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Department of Veterinary Clinical Sciences, Leahurst, University of Liverpool, United Kingdom
MRSA AND ANTIBIOTIC-RESISTANT
STAPHYLOCOCCI IN DOGS AND HORSES
Thomas W. Maddox BVSc CertVDI MRCVS
BACKGROUND
Antimicrobial-resistant bacteria (especially MRSA) are
recognised as an important and increasing problem in both
human and veterinary medicine
BACKGROUND
Antimicrobial-resistant bacteria (especially MRSA) are
recognised as an important and increasing problem in both
human and veterinary medicine



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Infections resulting from resistant organisms increasingly reported
Limited range of antibiotics available for animal use (especially for
horses)
New antibiotics likely slower in development (especially for horses)
Zoonotic potential of antibiotic resistant bacteria
Future restrictions on antibiotic use possible?
BACKGROUND
Antimicrobial-resistant bacteria (especially MRSA) are
recognised as an important and increasing problem in both
human and veterinary medicine
BACKGROUND
Antimicrobial-resistant bacteria (especially MRSA) are
recognised as an important and increasing problem in both
human and veterinary medicine
BACKGROUND


228 cases reported of MRSA in horses
Variety of sites of infection (similar to other animals and humans)
Figure courtesy of C. Scantlebury
BACKGROUND


228 cases reported of MRSA in horses
Variety of sites of infection (similar to other animals and humans)
STAPHS IN ANIMALS
Staphyloccus aureus

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Most common Staph. of people with 28-39% nasal colonisation (also
throat, axilla, perineum and groin)
Unknown prevalence in animals but likely lower
Other staphylococci
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
S. (pseud)intermedius in dogs, coagulase-negatives (S. epidermidis)
Coagulase-negatives more common in horses (S. scuiri)
80% of Staphylococcus aureus strains produce a
b-lactamase enzyme

Methicillin-resistance mediated by production of alternative penicillin
binding protein (PBP2a) which is the normal target for b-lactam
antibiotics
RESISTANCE IN STAPHS
SCCmec cassette



Gene cassette (mobile genetic element)
Codes for PBP2a on the mecA gene (plus several other genes).
7 types of varying size and composition (and sub-types)
MRSA TYPING
Macro restriction pulsed-field gel electrophoresis


Chromosomal DNA extracted and then digested into many (large)
fragments by restriction enzymes (Sma1)
Band pattern of various sizes produced on PF gel
Compare bands to
identify MRSA type
Gold standard for strain
typing MRSA
BUT... inter-laboratory
comparison difficult
MRSA TYPING
Multi-locus sequence typing


PCR amplification of housekeeping gene fragments at 7 loci on
chromosome
These then sequenced and submitted to database to compared with
known alleles
Each isolate will then have a 7
integer “profile” which can be
used to identify its sequence
type (eg 4-7-6-1-5-8-8-6).


Sequence types (ST) named arbitrarily
Grouped into clonal complexes (CC)
named after first ST identified (eg
ST30-MRSA was first member of CC30)
MRSA TYPING
spa gene typing


Amplification and sequencing of a single gene loci
Target is region X of the (serum) protein A gene

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Region X has varying number of 24 base pair repeats
Highly polymorphic between MRSA types
From size of fragments produced by PCR can estimate number of
repeats (not actually interested in sequence itself)
Normally compared with MLST-types as easier way to identify ST
MRSA TYPING
SCCmec typing


PCR amplification of several genes within the
SCCmec to differentiate between types and allow
sub-typing
Simple band pattern produced on gel
Variable Number Tandem Repeat Typing

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Regions of short repeating sequences of differing length found at
various loci throughout chromosome
Amplified by PCR and run on gel to produce banding pattern
PCR of mecA and femA/nuc genes

PCR amplify these genes for identification/molecular confirmation of
methicillin-resistance and S. aureus respectively
MRSA IN ANIMALS
Epidemiology
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Small animals MRSA isolates in UK are mostly EMRSA 15 (same as
predominant health care strain)1,2
EMRSA type 16 seen to a lesser extent
Mostly SCCmec IV
Same types seen in small animal vets and owners.
MRSA carriage in vets approx 10% (10 x public)
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Equine isolates are more variable, rarely EMRSA 15 or 16 and have
greater range of reistance3
In-contact humans may have same types (unusual for humans)
Rarely other than SCCmec IV4
1.
2.
3.
4.
Baptiste, K.E. et al (2005) Emerg Inf Diseases, 11(12,)
Loeffler, A., et al(2005) J Antimicrobial Chemotherapy, 56(4)
Cuny, C., et al (2008) Microbial Drug Resistance, 14(4)
Weese, J.S. (2007) Vet Rec, 161(10)
MRSA IN ANIMALS
Epidemiology

