ArticlePDF Available

Antimicrobial resistance and virulence factors profile of Salmonella spp. and Escherichia coli isolated from different environments exposed to anthropogenic activity

Authors:
  • Food and Agriculture Organization of the United Nations

Abstract and Figures

Objective The study aimed to identify the antimicrobial resistance (AMR) determinants and virulence factors in Salmonella spp. and Escherichia coli recovered from different anthropogenic areas in North Carolina. Methods Soil samples were collected from different anthropogenic areas: urban and natural. Minimum inhibitory concentration (MIC) was determined by using the broth microdilution method. Whole genome sequencing and analysis were done to identify the AMR determinants and virulence factors. Results A higher prevalence of Salmonella spp. and E. coli was detected in the urban environment. The Salmonella spp. isolates showed resistance to Sulfisoxazole and Streptomycin, while E. coli was resistant to Sulfisoxazole, Cefoxitin, and Ampicillin. Salmonella serotypes Schwarzengrund and Mississippi were identified based on WGS analysis. Aminoglycoside resistance genes and IncFIB and IncFIC(FII) plasmids were detected among Salmonella spp. In general, E. coli was predominated by isolates from phylogroup B1, B2, and D. Multidrug transporter mdfA gene was detected in majority of the E. coli from both the urban (100%) and natural (84.5%) environment. FosA7 gene was detected in an isolate from a residential yard. The pCoo and pB171 plasmids were detected in urban, while col(156) and pHN7A8 plasmids were detected in natural environments. Conclusion The detection of AMR determinants and virulence factors in these bacteria is significant in understanding the occurrence and even the development of AMR. The presence of these determinants in different anthropogenic areas suggests the need to conduct longitudinal studies for comparing the profile of pathogens across different environments.
Content may be subject to copyright.
Short
Communication
Antimicrobial
resistance
and
virulence
factors
prole
of
Salmonella
spp.
and
Escherichia
coli
isolated
from
different
environments
exposed
to
anthropogenic
activity
Michelle
M.
Balbin
a,d
,
Dawn
Hull
a
,
Chloe
Guest
b
,
Lauren
Nichols
c
,
Robert
Dunn
c
,
Dawn
Hull
a
,
Siddhartha
Thakur
a,b,
*
a
Department
of
Population
Health
and
Pathobiology,
College
of
Veterinary
Medicine,
North
Carolina
State
University,
Raleigh,
NC,
USA
b
Faculty
of
Health
and
Medical
Sciences,
University
of
Surrey,
Guildford,
Surrey
GU2
7XH,
UK
c
Department
of
Applied
Ecology,
College
of
Agriculutre
and
Life
Sciences,
North
Carolina
State
University,
Raleigh,
NC,
USA
d
Comparative
Medicine
Institute,
North
Carolina
State
University,
NC,
USA
A
R
T
I
C
L
E
I
N
F
O
Article
history:
Received
11
September
2019
Received
in
revised
form
27
May
2020
Accepted
29
May
2020
Available
online
10
June
2020
Keywords:
Anthropogenic
areas
Whole-genome
sequencing
FosA7
gene
pHN7A8
(F33:A-:B-plasmid)
A
B
S
T
R
A
C
T
Objective:
The
study
aimed
to
identify
the
antimicrobial
resistance
(AMR)
determinants
and
virulence
factors
in
Salmonella
spp.
and
Escherichia
coli
recovered
from
different
anthropogenic
areas
in
North
Carolina.
Methods:
Soil
samples
were
collected
from
different
anthropogenic
areas,
urban
and
natural.
The
minimum
inhibitory
concentration
(MIC)
was
determined
by
using
the
broth
microdilution
method.
Whole-genome
sequencing
(WGS)
and
analysis
were
done
to
identify
the
AMR
determinants
and
virulence
factors.
Results:
A
higher
prevalence
of
Salmonella
spp.
and
E.
coli
was
detected
in
the
urban
environment.
The
Salmonella
spp.
isolates
showed
resistance
to
sulsoxazole
and
streptomycin,
whereas
E.
coli
was
resistant
to
sulsoxazole,
cefoxitin
and
ampicillin.
Salmonella
serotypes
Schwarzengrund
and
Mississippi
were
identied
based
on
WGS
analysis.
Aminoglycoside
resistance
genes
and
IncFIB
and
IncFIC(FII)
plasmids
were
detected
among
Salmonella
spp.
In
general,
E.
coli
was
predominated
by
isolates
from
phylogroups
B1,
B2
and
D.
The
multidrug
transporter
mdfA
gene
was
detected
in
most
of
the
E.
coli
from
both
the
urban
(100%)
and
natural
(84.5%)
environments.
The
FosA7
gene
was
detected
in
an
isolate
from
a
residential
yard.
The
pCoo
and
pB171
plasmids
were
detected
in
an
urban
environment;
col(156)
and
pHN7A8
plasmids
were
detected
in
natural
environments.
Conclusions:
The
detection
of
AMR
determinants
and
virulence
factors
in
these
bacteria
is
signicant
in
understanding
the
occurrence
and
even
the
development
of
AMR.
The
presence
of
these
determinants
in
different
anthropogenic
areas
suggests
the
need
to
conduct
longitudinal
studies
for
comparing
the
prole
of
pathogens
across
different
environments.
©
2020
The
Author(s).
Published
by
Elsevier
Ltd
on
behalf
of
International
Society
for
Antimicrobial
Chemotherapy.
This
is
an
open
access
article
under
the
CC
BY-NC-ND
license
(http://creativecommons.
org/licenses/by-nc-nd/4.0/).
1.
Introduction
The
emergence,
persistence
and
continuous
spread
of
antimi-
crobial
resistance
(AMR)
is
considered
as
one
of
the
greatest
threats
to
humans
[13].
Currently,
the
annual
deaths
due
to
AMR
are
estimated
to
be
700
000
[4];
by
2050,
it
is
projected
that
this
number
could
reach
up
to
10
million
[4]
and
an
economic
loss
amounting
to
$100
trillion.
Recognising
this
urgent
problem,
the
World
Health
Organization
(WHO)
issued
a
Global
Action
Plan
on
AMR
to
ensure
the
continuity
of
successful
treatment
and
prevention
of
infectious
diseases
through
the
responsible
use
of
quality,
effective
and
safe
medicines
[4].
The
environment
plays
a
signicant
role
in
the
emergence
and
transmission
of
AMR
determinants
and
pathogenic
bacteria
[2].
The
constant
interactions
of
humans,
animals
and
the
environ-
ment
can
give
rise
to
selection
pressures
leading
to
changes
that
would
help
an
organism
to
survive,
such
as
mutations,
horizontal
transfer
of
AMR
genes,
plasmids
and
virulence
factors
[1,3,5].
Soil
is
a
huge
source
of
AMR
determinants
considering
its
diverse
microbial
composition,
which
varies
depending
on
the
geograph-
ical
and
biochemical
gradient
[6]
as
a
result
of
constant
selective
pressures
exerted
in
the
environment.
