ESCMID Online Lecture Library © by Author
Biofilms : how to handle them?
Chris ne IMBERT © by author Université de Poi ers - UFR Médecine et Pharmacie Laboratoire EBI UMR CNRS 7267– Poi ers – France ESCMIDchris ne.imbert@univ-poi ers.fr Online Lecture Library
Bringing PK and PD in fungal infections into the clinic Nijmegen 4-6 July 2014 Background: 2 opposite life-styles: « planktonic » ou « sessile » ?
« Planktonic » « sessile » Passively floa ng Adherent to a substrate
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Biofilm 2 Biofilms are on and in us
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Candida spp. can form biofilms on ssues and on almost any medical device!3 Biofilms : single-species or mixed ?
Single-species biofilms
Vascular catheters, Staphylocoques coagulase neg Prosthe c cardiac Staphylococcus aureus valve … Pseudomonas aeruginosa Enterobacteries Candida sp
Denture, Tissues : skin, Urinary catheter, vaginal mucosa, Contact lenses… © by author lung…
• C. albicans + P. aeruginosa ESCMID •Online A. fumigatus Lecture+ P. aeruginosa Library • C. albicans + Streptoccocus Mixed biofilms • C. albicans + Staphylococcus (S. epidermidis, S. aureus) • C. albicans + Gardnerella vaginalis 4 • Fusarium + free living amoebae C Imbert EBI picture
A. fumigatus biofilm Ramage et al., 2011
C. albicans biofilm
Douglas, 2003 © by author ESCMID Online Lecture Library
Bjarnsholt et al, Nature Reviews Drug Discovery 2013 C. neoformans biofilm (polystyrene) 5 Martinez et al, 2010 Main mechanisms explaining an fungal resistance associated to C. albicans biofilms
Decreasing Decreasing role role
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6 Ramage et al, 2012, Interna onal Journal of Microbiology Human consequences : high risk of infec on
• Biofilms are associated with 90% of catheter–related infec ons • Device-associated Candida sp infec ons ðmortality rates ≥ 30%
Biofilm
Biofilm © by author = Infec on Reservoir of infection ESCMID Online Lecture Library
Viudes et al., 2002; Finkel & Mitchell, 2011 ; DiDone et al., 2011; Ne et al., 2007 7 So...Comba ng fungal biofilms: strategies and goals?
Remove and eradicate the biofilm Inhibit development of biofilm Mechanically disrupt the biofilm
Preven on Systemic therapy
Ethanol lock solu ons
Remove + replace the device Ethanol lock solu ons Biofilm © by author Control ESCMIDEradica on Online Lecture Library
lock solu ons Other? Other? 8 An fungal used as Management of a catheter associated candidemia
ESCMID guidelines. Cornely et al., CMI 2012
ó Where possible the catheter should be removed
Study of Garnacho-Montero et al, JAC 2013 • 188 pa ents : with posi ve blood culture for Candida spp. + CVC • Confirmed that : © by author • catheter removal is a protec ve factor •ESCMID and adequacy of an fungal therapy in the first 48h seemed to Online Lecture Library reduce the mortality rate
9 Comba ng a biofilm already formed
BUT…. removal of the CVC is not always possible
Antifungal arsenal
SYSTEMIC ANTIFUNGAL OTHER ANTIFUNGAL © by authorLOCK APPROACHES TREATMENT TREATMENT ? ESCMID Online Lecture Library
10 Systemic an fungal therapy
ð ATF ac ve against Candida biofilms: • Lipid formula ons of amphotericin B – L-AmB and ABLC • Echinocandins – caspofungin, micafungin and anidulafungin
Flu
Cocuaud et al, JAC 2005 ; Cateau et al, ©CMI 2007by ; author Cateau et al, JAC 2008 ; Cateau et al, IJAA 2011 ; Toulet et al, JAC 2012 ; Bachmann et al, AAC 2002 ; Katragkou et al, AAC 2008 ESCMID Online Lecture LibraryCasp, AMB, Abelcet
