Gram-Positive Bacilli Lecture Notes PDF

Gram-Positive Bacilli PATH 3100 Prof. Mal Leetham  2 Objectives Separate common genera of Gram-positive bacilli based on the presence or absence of spore production Describe the co...

Gram-Positive Bacilli PATH 3100 Prof. Mal Leetham  2 Objectives Separate common genera of Gram-positive bacilli based on the presence or absence of spore production Describe the common diseases associated with Gram- positive bacilli and the associated bacteria Differentiate normal microbiome and environmental Gram-positive bacilli from the pathogens  3 Classify according to spore production No Spores: Spores:  4 Classify according to spore production Medically important Gram-positive bacilli: No Spores: Spores: Listeria Clostridium/ Corynebacterium Clostridioides Gardnerella Bacillus spp. Nocardia Lots more!  5 Differentiate Gram-positive bacilli Opportunistic pathogens: Primary pathogens: Corynebacterium Listeria monocytogenes jeikeium Corynebacterium Gardnerella vaginalis diphtheriae Bacillus spp. Nocardia spp. Clostridium spp. Bacillus anthracis Clostridium botulinum NON-SPORE FORMERS  7 Listeria monocytogenes Characteristics: Motile at 20-25 °C, facultatively anaerobic, non-spore-forming Gram-positive bacilli Frequently found in the environment Psychrophile – can grow at refrigerated temperatures Causes significant disease in neonates, pregnant persons, immunocompromised Foodborne illness. Causes listeriosis.  8 Listeria Transmission Common soil contaminant Most human infections are a result of the consumption of contaminated food: – uncooked meats and vegetables – soft cheeses, processed meats – Unpasteurized milk and cheese https://www.cdc.gov/listeria/outbreaks/delimeats-7- https://www.fda.gov/food/outbreaks-foodborne-illness/outbreak-investigatio 24/index.html accessed: 07/30/24 listeria-ice-cream-august-2023 accessed: 09/14/23  10 WARNING!  11 Case Study A 21-year-old female presents to the clinic 27 weeks pregnant. She presented with complaints of fever, headache, chills, frequency and urgency on urination, decreased appetite, and a 1-day history of diarrhea and decreased fetal movement. She had a temperature of 38.3ºC, abdominal tenderness, and tachycardia. Her chest was clear, and no cervical discharge or tenderness was seen on the pelvic exam. Her WBC was 21,300. The next morning the patient complained of abdominal pain. On ultrasound, there was no fetal movement, and with labor induced, a stillborn infant was delivered vaginally.  12 Listeriosis in humans Listeriosis is a severe infection caused by eating food contaminated with Listeria monocytogenes Healthy adults may experience self-limited acute febrile gastroenteritis following high-dose Listeria exposure. Other symptoms may include watery diarrhea, fever, nausea, headache, myalgia, arthralgia Older adults and immunocompromised people may experience invasive infections: sepsis, meningitis, and meningoencephalitis. focal infections: septic arthritis, osteomyelitis, prosthetic graft infections  13 Listeriosis in pregnant people and neonates Pregnant people may experience mild “flu-like” illness or have no symptoms of listeriosis Infection during pregnancy can result in: Miscarriage Stillbirth Preterm labor Sepsis or meningitis in the neonate  14 Listeria epidemiology In the United States, an estimated 1,600 people become seriously ill with listeriosis each year, and 260 die. Severe Listeriosis infection has a higher mortality rate than most other foodborne illnesses (~20-30%). Nearly one-quarter of pregnancy-associated cases result in fetal loss or death of the newborn. https://www.cdc.gov/listeria/technical.html#clinical-features  15 Listeria Lab Identification Specimen: GI illness: will not grow in a standard stool culture Meningitis: CSF or blood Methods: Culture Molecular detection (such as PCR) Serotyping Mahon, C. R., & Lehman, D. C. (2019). Textbook of Diagnostic Microbiology. St. Louis, MO: Elsevier Saunders.  