Aerobic Gram Negative Bacilli (GNB) 1 PDF - RCSI 2024

RCSI Royal College of Surgeons in Ireland Coláiste Ríoga na Máinleá in Éirinn SESSION ID: GIHEPMicroL1 Aerobic Gram Negative Bacilli (GNB) 1 *Coliforms, *Proteus, Pseudomonas Class Year 2 Course Undergraduate Medicine Lectu...

RCSI Royal College of Surgeons in Ireland Coláiste Ríoga na Máinleá in Éirinn SESSION ID: GIHEPMicroL1 Aerobic Gram Negative Bacilli (GNB) 1 *Coliforms, *Proteus, Pseudomonas Class Year 2 Course Undergraduate Medicine Lecturer Dr. Rachel Grainger Date 10th September 2024 * The term “Enterobacterales” includes these  Learning Outcomes By the end of the lecture, you will be able to: 1. Outline the basic laboratory features of clinically important Enterobacterales & Pseudomonas species and explain the biological role of each in the pathogenesis of infection 2. Discuss the epidemiology of clinically important Enterobacterales & Pseudomonas species 3. Describe the pathogenesis of infections caused by clinically important Enterobacterales & Pseudomonas species 4. Recognise and describe the clinical features and complications of infections caused by clinically important Enterobacterales & Pseudomonas species.  Learning Outcomes By the end of the lecture, you will be able to: 5. Outline the laboratory diagnosis of infections caused by clinically important Enterobacterales & Pseudomonas species and describe their laboratory features e.g. Gram stain appearance etc. 6. Choose the appropriate antimicrobial agents to treat infections caused by clinically important Enterobacterales & Pseudomonas species 7. Use the appropriate measures to prevent the acquisition and spread of infections caused by clinically important Enterobacterales & Pseudomonas species Introduction gram +: pink/ red gram -: blue, purple 1. Enterobacterales a. A family of gram-negative (pink) bacilli (rod-shapped) b. ‘Enteric’ = relating to or occurring in the intestines (GIT is their habitat) c. Common causes of intra-abdominal, respiratory tract & bloodstream infections d. Increasing antibiotic resistance with these organisms i. Beta-lactamase production including extended-spectrum beta-lactamases (ESBLs) 2. Pseudomonas spp. & related genera ENTEROBACTERALES: CLASSIFICATION Normal intestinal flora Escherichia coli Important organisms, Klebsiella spp. covered in this Proteus spp. lecture, revise GNB lecture in FFP2 also Others Serratia spp. Enterobacter spp. Less important Citrobacter spp. Pathogens (not normal flora) associated with the exposure to antibiotics Salmonella spp. Important, covered in next Shigella spp. lecture Yersinia spp. Important, covered in Enteric Toxin-producing E.coli infections normal flora of the bile ENTEROBACTERALES: MICROBIOLOGY Most are motile with flagellae Facultative anaerobes distinguish between the lactose fermenters and non-lactose fermenters can grow in both the presence or absence of oxygen (fermenters turn pink, non-lactose fermenters are pale or colourless) (thy prefer using oxygen to produce energy but when there is no oxygen they switch to the anaerobic mechanisms or fermentation to generate the energy) Ferment glucose and other carbohydrates May be lactose fermenters or non-lactose fermenters lactose fermenters (break down the lactose)  ENTEROBACTERALES: STRUCTURE Cell wall (contains Capsule (contains lipopolysaccharide O-antigen) K-antigen) evade the immune system - help the bacteria hide from the immune system (prevent the phagocytosis of the bacteria by the immune cells) Fimbriae Flagella (H-antigen)  ENTEROBACTERALES: STRUCTURE (THE GRAM- NEGATIVE CELL WALL) endotoxin: gram negative, only released when the bacteria dies Composed of: to avoide the immune system exotoxins: both gram positive and negative,, released when the bacteria is O-side-chains alive.