11 Kuhn et al, 2002 11 However…
ð ATF poorly ac ve on Candida biofilms • Amphotericin B deoxycholate Lewis et al, AAC 2002 ; Ramage et al, AAC 2002 • Azoles Ramage et al, AAC 2001 ; Cateau et al, IJAA 2011
Flu However they may prevent : © by author - dispersion Casp, AMB, Abelcet - propaga on ESCMID Online Lecture Library
Kuhn et al, 2002 12 Cornely et al, EFISG, CMI
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13 13 An fungal combina ons against Candida biofilms? Chatzimoschou et al, AAC 2011
In vitro study ; C. albicans and C. parapsilosis biofilms
ð When used serially, triazole-echinocandin combina ons were marginally effec ve against biofilms of Candida
ð When used simultaneously, triazole-echinocandin demonstrated indifferent interac ons against © by author biofilms of Candida ESCMID Online Lecture Library ðInterest not really demonstrated
14 A. fumigatus biofilms Seidler et al, AAC 2008
In vitro results – Mature biofilms
© by author HBE : Human Bronchial Epithelial ; CFBE : Cys c Fibrosis Bronchial Epithelial ; PS : Polystyrene ESCMID Online Lecture Library Fiori et al, AAC 2011 : - Amphotericin B, voriconazole, Caspofungin and anidulafungin : poorly active against biofilms - Amphotericin B > voriconazole > echinocandin 15 A. fumigatus biofilms – an fungal combina on Liu et al, AAC 2012
Synergis c an fungal ac vity with a combina on of : - CAS and AmB - CAS and VORI - no observed antagonism © by author ESCMID Online Lecture Library
16 Preven on of fungal infec ons associated to biofilms
Antifungal used as lock solutions ? Antimicrobial coating ?
Ethanol, Taurolidine, EDTA, etc used as lock solutions ?
Natural© anti-adherentby author components ? ESCMID Online Lecture Library
17 Preven on of fungal infec ons associated to biofilms
Antifungal used as lock solutions ? Antimicrobial coating ?
Ethanol, Taurolidine, EDTA, etc used as lock solutions ?
Natural© anti-adherentby author components ? Guidelines ? ESCMID Online Lecture Library - EFISG guidelines 2012 - ESGB guidelines : soon… BUT recommandations for bacterial biofilms/ infections will be more documented than for fungal ones… 18 Too few available guidelines for preven on of candidiasis biofilm-related
Cornely et al, EFISG
C: marginally supports a © by author recommenda on for use ESCMID Online Lecture Library
ððThe main issue : deficiency of clinical data 19 Preven on of fungal infec ons associated to biofilms
Antifungal used as lock solutions ? Antimicrobial coating ?
Ethanol, Taurolidine, EDTA, etc used as lock solutions ? © by author Natural components ? ESCMID Online Lecture Library Other?
20 An microbial coa ng to prevent biofilm forma on
Meta-analysis: Impact of catheter an microbial coa ng on risk of catheter Novikov et al, 2012 coloniza on : an microbial- versus standard- CVC ð Increase of Candida sp coloniza on among minocycline-rifampicin CVCs
ð Efficiency of silver-sulfadiazine coa ng (960 adult pa ents: Walz et al, Crit Care Med, 2010)
© by author The use of coating that have limited antifungal activity may serve to increase colonization / infection due to Candida sp ESCMID Online Lecture Library ð The choice of an an microbial CVC may be influenced by a previous history of infec on with Candida species 21 Antifungal used as lock solutions ?
Antimicrobial coating ?
Ethanol, Taurolidine, EDTA, etc used as lock solutions ? © by author Natural anti-adherent components ? ESCMID Online Lecture Library Other?
