Convegno "X Hot Topics in Gastroenterologia 2015" - 07/11/2015 - Sala Rita Levi Montalcini - Ospedale S. Eugenio - Roma

1. L’infezione da Clostridium difficile
- quello che bisogna sapere -
Gianpiero Manes
Unità Operativa Complessa di Gastroenterologia ed
Endoscopia Digestiva
Azienda Ospedaliera G. Salvini
Garbagnate Milanese, Rho e Bollate

2. Historical Perspective
• In the 1960s it was noted that patients on antibiotics
developed diarrhea1
– “staphylococcal colitis”
• Originally thought to be caused by S. aureus and
treated with oral bacitracin
• Stool cultures routinely ordered for S. aureus
• Early 1970s, a new explanation
– “clindamycin colitis”
• Severe diarrhea, pseudomembranous colitis, and
occasional deaths documented in patients on
clindamycin
1. Gorbach SL. NEJM. 1999;341:1689-1691.

3. “Antibiotic Associated
Pseudomembranous Colitis Due to
Toxin-Producing Bacteria”
• Bartlett and co-workers1
demonstrated cytotoxicity in
tissue culture and enterocolitis in hamsters from stool
isolates from patients with pseudomembranous colitis
– Isolate was C. difficile
• Bacillus difficilis (now confirmed as C. difficile) was
cultured from healthy neonates (with difficulty, hence the
name) in 19352
1. Bartlett JG, et al. NEJM. 1978;298: 531-534.
2. Hall JC and O’Toole E. Am J Dis Child. 1935;49:390-402.

4. Clostridium difficile
• Gram-positive, anaerobic, spore-forming bacillus
• Vegetative cells die quickly in an aerobic environment
• Spores are a survival form and live for a very long time in
the environment
• Grows on selective media in 2 days and smells like
horse manure
Classificazione
Dominio Prokaryota
Regno Bacteria
Phylum Firmicutes
Classe Clostridia
Ordine Clostridiales
Famiglia Clostridiacea
e
Genere Clostridium
Specie C. difficile

5. Importance of Spores
• Resistant to heat, drying, pressure, and many
disinfectants
• Resistant to all antibiotics because antibiotics only kill or
inhibit actively growing bacteria
• Spores survive well in hospital environment: 50% of
rooms of C diff (+) pts have culture-positive fomites
(bedpans, toilets, scales, furniture)
• Spores are not a reproductive form, they represent a
survival strategy

6. Source of Infections
• Spores in hospital, nursing home, or long-term care
environment associated with ill patients
– Large numbers of spores on beds, bed-rails, chairs,
curtains, medical instruments, ceiling, etc.
• Asymptomatic carriers in those same environments
– Low risk compared to patients with active disease
• Unknown in community based infections, but food has
been implicated1
• About 30% of hospitalized patients become colonized
(13% in 2 wks 50% in 4 wks)
1. Jhung MA, et al. Emerg Infect Dis. 2008;14:1039-1045.

7. Role of Antibiotics
• All antibiotics (including metronidazole and vancomycin)
are associated with CDI
• High-risk group: Clindamycin; cephalosporins, amoxicill;
ampicillin; fluoroquinolones
• Low-risk group: macrolides and other penicillins.
• Occasionally reported aminoglycosides, trimethoprim-
sulfamethoxazole, metronidazole, chloramphenicol,
tetracycline, imipenem, and meropenem
• Alteration of normal colonic flora thought to favor growth
of C. difficile

8. C. difficile: Role of antibiotics
• Canadian cohort study 2002-2003
• 1187 hospital pts prescribed Abx for >3d
• C diff diarrhea:
– Medical wards: 10.9% (OR 4.1)
– Surgical wards: 2.9%
– > 3 Abx: 12.1% (OR 2.1)
– 1-2 Abx: 5.1%
Dial S. CMAJ 2004;171:33

11. Other factors
• FDA in 2012 described a possible association between
the use of PPIs and the development of Clostridium
difficile –associated diarrhea, mainly in patients who
were elderly, had chronic and/or concomitant underlying
medical conditions, or were taking broad-spectrum
antibiotics.
• 23 of 28 observational studies showed a higher risk of C
difficile infection or disease associated with PPI
exposure, compared with no PPI exposure.
• Possible role for mirtazapine and fluoxetine
US Food and Drug Administration. FDA Drug Safety Communication:
Clostridium difficile-associated diarrhea can be associated with stomach acid
drugs known as proton pump inhibitors (PPIs). Available at:
http://www.medscape.com/viewarticle/777772.

