Lecture 7: Gram Negative Bacteria

Medically Important Gram-Negative Bacteria

• These bacteria are a diverse group, crucial in various medical contexts.
• Learning objectives focus on key characteristics, diseases, and virulence factors.
• Neisseria spp. and Moraxella spp. exhibit distinct morphologies, features, and common diseases, also outlining their key virulence factors.
• Enterobacteriaceae family members are noted for their differing oxidase and lactose fermentation tests, distinguishing opportunistic from primary pathogens and serological classification (O, H, and K antigens).
• Gram-negative organisms like Pseudomonas aeruginosa, Burkholderia cepacia, Acinetobacter baumannii, Vibrio spp., Campylobacter spp., Helicobacter pylori, Haemophilus spp., and Bordetella pertussis each possess specific morphologies, virulence factors, and related illnesses.
• These organisms' features and classifications are covered in Murray's Medical Microbiology (9th edition), specifically chapters 23 through 29.

Gram (+ve) and Gram (-ve) Bacteria

• Gram-positive bacteria have a thick peptidoglycan layer, while gram-negative bacteria have a thin peptidoglycan layer.
• Their contrasting structures affect how antibiotics affect them, leading to differential susceptibility.
• Gram-positive cell walls contain lipoteichoic acid and are sensitive to certain antibiotics.
• Gram-negative cell walls contain lipopolysaccharides (endotoxins), endotoxins are released when the bacteria die, leading to inflammation and possible shock.

Neisseria species

• Bean-shaped gram-negative diplococci (pairs).
• Non-spore-forming, intracellular pathogens.
• Aerobic, requiring carbon dioxide for optimal growth.
• Most are oxidase and catalase positive.
• All metabolize glucose, but only N. meningitidis metabolizes specifically maltose.
• Neisseria species grow on modified Thayer Martin Agar.

Neisseria meningitidis

• Second most common cause of community-acquired meningitis.
• Transmitted via respiratory droplets.
• Encapsulated, capnophilic (5% CO2).
• Metabolizes glucose and maltose.
• A significant percentage of humans carry it in their nasopharynx.
• Key virulence factors include a polysaccharide capsule and pili for attachment.

Virulence and Pathogenesis of N. meningitidis

• Polysaccharide capsule: Antiphagocytic, protecting the bacteria from attack.
• Various serotypes (A, C, W-135, Y, B). Serotype B is most common and least immunogenic.
• Pili: Attachment to respiratory epithelium.
• IgA protease: Colonization of mucosal membranes.
• Lipooligosaccharide (LOS) endotoxin: Fever, shock.
• Complement deficiency (C6-C9): Increased risk of Nisseria infections.

Clinical Signs and Manifestations of N. meningitidis

•  Meningitis: Fever, headache, and stiff neck ("nuchal rigidity").
• Vomiting is a common symptom.
• Complications: Septicemia (Waterhouse-Friderichsen syndrome), Adrenal insufficiency, and Intravascular coagulation.
• Diagnosis using Gram stain and culture, or possibly rapid PCR of CSF.

Neisseria gonorrhoeae

• Gram-negative diplococci
• Intracellular pathogen
• Does not metabolize maltose
• Fastidious
• Symptomatic in 90% of males and 50% of females.
• More prevalent in southern U.S. states.
• Infections are frequently underreported.
• Key virulence factors are pili for attachment and IgA protease to degrade the immune response.

Virulence and Pathogenesis of N. gonorrhoeae

• Pili: Attachment, antigenic variation.
• Outer membrane proteins: Antigenic variation.
• Lipo-oligosaccharide (LOS): Modified endotoxin.
• IgA protease: Hydrolyzes secretory IgA.
• Invades mucosal surfaces.

Clinical Signs and Manifestations of N. gonorrhoeae

• Males: Urethritis with dysuria (painful urination) and purulent discharge.
• Females: Endocervicitis, salpingitis, and possible rectal infections in homosexual males.
• Some patients with conjunctivitis.
• Diagnosis: NAAT (Nucleic Acid Amplification Test) and culture.
• Treatment: Ceftriaxone (IM) and Azithromycin (oral).

Gram-Negative Bacilli

• Diverse group of gram-negative rod-shaped bacteria.
• Key classifications: non-enteric and enteric.
• Pseudomonas aeruginosa, Burkholderia cepacia, and Bordetella pertussis are significant non-enteric varieties.
• Haemophilus influenzae and Yersinia pestis are facultative anaerobes, important non-enteric bacilli. Specific pathogens are noted as aerobic and facultative anaerobes respectively.

Pseudomonas aeruginosa

• Ubiquitous, water-inhabiting gram-negative rod with a flagellum.
• Frequent in hospital environments.
• Strict aerobe, opportunistic.
• Common in cystic fibrosis (CF) patients, leading to pneumonia.
• Resistant to multiple antibiotics.
• Some factors leading to tissue damage (phospholipase C and Exotoxin A) are discussed.

