Veterinary Master Thesis DefenseEmily Clare Nightingale stud.med.vet. V9089 Methicillin-resistant Staphylococcus aureus (MRSA) Colonization of Danish Veterinary Staff and Companion Animals; Insight into Prevalence and Zoonotic Potential Wednesday 14th March, 13:00
Seminar Room B, Stigbøjlen 7, LIFE campus
Supervisors: Censor:
Luca Guardabassi Dr. Robert Skov Prof. Niels Høiby
Associate Professor Head of the Staphylococcal Dept. Clinical Microbiology
Dept. Veterinary Pathobiology Ref
Veterinary Master Thesis Defense
Emily Clare Nightingale stud.med.vet. V9089 Methicillin-resistant Staphylococcus aureus (MRSA) Colonization of Danish Veterinary Staff and Companion Animals; Insight into Prevalence and Zoonotic Potential Wednesday 14th March, 13:00
Seminar Room B, Stigbøjlen 7, LIFE campus
Supervisors: Censor:
Luca Guardabassi Dr. Robert Skov Prof. Niels Høiby
Associate Professor Head of the Staphylococcal Dept. Clinical Microbiology
Dept. Veterinary Pathobiology Reference Laboratory Rigshospitalet
Faculty of Life Science (LIFE) Statens Serum Institut Copenhagen
Copenhagen University Copenhagen
Welcome to this Master Thesis defence
It’s a pleasure seeing so many of you here today
Firstly: Thank you LG and RLS and NH
Title of thesis is Methicillin-resistant Staphylococcus aureus (MRSA) Colonization of Danish Veterinary Staff and Companion Animals; Insight into Prevalence and Zoonotic Potential
Specific objectives of this thesis
Brief background information on MRSA
Account of the three parts of the thesis, including results and conclusions
To end a brief summary.
Specific Objectives
Part I
To establish the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) carriage amongst veterinary staff in Denmark
Part II
To investigate the MRSA carrier status in animals owned by individuals currently or historically tested MRSA positive in the county of Vejle, Denmark
Part III
To compare the use of Oxacillin Resistance Screening Agar Base and MRSA-ID agar as commercially available tools for the screening of MRSA
Two prevalence studies and one comparison study
Part I – DVA AGM in Autumn 06
Prompted further investingation
Three conferences + negative control group
Part II - SSI and Vejle County Hospitals Microbiology Department
Epidemic since 2002 Vejle
Surveillance program
Part III - Comparison study
SSI / LIFE
Background
Source: online CDC/Janice Carr S. aureus infection can be fatal Etiology of Staphylococcus aureus
Gram positive cocci 1µm
Coagulase positive
Commensal on 30% of world human population
Opportunistic pathogen
Dermatitis Osteomyelitis
Pneumonia Abcesses
Acute endocarditis UTI
Myocarditis Post operative infections
Pericarditis Meningitis ......
Commensal: 1/3 of you in here are likely to be carrying S. Aureus, most likely in your noses, armpits or groins..
Background
PBP2a (mecA gene) Methicillin Source: www.wikipedia.org Source: www.bakteriologie.de Methicillin molecule History
Anno 1961 - First case of methicillin-resistant S. aureus (MRSA)
B lactamase stable
Side chains
Low affinity
Chromosomal mecA gene
Background
Source: EARSS Annual Report 2005
Non healthy individuals:
percentage of MRSA amongst S. aureus isolates in European countries
Holland and Scandinavia LOW Europe and USA
Healthy individuals:
percentage MRSA amongst population
USA ≤ 3.0%
Holland ≤ 1%
Healthy individuals
Non healthy individuals
DK = Holland
Same stringent treatment and surveillance policies
Background
Source = Epi-Nyt 2005, SSI Number of MRSA cases per year in DK
Exponential rise
Mortality rate difficult- other concurrent conditions
Increase Mortality
Morbidity
Length of hospitalisation
cost of treatment
compared to M Susceptible SA infections
Part I
Aim
To establish the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) carriage amongst veterinary staff in Denmark
Hypothesis (H1)
Prevalence of MRSA amongst veterinary staff ≠ MRSA prevalence in control
Null hypothesis (H0)
Prevalence of MRSA amongst veterinary staff = MRSA prevalence in control
England, Canada and the USA
MRSA has been found in veterinary staff and companion animals
Pertinent to investigate veterinary staff in DK.
A negative control group was also investigated.
Part I - Method
Data collection
Conferences
Danish Pig Veterinary Society, Kolding
Danish Veterinary Association – Horse Section, Ebeltoft
Danish Small Animal Veterinary Association, Aarhus
Negative control group
Copenhagen Business School, Frederiksberg
Questionnaire
Nasal Sample
Criteria
- national participation
- A large proportion vets: daily contact with animals
- Good health
The Negative control group consisted of students from the CBS.
Criteria
- No contact to animals, antibiotics or hospital environments
Minimum 125 samples
Questionnaire
demographics
postal code for home and work address
occupation
Pets
For veterinary staff
small animals/horses/cattle/pigs/othe
Handled antibiotics
Gloves when doing so
Nasal sample
Part I - Method
Sample Enrichment ORSAB MRSA-ID SSI LIFE 5% blood
agar PCR
- mecA
- spa
Coagulase test 5% blood
agar Laboratory analysis
2 methods
ORSAB
oxacillin kills G+
polymixin B kills G- >>> G+
Aniline blue
Enrichment
aztreonam kills G+
cefoxitin 2nd gen. Cephalosporin: kills G+ and some G-
MRSA-ID
cefoxitin
alpha glucosidase
Part I - Method
Source: www.langara.bc.ca/biology/mario/Assets/PCR.jpg PCR
Denaturing
Annealing
Polymerisation
Amplify DNA target sequence ie. Meca and spa
Detectable end products
Primers locate target sequence
Products -> gel electrophoresis
Voltage applied to amplified DNA fragments
Position of fragments in the gel -> contains the meca or spa gene.