12. Acid Suppression: New Risk Factor?
• UK case:control studies 1994-2004
• 1672 cases C. difficile matched to 10 controls
• Incidence: 1/100,000 (1994) →22/100,000 (2004)
• Adjusted Rate Ratios:
– Current PPI use: 2.9 (2.4-3.4)
– Current H2RA use: 2.0 (1.6-2.7)
– Current NSAID use: 1.3 (1.2-1.5)
Dial S et al. JAMA 2005;294:2989

13. Other factors
• Severe illnesses
• Immune suppression
• Gastric acid suppression (bypassing gastric acid via enteral feeds)
• inflammatory bowel disease (IBD)
• Antineoplastic agents, principally methotrexate
• Hemolytic-uremic syndrome
• Malignancies
• Intestinal ischemia
• Chronic kidney disease
• Necrotizing enterocolitis
• Hirschsprung disease
• Nonsurgical gastrointestinal procedures, including placement of
nasogastric tubes

14. Special situations
• Pregnancy and breastfeeding (use oral Vancomycin)
• IBD
– All patients with IBD flare need testing for C.difficile
– Highest risk with corticosteroid use > 3-fold
– Immunosuppression can be maintained but escalation
should be avoided
– Initiation of anti-TNF 72-hrs after starting therapy for
CDI
• C. difficile can cause enteritis and pouchitis!

15. Clinical pictures
– Asyntomatic carriage (60-65%, about 2-5% of health
adults, 70% of healthy infants)
– Diarrhea without colitis (antibiotic associated diarrhea
AAD)
– Colitis without pseudomenbranes
– Pseudomembranous colitis
– Fulminant colitis (2-3%)
– Relapsing infection (15-20%)- Most CDI are mild
- Diarrhea is the main symptom
- Pseudomembranous colitis and toxic megacolon are rare
- Discontinuing antibiotics worked in many cases
- High response rate to metronidazole and vancomycin

16. Antibiotic-Associated Diarrhea (AAD)
• 10-20% of all hospitalized patients treated with
antibiotics will develop diarrhea (AAD)
• Only 15-20% of AAD is due to C. difficile
• Most frequently implicated antibiotics include
penicillin, clindamycin, and cephalosporins
• Prolonged use of antibiotics alters colonic flora,
especially fecal anaerobes which normally
metabolize malabsorbed carbohydrates and break
down primary bile acids; this can result in osmotic
(carbohydrate) or secretory (bile acid) diarrhea

17. Epidemiology of CDI
• In Usa
– 9th USA cause of death for GI pathology
– High case mortality: 2195 in 2002, 7251 in 2012 (+230% in 10 yrs)
• In Europa
– 3 mld €/yr (457 mln people - each sigle infection costs 5000-15000 €)
– Double in the next 40 yrs
• The worldwide increased incidence of CDI has been attributed to
– more elderly patients in the population
– treatment resistance to fluoroquinolones
– the emergence of more virulent strain of C difficile (BI/NAP1/027).
– increased use in the total number of antibiotics in the community (in
USA in 2009 3 mln di Kg antibiotics used).
CDI = C. difficile infection.

18. Pathogenesis
• Toxigenic strains produce 2 large protein exotoxins that
are associated with virulence
– Toxins A and B
– Mutants strains that do not make toxins A and B are
not virulent
– Some strains make a third toxin known as Binary
Toxin
• By itself, not pathogenic
• May act synergistically with toxins A and B in
severe colitis
• More common in animal strains

19. Epidemic Strain
• Strain typed BI/NAP1/0271,2
• Is highly resistant to fluoroquinolones2,4
• Binary toxin genes are present
• Produces large quantities of toxins A and B1,3
• Has a tcdC gene deletion1
1. Warny M, et al. Lancet. 2005;366:1079-1084.
2. Hubert B, et al. Clin Infect Dis. 2007;44:238-244.
3. CDC Fact Sheet. July 2005.
4. McDonald LC, et al. N Engl J Med. 2005;353:2433-2441.
Adapted from McDonald LC, et al. N Engl J Med. 2005;353:2433-2441; with
permission.