Difference of colonisation versus clinical infection
Also transient carriage vs persistent colonisation
MRSA IN HORSES
Epidemiology
 Varying prevalence of nasal colonisation reported of 0-12% for
horses in the community1-3
Colonisation of hospitalised/unwell horses ranges
from 5.3-16%4,5 (3.5% at PLEH)
 Resistance patterns variable (usually gentamicin, sometimes
tetracyclines and/or TMS, occasionally fluoroquinolones)
 Usually spa or MLST typed as belonging to CC8 (eg ST8 or
ST254)
 In-contact humans may have same types (unusual for humans)
1.
2.
3.
4.
5.
Burton et al (2008) Can. Vet. J. 49(8)
Vengust et al (2006) Let Appl Microbiology 43(6)
Weese (2005) JAVMA 226(4)
Van den Eede et al (2009) Vet. Microbiology 133(1-2)
Weese et al (2006) JVIM 20(1)
MRSA IN DOGS
Epidemiology
 MRSA colonisation has not been identified in healthy dogs in the
community1-2
Colonisation of hospitalised/unwell dogs reported at
9-23%3,4 (3% at SATH)
 Resistance patterns more consistent (usually fluoroquinolones,
occasionally tetracyclines and/or TMS, rarely gentamicin)
 SCCmec type IV (occasionally type II or V5)
 Usually spa or MLST typed as ST22 or ST36
 In-contact humans may have similar types (common to humans)
1.
2.
3.
4.
5.
Murphy et al (2005) J Vet Int Med. 19
Bagcigal et al (2007) Vet Microbiol 121 (3-4)
Loeffler (2005) J Antimicr Chemotherapy 56 (4)
Baptiste (2005) Emerg Inf Disease
Witte et al 2007 Emerg Inf Disease 13 (2)
MRSA IN ANIMALS
ST398-MRSA
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New “untypeable” (spa t011) strain of MRSA first identified in 2005
Cause of disease in humans and appears readily transferred from
animals (pig farmers in Netherlands)
Recently reported causing disease in horses1-3 and dogs4
Some
human
cases
reported in UK (Scotland)
None reported in dogs,
just reported in 2 horses
from UK5
1.
2.
3.
4.
5.
Van den Eede et al (2009) Vet. Microbiology 133(1-2)
Cuny et al (2008) Microbial Drug Resistance14(4)
Hermans et al (2008) Vlaams Dierg Tijdschrift 77(6)
Witte et al 2007 Emerg Inf Disease 13 (2)
Loeffler et al (2009) Hosp Inf Soc 72 (3)
CA-MRSA
Community associated MRSA
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MRSA but without known risk factors (immunosuppressed/
hospitalisation/antibiotics etc)
Not nosocomial like HA-MRSA.
Carry smaller SCCmec types IV and V (hence survival?)
Different antibiotic susceptibility (resistance to fewer antibiotics- often
just b-lactams)
Frequently produce PVL toxin
More virulent

Small animal prevalence unknown but has been reported1, not
definitively identified in horses2
1.
2.
Rankin et al (2005) Vet Microbiol 108
Maeda et al (2007) Vet Rec 161
MRSA AS A ZOONOSIS
Zoonotic Potential

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Transmission between people and animals has been reported
Usually from humans to animals (unsurprising given respective
prevalence) 1-3
Some cases of animals transmitting to people4,5
Co-colonisation of animals and in-contacts may be
relatively common but actual cross-infection seems to
require normal risk factors
1.
2.
3.
4.
5.
Rutland et al (2009) Emerg Inf Disease 15 (8)
van Duijkeren (2005) J Clin Microbiol 43 (12)
van Duijkeren (2004) Emerg |Inf Disease 10 (12)
Sing (2008) New Eng J Med 358 (11)
Weese et al (2006) Vet Microbiol 114
MRSA AS A ZOONOSIS
Zoonotic Potential
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Transmission between people and animals has been reported
Usually from humans to animals (unsurprising given respective
prevalence) 1-3
Some cases of animals transmitting to people4,5
Survey of 274 veterinary personnel at equine conference in 20066
 22 people identified with nasal carriage of MRSA (8.0%)
 9 isolates typical human strains
 Remainder were non-human strains more commonly seen in horses
(EMRSA-10, ST8, ST254)
6.
Scantlebury (2007) BEVA Conf Proceedings
STUDIES AT LIVERPOOL


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
Nationwide
cross-sectional
study
on
the
microbiology and epidemiology of antimicrobialresistant E. coli and staphylococci in dogs and
horses
Longitudinal study on the microbiology and
epidemiology of antimicrobial-resistant E. coli in
horses in the community
All studies on-going currently
Some preliminary results will be summarised
CROSS-SECTIONAL STUDY
Study Design