16 Corynebacterium species (Diphtheroids) Gram-positive bacilli pleomorphic palisading “L’s” and “V’s” C. diphtheriae and the non- diphtherial corynebacteria are known as diphtheroids Photo: Becky Buxton  17 Corynebacterium species (Diphtheroids) Characteristics of the genus: Found in the environment Normal microbiota on skin, in the oral cavity, and in the genitourinary tract of humans Opportunistic pathogen Usually considered a contaminant in clinical cultures  18 Corynebacterium diphtheriae Causes the disease diphtheria: pharyngitis with a thick, leathery membrane Vaccine-preventable disease Widespread use of diphtheria vaccination since 1930s Last reported case of diphtheria in the US was in 2003 https://www.cdc.gov/diphtheria/index.html  19 Corynebacterium diphtheriae Only toxigenic strains of C. diphtheriae secrete the potent exotoxin that causes diphtheria The bacteria must be infected with a phage (virus) that carries the tox gene to produce the exotoxin Strains of C. diphtheriae that are not infected with the tox phage do not cause diphtheria Image: https://www.technologynetworks.com/immunology/ar ticles/lytic-vs-lysogenic-understanding-bacteriophage- life-cycles-308094 accessed: 07/30/24  20 Acute Diphtheria Communicable disease Respiratory diphtheria ü Nasal diphtheria ü Pharyngeal and tonsillar diphtheria ü Laryngeal diphtheria Cutaneous diphtheria caused by non-diphtheria Corynebacterium spp. Transmission is primarily person-to-person via respiratory droplets Portals of entry for Corynebacterium diphtheriae are nose, tonsils, and throat The incubation period is 2–5 days https://www.cdc.gov/diphtheria/clinicians.html  21 Diphtheria - Pseudo-Membrane Symptoms: Mild fever, sore throat, difficulty swallowing, malaise, loss of appetite Pseudo-membrane is the hallmark of respiratory diphtheria Pseudo-membrane appears within 2–3 days of illness over the mucous lining of the tonsils, pharynx, larynx, or nares. Potential for fatal airway obstruction if the pseudo-membrane extends into FIGURE 21-5 Pharynx of a 39-year-old woman with the larynx or trachea. bacteriologically confirmed diphtheria. Murray, et al. Medical Microbiology, 8th ed. Accessed 6/21/17  22 Reported Cases of Diphtheria in US: Treatment: – Diphtheria antitoxin – Antibiotics (e.g., erythromycin orally or by injection (40 mg/kg/day; maximum, 2 gm/day) for 14 days  23 C. diphtheriae Lab Identification Specimen Respiratory diphtheria: exudate from nasopharynx or throat Cutaneous illness: wound material Blood for serology testing Culture Only performed in state health labs Isolate on: Tellurite or Tinsdale (black colonies) Sheep blood agar ELEK test for toxin production Toxin production must be demonstrated to confirm disease  24 Corynebacterium jeikeium (CDC JK group) Normal flora of the skin Infects mostly immunosuppressed hosts prosthetic heart valve Septicemia Skin infections Bone marrow transplant patients Often resistant to multiple antibiotics  25 Nocardia spp. (asteroides) Characteristics: Beaded branching GPB Strict aerobe Partially acid-fast Unusual colony appearance Present in the environment (soil, water) Opportunistic pathogen, not part of the human microbiome Some species can cause pulmonary infections and brain abscesses  26 Nocardia Gram stain  27 Nocardia acid fast stain  28 Nocardiosis Immunocompromised patients are most at risk for acquiring infection Causes respiratory and cutaneous infections Respiratory infection symptoms: Fever Weight loss Night sweats Cough Chest pain Pneumonia When the bacterium disseminates, it most commonly goes to the CNS  29 Nocardia spp. Lab Identification Specimen: Respiratory illness: Sputum, BAL Wound infection: wound material Methods: Culture  30 Gardnerella vaginalis Characteristics: Only one species GRAM VARIABLE – still technically a GPR