Antibiotic Resistance and Virulence PDF

Microbial communities & Antimicrobial Resistance J. Penders, PhD Medical Microbiology, MUMC Bacterial structure (1) genetic material One chromosome: - dsDNA - circular - > 1200 µM (folded) Plasmides:...

Microbial communities & Antimicrobial Resistance J. Penders, PhD Medical Microbiology, MUMC Bacterial structure (1) genetic material One chromosome: - dsDNA - circular - > 1200 µM (folded) Plasmides: - Extrachromosomal DNA fragment - circular - non-essential Bacterial structure (2) External structures Pili/Fimbria (protein): - Adherence - Sex pili: conjugation Capsule (sugar): - Adherence - Protection against phagocytoses - Protects against antibiotics Flagellum (protein): - Movement  The gut microbiome: a highly diverse ecosystem 100 billion inhabitants 1,000-2,000 species Unique composition  6 Trivia about our microbiome Microbes feed on ~60 g. sugars/day Gas released from microbial METABOLISM 1-4 liters/day Microorganisms as symbionts: 7 co-evolution & co-adaptation The “sushi” factor gut bacterium Bacteroides plebeis Marine Porphyranases, Bacteroidetes agarases breakdown of polysaccaharides of Porphyra spp. (nori) Transfer of carbohydrate-degrading enzymes from marine bacteria to gut bacteria of Japanese human population Department of Medical Microbiology – Maastricht University Hehemann et al. Nature 464, 908-912 (8 April 2010)  8 How bacteria talk: The bobtail squid and its Vibrio fischerii 8  9 Counter-illumination 9  10 Bacterial communication: quorum-sensing 10 Department of Medical Microbiology – Maastricht University What are important functions of microbes? - In our body? - In food processing? - In nature? 11  12 Functions of ‘The Forgotten Organ’ Department of Medical Microbiology – Maastricht University Gutmicrobiotaforhealth.com  13 Ecosystem perturbations and westernization Dysbiosis of ‘The Forgotten Organ’ 14 Department of Medical Microbiology – Maastricht University Gutmicrobiotaforhealth.com Dysbiosis affects different organs 15 Anxiety Diabetes (T1 & T2) Depression & Infections Atopy & Allergies Department of Medical Microbiology – Maastricht University Thaiss, 2016  Complex & diverse Daily exposure to AMR bacteria High population density → HGT Important AMR reservoir Microbiome ClinicalResistome isolates Indicator organisms What kind of diseases are caused by bacterial infections? Pathogenesis of Infections Disturbance of the natural balance Attack Defense guest host microorganism human  The microorganism Number: Minimally infectious dose = minimal number of microorganisms that are needed to cause an infection E.g. Vibrio cholera 106 bacterial cells, Shigella 102 bacterial cells Pathogenicity: primary pathogen vs. opportunistic pathogens Route of contamination Virulence Virulence = the relative capacity of a pathogen to overcome body defenses – Adherence → pili – Direct damage → toxins – Invasion – Immune evasion  Virulence factors Adhesion factors Protection against peristalsis Initial process for colonisation/local infection/invasion Fimbriae (adhesins) most important component for adhesion Invasion factors (e.g. type III/IV secretion systems) Capsules Protection against phagocytosis (escape from immune system) 21  Virulence factors: toxins Endotoxins LPS (lipid A) prototype endotoxin Elicits strong immune response (a.o. production of cytokines) Fever, other symptoms and in worst case septic shock Exotoxins Very toxic Secreted by bacteria – A & B part Superantigens Proteins existing of several (binding & enzymatic) Examples: cytotoxins, neurotoxins, enterotoxins 22 23 From commensal to pathogen 26  ANTIBIOTICS The term antibiotic (Selman Waksman 1942): Any substance produced by a microorganism (fungi) that is antagonistic to the growth of other microorganisms (bacterials) in high dilution. 1928: Alexander Fleming discovers penicillin How do antibiotics work? Antibiotics utilize the difference between the bacterial cell and the eukaryotic host cell Antibiotica are: Bacteriostatic: prevent replication of bacteria, giving the host immune system an opportunity to fight the infection (reversible action) or Bactericidal: kill bacteria (irreversible action) Antibiotics: mechanisms of action Broad- and narrow spectrum antibiotics Antibiotic development Antibiotics innovation gap between 1962 and 2000 Fischbach MA and Walsh CT Science 2009 What are the reasons for the lack of development of new antibiotics?  Pharmaceutical industry Aim to earn money Noncommunicable diseases Life style drugs Chronic suppressive therapy  Old & New strategies Chemical engineering of existing drugs Identification of novel antibiotics -> searching in unexplored ecosystems (aquatic ecosystems, soil ecosystems) New drugs…-> interfering with quorum sensing Quorum sensing as drug target Control of bacterial behaviours: Bioluminescence production Sporulation Swarming motility Biofilm formation Secretion of virulence factors Conjugation Target New therapeutic options for fighting bacterial diseases Antibiotic Resistance Antibiotic resistance Emergence of antibiotic resistance Worldwide In Europe (ECDC Surveillance rapport - 2011): Stabilisation of resistant Gram positive bacteria Emergence of resistant Gram negative bacteria 2002 Epidemiological