A
large
number
of
*
Corresponding
author
at:
1060
William
Moore
Drive,
Raleigh,
NC
27606,
USA.
E-mail
address:
sthakur@ncsu.edu
(S.
Thakur).
http://dx.doi.org/10.1016/j.jgar.2020.05.016
2213-7165/©
2020
The
Author(s).
Published
by
Elsevier
Ltd
on
behalf
of
International
Society
for
Antimicrobial
Chemotherapy.
This
is
an
open
access
article
under
the
CC
BY-
NC-ND
license
(http://creativecommons.org/licenses/by-nc-nd/4.0/).
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
Contents
lists
available
at
ScienceDirect
Journal
of
Global
Antimicrobial
Resistance
journal
home
page
:
www.e
lsevier.com/loca
te/jgar
antimicrobials
being
discharged
into
the
environment,
along
with
continual
interactions
of
humans
and
animals,
not
only
alters
the
microbial
community
but
also
hastens
and
sustains
the
selection,
proliferation
and
spread
of
AMR
determinants
[1,5,7].
As
for
pathogenic
bacteria
such
as
Escherichia
coli
O157:H7,
the
increased
virulence
occurs
when
there
is
gene
loss
or
silencing,
insertion
and
rearrangement,
which
occurs
via
the
same
mechanisms
associated
with
the
acquisition
of
AMR
determinants
in
the
environment
[5].
Accordingly,
AMR
is
commonly
seen
in
environments
where
microbes
routinely
come
into
contact
with
antimicrobials,
such
as
in
hospitals
and
livestock
farms
[3,5].
Most
of
the
previously
conducted
studies
focused
on
clinical
and
agricultural
environ-
ments.
This
left
a
knowledge
gap
regarding
AMR
determinants
and
virulence
factors
[1,3,7,8]
found
in
the
natural
environment,
as
well
as
in
areas
subjected
to
anthropogenic
impacts.
Salmonella
spp.
and
E.
coli
are
two
of
the
most
commonly
reported
bacterial
pathogens
affecting
millions
of
humans
annually
[2].
Although
some
serotypes
or
strains
of
these
bacteria
are
host
specic,
others
that
can
be
transmitted
from
animals
to
humans
and
to
the
environment.
These
bacteria
are
known
to
acquire
and
dissemi-
nate
AMR
determinants
and
virulence
factors,
allowing
them
to
thrive
in
the
environment
[1].
Often,
the
reintroduction
of
these
resistant
and
pathogenic
bacteria
into
their
primary
host
is
a
health
concern
due
to
their
new
properties,
which
could
cause
serious
diseases
[3].
E.
coli
O157:H7,
for
example,
is
a
pathogenic
strain
that
is
well
adapted
in
the
environment
[5,9].
For
these
reasons,
Salmonella
spp.
and
E.
coli
are
often
used
as
indicator
bacteria
in
AMR
surveillance
programs
and
other
public
health
related
studies
[1].
In
our
study,
we
isolated
Salmonella
spp.
and
E.
coli
from
different
anthropogenic
areas
in
North
Carolina
and
identied
the
AMR
determinants
and
virulence
factors
through
antimicrobial
susceptibility
testing
(AST)
and
whole-genome
sequencing
(WGS).
The
information
gathered
from
this
study
is
important
in
understanding
the
occurrence
and
persistence
of
resistance
determinants
and
pathogenic
bacteria
in
the
environment.
2.
Materials
and
methods
2.1.
Sample
collection
and
bacteria
isolation
A
total
of
70
soil
samples
were
collected
from
different
anthropogenic
areas
in
North
Carolina,
urban
(n
=
29)
and
natural
(n
=
41).
The
urban
environment
includes
garden
landscapes
(n
=
3),
residential
yards
(n
=
14)
and
indoor
potted
plants
(n
=
12);
the
natural
environment
includes
Forest
A
(n
=
14),
Forest
B
(n
=
8),
Forest
C
(n
=
5),
Forest
D
(n
=
5)
and
potting
soil
mix
(n
=
9).
Prior
to
bacterial
isolation,
each
of
the
soil
samples
was
sieved
and
mixed.
Salmonella
spp.
were
isolated
using
xylose-lysine-
tergitol
(XLT-4)
selective
media.
Subsequently,
ve
colonies
from
each
positive
plate
were
subjected
to
biochemical
tests
using
triple
sugar
iron
agar
slant
(TSI
agar),
lysine
iron
agar
(LIA)
and
urea
agar
slant
[10,11].
For
E.
coli,
each
sample
was
streaked
into
MacConkey
agar
[11,12].
From
each
plate,
multiple
(n
=
3)
colonies
were
picked
for
further
isolation
until
a
pure
culture
was
obtained.
Salmonella
spp.
isolates
were
conrmed
through
amplication
of
the
invA
gene
[10];
the
16s
rRNA
gene
was
used
to
identify
E.
coli
[13].
2.2.
Resistance
determination
The
AST
was
performed
by
the
broth
microdilution
method
in
a
96-well,
Gram-negative
sensititre
plate
containing
a
panel
of
14
antimicrobials
(CMV3AGNF
Gram-Negative
NARMS
plate,
Trek
Diagnostic
System,
Cleveland,
OH).
The
minimum
inhibitory
concentration
(MIC)
was
interpreted
based
on
Clinical
and
Laboratory
Standards
Institute
(CLSI)
guidelines
[10,11].
2.3.
Analysis
of
antimicrobial
resistance
genes
The
presence
of
antimicrobial
resistance
genes
(ARGs)
among
the
isolates
was
determined
through
the
amplication
of
ARGs
by
the
polymerase
chain
reaction
(PCR)
assay.
The
presence
of
integron
was
also
determined
by
amplication
of
the
integron
gene-1
(intI
1)
[10]
(Supplementary
Table
S1).
2.4.
Whole-genome
sequencing
The
bacterial
isolates
that
showed
resistance
to
antimicrobials
based
on
the
results
of
AST
by
the
broth
microdilution
method
and
that
carried
ARGs
based
on
the
results
of
PCR
were
selected
and
subjected
to
WGS.
DNA
extraction
was
carried
out
using
the
Qiagen
DNeasy
Blood
and
Tissue
Kit
(Qiagen,
Hilden,
Germany)
following
the
manufacturers
instructions.
The
quality
and
concentration
of
the
extracted
DNA
were
determined
using
the
NanoDrop
TM
2000/
2000c
Spectrophotometer
(Thermo
Fisher
Scientic,
Waltham,
Massachusetts)
and
Qubit
3.0
Fluorometer
(Thermo
Fisher
Scientic).
DNA
libraries
were
prepared
using
the
Nextera
XT
DNA
Library
Preparation
Kit
(Illumina,
San
Diego,
CA)
following
the
manufacturers
instructions.
The
resulting
DNA
libraries
were
puried
using
AMPure
XP
beads
(Beckman
Coulter,
Sharon
Hill,
PA)
and
requantied
using
the
Qubit
3.0
Fluorometer
(Thermo
Fisher
Scientic).