Fastidious Associated with bacterial vaginosis CLUE cells  31 Gardnerella vaginalis Lab Testing Specimen: Vaginal swab Clue Cell Normal Epithelial Cell Methods: NAAT Antigen detection Wet prep Gram stain  32 Lactobacillus spp. Normal flora of gastrointestinal and genital tracts Decrease in number with bacterial vaginosis Rarely cause disease  SPORE FORMERS  34 Bacillus species Characteristics of the genus: Spore-forming Gram-positive bacilli Aerobic or facultative anaerobe Most are low-virulence saprophytes found in soil, water, dust, and animal products Common laboratory contaminants  35 Bacillus spp. continued Gram stain: large, box car shaped Gram-positive rods Produces spores  36 Bacillus anthracis Primary pathogen in the Bacillus genus Causes Anthrax Pulmonary Cutaneous GI Infection can be acquired via contact with infected animals or animal products Small blisters or bumps that may itch. Most often on the face, neck, arm, or hands. Painless skin ulcer with a black center Swelling can occur around the sore https://www.mayoclinic.org/diseases- conditions/anthrax/symptoms-causes/syc-20356203 accessed: 09/04/20  37 Pulmonary Anthrax Inhalation exposure Symptoms may include: Fever, chills, body aches Chest pain and shortness of breath Altered cognition Cough Nausea, vomiting, and/or stomach pains. Headache Drenching sweats Extreme fatigue https://www.cdc.gov/anthrax/symptoms/index.html  38 Bacillus anthracis Bacillus anthracis is one of the most likely agents to be used in a biological attack Virulence factors: Capsule Two toxins comprised of 3 proteins https://www.mayoclinic.org/diseases- conditions/anthrax/symptoms-causes/syc-20356203 accessed: 09/04/20  39 https://www.npr.org/sections/goatsandsoda/2016/08/03/488400 947/anthrax-outbreak-in-russia-thought-to-be-result-of-thawing- permafrost  40 Bacillus cereus Food poisoning (rice, meat and vegetable dishes, sauces, pasta, desserts, and dairy products) Opportunistic human pathogen Spores survive cooking/pasteurization then germinate and multiply when the food is inadequately refrigerated.  41 Bacillus cereus Symptoms are caused by a toxin The diarrheal type: diarrhea & abdominal pain 8-16 hrs after eating contaminated food (meat and veggie dishes, sauces, pasta, desserts, and dairy products) The emetic disease: nausea and vomiting 1- 5 hrs after eating contaminated food (rice).  42 Bacillus Lab Identification Specimen depends on the site of infection Several molecular assays available for detecting B. anthracis – these are time critical! Both B. anthracis and B. cereus may be recovered in culture B. cereus is often considered a contaminate “Ground glass” appearance of B. cereus on SBA  43 Clostridium species Characteristics of all Clostridium and Clostridioides species: Gram stain shows large, box-car Gram-positive bacilli (often stain Gram variable) Produce spores Anaerobes! Inhabits soil, water, vegetation, large bowels of humans and other animals  44 Clostridium perfringens Characteristics: Produces many exotoxins Growth is rapid Growth produces high amounts of gas Disease: Gas gangrene One of the most common causes of food poisoning in US  45 Case study The patient was an 85-year-old female with advanced squamous cell carcinoma of the bladder, who was admitted for treatment of a presumed cellulitis of her right lower leg. On exam, the patient appeared chronically ill. The right heel had a bluish hematoma and blister with approximately 6 by 6 cm of erythema. Her WBC was 35,000/mm3. The patient was treated with IV antibiotics for 2 days but showed no improvement.  