Animals seen by vets from 65 equine
and 87 small animal randomly selected
veterinary practices across the UK
Nasal swab obtained from each animal,
with owner completed questionnaire on
veterinary history and
treatment,
housing and management
Majority of animals (88%)
seen for routine reasons
CROSS-SECTIONAL STUDY
Microbiological Methods
Nasal swabs enriched in 6% NaCl nutrient broth.
Streaked onto mannitol salt agar (MSA) and oxacillinresistance screening agar (ORSA)
Typical isolates selected and characterised by Gram
stain, catalase, coagulase and stapylase testing
Staphylococci subjected to antibiotic susceptibility
testing in accordance with BSAC guidelines 1.
mecA PCR for methicillin-resistance confirmation and
femA and nuc PCR.
(MRSA isolates: SCCmec typing and spa gene typing)
1.
Antimicrobial
Potency
Ciprofloxacin (CIP)
1mg
Fusidic acid (FS)
10mg
Gentamicin (GM)
10mg
Methicillin (MET)
5mg
Mupirocin (MUP)
5mg
Rifampicin (RIF)
2mg
Tetracycline (TET)
30mg
Co-trimoxazole(TMS)
2.5mg
Teicoplanin (TEC)
30mg
Vanocmycin (VAN)
5mg
British Society for Antimicrobial Chemotherapy (2007), Methods for Antimicrobial
Susceptibility Testing
STAPHYLOCOCCI IN HORSES
Nasal samples from 677 horses returned
 617 horses (91.1%; 89.0-93.3%) had at least one Staphylococcus spp
 215 horses (31.8%; 28.3-35.3%) had at least one methicillin-resistant
Staphylococcus (mostly coagulase-negative)

Low prevalence of MRSA (0.6%; 0.0-1.2%)
STAPHYLOCOCCI IN DOGS
Nasal samples from 672 dogs returned
 394 dogs (58.6%; 54.9-62.4%) had at least one Staphylococcus spp
 42 dogs (5.3%; 3.7-7.1%) had at least one methicillin-resistant
Staphylococcus (more coagulase-negative)
Data

Low MRSA prevalence of 6 dogs (0.9%; 0.2-1.6%)
courtesy
Wedley
of
A.
STAPHYLOCOCCI IN HORSES
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Varying levels of resistance to all antibiotics except teicoplanin and
vancomycin
High levels of resistance to fusidic acid, as well as to mupirocin,
tetracycline and co-trimoxazole
78.4% (71.4-82.6%) of MR-staphylococci were multidrug resistant
MRSA IN HORSES

4 confirmed MRSA isolates (mecA, fem and nuc positive)

Variable resistance pattern seen in the four isolates (isolate 060
slightly unusual pattern for horse MRSA)
All multidrug-resistant (to three or more antimicrobial classes)

Isolate
Cip
F. Acid
GM
Meth
Mup
Rif
Tet
TMS
Teic
Vanc
060
R
R
S
R
S
S
S
S
S
S
206
S
R
R
R
S
R
R
R
S
S
846
S
R
S
R
R
R
S
S
S
S
953
S
R
R
R
S
R
R
R
S
S
MRSA IN HORSES
 All 4 equine isolates confirmed MRSA SCCmec
type IV by PCR
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
spa typing carried out for all isolates (one failed)
Isolate
spa type
Presumptive
Sequence Type
Presumptive
Clonal Complex
060
unidentified
N/A
N/A
206
t064
ST8
CC8
846
t064
ST8
CC8
953
t451
?ST8
?CC8
spa types represent common equine strains (no ST398 identified)
CONCLUSIONS

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
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Animals can carry antibiotic-resistant staphylococci
Carriage of MRSA appears rare, especially in animals in the general
community
The epidemiology of MRSA carriage is distinct in different species
-Dogs carry common (local) human epidemic strains
-Horses carry strains uncommon in humans
Animal-related MRSA strains can be found in humans in close contact
with animals
Transmission may occur both ways between animals and humans
Companion animals may act as a reservoir for infection of humans in
close contact
AKNOWLEDGEMENTS
PhD colleague
 Amy Wedley
MSc Student
 Andrew O’Donnell
Ph.D Supervisors
 Dr. Nicola Williams
 Prof. Pete Clegg
 Dr. Gina Pinchbeck
 Dr. Susan Dawson
 Dr. Tim Nuttall
Colleagues in the lab…
 Ruth Ryvar
 Gill Hutchinson
Antimicrobial resistance in companion
animals project
 DEFRA
Bransby Home of Rest for Horses
 PhD funding