Sequencing
was
performed
on
the
MiSeq
System
using
v2
sequencing
reagent
kits
(Illumina)
[10].
2.5.
Data
assembly
and
analysis
Raw
sequences
were
assembled
using
the
CLC
Genomic
Workbench
(Qiagen
Bioinformatics,
Redwood
City,
CA).
Draft
genomes
were
annotated
with
Rapid
Annotation
using
Subsys-
tem
Techn ology
(RAST)
(http://rast.theseed.org/FIG/rast.cgi).
The
following
provided
further
bioinformatic
analyses:
pubMLST
(https://pubmlst.org/bigsdb),
SeqSero
(http://den-
glab.info/SeqSero),
SISTR
via
EnteroBase
(https://enterobase.
warwick.ac.uk),
PlasmidFinder
(https://cge.cbs.dtu.dk/services/
PlasmidFinde/),
VirulenceFinder
(https://cge.cbs.dtu.dk/ser-
vices/VirulenceFinder/),
Virulence
Finder
Database
(http://
www.mgc.ac.cn/VFs/main.htm),
ResFinder
(https://cge.cbs.dtu.
dk/services/ResFinder)
[9],
Comprehensive
Antibiotic
Resis-
tance
Database
(CARD)
(https://card.mcmaster.ca),
GrapeTree
(https://enterobase.warwick.ac.uk/)
and
visualisation
through
iTOL
(https://itol.embl.de).
The
Salmonella
Schwarzengrund
str.
CVM19633
(NC_011094)
and
E.
coli
str.
K-12
substr.
MG1655
(NC_000913)
were
used
as
references
for
Salmonella
spp.
and
the
E.
coli
phylogenetic
tree
analysis,
respectively.
3.
Results
3.1.
Prevalence
of
Salmonella
spp.
and
E.
coli
The
prevalence
of
Salmonella
spp.
in
soil
samples
from
the
urban
and
natural
environments
were
10.34%
and
2.44%,
respectively.
In
E.
coli,
the
prevalence
was
observed
to
be
62.1%
in
the
urban
environment
and
26.8%
in
the
natural
environment.
Interestingly,
two
soil
samples
from
RY
and
a
single
soil
sample
each
from
IPP
and
Forest
C
were
positive
for
both
bacteria.
From
these
positive
soil
samples,
a
total
of
20
Salmonella
spp.
(urban,
n
=
15;
natural,
n
=
5)
and
66
E.
coli
(urban,
n
=
41;
natural,
n
=
25)
isolates
were
obtained
and
analysed.
3.2.
AMR
in
Salmonella
spp.
and
E.
coli
The
Salmonella
spp.
from
the
urban
environment
were
resistant
to
streptomycin
(66.67%)
and
sulsoxazole
(46.47%)
whereas
those
M.M.
Balbin
et
al.
/
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
579
from
the
natural
environment
were
resistant
to
sulsoxazole
(100%)
(Supplementary
Table
S2).
Similarly,
E.
coli
isolated
from
the
urban
environment
showed
the
highest
resistance
to
sulsoxazole
(78.05%),
followed
by
ampicillin
(7.32%)
and
cefoxitin
(4.88%).
Isolates
from
the
natural
environment
only
showed
resistance
to
sulsoxazole
(80%)
(Table
1).
3.3.
ARGs
detection
using
the
PCR
Salmonella
spp.
from
the
urban
environment
carry
fox
(80%),
strA
(66.67%)
and
strB
(66.67%)
genes,
whereas
all
the
isolates
from
the
natural
environment
encoded
for
the
fox
(100%)
gene.
With
E.
coli,
the
ARGs
sul1
(4.89%)
and
bla
CMY-2
(4%)
were
detected
from
the
urban
and
natural
environments,
respectively.
Class
I
integron
was
not
detected
in
both
bacteria.
3.4.
WGS
analysis
Based
on
the
WGS
analysis,
we
identied
Salmonella
serotypes
Schwarzengrund
(64.28%)
and
Salmonella
Mississippi
(35.71%)
from
the
urban
environment
(n
=
14);
the
isolates
from
the
natural
environment
(n
=
5)
were
Mississippi
(100%).
All
S.
Mississippi
carried
aaa(6)-Iaa
gene;
S.
Schwarzengrund
harboured
aac(6)-
Iaa,
aph(6)-Id
and
aph(3)-Ib.
All
Salmonella
isolates
also
contained
mdtK
(multidrug
and
toxic
compound
extrusions
[MATEs]),
mdsABC
(metal
and
antibiotic
efux
pump)
and
golS
(mdsABC
efux
pump
regulator.
Only
S.
Schwarzengrund
isolates
were
found
to
carry
IncFIB
and
IncFIC(FII)
plasmids.
Virulence
factors
for
Salmonella
pathogenicity
island
(SPI)-1
and
SPI-2,
mig-14,
mgtB-C,
adherence
factors
sinH,
misL,
csgA-F
and
m
type
I
mbriae
genes
were
present
in
all
20
Salmonella
isolates.
All
S.
Schwarzengrund
contained
the
cdtB
(cytothethal-distending
toxin)
and
iucB-Di;
S.
Mississippi
serotypes
carried
grvA
and
sodCI
at
>95%
identity
[14]
(Supplementary
Fig.
S1).
Similarly,
WGS
analysis
of
E.
coli
isolates
from
the
urban
environment
predominantly
belonged
to
phylogroups
A
(26.67%),
B1
(21.67%),
D
(20%)
and
B2
(13.33%).
The
E.
coli
from
the
natural
environment
(n
=
13)
were
determined
to
be
from
phylogroups
B1
(38.46%),
B2
(38.46%),
D
(7.69%)
and
E
(7.69%).
There
were
23
virulence
genes
identied
among
the
isolates;
only
9
were
detected
at
a
frequency
>10%.
These
genes
include
gad
(glutamate
decarboxylase),
lpfA
(long
polar
mbriae),
iss
(increased
survival
serum),
air
(enteroaggregative
immunoglobulin
repeat
protein)
and
eilA
(Salmonella
HilA
homolog)
among
others.
Also,
virulence
factors
such
as
iroN
(Salmochelin),
espA
(secreted
proteins)
and
tsh
(temperature-sensitive
hemagglutina-
tion)
were
identied.
The
mdfA
gene
was
detected
in
100 %
and
84.5%
of
E.
coli
isolates
from
the
urban
and
natural
environment,
respectively
(Fig.
1).
A
new
gene,
FosA7,
conferring
resistance
to
fosfomycin
was
detected
in
an
isolate
from
a
residential
yard.
The
activity
of
the
FosA7
gene
was
assessed
by
amplifying
and
cloning
the
segment
into
E.
coli
(TOP10).
Using
the
Etest
(Liolchem,
Thermo
Fisher
Scientic),
the
donor
isolate
and
the
transformed
E.
coli
showed
resistance
to
fosfomycin
(MIC
>
256
m,g/mL).
Its
relatedness
to
other
fosfomycin
resistance
genes
was
analysed
using
MAFFT
(https://mafft.cbrc.jp/alignment/software)
(Fig.