surveillance Surveillance programme Fast identification of an outbreak Infection control measurements WIP guidelines 2011  42 Global rise in AMR prevalence ESBL: Extended Spectrum Beta- Woerther PL. Clin Microbiol Rev. 2013. Lactamases  43 http://www.bnr.nl/incoming/2011/04/05/schippers-hoopvol-over-europese-zwarte-lijst.jpg/ALTERNATES/i/Schippers-hoopvol-over-Europese-zwarte-lijst.jpg  44 Antibiotic resistance in perspective – anno 2050 Antibiotic deployment and development of resistance 46 Effect of selective antibiotic pressure Darwinian selection 47 Mulvey M R , Simor A E CMAJ 2009;180:408-415 Important mechanisms for resistance Enzymatic breakdown/alteration of the antibiotic Decreased permeability/uptake of the antibiotic Mutation of antibiotic target Active export (efflux pump) of antibiotic 48 Enzymatic breakdown of antibiotics: Beta-lactamase Acquisition of genetic information by bacteria: 3 different mechanisms 50  Mobile genetic elements Phages Plasmids Transposons 51  52 Antibiotic resistance in perspective - present Important mechanisms for resistance Mulvey M R , Simor A E CMAJ 2009;180:408-415  Multiresistant bacteria are a threat Problem is growing worldwide Therapy failure Increased healthcare costs Modern medicin: antibiotics to prevent and treat infectious diseases 53  Multiresistant bacteria = insensitive to various types of antibiotic Vancomycin resistant enterococci (VREs) G+ Methicillin resistant Staphylococcus aureus (MRSA) Extended-spectrum beta-lactamase (ESBL) producing gram-negative bacteria 54  Beta-lactamases: June 1964 – ampicillin markted in Europe December 1964 – 1st case of ampicillin resistant E. coli Ms. Temoneira (Athens, Greece) – E. coli isolate of urine culture – Producing beta-lactamase (TEM-1) – Genes coding for beta-lactamases located on plasmid X Penicillin Beta-lactam antibiotics  Cephalosporines Discovered in Italy 3rd gen. developed after world-wide spread of beta- lactamases against ampicillin and first generations of cephalosporins Extended-spectrum beta-lactamases X 3rd gen. cephalosporins X 1-2nd gen. cephalosporins X Penicillin Beta-lactam antibiotics  Extended-spectrum beta-lactamases Group of enzymes capable of inactivating penicillins and 1st, 2nd, 3rd generation cephalosporins Originated through: Mutations in penicillinases (e.g. TEM-1, TEM-2, SHV-1) Horizontal gene transfer of ESBLs present in environmental bacteria (CTX-M-type ESBLs) Last resort treatment Carbapenems Carbapenems X 3rd gen. cephalosporins 1-2nd gen. X cephalosporins X Penicillin Beta-lactam antibiotics ESBL: mutations in classical TEM and SHV enzymes (penicillinases) ESBL: acquisition through HGT from environmental bacteria CTX-M-type ESBLs: - Appear to originate from Kluyvera species (on chromosome) - Transferred to plasmids in o.a. E. coli & Klebsiella spp. Resistance – Europe Proportion 3rd gen. cephalosporin resistant Escherichia coli 2005 2011 60  Carbapenems 3rd gen. cephalosporins 1-2nd gen. cephalosporins Penicillin Carbapenemases (NDM-1, OXA-48, VIM…) 61 Laboratory diagnostics of antibiotic resistance  Antimicrobial Susceptibility: E-test & Disk Diffusion MIC Etest Disk Diffusion 63 Photo courtesy of Dr. Lesley McGee, CDC  Broth Microdilution Method Low High concentration Clindamycin MIC >32 µg/ml Penicillin MIC 0.06 µg/ml Erythromycin MIC 8 µg/ml Sterile control (MIC) Minimum Inhibitory Concentration Growth Control Photo courtesy of Dr. Lesley McGee, CDC Cause of emerging antibiotic resistance Effect of selective antibiotic pressure Darwinian selection 66 Mulvey M R , Simor A E CMAJ 2009;180:408-415  Where do resistance genes come from ? - Spontaneous mutations in genes with originally different functions - Present in environmental bacteria (a.o. soil) way before antibiotics were deployed in medicine Buhllar, Plos ONE 2012 lechuguilla cave, new mexico Resistome en antibioticaresistentie | J. Penders | Boerhave 22 Januari 2016 Knapp CW et al., Environ Sci Technol. 2010 Relation with environment - Antibiotic use in humans Relation with environment – veterinarian antibiotic use Relation with environment – veterinary antibiotic use Relation with environment – travel ?  First evidence of international travel as a risk factor for the dissemination of ESBLs: Population-based research (Calgary Health Region): -Identification of patients with UWI’s caused by ESBL-producing E. coli -247 patients included - Known risk factors (female gender, elderly, co-morbidities) - Travel as a newly identified risk factor: - India: RR = 147 - Middle-East RR = 18, Africa RR = 8, etc.  76 David Ricci 19 years-old Volunteer in orphanage in Kolkata, India Train accident Hit by a train, his leg crushed between the wheels Amputation in Indian hospital Back in US Infection of wound with 4 different multiresistant bacteria all producing NDM-1 a carbapenemase Treatment: - Further amputation of his leg (cutting away the infected tissue) - Colistin, an antibiotic that was no longer in use because of toxicity - Tigecycline, an antibiotic associated with increased mortality Almost died due to organ failure Microbiota in health and disease: lessons from longitudinal cohort studies| John Penders | 22nd January 2015 77 ESBL-E acquisition according to UN subregions What to do with these results?