1. Cystic Fibrosis Microbiology: Away Day October 2017
Dr David Garner
Consultant Microbiologist
Frimley Park Hospital

2. Aims & Objectives Discuss current issues in CF microbiology
To understand how a sputum sample is processed
To know how to interpret a sputum result
To understand why antibiotic sensitivities are of limited use in CF
To know the problem microorganisms in CF
To be aware of the infection control issues in CF
To make microbiology “interesting” for all of you!

3. Mark 27 years old with cystic fibrosis
Initial problems as a child with bacteria such as:
Streptococcus pneumoniae
Haemophilus influenzae
Staphylococcus aureus
More recent problems with:
Pseudomonas aeruginosa
Admitted with an exacerbation of his cystic fibrosis
What are you going to do?

4. Mark Check what he has grown before
Admit him to a side room on the CF unit
Send a sputum sample to Microbiology
Intensify physiotherapy
Start antibiotics
Review compliance with other medications

5. Requesting sputum cultures?
In ICE select “Respiratory and AAFB Acute

6. Requesting sputum cultures?
Top box is a drop down menu for the test, next box is for clinical details (don’t leave it blank!)
This affects the tests the laboratory do

7. Requesting sputum cultures
In ICE common tests tab select CFU tab

8. What tests does the lab do?
Chronic Pseudomonas Biannual review
sent to Ref. Lab for Typing and MIC testing
Only select twice yearly (the reference laboratory will not test samples which are sent more frequently)
Chronic pseudomonas NOT annual review
Identification of Pseudomonas, no sensitivities
NOT chronic pseudomonas
Full identification and sensitivities of microorganisms

9. What does a lab look like?

10. What does a lab look like?

11. What does a lab look like?

12. Respiratory laboratory

13. Respiratory laboratory

14. Culture: how is sputum processed?
Plated to mixture of selective and non-selective agar depending on clinical details
E.g. Cystic Fibrosis = B. cepacia agar
Incubated for 48 hours before reporting
Sensitivities take a further hours
Total time hours after receipt.

15. How to interpret a sputum result?
Appearance
Mucoid, Salivary, Purulent, Blood Stained…
Microscopy
Gram’s stain, Ziehl Neelsen (ZN) stain…
Culture
Is the organism consistent with the clinical picture?

16. Appearance of sputum Salivary Mucoid Purulent Blood stained
Spit not phlegm, risk of contamination
Mucoid
Upper respiratory tract specimen, no evidence of inflammation
Purulent
Pus, indicates inflammation
Blood stained
May indicate infection

17. Gram’s stain

18. Ziehl Neelson Stain

19. Culture: classification of bacteria
Causes of pneumonia usually originate in the upper respiratory tract

20. Classification of Gram-positive cocci

21. Bacterial Identification: Gram-negative bacilli

22. Normal Flora

23. CF “Normal” Flora Eventually also:
MDR P. aeruginosa, Burkholderia spp., Stenotrophomas maltophilia, Achromobacter spp., Non-tuberculous mycobacteria…!

24. Factors Affecting Normal Flora
Exposure to antibiotics provides a selective pressure
e.g. previous antibiotics for CAP
Increased antimicrobial resistant organisms in the environment
e.g. Pseudomonas in critical care units
Easily transmissible organisms
e.g. Staphylococcus aureus
Failure to clear colonising bacteria
E.g. Pseudomonas aeruginosa
Immunosuppressants
e.g. steroids

25. Do patients need antibiotics?
CF patients can get the same types of infection everyone gets e.g. UTI, tonsillitis, cellulitis
Viruses do not respond to antibacterials!
However there are antivirals e.g. aciclovir, oseltamivir
The presence of bacteria does not necessarily mean there is an infection!
Bacteria colonise, such as upper respiratory tract, surgical wounds, ulcers

26. How do you choose an antibiotic?
What are the common micro-organisms causing the infection?
Is the antibiotic active against the common micro-organisms?
Do I need a bactericidal antibiotic rather than bacteriostatic?
Does the antibiotic get into the site of infection in adequate amounts?
How much antibiotic do I need to give?
What route do I need to use to give the antibiotic?

27. …you choose what worked before…
In reality…
…you choose what worked before…

28. Why do exacerbations of cystic fibrosis occur?

29. Exacerbation of CF Caused by:
New bacteria
New strain of bacteria or change in phenotype
Non-cultured bacteria or viruses
Increased exacerbations  decreased survival
Decreased symptoms  increased FEV1
Risk factors for treatment failure:
Female
Low BMI
Cepacia
Time delay to starting treatment

30. Treatment of Exacerbation
High doses of Beta-lactam PLUS aminoglycoside
Choice of antibiotic
Identification of organism
Previous successful therapy
Allergies
Susceptibility testing not helpful
Too variable
Not predictive of clinical response
No value in synergy testing
Duration
Good lung function = 7-14 days
Poor lung function = longer (guided by response)

31. Why sensitivities don’t help
At least 4 different appearances of
P. aeruginosa

32. Why sensitivities don’t help
At least 3 different sensitivity patterns for each appearance of
P. aeruginosa
Is this 12 different bacteria?

