University:
West Coast University
Course:
PHAR 837 | IPC Infectious Diseases I
Academic year:

2024
Views:

371

Pages:

43
Author:

Asal A.

1 MICROBIOLOGY OVERVIEW 2 Antibacterials 3 Objectives • Classify the bacteria • Characteristics • gram (+) vs. (–) • anaerobes vs. aerobes • cocci vs. bacilli • Laboratory testing • Coagulase (+) vs. (-) • Glucose-fermenter vs. non- fermenter • Identify Normal flora and Common bacterial pathogens • Respiratory Inf. • Urinary Inf. • GI/Intra-abdominal Inf. • Skin Inf. • Describe Microbiology Lab Test • Culture • MIC/Susceptibility/sensitivity Test • Rapid Molecular Methods for Organism Identification: Nucleic Acid Based Test • Complete UCSF Microbiology Module 4 HR, 61 YOF • Admission: 8/19/2024 • CC: fever (103.6oF) • HPI: Brought by husband at 10 AM for evaluation with fever, pain in urination, and possible • • • • sepsis. PMH : Morbidly obese, HTN, DM II Allergy : NKDA Laboratory Data: • U/A: blood, protein, high leukocyte esterase • CBC/CMP : WBC 21K, glucose 297 Assessment and Plan: • Urosepsis • Order urine and blood cultures • Levofloxacin 500 mg IV Daily : EMPIRIC THERAPY • Empiric (pathogen not known but treat based on common bacteria at the site of infection) vs. Definitive (Pathogen & sensitivity known) Therapy 5 ID Consult: 8/21/2024 antibiotics resistance to high no • Blood culture: (2/2) positive for gram negative bacilli (GNB, collected 8/19/24 7:55) • Klebsiella pneumoniae • Urine culture: GNB (collected 8/19/24 22:00) • Klebsiella pneumoniae 6 LABORATORY TEST É Culture Susceptibility/Sensitivity Test •a Disk Diffusion including E-test • • Microdilution Methods Automated Tests so They usetesttubes 7 solidoglerplates as asabsrate per Disk Diffusion Susceptibility Testing—Kirby Bauer Method • A “standardized” suspension of the organism is used to inoculate agar (generally Mueller-Hinton) surface • FilterOkay paper disks containing antibiotics are placed on surface of agar • concentration gradient • incubated 16-24 hrs • Zones of inhibition are measured (mm) • CLSI (Clinical Laboratory Standard Institute) interpretive chart • Qualitative (categorical) results: S (susceptible), I (intermediate), R (Resistant) 0 8 Etest Diffusion Susceptibility Testing • Variation of disk diffusion method • Etest strips replace filter paper disks • MIC is read where “ellipse” (growth inhibition) intersects the strip at • Allows quantitative determination of MIC on agar • More expensive has eachpart concentration different 2 M 9 mininhibitory concentration MIC Dilution Susceptibility Testing be cleanesttesttabwilovastroncentration cleanesttesttube w loves Contentration B MIC • A range of antibiotics concentrations is established • • • • • • in agar or broth The antibiotic concentration is reduced by half with each successive dilution onor sowell is inoculated with Each plate, tube, approximately the same amount of bacteria (inoculums, usually 105 – 106 CFU/mL) and incubated Examine all plates or tubes in the series for growth (colonies or turbidity) The MIC is the lowest concentration of antibiotic(mg/L) that inhibits the growth of the bacteria Reliable, standardized reference method Quantitative results McCoy MEE • MBC(Minimum Bactericidal Concentration) testing may be performed with broth dilution method • Remove an aliquot from “clear” tubes or wells and transfer to an agar plate; incubate • Each colony represents one viable bacterium • MBC is the concentration of antibiotic that kills 99.9% or 3-logs of the original inoculum 10 3 Automated Susceptibility Tests • Utilizes robotics, micro-processors, micro- computers to provide results • VITEK 2TM, MicroscanTM, SensititreTM, PhoenixTM • Rapid results • May not detect subtle resistance, but results improve as technology improves • VITEK 2TM • Uses “test cards” • Micro-wells (64 wells/card) contain antimicrobials for susceptibility testing • Wells are inoculated with bacterial suspension • Photometer measures turbidity changes over time E • Computer compiles data and calculates MIC • Can also be used for ID purposes 11 Rapid Molecular Methods for Organism Identification : Nucleic Acid Based Test Name/Time Pathogens detected Limitations less accurate in detecting all organisms within polymicrobial infections https://www.luminexcorp.com/theverigene-system/#overview Staph, Strep, Enterococcus, Listeria, E.coli, Citrobacter, proteus, Acinetobacter, Serratia, PSA, Klebsiella, Enterobacter, Fungal, Viral, Resistant ones PNA FISH (Peptide Nucleic Acid l Fluorescence In Situ Hybridization)/1-2 hrs S. aureus, Enterococcus, E.coli, K pneumoniae, PSA, Candida Staph is the only one with resistance gene detection (mec A) Bacteria, Fungi, Mycobacteria Identification of resistance marker is not currently available o Multiplex PCR (Verigene or FilmArray)/12.5 hrs 3MALD-TOF MS (Matrix Assisted Laser Desorption Ionization Time of Flight Mass Spectrometry) /0.2 hr Perez KK, Microbiology Rapid Diagnostics, in 2018 ACCP BCIDP Prep lb24hr tosee ifthere'ssomething 12 BACTERIAL CLASSIFICATION 13 Gram Stain • Quick, easy diagnostic tool • Detects peptidoglycan in thicker layer of G(+) 14 Gram Stain • Divides bacteria into 2 groups based on cell walls: a • Gram Stain (+) : Stain adhering to rigid peptidoglycan layer  Purple/Blue Who www.n 2 • Gram Stain (-) : Lipopolysaccharides blocks the stain from adhere peptidoglycan  Pink/Red • Determines cell morphology • Shape : cocci or bacilli • Size • Arrangement : cluster vs. chain 15 What are they?gram t y Cosas by Y Tambe 2010 16 Oxygen Requirements 8133 • Anaerobic—intolerant of oxygen; require proper handling and special conditions for growth A's • Aerobic—utilizes oxygen; oxygen is required for growth •aFacultative (optional) anaerobic—will grow in presence of oxygen; however, if oxygen is not present, can grow in its absence 17 r 18 0000 O Streptococcus spp. • Group A Strep (GAS): e.g., Streptococcus pyogenes • Can cause pharyngitis, scarlet fever, skin and soft tissue infections • Invasive GAS diseases = necrotizing fasciitis, streptococcal toxic shock syndrome • Group B Strep : e.g., Streptococcus agalactiae tf • Most common cause of neonatal sepsis and meningitis • CDC recommendation sis • screen pregnant women 35-37 weeks gestation (~10-30% are asymptomatic carriers) • If Positive, IV Penicillin during delivery • If allergic to penicillin, cefazolin or vancomycin 19 Streptococcus spp. respiratory upper Issus Infections • Streptococcus pneumoniae t • Pneumococcus : pneumococcal vaccine target Middleearinfection no • Can cause community-acquired pneumonia, otitis media, sinusitis, meningitis • Estimated that up to 45% of strains (14% in CA) are penicillin resistant • Sputum gram stain and culture (57% and 79% sensitive) • Streptococcus viridans • Miscellaneous Strep group; part of normal flora of oral, respiratory, and Gastrointestinal mucosa • Opportunistic pathogen; low virulence • One of major etiological agents of endocarditis in the U.S 20 Enterococcus spp. be insidethebody • Normal Flora of the GI tract hospital was • Can cause UTIs, nosocomial infections, wound infections e • Highly resistant to many antibiotics • Enterococcus faecalis Grant Moe Resistant Enterococcus • Enterococcus faecium—Vancomycin 21 Staphylococci spp. animals and infections • Arranged in clusters resembling grapes inhares cause me Coagulase positive = S. aureus Coagulase negative Staphylococcus (CNS) Example, S. epidermis • Normal flora of surface epithelium • Catalase positive 22 Staphylococcus aureus (SA) t • Colonizes nasal passages a • Virulent pathogen causes • skin/wound infections, toxic shock syndrome • Pneumonia • Osteomyelitis thathappensinthebond Informationorsaying • food poisoning • Methicillin Resistant Staphylococcus aureus (MRSA) • common cause of nosocomial(=Hospital acquired) infection 23 Coagulase Negative Staphylococci (CoNS) • Staphylococcus epidermidis • 75% of CoNS • Normal flora of the skin • Opportunistic pathogen • Nosocomial infections • Catheter sites, prosthetic joint infections • Other CoNS • S. saprophyticus : UTI in young adult women • S. lugdunensis : Endocarditis or osteomyelitis • S. hominis, S. haemolyticus, S. capitis, etc : infrequently isolated 33 0 needto Dowe