2).
The
pCoo
and
pB171
plasmids
were
detected
in
isolates
from
a
residential
yard
and
garden
landscape,
respectively,
whereas
col
(156)
and
pHN7A8
plasmids
were
detected
from
the
natural
environment
Forests
A
and
D.
Table
1
Resistance
and
MIC
distribution
(squashtogram)
a
of
E.
coli
isolates
from
urban
(n
=
41)
and
natural
(n
=
25)
environments.
AM
Anthropogenic
area
%
R
b
Distribution
of
MICs
in
m
g/mL
(%)
0.015
0.03
0.06
0.125
0.25
0.5
1
2
4
8
16
32
64
128
256
512
GEN
Natural
0
0.00
16.00
36.00
4.00
40.00
4.00
0.00
Urban
0
0.00
46.34
39.02
4.88
2.44
7.32
0.00
STR
Natural
0
0.00
0.00
56.00
36.00
8.00
0.00
Urban
0
0.00
29.27
58.54
7.32
4.88
0.00
AUG2
Natural
0
0.00
8.00
64.00
28.00
0.00
0.00
Urban
0
2.44
24.39
68.29
4.88
0.00
0.00
FOX
Natural
0
0.00
0.00
12.00
44.00
40.00
4.00
0.00
Urban
4.88
0.00
0.00
0.00
53.66
41.46
0.00
4.88
XNL
Natural
0
0.00
8.00
84.00
8.00
0.00
0.00
0.00
Urban
0
0.00
14.63
78.05
2.44
0.00
4.88
0.00
AXO
Natural
0
100.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Urban
0
100.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
FIS
Natural
80
0.00
0.00
16.00
0.00
4.00
80.00
Urban
78.05
0.00
0.00
9.76
9.76
2.44
78.05
SXT
Natural
0
60.00
24.00
12.00
4.00
0.00
0.00
Urban
0
82.93
12.20
2.44
2.44
0.00
0.00
AZI
Natural
0
0.00
0.00
0.00
0.00
8.00
76.00
16.00
0.00
Urban
0
0.00
0.00
0.00
0.00
19.51
65.85
14.63
0.00
AMP
Natural
0
0.00
24.00
64.00
12.00
0.00
0.00
Urban
7.32
4.88
34.15
48.78
4.88
0.00
2.44
4.88
CHL
Natural
0
0.00
4.00
24.00
64.00
8.00
Urban
0
0.00
0.00
14.63
85.37
0.00
CIP
Natural
0
44.00
56.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
Urban
0
78.05
19.51
2.44
0.00
0.00
0.00
0.00
0.00
0.00
NAL
Natural
0
0.00
0.00
0.00
44.00
56.00
0.00
0.00
Urban
0
0.00
0.00
68.29
26.83
4.88
0.00
0.00
TET
Natural
0
100.00
0.00
0.00
0.00
Urban
0
100.00
0.00
0.00
0.00
AM,
antimicrobial:
AMP,
ampicillin
(132
m
g/mL);
AUG2,
amoxicillin-clavulanic
acid
(1/0.532/16
m
g/mL);
AXO,
ceftriaxone
(0.2564
m
g/mL);
AZI,
azithromycin
(0.1216
m
g/mL);
CHL,
chloramphenicol
(232
m
g/mL);
CIP,
ciprooxacin
(0.0154
m
g/mL);
FIS,
sulsoxazole
(16256
m
g/mL);
FOX,
cefoxitin
(0.532
m
g/mL);
GEN,
gentamycin
(0.25
16
m
g/mL);
MIC,
minimum
inhibitory
concentration;
NAL,
nalidixic
acid
(0.532
m
g/mL);
STR,
streptomycin
(3264
m
g/mL);
SXT,
trimethoprim-sulfamethoxazole
(0.12/2.38
4/76
m
g/mL);
XNL,
ceftiofur
(0.128
m
g/mL);
TET,
tetracycline
(432
m
g/mL.
a
Areas
with
white
background
and
numbers
(%)
indicate
the
range
of
dilutions
tested
for
each
antimicrobial.
Areas
with
a
solid
white
background
fall
outside
the
range
of
tested
concentrations.
Numbers
(%)
in
boldface
indicate
the
percentages
of
isolates
with
resistance
measured
on
the
broth
microdilution
plates.
b
%
Resistant
isolates
to
each
of
the
antimicrobials.
580
M.M.
Balbin
et
al.
/
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
4.
Discussion
The
development
and
transmission
of
resistant
bacteria
have
long
been
recognised
[2,8,15],
especially
in
clinical
and
agricultural
settings
where
selective
pressure
and
AMR
determinants
are
expected
to
be
high
[15].
This
study
identied
AMR
determinants
and
virulence
factors
in
Salmonella
spp.
and
E.
coli
from
different
anthropogenic
areas
in
North
Carolina
using
WGS.
S.
Swarzengrund
and
S.
Mississippi
were
the
only
serotypes
detected
in
our
isolates
from
the
soil
samples.
This
might
relate
to
the
ability
and
adaptability
of
some
serotypes
to
survive
outside
their
host
for
an
extended
period.
S.
Schwarzengrund
was
detected
in
our
isolates
from
the
urban
environment.
S.
Schwarzengrund
was
identied
as
one
of
the
frequently
detected
serotypes
in
nonhuman
and
nonclinical
sources
in
the
USA
[16].
There
have
been
reports
of
disease
outbreaks,
food
contamination
[16]
and
Fig.
1.
Phylogenetic
tree
and
genotypic
characteristics
of
E.
coli
isolated
in
the
urban
a
and
natural
b
environments.
The
phylogenetic
tree
was
generated
based
on
single-
nucleotide
polymorphisms
(SNPs)
found
in
the
isolates.
The
E.
coli
str.
K-12
substr.
MG1655
(NC_000913)
was
used
as
a
reference.
The
phylotypes
of
the
isolates
were
determined
using
ClermonTyping
(EnteroBase);
the
virulence
factors,
antimicrobial
resistance
genes
(ARGs),
and
plasmids
were
analysed
using
VirulenceFinder,
ResFinder
and
PlasmidFinder,
respectively.
Fig.
2.
Phylogenetic
relationship
of
fosfomycin
resistance
genes.
The
fosfomycin
resistance
gene
has
been
detected
in
different
species
of
bacteria
from
different
countries.
The
sequences
from
this
study,
FosA7_WGS
and
FosA7_
PCR,
clustered
with
the
FosA7
genes
from
Canada
and
Brazil.
The
fosfomycin
determinant
accession
numbers
are
FosA1,
FJ829469;
FosA2,
EU487198;
FosA3,
AB522970;
FosA4,
AB908992;
FosA5,
KP143090;
FosA6,
KU25459;
FosA7,
LAPJ01000014;
FosA,
MK043330;
FosB,
ABS73480;
FosB3,
NG_050412;
FosC,
DQ112222;
FosC2,
NG_047891;
FosK,
NG_047898;
FosX,
LT795756;
and
FosA7
from
this
study,
SAMN10396967.