33. Why sensitivities don’t help
Genetic finger printing shows only 1
P. aeruginosa
= clever bug!!!!

34. Problem microorganisms

35. Mycobacterium abscessus

36. Decreased macrophage activity
Who gets M. abscessus?
Rates are increasing: highest in year olds
Transmission
Indirect patient to patient
Surface contamination
Airborne? Pseudomonas detectable in air 3 hours after room vacated
Vitamin B12 deficiency
Azithromycin
Decreased autophagy
Decreased macrophage activity
Increased M. abscessus

37. Why does it matter? Faster rate of decline in lung function (FEV1)
2.5% vs. 1.6% per year
Absolute contraindication to transplant (unless able to eradicate?)
Infection control restrictions
Stigmatisation?
Difficult to diagnose in patients on antibiotics that interfere with culture e.g. macrolides, aminoglycosides

38. Combination therapy
Approx. 20% response (off Abx and culture negative)
Intensive phase (1-3 months)
Clarithromycin (if sensitive)
PLUS IV Amikacin
PLUS Tigecycline AND Imipenem
Continuation phase (until culture negative for 12 months on therapy!)
PLUS nebulised Amikacin
PLUS 1-3 of Minocycline OR Clofazamine OR Linezolid OR Moxifloxacin OR Cotrimoxazole

39. Rhinovirus

40. Rhinovirus 1 in 3 patients have a virus in an exacerbation
Two fold increased risk of exacerbation
Less effect on FEV1 than other causes
Associated with increased Abx use
Mainly found in summer and autumn
Increase bacterial release from biofilms
Individual response to Rhinovirus
Immune mediated?
Worse with type A than B or C
Persists for ≥ 2 months in some patients

41. Burkholderia cepacia complex

42. B. cepacia complex 2010 = 17 species
2014 = 18 species (B. pseudomultivorans)
Previous Genomovars now have distinct names
Most important
B. cepacia
B. multivorans
B. cenocepacia (contraindication to transplant)
Mortality
2003 >50% at 5 years (Manchester, Vancouver)
2010 <10% at 5 years (Prague) – Why?
Not part of the normal human flora

43. Epidemiology Infection control precautions work
Prevent spread from patient-to-patient
Global epidemic B. cepacia strains
How do they spread internationally?
B. multivorans
Increasingly common
Rare patient-to-patient transmission
Can be replaced by B. cepacia (but not the reverse)

44. Non B. cepacia complex strains
B. gladioli
3rd most common strain in USA
Clinical significance unclear

45. Infection Control

46. Transmission Main problems P. aeruginosa and B. cepacia complex Routes
Environment-to-patient
Patient-to-patient
Mainly droplet spread
Within few metres
Potential droplet nuclei spread
Remain suspended in air up to 3 hours after patient leaves

47. Recommendations Surveillance Standard precautions
Identify organisms
Refer for PFGE
Standard precautions
For patients as well as staff
Consider H2O2 but no evidence
Segregate ALL patients
No common areas
Only patients from same household can share
No B. cepacia complex or M. abscessus patients in CF wards or outpatient clinics

48. M. abscessus CF Trust UK and American CF Guidelines
Separate clinic in different building
No shared equipment AT ALL!
No admission to CF unit
Negative pressure side-room
Sporicidal cleaning then leave room empty for 24 hours

49. Recommendations
Gloves and aprons for contact with patient and their environment (UK CF Trust)
20% of HCW contamination from environment
No routine use of masks for inpatients
Ventilation
Wait 60 minutes before final disinfection of room after patient discharged
Negative pressure room

50. After all that… Mark Previous results checked Admitted to CF unit
No epidemic P. aeruginosa, B. cepacia complex, M. abscessus
Admitted to CF unit
Sputum sample sent (requested as exacerbation of CF not chronic Pseud)
Started on Meropenem PLUS Tobramycin
Sputum result confirmed P. aeruginosa sensitive to Meropenem and Tobramycin
Mark improved quickly and went home after a week in hospital

52. Conclusion Infections in cystic fibrosis are very difficult to manage
Multiple factors cause problems:
Patient factors
Allergies
Antibiotic resistance
Laboratory issues
Odd bacteria!
Successful management requires team work!

53. Any Questions?
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