knowtis 24 Clostridioides spp. (previously Clostridium) • Anaerobic bacilli • Commonly found in soils and Gastrointestinal (GI) tracts of humans and animals • Produce highly potent toxins that are often responsible for symptoms • Clostridioides difficile infections (CDI) • antibiotic-associated diarrhea and pseudomembraneous enterocolitis • Colonization is common in hospitals and nursing homes • Spores, wash with soap and water (not alcohol) • Vancomycin PO, fidaxomicin PO (DIFICID) 25 Listeria monocytogenes • Causes Listeriosis, ~2500 cases/yr in U.S. • Seen almost exclusively in pregnant women and in immunocompromised individuals • Untreated/trans-placental infection can result in premature labor, septic abortion, neonatal meningitis • Can grow in refrigerator • Mortality rate for listeriosis is around 15-20% • Drug of Choice : Ampicillin 26 Gram Positive – Acid Fast Mycobacterium spp. • Classified as Gram positive because they lack an outer membrane • Very slow-growing (12-18 hrs to double); colonies not visible in lab for 3- 6 weeks • Stain acid fast (not easily decolorized by acid) with Ziehl-Neelson stain • Drug resistance due to mutations and unique cell wall composition: Combination of multi agents required 27 Gram Positive – Acid Fast Mycobacterium tuberculosis (Tb) • Humans are only reservoir 3 • Tb is a leading cause of worldwide death from bacterial infectious disease • Multiple drug treatment due to resistance Nocardia spp. • Commonly found in soil • Opportunistic pathogen in the immunocompromised, typically pneumonia 28 30 31 Neisseria spp. • G(-) Diplococci • Neisseria meningitidis • Meningococcus, meningococcal vaccine target • Colonizes nasopharynx É and cause meningitis • Can disseminate • Bacteremia, petechial rash, pneumonia • Neisseria gonorrhoeae • Causes gonorrhea (Sexually Transmitted Infection) 32 Moraxella catarrhalis (=M. Cat) • Acute, localized community acquired respiratory infections = otitis media, sinusitis, conjunctivitis, bronchopneumonia • Systemic diseases = endocarditis, meningitis • most cases are in elderly patients • >90% beta-lactamase positive (penicillin-resistant) • ampicillin, amoxicillin, or penicillin do not work against M. cat 33 Pseudomonas aeruginosa(PSA) hospital Major cause of nosocomial infections • Pneumonia (18%) • UTI (16%) • Surgical site infections (9.5%) • Blood stream infections (3.4%) • Skin and soft tissue • Others Lactose (glucose) non-fermenter • Not part of normal flora in healthy individuals • Environmental organism in water, soil, plants • Well-adapted to harsh environmental conditions, able to grow in distilled water! 34 Pseudomonas aeruginosa • Classic opportunistic pathogen • Typically infects pulmonary, urinary tracts, burns and wounds • Causes a broad spectrum of diseases • Maintains antibiotic resistance plasmids 0g• Mutate easily, develop resistance quickly in monotherapy • Pigmented and fluoresces under UV light 35 Other glucose NON-fermenters • Acinetobacter baumannii • • • • Noted increase in antibiotic resistance Will be discussed later more detail Drug of Choice (DOC): Sulbactam Combination Tx • Stenotrophomonas maltophilia • Low in virulence • Often colonizes more frequently than acts as an infectious agent • • • • May cause septicemia, pneumonia, wound infections in ICU Often multi-drug resistant DOC: TMP/SMX Combination Tx see thisbacteriain UI 36 Enterobacterales (=Previously Enterobacteriaceae) • Glucose Fermenters • Many are normal gastrointestinal flora in humans and other animals, also found in soil, water, and on plants • Often referred to as ‘enterics’ stop • Numerous pathogens are in this family • Urinary tract infections (UTI), gastrointestinal (GI), or intra-abdominal infections (IAI) • Escherichia coli • Common cause of Urinary Tract Infections (UTI), Gastrointestinal/Intraabdominal infections • O157:H7 is most infamous pathogenic strain of Gastrointestinal infections 37 Enterobacterales • Klebsiella pneumoniae, K. oxytoca a skinandsofttissueinfection • Normal flora, opportunistic UTI, pneumonia, SSTI, etc • Enterobacter spp. • Several pathogenic