M.M.
Balbin
et
al.
/
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
581
multidrug
resistance
associated
with
this
serovar
[17].
Moreover,
we
detected
IncFIB
and
IncFII
plasmids,
which
were
reported
to
be
signicantly
associated
with
S.
Schwarzengrund
from
humans,
animals
and
manure-treated
environments
[17].
All
Salmonella
isolates
contained
virulence
factors
in
SPI-1
and
SPI-2,
which
are
well
known
to
code
for
type
III
protein
secretion
systems
that
lend
pathogenic
Salmonella
the
ability
to
invade
intestinal
cells
[18]
and
proliferate
by
intracellular
replication
[18,19].
The
E.coli isolates predominantly
belonged
to
phylogroups
B1,
B2
and
D,
which
are
known
to
thrive
in
various
ecological
niches,
including
soil
[9].
Several
strains
from
these
phylogroups
are
commensals
and
pathogenic
and
often
have
large
genomes
that
code
for
AMR
determinants
and
virulence
factors,
allowing
them
to
adapt
and
survive
in
different
environments
[5,7,9].
The
plasmids
pCoo
and
pB171
were
detected
in
the
urban
environment;
col(156)
and
pHN7A8
were
identied
in
the
natural
environment.
Both
pCoo
and
pB171
arevirulence
plasmids
associated
with
enterotoxigenic
E.
coli
(ETEC)
and
enteropathogenic
E.
coli
(EPEC),
respectively,
and
are
often
detected
in
cases
of
traveler
and
infantile
diarrhoea
[20].
The
plasmid
col(156)
is
associated
with
extraintestinal
pathogenic
E.
coli
(ExPEC),
an
infectious
strain
affecting
both
humans
and
animals,
particularly
birds
[20].
The
pHN7A8,
a
multiresistance
plasmid
from
E.
coli
of
animal
origin
in
China
and
recently
in
Bolivia,
was
detected
in
our
isolate
from
a
forest.
Although
we
did
not
discover
ARGs,
it
is
essential
to
note
that
this
plasmid
is
classied
as
an
F33:A-:B-
plasmid,
which
is
a
crucial
vector
of
bla
CTX-M-55/-65
,
bla
NDM-1
,
FosA3
and
rmtB
resistance
genes
and
has
the
capability
to
acquire
markers
involved
in
plasmid
replication
or
stability
[20].
E.
coli
has
intrinsic
and
acquired
genes
that
degrade
and
resist
toxic
compounds
such
as
biocides
and
metals
[3,4],
allowing
their
survival
in
the
environment.
The
mdfA
is
a
multidrug
efux
protein,
and
its
overexpression
results
in
resistance
to
several
antimicrobials
and
organic
cations
[3].
The
FosA7
gene
is
a
new
ARG
that
was
detected
from
S.
Heidelberg
from
broiler
chickens
in
Canada
and
was
discovered
in
our
E.
coli
isolate
from
a
residential
yard.
FosA7
confers
resistance
to
fosfomycin,
a
broad-spectrum
antibiotic
that
is
used
to
treat
uncomplicated
urinary
tract
infections
(UTIs)
and
extensively
drug-resistant
(XDR)
Gram-
negative
bacteria.
Fosfomycin
resistance
has
been
reported
in
bacterial
isolates
of
human
and
animal
origins
in
China,
Japan,
France
and,
recently,
the
USA
and
Canada
[21],
but
was
not
reported
in
isolates
from
the
environment.
Most
antimicrobials
being
used
today
were
initially
isolated
from
the
natural
environment,
particularly
from
soil
Actinomyces
spp.
[68].
The
antimicrobial-producing
organisms
have
determi-
nants
that
would
help
resist
the
action
of
the
antimicrobial(s)
they
produce,
along
with
the
other
microbes
found
in
the
same
environment
[3,6,7].
Therefore,
it
is
not
surprising
to
detect
AMR
determinants
in
the
environment
[3].
However,
the
proximity
of
humans
and
their
activities
in
the
environment
greatly
inuence
the
dynamics
in
the
microbial
community,
genetic
variation,
resistance
selection
and
possible
emergence
of
novel
mechanisms
of
resistance
[7].
In
this
study,
we
found
a
higher
prevalence
of
both
Salmonella
spp.
and
E.
coli
in
the
urban
environment.
The
AMR
pattern
based
on
the
results
of
AST
was
also
observed
to
be
more
diverse
among
isolates
from
the
urban
environment.
This
can
be
explained
by
the
continuous
exposure
of
urban
environments
to
different
anthropogenic
activities,
making
it
more
vulnerable
to
changes
that
may
shape
the
composition
of
the
bacterial
community
and
AMR
determinants.
The
presence
of
human
activities
in
an
environment
increases
the
chance
of
contamination
[1,9],
such
as
spillage
of
antimicrobials,
heavy
metals,
biocides
and
even
resistant
bacteria
[1,3,7].
Such
contami-
nation
creates
a
selective
pressure
or
environmental
hotspot
for
the
development
and
dissemination
of
AMR
[1,5].
Salmonella
spp.
and
E.
coli
are
commensal
bacteria
in
humans
and
animals,
but
these
bacteria
are
also
well
adapted
in
the
natural
environment.
Aside
from
the
presence
of
antimicrobial-
producing
microorganisms,
the
role
of
wildlife
was
also
cited
in
regard
to
the
dissemination
of
resistance
and
pathogenic
bacteria
[3].
Several
reports
of
AMR
contamination
in
the
natural
environments
have
been
associated
with
wildlife,
such
as
migratory
birds
and
foxes
[2,3,7].
It
is
essential
to
understand
the
occurrence
and
develop-
ment
of
AMR
in
the
environment
because
there
has
been
a
continuous
rise
of
resistant
and
pathogenic
bacteria.
The
presence
of
these
determinants
in
different
anthropogenic
areas
suggests
the
need
for
constant
surveillance.
Accession
numbers
The
paired-end
reads
used
in
this
study
were
deposited
in
the
National
Center
for
Biotechnology
Information
(NCBI)
under
the
Bioproject
accession
numbers
PRJNA293224
and
PRJNA293225
for
Salmonella
spp.
and
E.
coli
isolates,
respectively.
Funding
This
project
was
supported
by
the
Duke
Triangle
Center
for
Evolutionary
Medicine
(TriCEM).
The
whole-genome
sequencing
was
completed
by
the
FDAGenomeTrakr
program
funded
grant
1U18FD00678801.
Ethical
approval
This
was
not
required.
Competing
interests
All
authors
declare
no
conict
of
interest.
Acknowledgements
The
authors
acknowledge
the
support
and
assistance
of
the
research
staff
and
students
in
the
Thakur
Laboratory
and
Rob
Dunn
Laboratory,
North
Carolina
State
University,
Raleigh,
NC,
USA.
Appendix
A.
Supplementary
data
Supplementary
material
related
to
this
article
can
be
found,
in
the
online
version,
at
doi:https://doi.org/10.1016/j.jgar.2020.05.016.
