MRSA AND ANTIBIOTICRESISTANT STAPHYLOCOCCI IN DOGS AND HORSES

1
MRSA AND ANTIBIOTIC-RESISTANT STAPHYLOCOCCI IN
DOGS AND HORSES
Department of Veterinary Clinical Sciences,
Leahurst, University of Liverpool, United Kingdom

• Thomas W. Maddox BVSc CertVDI MRCVS

2
BACKGROUND

• Antimicrobial-resistant bacteria (especially
  MRSA) are recognised as an important and
  increasing problem in both human and veterinary
  medicine

3
BACKGROUND

• Antimicrobial-resistant bacteria (especially
  MRSA) are recognised as an important and
  increasing problem in both human and veterinary
  medicine

• 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?

4
BACKGROUND

• Antimicrobial-resistant bacteria (especially
  MRSA) are recognised as an important and
  increasing problem in both human and veterinary
  medicine

5
BACKGROUND

• Antimicrobial-resistant bacteria (especially
  MRSA) are recognised as an important and
  increasing problem in both human and veterinary
  medicine

6
BACKGROUND

• 228 cases reported of MRSA in horses
• Variety of sites of infection (similar to other
  animals and humans)

Figure courtesy of C. Scantlebury
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BACKGROUND

• 228 cases reported of MRSA in horses
• Variety of sites of infection (similar to other
  animals and humans)

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STAPHS IN ANIMALS

• Staphyloccus aureus
• 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
• 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

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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)

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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
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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)

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MRSA TYPING

• spa gene typing
• Amplification and sequencing of a single gene
  loci
• Target is region X of the (serum) protein A gene
• 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

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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
• 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/molecul
  ar confirmation of methicillin-resistance and S.
  aureus respectively

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MRSA IN ANIMALS

• Epidemiology
• 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
• 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

• 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)

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MRSA IN ANIMALS

• Epidemiology
• Difference of colonisation versus clinical
  infection
• Also transient carriage vs persistent
  colonisation

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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 5.3-164,5 (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)

• 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)

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MRSA IN DOGS

• Epidemiology
• MRSA colonisation has not been identified in
  healthy dogs in the community1-2
• Colonisation of hospitalised/unwell dogs
  reported at
• 9-233,4 (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)

• 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)

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MRSA IN ANIMALS

• ST398-MRSA
• 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

• 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)

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CA-MRSA

• Community associated MRSA
• 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)
• Small animal prevalence unknown but has been
  reported1, not definitively identified in
  horses2

• Frequently produce PVL toxin
• More virulent

• Rankin et al (2005) Vet Microbiol 108
• Maeda et al (2007) Vet Rec 161

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MRSA AS A ZOONOSIS

• Zoonotic Potential
• 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

• 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

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MRSA AS A ZOONOSIS

• Zoonotic Potential
• 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
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STUDIES AT LIVERPOOL

• Nationwide cross-sectional study on the
  microbiology and epidemiology of
  antimicrobial-resistant 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

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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

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CROSS-SECTIONAL STUDY

• Microbiological Methods

Nasal swabs enriched in 6 NaCl nutrient broth.
Streaked onto mannitol salt agar (MSA) and
oxacillin-resistance 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)

• British Society for Antimicrobial Chemotherapy
  (2007), Methods for Antimicrobial Susceptibility
  Testing

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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)

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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)
• Low MRSA prevalence of 6 dogs (0.9 0.2-1.6)

Data courtesy of A. Wedley
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STAPHYLOCOCCI IN HORSES

• 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

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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)

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MRSA IN HORSES

• All 4 equine isolates confirmed MRSA SCCmec
• type IV by PCR
• spa typing carried out for all isolates (one
  failed)
• spa types represent common equine strains (no
  ST398 identified)

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CONCLUSIONS

• 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

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AKNOWLEDGEMENTS

• PhD colleague
• Amy Wedley
• MSc Student
• Andrew ODonnell
• 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