22 Lock therapy
• To prevent or control or eradicate fungal biofilms? • Method: sta c ins lla on ; 8 – 12h into the catheter lumen • Advantages : • conserva ve strategy • Cost effec ve method • Safe for pa ents © by author
ESCMIDEthanol Lock therapy Online LectureAn fungal Lock therapy Library concentrated solu on (>100 MIC) Role = s ll not well defined 23 Ethanol Lock therapy Preven on / control / eradica on ? Numerous In vitro models • Rane et al, AAC 2012; Peters et al, AAC 2013, … ð 30% to 35% EtOH for 4h – >99% reduc on of mature C. albicans biofilms – and prevented the regrowth of C. albicans on silicone • Oncu, Clin Nutri on 2014 © by author ð 40% EtOH for 30 min is enough to sterilize the catheters ESCMID Online Lecture Library • Wales et al, J Ped Surgery 2011 ; Ghannoum et al, AAC 2011 ð Suggested the interest of EtOH lock combined with EDTA 24 Promising results using Taurolidine lock solu ons Preven on + eradica on ?
Olthof et al, Clinical Nutri on 2014 • In vitro interest of taurolidine containing lock solu ons against fungal, Gram- and Gram+ pathogens
Diaman et al, Clin Nutri on 2014 © by author • 1 pa ent with repeated and severe C. glabrata central ESCMIDline sepsis was successfully treated with taurolidine Online Lecture Library locks
25 Echinocandins used as lock solu ons Control + eradica on ? Cateau et al, JAC 2008+IJAA 2011 : silicone catheters ; in vitro • CAS, MFG : 5 to 25 mg/L (100-500 x MIC); lock12h, biofilm: 12h/5d ; silicone - C. albicans and C. glabrata biofilms ó ac vity ++ - BUT : low persistence of CAS against C. glabrata biofilms
Lazzell et al, 2009 ; Shuford et al, 2006 ; Kucharikova et al, 2010:
• CAS, MFG = ac ve using in vivo models © by author ESCMID Online Lecture Library
26 Echinocandins used as lock solu ons Control + eradica on ? Cateau et al, JAC 2008+IJAA 2011 : silicone catheters ; in vitro • CAS, MFG : 5 to 25 mg/L (100-500 x MIC); lock12h, biofilm: 12h/5d ; silicone - C. albicans and C. glabrata biofilms ó ac vity ++ - BUT : low persistence of CAS against C. glabrata biofilms
Lazzell et al, 2009 ; Shuford et al, 2006 ; Kucharikova et al, 2010:
• CAS, MFG = ac ve using in vivo models © by author Simitsopoulou et al, AAC 2014 ; lock 24h ; polystyrene • Eradica on + persistence: ESCMID OnlineCAS lock solu ons at 512-2048 mg/L Lecture Library • CAS = more ac ve than ANID and MFG against C. lusitaniae and C. guillermondii biofilms 27 Lipid formula ons of AmB used as lock solu ons Control + eradica on ? Toulet et al., JAC2012: silicone catheters; in vitro • L-AmB (0.2mg/mL or 1mg/mL), 12h-5d old biofilms, single lock : 4 or 12h ðInhibi on ≥ 70% of : ð young biofilms ( C. albicans, C. glabrata, C. parapsilosis) ð mature biofilms of C. albicans, C. glabrata ðReduced efficacy on mature C. parapsilosis biofilm: inhibi on ≥ 40% Mukherjee et al, IJAA 2009 ; in vivo (rabbit model) ðefficacy of ABLC : C. albicans ©biofilms by author, locks :4h or 8 h, repeated for 7 days ESCMID Online Lecture Library
28 Lipid formula ons of AmB used as lock solu ons Control + eradica on ? Toulet et al., JAC2012: silicone catheters; in vitro • L-AmB (0.2mg/mL or 1mg/mL), 12h-5d old biofilms, single lock : 4 or 12h ðInhibi on ≥ 70% of : ð young biofilms (C. albicans, C. glabrata, C. parapsilosis) ð mature biofilms of C. albicans, C. glabrata ðReduced efficacy on mature C. parapsilosis biofilm: inhibi on ≥ 40% Mukherjee et al, IJAA 2009 ; in vivo (rabbit model) ðefficacy of ABLC : C. albicans ©biofilms by author, locks :4h or 8 h, repeated for 7 days SimitsopoulouESCMID et al, AAC 2014 : polystyrene ; Online Lecturein vitro Library • L-AmB (0.25 to 2 mg/mL), 24h lock: ðac ve on biofilms of C. lusitaniae (eradica on: 74%) and C. guillermondii (67%) 29 Echinocandin and lipid forms of AmB lock solu ons may be an alterna ve solu on to manage Candida biofilms
To be efficient, this approach would need: - to repeat locks - be used as an adjunct to systemic an fungal therapy
Dimondi et al, 2014, Pharmacotherapy: good results with L-AmB lock administered in combina on © by author with IV MFG (6 days) in an adult receiving hemodialysis to manage ESCMIDC. albicans Onlineinfec on Lecture Library ððOther clinical data are needed !