References
[1]
Berendonk
TU,
Manaia
CM,
Merlin
C,
Fatta-Kassinos
D,
Cytryn
E,
Walsh
F,
et
al.
Tackling
antibiotic
resistance:
the
environmental
framework.
Nat
Rev
Microbiol
2015;13:3107.
[2]
Huijbers
PMC,
Blaak
H,
de
Jong
MCM,
Graat
EAM,
Vandenbroucke-Grauls
CMJE,
Husman
AM.
Role
of
the
environment
in
the
transmission
of
antimicrobial
resistance
to
humans:
a
review.
Environ
Sci
Technol
2015;49:119932004.
[3]
BengtssonPalme
J,
Kristiansson
E,
Larsson
DGJ.
Environmental
factors
in
inuencing
the
development
and
spread
of
antibiotic
resistance.
FEMS
Microbiol
Rev
2018;fux53:6880.
[4]
ONeill
J.
Antimicrobial
resistance:
tackling
a
crisis
for
the
health
and
wealth
of
nations.
Review
on
antimicrobial
resistance.
London:
Wellcome
Trust;
2014.
[5]
Gyles
C,
Boerlin
P.
Horizontally
transferred
genetic
elements
and
their
role
in
pathogenesis
of
bacterial
disease.
Vet
Pathol
2014;51:32840.
[6]
Aminov
RI.
A
brief
history
of
the
antibiotic
era:
lessons
learned
and
challenges
for
the
future.
Front
Microbiol
2010;1:17.
[7]
Martinez
JL.
Environmental
pollution
by
antibiotic
resistance
determinants.
Environ
Pollut
2009;157:2893902.
[8]
Larsson
DGJ,
Andremont
A,
Bengtson-Palme
J,
Brandt
KK,
Husman
AM,
Fegerstedt
P,
et
al.
Critical
knowledge
gaps
and
research
needs
related
to
the
environmental
dimensions
of
antibiotic
resistance.
Environ
Int
2018;117:1328.
[9]
Elsas
JD,
Semenov
AV,
Costa
R,
Trevors
JT.
Survival
of
Escherichia
coli
in
the
environment:
fundamentals
and
public
health
aspects.
ISME
J
2011;5:17383.
582
M.M.
Balbin
et
al.
/
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
[10]
Pornsukarom
S,
Thakur
S.
Assessing
the
impact
of
manure
application
in
commercial
swine
farms
in
the
transmission
of
antimicrobial
resistant
Salmonella
in
the
environment.
PLOS
One
2016;11:e0164621.
[11]
US
Food
and
Drug
Administration.
The
National
Antimicrobial
Resistance
Monitoring
System
(NARMS)
manual
of
laboratory
methods.
Rockville,
MD:
US
Department
of
Health
and
Human
Services;
2016.
[12]
Gutierez-Rodriguez
E,
Gundersen
A,
Sbodio
AO,
Suslow
TV.
Variable
agronomic
practices,
cultivar,
strain
source,
and
initial
contamination
dose
differentially
affect
survival
of
Escherichia
coli
on
spinach.
J
Appl
Microbiol
2011;112:10918.
[13]
Fratamico
PM,
DebRoy
C,
Liu
Y.
The
DNA
sequence
of
the
Escherichia
coli
O22
antigen
gene
cluster
and
detection
of
pathogenic
strains
belonging
to
E.
coli
serogroups
O22
and
O91by
multiplex
PCR
assays
targeting
virulence
genes
and
genes
in
the
respective
Oantigen
gene
clusters.
Food
Anal
Methods
2009;2:16979.
[14]
Yang
J.
Virulence
factors
of
pathogenic
bacteria.
2020.
http://www.mgc.ac.cn/
cgi-bin/VFs/vfs.cgi?Genus=Salmonella&Keyword=Toxin.
[15]
Maz
AI,
Perera
LN,
He
Y,
Zhang
W,
Xiao
S,
Hao
W,
et
al.
Case
study
on
the
soil
antibiotic
resistome
in
an
urban
community
garden.
Int
J
Antimicrob
Agents
2018;52:24150.
[16]
Akiyama
T,
Khan
AA.
Molecular
characterization
of
strains
of
uoroquinolone-
resistant
Salmonella
enterica
serovar
Schwarzengrund
carrying
multidrug
resistance
isolated
from
imported
foods.
J
Antimicrob
Chemother
2012;67:10110.
[17]
Pornsukarom
S,
van
Vliet
AHM,
Thakur
S.
Whole
genome
sequencing
analysis
of
multiple
Salmonella
serovars
provides
insights
into
phylogenetic
relatedness,
antimicrobial
resistance,
and
virulence
markers
across
humans,
food
animals,
and
agriculture
environmental
sources.
BMC
Genomics
2018;19:801.
[18]
Lou
L,
Zhang
P,
Piao
R,
Wang
Y.
Salmonella
Pathogenicity
Island
1
(SPI-1)
and
its
complex
regulatory
network.
Front
Cell
Infect
Microbiol
2019;9:270.
[19]
Jennings
E,
Thurston
TL,
Holden
DW.
Salmonella
SPI-2
type
III
secretion
system
effectors:
molecular
mechanisms
and
physiological
consequences.
Cell
Host
Microbe
2017;9:21731.
[20]
Johnson
TJ,
Nolan
LK.
Pathogenomics
of
the
virulence
plasmids
of
Escherichia
coli.
Microbiol
Mol
Biol
Rev
2009;73:75074.
[21]
Rehman
MA,
Yin
X,
Persaud-Lachmann
MG,
Diara
MS.
First
detection
of
Fosfomycin
resistance
gene,
FosA7,
in
Salmonella
enterica
serovar
Heidelberg
isolated
from
broiler
chickens.
Antimicrob
Agents
Chemother
2017;61:
e004107.
M.M.
Balbin
et
al.
/
Journal
of
Global
Antimicrobial
Resistance
22
(2020)
578583
583
... This is somewhat expected when referring to the gut microbiota or the hospital environment, due to the regular use of antibiotics (Kolář et al., 2001;Sib et al., 2020;Witte, 2000). However, in the past years, an increased AMR has been observed in bacteria isolated from natural environments (water bodies, soil, wildlife, etc.) (Balbin et al., 2020;Marinho et al., 2016;Sourenian et al., 2020;Tello et al., 2012). ...
... Environmental achievement of AMR has been monitored through identification of ARGs in water bodies worldwide (Arsand et al., 2020;Balbin et al., 2020;Böger et al., 2021;Liu et al., 2020;Voigt et al., 2020;Xu et al., 2020a). In some cases, the identified ARGs are well correlated with the antibiotic concentrations detected in the environment. ...