20. In Vitro Production of Toxins
in Epidemic Strain
From Warny M, et al.
Lancet. 2005;366:1079-
1084, with permission.

21. Should You Treat the Patient or Treat the
Strain?
PCR ribotypes 018 and 056, identified in Europe, have
been also associated with more severe C.difficile colitis
•Routine diagnostics laboratory tests do not provide strain
type
•Routine tests not always reliable
•Always treat the patient based symptoms, history, risk
factors and markers of severe disease

22. Symptoms of CDI
• Asymptomatic colonization
• Diarrhea
mild → moderate → severe
• Abdominal pain and distension
• Fever
• Pseudomembranous colitis
• Toxic megacolon
• Perforated colon → sepsis → death

23. Markers of Severe Disease
• Leukocytosis
– Prominent feature of severe disease
– Rapidly elevating WBC
– Up to >100 K
• >10 BM/day
• Albumin < 2.5
• Creatinine 2x baseline
• Hypertension
• Pseudomembranous colitis
• Toxic megacolon
• Severe distension and abdominal pain

24. When should we suspect C.difficile?
• The diagnosis of C difficile colitis should be suspected in
any patient with diarrhea who has received antibiotics
within the previous 3 months, has been recently
hospitalized, and/or has an occurrence of diarrhea 48
hours or more after hospitalization.
• But, C difficile can be a cause of diarrhea in community
dwellers without previous hospitalization or antibiotic
exposure.

25. Laboratory Diagnosis of
C. difficile Infection (CDI)

26. Which Test Should I Use?
• Considerations
– Accuracy
– Time to detection
– Prevalence in your population
• Screening tests followed by confirmatory tests
• In a low prevalence population, a screening test with a high
sensitivity is useful (no/few false negatives)
– Cost
– Ease of use
• At this time, there is no perfect test for the
diagnosis of CDI

27. The Specimen
• Fresh is best (test within 2 hours)
• Liquid or loose, not solid
• If unable to test within 2 hours, refrigerate at 4°C
for up to 3 days
• Freeze at -70°C (not -20°C) if testing will be
delayed
• Specimen quality will influence test results
In: Manual Clin Micro. 9th ed. 2007;p. 897.

28. Laboratory Diagnosis of CDI
LaboratoryLaboratory
DiagnosisDiagnosis
Enzyme Immunoassay (EIA)Enzyme Immunoassay (EIA)
GlutamateGlutamate
Dehydrogenase (GDH)Dehydrogenase (GDH)
Cell CultureCell Culture
NeutralizationNeutralization
Assay (CCNA)Assay (CCNA)
Toxigenic CultureToxigenic Culture
(Culture and CCNA)(Culture and CCNA)
Molecular BasedMolecular Based
(PCR Or LAMP)(PCR Or LAMP)
Stool CultureStool Culture

29. Stool culture/toxigenic culture: The most sensitive test (sensitivity, 90-100%; specificity,
84-100%), but the results are slow and may lead to a delay in the diagnosis if used alone
Glutamate dehydrogenase enzyme immunoassay (EIA): This is a very sensitive test
(sensitivity, 85-100%; specificity, 87-98%); it detects the presence of glutamate
dehydrogenase produced by C difficile
NAAT (nucleic acid amplification tests , LAMP or PCR): This test is an alternative gold
standard to stool culture (sensitivity, 86%; specificity, 97% ); it may be used to detect
the C difficile gene toxin
Stool cytotoxin test: A positive test result is the demonstration of a cytopathic effect that is
neutralized by a specific antiserum (sensitivity, 70-100%; specificity, 90-100%)
EIA for detecting toxins A and B: This test is used in most laboratories (moderate
sensitivity, 79-80%; excellent specificity, 98%)
Latex agglutination technique: Another means of detecting glutamate dehydrogenase;
however, the sensitivity of this test is poor (48-59%), although the specificity is 95-96%
Stool assays for C. difficile (from the most to the least sensitive)