strains, causing mainly opportunistic infections of the urinary and respiratory tract in immunocompromised • Often AmpC carrier, resistant gene • Proteus mirabilis to B lactamases • UTIs, an alkaline urine sample • Serratia spp. • Citrobacter spp. • Salmonella enteriditis • Foodborne illness associated with raw eggs 38 Other Gram Negatives metronidazole Bacteriodes fragilisas • Gram negative anaerobes causing intra-abdominal infections(IAI)s sites Haemophilis influenzae • NOT related with flu or influenza (by virus) • Fastidious BACTERIA in labs; grows on chocolate agar • Non-encapsulated strain: normal flora in nasopharynx of ~75% of healthy children and adults • Can cause meningitis, conjunctivitis, otitis media (Community Acquired Respiratory Infections) • Hib vaccine for protection against strain belonging to serotype B, capsulated strain • 30% produce beta-lactamase (resistant to penicillin), ampicillin and amoxicillin should only be used if susceptibility confirmed I'm 39 Other Gram Negatives Legionella pneumophila • Widespread in environment; found often in warm water (hot tubs, hot water heaters, humidifiers, and air conditioning systems) • Aerosol transmission, not person-to-person • Cause Legionnaire’s Disease Bordetella pertussis • Resides on mucous membranes in respiratory tracts of humans and animals • “P” in DPT vaccine • Can cause “whooping cough” Helicobacter pylori • “helical” shaped microaerophile • Catalase, oxidase, and urease positive • Can thrive in acidic environment of stomach; major habitat is gastric mucosa • Etiological agent of gastritis and gastric ulcers High resistance Gram-positive Gram-negative Enterobacterales Strep & Enterococci Enterococcus faecium (VRE) Serratia Klebsiella Atypical resp pneumoniae pathogens E. coli Proteus Legionella/Mycoplasma/ mirabilis Chlamydophila Enterococcus faecalis S. pneumoniae Strep pyogenes Low resistance Staph aureus (MSSA) Staph aureus (MRSA) H. influenzae Respiratory GNR Enterobacter spp Bacteroides fragilis Pseudomonas AmpC producers Staphylococci anaerobes 43 in so 44 Predominant Microbiota at Various Body Sites (Sterile site?) Body site Microbiota Skin Streptococci, Staphylococci, Propionibacterium, Corynebacterium(diphtheroids), Coagulasenegative staphylococci Oral cavity (teeth, mucous membrane) Candida albicans, Neisseria spp., Moraxella, Streptococci, Peptostreptococcus Upper respiratory tract (nares, pharynx) Streptococci, Staphylococci, Neisseria, group A streptococci, Gram (-) (M. cat, H. influenzae) Gastrointestinal tract Stomach Small intestine Colon e Anterior urethra H. Pylori Lactobacilli Enterobacterales (E. coli) Bacteroides fragilis, Candida, Clostirioides, Eubacterium, Fusobacterium, Enterococcus Staphylococci, Corynebacterium, Enterics (Enterobacterales, Enterococcus) Hi s Vagina C. albicans, Diphtheroids, Staphylococci, Enterobacterales, Lactobacilli, Streptococci dit g show clean 45 Summary • Bacterial Classification • (Gram positive vs. gram negative, anaerobes vs. aerobes, cocci vs. bacilli, non-glucose fermenters vs. fermenters) • Gram Positive Bacteria • Cocci in chain : Strep, skin, upper respiratory infections • S. pneumoniae, upper respiratory infections • Group A strep, S. pyogenes • Cocci in clusters : Staphylococcus • Coagulase (-) : S. epidermidis, skin or catheter, devices • Coagulase (+) : S. aureus, MSSA, MRSA, skin, other infections (MRSA, hospital related infections) • C. difficile : nosocomial antimicrobial related diarrhea • Gram Negative Bacteria • Gram negative bacilli glucose/lactose non-fermenter causing nosocomial infections : P. aeruginosa • Gram negative bacilli glucose fermenter causing UTI : E. coli • Anaerobes causing IAI : B. fragilis 46 Summary • Normal flora • Pathogens for each infectious disease status (respiratory, urinary, intra-abdominal, skin/line infection), sterile site? • Laboratory Test • MIC/Susceptibility/sensitivity Test • Disc diffusion vs. Microdilution test • Rapid Molecular Methods for Organism Identification: Nucleic Acid Based Test Mtn inhibitoryconcentration

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