46 Case study cont. A radiograph of her right ankle and foot showed extensive gas in the soft tissues. She was taken for surgical debridement and ultimately a below-the-knee amputation. A Gram stain of the wound aspirate revealed rare PMNs and many boxcar-shaped, Gram-positive rods. No growth was seen on aerobic cultures. The patientʼs condition worsened, and she died.  47 C. perfringens - Gas Gangrene Gas gangrene can develop after certain types of surgery or injuries. Blisters with gas bubbles form near the infected area, accompanied by fever, rapid heartbeat and breathing, and often pain at the infection site. If untreated, quickly leads to death Treatment involves high doses of antibiotics and surgical removal of dead or infected tissue.  48 C. perfringens - Gas Gangrene A. Arm of a drug abuser with ulcers and swelling traced to needle tracks. B. Radiographs from the same patient demonstrating gas (clear spaces) in the tissues. Chapter 29 Clostridium, Bacteroides, and Other Anaerobes, Ryan KJ. Sherris Medical Microbiology, 7e; 2017.  49 C. perfringens Lab Identification Specimen: GI illness: will not grow in a standard stool culture Wound infection: wound material or blood Methods: Culture  50 Clostridioides (Clostridium) difficile Characteristics: May be present in the bowel of healthy persons or acquired Antibiotic therapy decreases the number of normal microbiota organisms in the bowel, allowing C. difficile to overgrow Easily transmitted between patients; extra contact precautions are required with “C. diff” positive patients Disease: “C. diff” - C. difficile produces toxins; causes bloody diarrhea - Pseudomembranous colitis in severe cases  51 Pseudomembranous Colitis Epidemiology Most cases of C. diff occur after antibiotic therapy >65 years or older Recent hospitalizations A weakened immune system Previous infection with C. diff or known exposure Symptoms Diarrhea, including loose, watery stools or frequent bowel movements for several days Fever Stomach tenderness or pain Loss of appetite, nausea  52 C. difficile Lab Identification Specimen: Stool Methods: Culture with reflex to cytotoxin assay NAAT for toxin gene Antigen detection for toxin or organism Smells like a horse stable (or elephant barn) Stool transplant has been recognized as an excellent up-and-coming treatment for refractory C. difficile infection.  53 Clostridium tetani Characteristics In soil, dust, human and animal excreta Spores enter the human host through a wound Spores then germinate to vegetative cells and produce toxin Disease Causes TETANUS. Symptoms are due to a potent neurotoxin. Testing is usually performed by public health laboratories.  54 Tetanus Generalized tetanus is the most common form of the disease, which presents with pain, headache, stiffness, rigidity, opisthotonus, laryngeal obstruction, and spasms. Rigid paralysis.  55 Tetanus Treatment Neutralize circulating toxin with antitoxin Relieve muscle spasms Keep airways open Remove infected tissue and treat with penicillin Immunizations!! Tetanus booster shot every 10 years!  56 Clostridium botulinum Characteristics: Spores in water, soil (surface of fresh vegetables) Tough endospores can withstand high temperatures The anaerobic environment in canned foods is a perfect habitat for C. botulinum to grow. Foods with a pH >4.6 are at most risk (think beans) When in doubt – throw it out! Any canned foods that are leaking, bulging, swollen, discolored, or foul-smelling should not be consumed. Testing is typically performed by public health laboratories.  57 Clostridium botulinum Disease: Botulism – caused by a potent neurotoxin Food botulism, neurotoxin ingested Wound botulism, spores contaminate wound Infant botulism, spores ingested (honey) Flaccid paralysis Treat with antitoxin and penicillin, give supportive therapy. The botulinum toxin is the most toxic substance known to humankind. Image: https://www.alamy.com/stock-photo-vial-of- botulinum-toxin-type-a-botox-25635528.html accessed: 07/31/24  September 13, 2023 https://www.npr.org/sections/thesalt/2017/05/23/529589579/n https://www.nytimes.com/2023/09/13/world/europe/france-botulism- acho-cheese-sauce-tainted-with-botulism-kills-california-man sardines.html