Article
Large amounts of antibiotics are produced and consumed worldwide, while wastewater treatment is still rather inefficient, leading to considerable water contamination. Concentrations of antibiotics in the environment are often sufficiently high to exert a selective pressure on bacteria of clinical importance that increases the prevalence of resistance. Since the drastic reduction in the use of antibiotics is not envisaged, efforts to reduce their input into the environment by improving treatment of contaminated wastewater is essential to limit uncontrollable spread of antibiotic resistance. This paper reviews recent progress on the use of non-thermal plasma for the degradation of antibiotics in water. The target compounds removal, the energy efficiency and the mineralization are analyzed as a function of discharge configuration and the most important experimental parameters. Various ways to improve the plasma process efficiency are addressed. Based on the identified reaction intermediates, degradation pathways are proposed for various classes of antibiotics and the degradation mechanisms of these chemicals under plasma conditions are discussed.
... The fosA7 is a new antimicrobial resistance gene against fosfomycin that was recently identified in S. Heidelberg from broiler chickens in Canada (Rehman et al., 2017). The gene fosA7 confers resistance to broad-spectrum antibiotic fosfomycin, which is extensively used to treat drug-resistant Gram-negative bacteria (Balbin et al., 2020). A total of 58.67% of the tested S. Telelkebir strains (71 strains) harbored the fosA7 gene. ...
Article
Full-text available
Non-typhoidal Salmonella (NTS) is a common cause for self-limiting gastroenteritis, representing a public health concern globally. NTS is one of the leading causes of foodborne illnesses in China; however, the invasive infection caused by NTS is largely underappreciated. Here, we reported an NTS invasive infection caused by an infrequently reported serovar Telelkebir (13,23:d:e,n,z15) strain FJ001 in China, which carries antimicrobial-resistant genes [fosA7 and aac(6′)-Iaa] and typhoid-toxin genes (cdtB, pltA, and pltB). By conducting the whole genomic sequencing, we also investigated the relatedness of this strain with an additional 120 global contextual Salmonella enterica serovar Telelkebir (S. Telelkebir) isolates, and assessed the antimicrobial-resistant determinants and key virulence factors using the available genomic dataset. Notably, all 121 (100%) of the S. Telelkebir strains possessed the typhoid toxin genes cdtB, pltA, and pltB, and 58.67% (71/121) of S. Telelkebir harbored antimicrobial-resistant gene fosaA7. The study by core genome multilocus sequence typing (cgMLST) and core single-nucleotide polymorphism (SNP)-based phylogenomic analysis demonstrated that the S. Telelkebir isolates from different sources and locations clustered together. This suggests that regular international travels might increase the likelihood of rapid and extensive transmissions of potentially pathogenic bacteria. For the first time, our study revealed the antimicrobial resistance, virulence patterns, and genetic diversity of the serovar S. Telelkebir isolate in humans and similar isolates over the world. The present study also suggests that genomic investigation can facilitate surveillance and could offer added knowledge of a previously unknown threat with the unique combination of virulent and antimicrobial-resistant determinants.
... Earlier, fosA7 was described in Salmonella isolated from broiler chickens in Canada, as well as from retail meat and clinical incidents in the United States (Rehman et al., 2017;Keefer et al., 2019). The gene was also noted in E. coli recovered from soil exposed to anthropogenic activities in North Carolina (Balbin et al., 2020). In Europe, the presence of fosfomycin resistance gene fosA3 in a Salmonella isolated from the migratory bird black kite was reported in Germany (Villa et al., 2015). ...
Article
Full-text available
Antimicrobial resistance (AMR) is one of the most important global health concerns; therefore, the identification of AMR reservoirs and vectors is essential. Attention should be paid to the recognition of potential hazards associated with wildlife as this field still seems to be incompletely explored. In this context, the role of free-living birds as AMR carriers is noteworthy. Therefore, we applied methods used in AMR monitoring, supplemented by colistin resistance screening, to investigate the AMR status of Escherichia coli from free-living birds coming from natural habitats and rescue centers. Whole-genome sequencing (WGS) of strains enabled to determine resistance mechanisms and investigate their epidemiological relationships and virulence potential. As far as we know, this study is one of the few that applied WGS of that number ( n = 71) of strains coming from a wild avian reservoir. The primary concerns arising from our study relate to resistance and its determinants toward antimicrobial classes of the highest priority for the treatment of critical infections in people, e.g., cephalosporins, quinolones, polymyxins, and aminoglycosides, as well as fosfomycin. Among the numerous determinants, bla CTX–M–15 , bla CMY–2 , bla SHV–12 , bla TEM–1B , qnrS1 , qnrB19 , mcr-1 , fosA7 , aac(3)-IIa , ant(3”)-Ia , and aph(6)-Id and chromosomal gyrA , parC , and parE mutations were identified. Fifty-two sequence types (STs) noted among 71 E. coli included the global lineages ST131, ST10, and ST224 as well as the three novel STs 11104, 11105, and 11194. Numerous virulence factors were noted with the prevailing terC , gad , ompT , iss , traT , lpfA , and sitA . Single E. coli was Shiga toxin-producing. Our study shows that the clonal spread of E. coli lineages of public and animal health relevance is a serious avian-associated hazard.
Article
Full-text available
Salmonella species can infect a diverse range of birds, reptiles, and mammals, including humans. The type III protein secretion system (T3SS) encoded by Salmonella pathogenicity island 1 (SPI-1) delivers effector proteins required for intestinal invasion and the production of enteritis. The T3SS is regarded as the most important virulence factor of Salmonella. SPI-1 encodes transcription factors that regulate the expression of some virulence factors of Salmonella, while other transcription factors encoded outside SPI-1 participate in the expression of SPI-1-encoded genes. SPI-1 genes are responsible for the invasion of host cells, regulation of the host immune response, e.g., the host inflammatory response, immune cell recruitment and apoptosis, and biofilm formation. The regulatory network of SPI-1 is very complex and crucial. Here, we review the function, effectors, and regulation of SPI-1 genes and their contribution to the pathogenicity of Salmonella.
Article
Full-text available
Background Salmonella enterica is a significant foodborne pathogen, which can be transmitted via several distinct routes, and reports on acquisition of antimicrobial resistance (AMR) are increasing. To better understand the association between human Salmonella clinical isolates and the potential environmental/animal reservoirs, whole genome sequencing (WGS) was used to investigate the epidemiology and AMR patterns within Salmonella isolates from two adjacent US states. Results WGS data of 200 S. enterica isolates recovered from human (n = 44), swine (n = 32), poultry (n = 22), and farm environment (n = 102) were used for in silico prediction of serovar, distribution of virulence genes, and phylogenetically clustered using core genome single nucleotide polymorphism (SNP) and feature frequency profiling (FFP). Furthermore, AMR was studied both by genotypic prediction using five curated AMR databases, and compared to phenotypic AMR using broth microdilution. Core genome SNP-based and FFP-based phylogenetic trees showed consistent clustering of isolates into the respective serovars, and suggested clustering of isolates based on the source of isolation. The overall correlation of phenotypic and genotypic AMR was 87.61% and 97.13% for sensitivity and specificity, respectively. AMR and virulence genes clustered with the Salmonella serovars, while there were also associations between the presence of virulence genes in both animal/environmental isolates and human clinical samples. Conclusions WGS is a helpful tool for Salmonella phylogenetic analysis, AMR and virulence gene predictions. The clinical isolates clustered closely with animal and environmental isolates, suggesting that animals and environment are potential sources for dissemination of AMR and virulence genes between Salmonella serovars. Electronic supplementary material The online version of this article (10.1186/s12864-018-5137-4) contains supplementary material, which is available to authorized users.