(more severe) Lipid Aacts as a toxin (endotoxin)  Pathogenesis Gets in – portal molecules that allow the of entry bacteria to attache to the Contact, epithelial cells environment, Adhesins, pili AMR antimicrobial resistance through the Gets out & Attaches to plasmids spreads further cells LPS, toxins Causes Defeats/evades Capsule LPS contain damage to host the immune antigen K helps avoid immune recognition the endotoxin cells system lipid A  Cystitis is inflammation of the bladder, usually caused by a bladder infection VIRULENCE FACTORS Adhesins – aid in binding to host cells, e.g. fimbriae or pili Capsules – help avoid phagocytosis but poor immunogens they do not elicit an immune response (the immune system recognize the molecules on the capsule as self) Lipopolysaccharide – potent inducer of host immune response via endotoxin release (lipid A) – Endotoxin  activation of complement, cytokines & disseminated intravascular coagulation WBCs  decrease in platelets  DIC  fever, hypotension, death Toxins – e.g. haemolysins of E. coli Antimicrobial resistance – e.g. via plasmid exchange the LPS activate many systems in the body including the clotting cascade. when this is activated, it will result in the formation of small blood clots in the small vessels which will consume a large amount of platelets and this will reduce the ability of the body to react in the bleeding cases (so, this will enhance the bleeding problems as well as the clotting problems - because there are still small clots forming in the small vessels and this will result in the blockage of these vessels leading to organ damage )  CASE SCENARIO 1 A 22-year-old female attends her GP C/o urinary frequency & dysuria Otherwise systemically well Has not recently been on antibiotics vaginal She has no PV discharge. WHICH ONE OF THE FOLLOWING IS THE MOST LIKELY DIAGNOSIS? A.Cervical carcinoma B.Chlamydia urethritis C.Cystitis inflammation of the bladder D.Pyelonephritis presents with flank pain E.Vaginal thrush  CASE SCENARIO 1 Empiric Treatment (www.antibioticprescribing.ie ): NITROFURANTOIN PO for 3 days TRIMETHOPRIM PO for 3 days FOSFOMYCIN single dose  CASE SCENARIO 1 Laboratory Report: Day 1: Urine white cell count >100/ µl Day 2: E. coli isolated from urine; susceptibilities pending Day 3: E. coli – SUSCEPTIBLE to NITROFURANTOIN / TRIMETHOPRIM no need for more than 3 days She completes a three day course of nitrofurantoin Symptoms resolve & she requires no further investigations or follow-up CASE SCENARIO 2 A 36-year-old female presents to the ED with vomiting, left flank pain and a temperature of 38.8oC. flank pain = pyelonephritis WHICH OF THE FOLLOWING IS THE MOST LIKELY SOURCE? A. Appendicitis B. Diverticulitis C. Pneumonia D. Pancreatitis E. Pyelonephritis kidneys WHICH OF THE FOLLOWING MICROBIOLOGY INVESTIGATIONS ARE MOST IMPORTANT INITIALLY? A. Aspirate from left kidney for Gram stain B. Blood cultures alone C. Faeces (stool) for culture/PCR D. MSU & Blood cultures MSU = midstream specimen urine E. Pro-calcitonin level CASE SCENARIO 2 Empiric Treatment (Guidelines App): IV CEFUROXIME +/- GENTAMICIN Nitrofurantoin effective only against the lower urinary tract infections (the upper ones like the pyelonephritis require a wider-spectrum antibiotic) also, the Nitrofurantoin is secreted throgh the kidney, if the kidneys are not working properly like in this case, Nitrofurantoin cannot be Laboratory Report administered) Day 1: Urine white cell count >100/ µl Day 2: Escherichia coli isolated from urine; susceptibilities pending Day 3: Escherichia coli – RESISTANT to TRIMETHOPRIM / CO-AMOXICLAV – SUSCEPTIBLE to NITROFURANTOIN / CEFUROXIME / GENTAMICIN N.B. Nitrofurantoin not suitable in this case  CASE SCENARIO 3 A 2-day-old baby presents with: – Tachypnoea – Feeding poorly – Irritable when handled Meningitis is suspected E.coli must have been transmitted from the mother during delivery (early neonatal meningitis) WHICH ONE OF THE FOLLOWING IS THE MOST LIKELY CAUSATIVE PATHOGEN? A. Bordetella pertussis B. E. coli C. Haemophilus influenzae D. Staphylococcus epidermidis E. Streptococcus pneumoniae  CASE SCENARIO 3 Empiric Treatment (Temple St. Guidelines): IV CEFOTAXIME / AMOXICILLIN / GENTAMICIN  CASE SCENARIO 3 Empiric Treatment (Temple St. Guidelines): IV CEFOTAXIME/AMOXICILLIN/GENTAMICIN Laboratory Report (CSF Microscopy, Culture and Susceptibilities) Day 1: CSF white cell count 600/µl (normally 100/ µl This organism is an ESBL- Blood cultures positive after 9 hours: GNB in both bottles producer & represents an Day 2: Klebsiella pneumoniae isolated from blood and urine; infection control Susceptibilities pendingrisk. Day 3: Klebsiella pneumoniae – RESISTANT to CO-AMOXICLAV/ CEFUROXIME/ CIPROFLOXACIN – SUSCEPTIBLE to PIPERACILLIN-TAZOBACTAM/ GENTAMICIN ESBL: EXTENDED-SPECTRUM BETA- LACTAMASE Enzymes carried by Enterobacterales (e.g. E.coli, Klebsiella spp.) which make them resistant to: MOA like penecillins (interfere with cell wall synthesis) – Cephalosporin antibiotics – Sometimes co-amoxiclav (“Augmentin”) – Sometimes piperacillin-tazobactam (“Tazocin”) Cause UTIs, intra-abdominal infections High prevalence of ESBL producers in nursing homes in Ireland  IS THERE A PROBLEM WITH MULTIPLE ANTIMICROBIAL RESISTANCE IN ENTEROBACTERALES?? Electron microscopic image by Charles C. Brinton, Jr. Plasmid The genes for CPE & ESBL enzymes are carried on PLASMIDS A plasmid is a piece of genetic material which can easily be transmitted from one bacterium to another  CASE SCENARIO 6 A 70yo female is transferred to your ICU from an ICU in Greece Suffered an intra-cerebral bleed while on holidays Has just completed a course of meropenem for VAP Currently has no symptoms or signs of infection.  CASE SCENARIO 6 Lab Report (Rectal Swab for CPE screen) on Day 4: Klebsiella pneumoniae isolated RESISTANT to AMOXICILLIN, CO-AMOXICLAV, CEFUROXIME, CEFOTAXIME, CIPROFLOXACIN, GENTAMICIN, PIPERACILLI-TAZOBACTAM, MEROPENEM This organism is a carbapenemase producer and represents and infection control risk. No antibiotic treatment indicated (colonisation only) N.B. Isolate with high-level contact precautions (contact plus) CPE: CARBAPENEMASE-PRODUCING Carbapenemase-Producing Enterobacteriaceae (CPE) are ENTEROBACTERALES considered particularly dangerous because they produce enzymes that can inactivate carbapenems, which are often used as last-resort antibiotics for treating severe infections when other antibiotics have failed. Enterobacterales (e.g. E.coli, Klebsiella spp.) resistant to meropenem (our “last-resort” antibiotic) – They produce enzymes called carbapenamases e.g. IMP, KPC, OXA-48, NDM Also resistant to many other classes of antibiotics Live in the bowel More & more evidence that CPE widespread in hospital environment Management of infection with these organisms is a challenge (e.g. UTIs, intra-abdominal infection) because the organisms are resistant to most antibiotics  CPE IN IRELAND 2013-2022 Source: HPSC – Enhanced surveillance of CPE in Ireland ENTEROBACTERALES: PROTEUS MIRABILIS Microbiology Non-lactose fermenter Characteristic “swarming” on agar: spreads out and takes over the plate Unpleasant fish-like smell Oxidase-negative oxidase test is used to distinguish between the types of the non-lactose fermenters bacteria  ENTEROBACTERALES: PROTEUS MIRABILIS Proteus mirabilis is most common Proteus species causing infection Normal GI flora; may be pathogenic at other sites May cause: – Urinary tract infection (Patients with urinary tract abnormalities/long-term catheters) – Bloodstream infection (Often related to urosepsis) Infection is often healthcare-associated  ENTEROBACTERALES: PROTEUS MIRABILIS associated with the urinary stone formation (kidney or bladder stones) staghorn calculus (large, branched kidney stone ENTEROBACTERALES: ENTEROBACTER SPP., SERRATIA SPP., CITROBACTER SPP. May form part of the normal intestinal flora but are typically found in the environment Not intrinsically pathogenic Cause healthcare-associated infections (HCAI) (e.g., pneumonia, intravascular catheter sepsis, UTI, abdominal wound), infection/abscess) in at-risk patients Often resistant to multiple antibiotics  CASE SCENARIO 7 A 25yo male with CF is admitted to hospital with: – Increased sputum production – Breathlessness A sputum sample is taken for culture. WHICH ONE OF THE FOLLOWING IS THE MOST LIKELY CAUSE? A.Enterococcus faecalis B.Haemophilus para- influenzae C.Listeria monocytogenes D.Pseudomonas aeruginosa E.Staphylococcus saprophyticus  PSEUDOMONAS SPP. EPIDEMIOLOGY Pseudomonas aeruginosa is the most important clinically Widespread in moist areas in the environment – Sinks, drains – Also soil, plants, animals Difficult to treat – Intrinsically resistant to many antibiotics – Acquires resistance quickly Forms biofilm e.g., in taps virulece factor (makes it harder for the antibiotic to reach the bacteria) http://ru.pall.com/main/medical/scientific-information-45031.page  P. AERUGINOSA: MICROBIOLOGY Non-lactose