30 Research for new an -biofilm agents
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31 An -biofilm molecules extracted from natural Substances?
Lichen Blueberry species Vaccinium myr llus
Cranberry V. macrocarpon
Essen al oils, terpenes, …
Raspberry © by author Rubus idaeus
Lichen Acerola ESCMID Online Malpighia punicifoliaLecture Library Red berry fruits: species • preven on of dental diseases by natural foodstuff sources
• fruits rich in tannins ; proanthocyanidins 32 Anti-adherent activity of berry fruits
PBS Vaccinium macrocarpon (Canneberge)- cranberry Rubus idaeus (Framboise)- raspberry Vaccinium myrtillus (Myrtille)- blueberry Malpighia punicifolia (Acérola)- acerola 0,4
0,35
0,3
0,25 * ** * * 0,2 * * * * 0,15 * (A450 nm) * * statistically from control * ≠ 0,1 variance analysis and Scheffe* test *p<0,05) * * * * 0,05 © by author
0 10 20 40 10 20 40 10 20 40 Metabolic ac vity of adherent cells Activité métabolique des levures adhérentes des levures adhérentes métabolique Activité ESCMIDC. albicans ATCC Online 3153 C. glabrataLecture IHEM 9556 LibraryC. parapsilosis ATCC 22019
Red berry extracts and PBS control concentrations (mg/ml)
33 Girardot et al, Pathogens and Disease, 2014 Ac vity of red berries against planktonic yeasts (MIC)
C. albicans C. glabrata C. parapsilosis ATCC 3153 IHEM 9556 ATCC 22019 (mg/ml) (mg/ml) (mg/ml) Vaccinium macrocarpon - Cranberry > 100 > 100 > 100 Rubus idaeus - Raspberry > 100 > 100 > 100 Vaccinium myr llus - Blueberry > 100 > 100 > 100 Malpighia punicifolia - Acerola > 100 > 100 > 100 Amphotericin B © by <0,002 author <0,002 <0,002
ESCMIDThe tested commercial red berry extracts displayed Online Lecture Library no relevant ac vity against planktonic Candida spp
34 Girardot et al, Pathogens and Disease, 2014 Other « experimental » strategies based on:
• Quorum sensing molecules – Fungal and bacterial QS • eDNA diges on • carbohydrate lysis • Surfactant © by author • Dispersing agents ESCMID• .... Online Lecture Library
35 Fungal Biofilms The way to go….
oups…!!!
© by author -For the most documented strategies : clinical data are needed ESCMID Online Lecture Library -For the other: the road will be s ll long but results are promising! 36 Laboratory Ecology and Biology of Interactions, UMR CNRS 7267 Poitiers – France Team « Microbiology » Collaborations on natural substances:
- Cranberry: Leslie Boudesocque (Tours, France) Cécile Enguehard-gueiffier (Tours)
Laurent Bazinet (Quebec, Canada)
- Lichens: © by author Marion Millot (Limoges, France) Angèle Lengo-Mambu (Limoges)
- EssentialESCMID oils: Online Lecture Library Nawel Chaftar (Sousse, Tunisia) Taoufiif Ghrairi (Sousse) Khaled Hani (Sousse) Thank you for your attention37 !