Questionnaire correlation
Prevalence of MRSA = no. + MRSA samples /total number samples
Part I - Results
98 Danish Small Animal Veterinary Association 225 Total 55 Danish Veterinary Association – Horse Section 72 Danish Pig Veterinary Society Number of samples Conference 128 Copenhagen Business School Number of samples collected for negative control group Number of samples collected at each conference
Target 125
Part I - Results
mecA spa - + 9 samples out of 225 were mecA, spa and coagulase positive 9 out of 225 samples = MRSA Result of the mecA and spa PCR
Gel to which a voltage is applied. Fragments move according to size
First well = negative control (water)
Second well = positive control
Each column represents each of the 17 samples
Markers at the side act as rulers so the bp size can established
Part I - Results
Prevalence of MRSA amongst Danish Veterinary Staff = 4.00%
Prevalence of MRSA in negative control group = 0.00%
Fishers Exact test p(0.05) value = 0.0292
The difference is significant
Part I - Results
Vet in practice A165 Vet in practice A213 Vet in practice A205 Vet Nurse A160 Vet in practice A241 Other K101 Vet in practice A172 Vet in practice K90 Vet in practice E20 Wear gloves Handle
Antimicrobials Occupation Sample no. Selected answers from questionnaire
9 positive samples in total
7/9 = vets in practice
1/9 = VN
1/9 not specified
8/9 handle Anti-M and of these 8 only 1 wore gloves when doing so.
Sample size too small
No risk analysis
Put together with results from DVA
Part I - Discussion
Why are 4.00% of Danish veterinary staff MRSA carriers?
Handling antimicrobials
Increased use of broad spectrum antimicrobials
Prophylactic antimicrobial use
Contact to animals that are reservoirs for MRSA?
MRSA transferred from animals to humans?
mecA gene in animal commensals transferred to human commensals?
Cephaloporins is increasing in both small and large animal
Fluoroquinoones LA -> legal restrictions. Used in human medicine.. Resistance here v. Undesirable.
Fluoroquinolones increase SA medicine
Predisposes humans to carriage or infection with MRSA.
This study: 10% of vet staff wear gloves when doing so -> reflects situation in DK
Part I - Limitations
Veterinary conferences
Sample size was relatively small.
Combine results from DVA AGM, the IPVS → MRSA risk analysis
Cattle specialists not included
Questionnaires were anonymous so MRSA positive individuals could be contacted
No treatment for MRSA colonization
No further epidemiological information available
Negative control group
A group of professionals would have been better i.e. conference for IT /Insurance professionals
Here information like postal codes and the number of hours of contact with animals per week will be analysed to see if there is a specific risk factor involved in MRSA colonisation ie contact to pigs for example
Conferences: pigs, small animals and equines.
Time constraints a cattle conference was not visited
More epidemiological info
Anonymity
6/9 = pig strain -> 5 no contact with pigs
Visited pig farm?
NCG – no permission, time constraints
MRSA notifiable disease so every sample collected will not be anonymous from now on.
Part I - Conclusion
What has this MRSA prevalence study shown?
MRSA carriers exist amongst Danish veterinary staff
Prevalence = 4.00%
Hypotheses
____________________________________________ H1 - Prevalence of MRSA amongst veterinary staff ≠ MRSA prevalence in control
H0 - Prevalence of MRSA amongst veterinary staff = MRSA prevalence in control
Accept the hypothesis and reject the null hypothesis
Part II
Aim
To investigate the MRSA carrier status in animals owned by individuals currently or historically tested MRSA positive in the county of Vejle, Denmark
Hypothesis (H1)
Animals owned by MRSA positive owners will have MRSA carrier status
Null hypothesis (H0)
Animals owned by MRSA positive owners will not have MRSA carrier status
Investigates companion animals as sources of MRSA.
Vejle hospital centre of a MRSA epidemic.
Patients admitted to the hospital or visiting the outpatients dept. And staff
MRSA positve -> treatment regime
Topical application of Bactroban
Daily washing with chlorhexidine
Retested after treatment
Some remained MRSA positive despite treatment.
Role of companion animals of MRSA reservoirs investigated as the possible source of reinfection.
Part II - Method
Source: www.biotrace.com Source: images.google.com Source: www.answers.com House visits in the county of Vejle
Members of household
Primary MRSA patient: Nasal mucosae, tonsils, urinary catheters, wounds.
Spouse/children: Nasal mucosae, tonsils
Household environment
Telephone handset, toilet, skirting board, chair, remote control
Pets
Questionnaire about previous history
Physical examination, nasal mucosae
perineum, wounds
5 separate days + Project Nurse from Vejle Hospital
Vejle County MRSA Hygiene regulations
- protective disposable gowns
- disposable footwear covers, gloves + mask
- Contact to any objects was limited
1ry MRSA patient : Nasal mucosae, tonsils, urinary catheters, wounds.
Other members of the household who were willing to provide nasal and tonsillar samples did so.
The household envrionment was also tested in 5 locations using a contact dip slide.
Part II - Results
17 Dog 3 Horse 1 Rabbit 1 Bird 13 Cat Number Species House visits in the county of Vejle
254 individuals contacted
27 households with pets consented
Four dogs showed aggressive or nervous behaviour and were not able to be sampled
All human samples processed at Vejle County Hospital
Environmental samples and pet samples processed at SSI
Animal samples processed on MRSA-ID, blood agar and PCR.
Comments