fermenter Oxidase positive Strict aerobe P. AERUGINOSA: MICROBIOLOGY Mucoid variants of Pseudomonas aeruginosa are seen in cystic fibrosis. These form biofilm in the respiratory tract. VIRULENCE FACTORS Pili Flagellae (adhesins), Polysaccharide capsule, Pyocyanin which impairs cilia Endotoxin, i.e. lipopolysaccharide  Pathogenesis Gets in – portal of entry Contact, environment, Adhesins, pili AMR Gets out & Attaches to spreads further cells Capsule, LPS, toxins Causes Defeats/evades biofilm, damage to host the immune pyocyanin cells system P. AERUGINOSA: CLINICAL PRESENTATIONS Bloodstream infection (e.g. in burns patients) Neutropenic sepsis (e.g., in neutropenic = low white blood cells count haematology/oncology patients) neonatal intensive care units Outbreaks in NICU - Preterm babies Pneumonia – Acute pneumonia in ventilated patients (VAP) – Chronic colonisation/infection in http://www.medscape.org/viewarticle/458771_5 cystic fibrosis, and bronchiectasis Urinary tract infection – Often complicated UTI, associated with urinary catheters P. AERUGINOSA: CLINICAL PRESENTATIONS eye and ears are moist areas -> perfect environment for P.Aerguinosa Eye infection – Post- trauma or surgery – Associated with contaminated fluids e.g., contact lens solutions – Can lead to destruction of eye loss of vision http://webeye.ophth.uiowa.edu/eyeforum /cases/171-pseudomonas-keratitis.htm Ear infections – Otitis externa in swimmers – May be severe in diabetics: malignant otitis externa the infection can move to the bones resulting in osteomylitis TREATMENT OF P. AERUGINOSA Fewer treatment options than coliforms as intrinsically resistant to many antibiotics - Piperacillin-tazobactam - Ceftazidime (3rd generation cephalosporin) - Ciprofloxacin (quinolone) - Aminoglycosides e.g. gentamicin - Carbapenems e.g. meropenem (reserved)  OPPORTUNISTIC PATHOGENS Organism of low intrinsic virulence which cause infection when the host defences are impaired Typically in patients whose immune system is compromised by disease or treatment of disease (e.g. malignancy, high dose corticosteroids)  BURKHOLDERIA CEPACIA A major cause of opportunistic infection in the lungs of patients with cystic fibrosis (CF) Easily transmitted by social contact Multi-resistant and difficult to treat Causes “cepacia syndrome” – Acute fatal necrotising pneumonia +/- BSI BURKHOLDERIA PSEUDOMALLEI Causes melioidosis, seen in SE Asia, AUS Reservoir: water e.g. paddy fields May cause severe pneumonia with BSI & high mortality even in normal hosts  STENOTROPHOMONAS MALTOPHILIA An opportunistic pathogen May be a coloniser in – Hospitalised patients – Respiratory tract of patients with cystic fibrosis Usually following prolonged broad- spectrum antibiotics – Bloodstream infections (often associated with central venous catheter) http://lib.jiangnan.edu.cn/ASM/112-19.jpg – Pneumonia Intrinsically resistant to many antibiotics  ACINETOBACTER BAUMANNII Epidemiology: Can survive on wet & dry surfaces in hospital environments Multi-resistant strains occur – e.g., seen in injured US soldiers returning from Iraq & Afghanistan in 2000s – Can be associated with hospital outbreaks, especially in ICU setting – Very limited treatment options  1 2 4 5 6 7 8 3 SAMPLE PSA MCQ: A, B, C are not the answer because she is allergic to penicillins Case presentation: A 22 year female attends her GP complaining of a 2 day history of urinary A. Amoxicillin 250mg frequency and dysuria, she is systemically well. twice daily Past medical history includes two previous B. Amoxicillin 500mg urinary tract infections. She is allergic to three times daily penicillin. C. Nitrofurantoin Investigations: Recent MSU cultured an immediate release organism with the following sensitivities: capsules 50mg four Amoxicillin: Sensitive times daily Trimethoprim: Resistant D. Piperacillin- Nitrofurantoin: Sensitive tazobactam 4.5 four Question: Select the most appropriate times daily prescription based on the most recent urine E. Trimethoprim 200mg culture result twice daily  SUMMARY There is increasing recognition of relatively non-pathogenic GNBs as a cause of opportunistic infection Emerging – and spreading – antibiotic resistance in these organisms (e.g., ESBL- producers, CPE) is a major challenge