37. C. difficile: Endoscopy
• Sigmoidoscopy/colonoscopy are not necessary for
patients with classic symptoms and a positive assay
• Indicated when a rapid diagnosis needed, and no stool
available due to ileus, or to rule out other confounding
diseases (i.e. ischemic colitis)
• Pseudomembranes virtually diagnostic of CD colitis
• Colonoscopy superior to sigmoidoscopy in detecting
pseudomembranes (10% right sided only)
• Invasive, increased risk perforation in toxic megacolon

38. Pseudomembranous Colitis

39. Treatment

40. C Difficile: Treatment
• Stop antibiotic(s) if medically reasonable
• Supportive measures
– Correct electrolytes
– Hydration
• Antiperistaltic agents (narcotics, Imodium) are
acceptable for mild diarrhea, but are to be avoided in
patients with evidence of active colitis (bloody diarrhea,
fecal leukocytes, systemic toxicity) due to risk of toxic
megacolon.

41. Initial Treatment Options for CDI
• Historical response (96%) and relapse rates (20%) similar
between metronidazole and vancomycin1
• More recently, efficacy of metronidazole for severe
disease called into question2-4
• Recent prospective trials report vancomycin to be superior
to metronidazole in severe CDI5-7
1. Aslam S, et al. Lancet Infect Dis. 2005;5:549-557.
2. Fernandez A, et al. J Clin Gastroenterol. 2004;38:414-418.
3. Gerding DN. Clin Infect Dis. 2005;40:1598-1600.
4. Musher DM, et al. Clin Infect Dis. 2005;40:1586-1590.
5. Lahue BJ, Davidson DM. The 17th ECCMID Meeting, March 31 to April 4, 2007; Munich, Germany.
Abstract 1732_215.
6. Zar FA, et al. Clin Infect Dis 2007;45:302-307.
7. Louie T, et al. The 47th Annual ICAAC Meeting, Sept. 17-20, 2007; Chicago, IL. Abstract k-425-a.

42. Initial Treatment Options for CDI
Metronidazole
250 mg QID or
500 mg TID
• May be administered PO or IV
• Development of resistance rare
• Historical first-line agent
Vancomycin
125 mg QID
• Effective in enteral (oral or rectal) form only
• Typically reserved for severe disease, those
failing to respond to metronidazole, or
cases in which metronidazole is
contraindicated
IV=intravenously; PO=orally.
Fekety R. Am J Gastroenterol. 1997;92:739-750.
Gerding DN, et al. Infect Control Hosp Epidemiol. 1995;16:459-477.
American Society of Health-System Pharmacists. Am J Health-Syst Pharm. 1998;55:1407-1411.

43. Treatment of Severe Disease
• Oral vancomycin drug of choice
– Dose varies based on severity
• Can add metronidazole (oral or IV)
• If patient is unable to tolerate oral medication,
intracolonic vancomycin instillation
– 0.5–1 g vancomycin (IV formulation) in 0.1 to 0.5 L of normal
saline via rectal (or Foley) catheter
– Clamp for 60 minutes
– Repeat every 4–12 hours

44. • Potential role of intravenous immunoglobulin G (IVIG)1-6
– Antitoxin A IgG predicts clinical outcome of CDI
– Serum antibodies to toxins A and B are prevalent in
healthy populations
• Recent studies in severe disease5,6
– Well tolerated in small numbers of patients
– Conflicting data regarding outcome improvement
(mortality and need for colectomy)
• Often administered when surgery is considered imminent
1. Salcedo J, et al. Gut 1997;41:366-370.
2. Beales ILP. Gut. 2002;51:456.
3. Kyne L, et al. N Engl J Med. 2000;342:390-397.
4. Kyne L, et al. Lancet. 2001;357:189-193.
5. McPherson S, et al. Dis Colon Rectum. 2006;49:640-645.
6. Juang P, et al. Am J Infect Control 2007;35:131-137.
Treatment of Severe Disease

