7AE
also show low-Intensity bands. (B) Analysis of blaSHV amplicon on 2% agarose gel ; Lane 1: the 100-bp ladder, lanes 2–7, the 6 samples, and lane
8 : the positive control. Two isolates showed high-intensity bands with sizes corresponded to blaSHV gene: 1BO [lane 3] & 3BO [lane 5], 700 bp.
All other samples also showed low-intensity bands. (C) Analysis of blaCTXM amplicon on 2% agarose gel. Lane 1: the 100-bp ladder; lanes 2–7: the
6 samples had bands with lengths that corresponded to two groups of CTXM genes; CTXMGp1 (700 bp) samples 1BE, 3AE,7AE and 9BE and
CTXMGp9 (561 bp) samples 1BO and 3BO. and lane 8 : the positive control.
ESBL bacteria are known
to carry genes such as
TEM, SHV, OXA & CTXM,
most commonly,
although there are
others (Shaikh et al.
2015). The Samples
collected are shown to
contain TEM, SHV and
CTXM.
figure 4: image of an apple,
strawberries and red-grapes,
the fruits where ESBL
bacteria were found
Results show that 10.3% of found bacteria were expressing ESBL genes including TEM,
SHV and CTXM. Samples 1BE, 1BO, 3AE, 3BO, 7AE & 9BE all contained high-intensity
bands of CTXM (fig. 6C) and 2 samples (1BO, 3BO) also show high intensity bands of
both TEM (fig. 6A) and SHV (fig. 6B). Other samples also contained low intensity bands of
TEM and SHV, although their primary gene
is the CTXM.
It has been shown in the past that fresh
produce can be the source of a food-borne
infection (Lynch et al. 2009). The study
conducted shows that unwashed fruits can
be a vessel for ESBL antibiotic resistant
bacteria to enter the human food chain via
consumption.