Article
Full-text available
There is growing understanding that the environment plays an important role both in the transmission of antibiotic resistant pathogens and in their evolution. Accordingly, researchers and stakeholders world-wide seek to further explore the mechanisms and drivers involved, quantify risks and identify suitable interventions. There is a clear value in establishing research needs and coordinating efforts within and across nations in order to best tackle this global challenge. At an international workshop in late September 2017, scientists from 14 countries with expertise on the environmental dimensions of antibiotic resistance gathered to define critical knowledge gaps. Four key areas were identified where research is urgently needed: 1) the relative contributions of different sources of antibiotics and antibiotic resistant bacteria into the environment; 2) the role of the environment, and particularly anthropogenic inputs, in the evolution of resistance; 3) the overall human and animal health impacts caused by exposure to environmental resistant bacteria; and 4) the efficacy and feasibility of different technological, social, economic and behavioral interventions to mitigate environmental antibiotic resistance.1.
Article
Full-text available
Antibiotic resistance and its wider implications present us with a growing healthcare crisis. Recent research points to the environment as an important component for the transmission of resistant bacteria and in the emergence of resistant pathogens. However, a deeper understanding of the evolutionary and ecological processes that lead to clinical appearance of resistance genes is still lacking, as is knowledge of environmental dispersal barriers. This calls for better models of how resistance genes evolve, are mobilized, transferred and disseminated in the environment. Here, we attempt to define the ecological and evolutionary environmental factors that contribute to resistance development and transmission. Although mobilization of resistance genes likely occurs continuously, the great majority of such genetic events do not lead to the establishment of novel resistance factors in bacterial populations, unless there is a selection pressure for maintaining them or their fitness costs are negligible. To enable preventative measures it is therefore critical to investigate under what conditions and to what extent environmental selection for resistance takes place. In addition, understanding dispersal barriers is not only key to evaluate risks, but also to prevent resistant pathogens, as well as novel resistance genes, from reaching humans.
Article
Full-text available
Land application of swine manure in commercial hog farms is an integral part of their waste management system which recycles the nutrients back to the soil. However, manure application can lead to the dissemination of bacterial pathogens in the environment and pose a serious public health threat. The aim of this study was to determine the dissemination of antimicrobial resistant Salmonella in the environment due to manure application in commercial swine farms in North Carolina (n = 6) and Iowa (n = 7), two leading pork producing states in the US. We collected manure and soil samples twice on day 0 (before and after manure application) from four distinct plots of lands (5 soil samples/plot) located at 20 feet away from each other in the field. Subsequent soil samples were collected again on days 7, 14, 21 from the same plots. A total of 1,300 soil samples (NC = 600; IA = 700) and 130 manure samples (NC = 60; IA = 70) were collected and analyzed in this study. The overall Salmonella prevalence was 13.22% (189/1,430), represented by 10.69% and 38.46% prevalence in soil and manure, respectively. The prevalence in NC (25.45%) was significantly higher than in IA (2.73%) (P
Article
Urban agricultural soils can be an important reservoir of antibiotic resistance and have great food safety and public health indications. This study was to investigate antibiotic-resistant bacteria and antibiotic resistance genes in urban agricultural soils using phenotypic and metagenomic tools. A total of 207 soil bacteria were recovered from 41 soil samples collected from an urban agricultural garden in Detroit, USA. The most prevalent antibiotic resistance phenotypes demonstrated by Gram-negative bacteria was the resistance to ampicillin (94.2%), followed by chloramphenicol (80.0%), cefoxitin (79.5%), gentamicin (78.4%), and ceftriaxone (71.1%). Gram-positive bacteria were all resistant to gentamicin, kanamycin, and penicillin. Genes encoding resistance to quinolone, β-lactam, and tetracycline were the most prevalent and abundant in the soil. qepA and tetA, both encoding efflux pumps, predominated in quinolone and tetracycline resistance genes tested, respectively. Positive correlation (p < 0.05) was identified among groups of antibiotic resistance genes and between antibiotic resistance genes and metal resistance genes. The data demonstrated a diverse population of antibiotic resistance in urban agricultural soils. Phenotypic determination together with soil metagenomics proved to be a valuable tool to study the nature and extent of antibiotic resistance in the environment.
Article
Serovars of Salmonella enterica cause both gastrointestinal and systemic diseases in a broad range of mammalian hosts, including humans. Salmonella virulence depends in part on its pathogenicity island 2 type III secretion system (SPI-2 T3SS), which is required to translocate at least 28 effector proteins from vacuolar-resident bacteria into host cells. Comparative genomic analysis reveals that all serovars encode a subset of “core” effectors, suggesting that they are critical for virulence in different hosts. An additional subset of effectors is found sporadically throughout different serovars, and several inhibit activation of the innate immune system. In this Review, we summarize the biochemical activities, host cell interaction partners, and physiological functions of SPI-2 T3SS effectors in the context of the selective pressures encountered by S. enterica in vivo. We also consider some of the remaining challenges to achieve a unified understanding of how effector activities work together to promote Salmonella virulence. The Salmonella SPI-2 type III secretion system transfers a large number of effector proteins to host cells. In this Review, Holden and colleagues summarize the biochemical activities, host cell interaction partners, and physiological functions of these effectors in the context of the selective pressures encountered by S. enterica in vivo.
Article
To establish a possible role for the natural environment in the transmission of clinically relevant AMR bacteria to humans, a literature review was conducted to systematically collect and categorize evidence for human exposure to extended-spectrum β-lactamase-producing Enterobacteriaceae, methicillin-resistant Staphylococcus aureus, and vancomycin-resistant Enterococcus spp. in the environment. In total, 239 datasets adhered to inclusion criteria. AMR bacteria were detected at exposure-relevant sites (35/38), including recreational areas, drinking water, ambient air, shellfish, and in fresh produce (8/16). More datasets were available for environmental compartments (139/157), including wildlife, water, soil, and air/dust. Quantitative data from exposure-relevant sites (6/35), and environmental compartments (11/139) were scarce. AMR bacteria were detected in the contamination sources (66/66) wastewater and manure, and molecular data supporting their transmission from wastewater to the environment (1/66) were found. The abundance of AMR bacteria at exposure-relevant sites suggests risk for human exposure. Of publications pertaining to both environmental and human isolates, however, only one compared isolates from samples that had a clear spatial and temporal relationship, and no direct evidence was found for transmission to humans through the environment. To what extent the environment, compared to the clinical and veterinary domains, contributes to human exposure needs to be quantified. AMR bacteria in the environment, including sites relevant for human exposure, originate from contamination sources. Intervention strategies targeted at these sources could therefore limit emission of AMR bacteria to the environment.