45. Multiple Recurrent CDI
• Rates of recurrent CDI
– 20% after first episode1
– 45% after first recurrence2
– 65% after two or more recurrences3
• Metronidazole or vancomycin resistance after treatment
not reported
• First recurrence can be treated in the same way as a first
episode according to disease severity
• Repeated, prolonged courses of metronidazole not
recommended (risk for peripheral neuropathy)
1. Aslam S, et al. Lancet Infect Dis. 2005;5:549-557.
2. McFarland LV, et al. Am J Gastroenterol. 2002:97:1769-1775.
3. McFarland LV, et al. JAMA. 1994;271:1913-1918.

46. Relapsing C Difficile : Treatment
• First Recurrence
– Confirm diagnosis
– Repeat 10-14 day course with metronidazole or vanco
• Second Recurrence: Vancomycin Pulse / Taper
– 125 qid for 7 days
– 125 bid for 7 days
– 125 qd for 7 days
– 125 qod for 7 days
– 125 tiw for 7 days
McFarland LV et al. Am J Gastro 2002;97:1769

47. Other Treatments
* Patients who produce antibody to toxins A and B usually do well so IVIG has been
tried.
Probiotics
Rifaximin
Chasers
Rifampin
Nitazoximide
IVIG*
FMT

48. Unproven Adjunctive Therapies
for Recurrent CDI
Probiotics
Saccharomyces boulardii
Lactobacillus GG
May reduce the likelihood of further recurrences in some
patients when added to and continued after treatment with
metronidazole or vancomycin1-3
Rifampin Efficacy in one series (n=7) when added to vancomycin4
Nitazoxanide Response demonstrated in patients (n=35) who failed
prior metronidazole therapy5
and similar response and
recurrence rates when compared with metronidazole for
initial therapy (n=110)6
Rifaximin “chaser” Effective when used for 14 days after vancomycin therapy
(n=8)7
1. McFarland LV, et al. JAMA. 1994;271:1913-1918.
2. McFarland LV. J Med Microbiol. 2005;54:101-111.
3. Surawicz CM, et al. Clin Infect Dis. 2000;31:1012-1017.
4. Buggy BP, et al. J Clin Gastroenterol. 1987;9:155-159.
5. Musher DM, et al. J Antimicrob Chemother. 2007;59:705-710.
6. Musher DM, et al. Clin Infect Dis. 2006;43:421-427.
7. Johnson S, et al. Clin Infect Dis. 2007;44:846-848.

49. Fidaxomicin
• New bacteriocidal antibiotic
• Poorly absorbed narrow-spectrum macrolide
• FDA approval for CDI in 2011

50. Fidaxomicin vs.Vancomycin
Louie TJ. NEJM. 2011;364:422-31

51. Relapsing C difficile: Bile Binders
• Cholestyramine and colestipol have the potential to
bind C diff toxins
• Lancet 1978: reduced toxin titers & exposure time in C
diff pts treated with bile binders
• Most often used to treat recurrent CD
• No new studies published in the last 25 years
• Vancomycin is bound by cholestyramine so doses
must be given three hours apart

52. Relapsing C Difficile: MAbs?
“Treatment with Monoclonal Antibodies against
Clostridium difficile Toxins”
• RDBPCT in 200 patients with symptomatic Clostridium
difficile infection
• Treatment with single infusion of two monoclonal
antibodies against C. difficile toxins A and B or placebo,
in addition to metronidazole or vancomycin
• Outcome: lab documented recurrence within 3 months of
infusion
Lowy, I et al. N Engl J Med 2010; 362:197-205

53. Infection Control
• Wash hands with warm soap and water
– Mechanical removal of spores
– Alcohol does not kill spores
– Stool is pre-treated with alcohol when growing C.
difficile
• Contact and barrier precautions
• Private room
• Antibiotic stewardship

54. Efficacy of Hand Hygiene Methods for Removal
of C. difficile Contamination from Hands
CFU = colony forming units
* Different from AHR (P<0.05).
** Different from AHR and AHW (P<0.05)
*
WWS = warm
water and soap
CWS = cold
water and soap
WWA = warm
water and
antibacterial
AHW = alcohol
hand wipe
AHR = alcohol
hand rub
Decrease in colony counts
compared with no wash
1.8 1.8
1.4
0.6
-0.1
-1
-0.5
0
0.5
1
1.5
2
2.5
Hand hygiene method
Decreaseincolonycounts
(logCFU/mL)
WWS CWS WWA AHW AHR
** ** *
Oughton M, et al. The 47th Annual ICAAC Meeting, 2007.

55. Isolation and Barrier Precautions
• Patients with CDI and incontinence should be in private
rooms or cohorted if private rooms are not available
• Contact precautions and isolation
– Gloves and gowns required for direct contact and
contact with environment
– Discontinuation of isolation when diarrhea resolves
• Dedicated equipment when possible
CDC Guideline for Isolation Precautions, 2007.
Gerding DN, et al. Infect Control Hosp Epidemiol. 1995;16:459-477.
Simor AE, et al. Infect Control Hosp Epidemiol. 2002;23:696-703.

56. Environmental Disinfection
• Removal/thorough cleaning of environmental sources can
decrease incidence
• Use chlorine-containing agents (at least
5000 ppm available chlorine 10 minutes contact time) for
environmental contamination, especially in outbreak areas
Poutanen SM, Simor AE. Can Med Assoc J. 2004;171:51-58.
CDC. Fact Sheet, July 2005.
McMullen KM, et al. Infect Control Hosp Epidemiol. 2007;28:205-207.
Mayfield JL, et al. Clin Infect Dis. 2000;31:995-1000.
Fawley WN, et al. Infect Control Hosp Epidemiol. 2007;28:920-925.

61. Il trapianto di microbiota fecale

62. La storia moderna
• Nel 1958 Eiseman, un chirurgo del Colorado, tratta e
cura 4 pazienti con colite pseudomembranosa
fulminante (il C.difficile non era ancora conosciuto)
usando clisteri di feci
• Al Centre for Digestive Diseases di Sydney Australia,
FMT è proposto da oltre 20 anni.
• Negli ultimi 45 anni descritti in letteratura oltre 700
pazienti
Eiseman B et al. Surgery 1958: 44 (5): 854–859
Borody TJ et al . Expert Rev Gastroenterol Hepatol
2011; 5(6): 653-655

63. Gough E et al. Clin Inf Dis 2011;
53: 994.

64. Che succede dopo il FMT?

65. Possibili effetti collaterali?
• Nessun caso di infezione trasmessa con FMT!!!
• Disturbi gastrointestinali
• Stipsi
• meteorismo

66. Brandt LJ et al. AJG 2012; 107: 1079-1087.
Follow up
3-68 months
-2 pazienti hanno avuto un miglioramento di una sinusite
ed artrite
-4 pazienti hanno sviluppato una nuova patologia: Sjogren;
neuropatia periferica, trombocitemia idiopatica, artrite
reumatoide

68. Lo studio !!!

71. Ricerche future sul FMT
• Cercare di capire il microbiota umano in condizioni di
normalità e malattia
• Valutare il rischio legato alla manipolazione del
microbiota nel ricevente
• Cercare di comprendere quale componente delle feci sia
responsabile del beneficio clinico
• Quali sono gli effetti a lungo termine del trapianto di
microbiota

73. e molto ancora….
• Autotrapianto per prevenire la diarrea da CD
• Creare banche di feci
• Capsule gastroresistenti contenenti estratti di
feci

74. C Difficile: Summary
• The bug is changing:
↑ risk in non-traditional (community, peripartum) pts
– Increased morbidity, mortality, and chronicity
– Epidemic strains w/ ↑ toxin production emerging
– Flouroquinolone use likely important
• We need to change:
– Increased monitoring, better infection control (gloves,
isolation, handwashing) and more antibiotic restraint
– Improved therapies are needed
– Extremely important to accurately detect and aggressively
treat severe disease

75. Grazie