Acinetobacter baumannii

• Gram-negative, aerobic rods.
• Common environmental organisms (soil, water, skin), commonly isolated from patients in hospitals.
• Rarely causes disease in healthy individuals, but strains in hospital settings are highly drug-resistant.
• Can cause multiple infections (Respiratory tract infections, Urinary tract infections, Wound infections, Septicemia)

Burkholderia cepacia

• Gram-negative rod.
• Can be found in water, soil, and hospital environments.
•  Opportunistic organism, frequently associated with respiratory infections, particularly in individuals with cystic fibrosis.
• Can spread via person-to-person contact, contact with contaminated surfaces, or exposure to Burkholderia in the environment.
• Many strains are antibiotic-resistant, posing clinical challenges.

Bordetella pertussis

•  Human-specific pathogen.
•  Highly contagious (person-to-person).
•  Non-motile, encapsulated gram-negative coccobacillus.
•  Fastidious, requiring specialized media (Bordet-Gengou or Regan-Lowe agar) for growth.
•  Associated with lymphocytosis – increased levels of white blood cells, and toxins (Adenylate cyclase toxin and Pertussis toxin).

Virulence and Pathogenesis of Bordetella pertussis

•  Polysaccharide capsule: Antiphagocytic.
• Adenylate cyclase toxin: Inhibits phagocytosis.
• Tracheal cytotoxin: Destroys ciliated epithelial cells, and induces apoptosis and potentially dermonecrotic lesions.
•  Pertussis toxin (PT): Increases cAMP levels.
•  Filamentous hemagglutinins (FHA): Bacterial attachment.

Clinical Signs and Manifestations of Bordetella pertussis

• Whooping cough, a specific paroxysmal cough, with periods of intense coughing.
• Other symptoms include fever, headache, sneezing, and anorexia.

Management of Bordetella pertussis

•  Vaccination is the primary method of prevention.
•  Diphtheria, tetanus, and pertussis vaccines (DTaP) are given in childhood.
• Tdap vaccines are recommended for adolescents and adults.

Haemophilus influenzae

• Gram-negative, pleomorphic (varying shape) coccobacilli
• Strict human pathogen, typically found in the nasopharynx.
• Facultative anaerobe

Campylobacter jejuni

• Gram-negative curved, rodshaped organism.
• Motile (polar flagellum), microaerophilic (needs a lower oxygen concentration).
• Common cause of bacterial diarrhea.
• Often associated with undercooked poultry.
• Virulence factors include adherence factors (that contribute to mucosal attachment), some O-antigens can cross react and result in demyelination.

Helicobacter pylori

• Gram-negative, microaerophilic, spiral-shaped bacterium.
•  Motile (flagella)
• Lactose non-fermenter.
• Urease positive
• Common cause of gastritis and peptic ulcers.
•  Can potentially lead to chronic/life-long infection if untreated.
•  Key virulence factors include urease, vacA, and type IV secretion system which neutralize gastric acid and facilitate mucosal colonization & attachment.

Clinical Signs and Manifestations of Helicobacter pylori

• H. pylori is associated with various digestive problems.
• These include gastritis (inflammation of the stomach lining), peptic ulcers, and potentially gastric adenocarcinoma. 

Vibrio cholerae

• Gram-negative, curved rods.
•  Facultative anaerobes.
• Single polar flagellum.
•  Oxidase positive
•  Human reservoir (colon)
• Outbreaks associated with contaminated water.
• 6 serotypes; only 01 & 0139 related to epidemics.

Virulence and Pathogenesis of Vibrio cholerae

• Heat-labile enterotoxin (composed of 6 subunits): ADP-ribosylation of G proteins, leading to increased cAMP levels, resulting in loss of water and electrolytes.

Escherichia coli

• Gram-negative, facultative enteric rods.
• Most common cause of UTIs and normal intestinal flora.
• Motile (some strains).
• Doesn't produce H2S, urease -, citrate - and indole + and lactose+. -Numerous pathotypes exist, including enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enterohemorrhagic E. coli (EHEC), and uropathogenic E. coli (UPEC).
•  Different types are associated with certain diseases (like EHEC causing hemorrhagic colitis and UPEC related to urinary tract infections).

Klebsiella pneumoniae

• Gram-negative encapsulated, non-motile rods.
• Lactose fermenter.
• Common GIT pathogen.
• Frequent nosocomial infection (hospital acquired infection).
• Patients with medical devices and/or underlying illnesses have a higher risk of infection.
• Significant antibiotic resistance (extended-spectrum beta-lactamases, fluoroquinolones, and aminoglycosides, and carbapenems).

Serratia marcescens

• Gram-negative, motile rods.
• Opportunistic pathogen.
• Lactose fermenter; produces red pigments, (such as prodigiosin).
• Associated with nosocomial infections, catheter-related septicemia, and burn infections.
• Drug resistance is increasing; antibiotic treatment success is decreasing.

Proteus spp.

• Gram-negative rods.
•   Urease positive, lactose non fermenters.
• Produce hydrogen sulfide (H₂S).
• Important in Struvite stone formation.
• Commonly found as normal microbiota in the human GI tract.
• P. mirabilis is most common; P. vulgaris are less common, but may be more associated with hospital settings.

Salmonella

• Gram-negative motile rods.
•  Lactose nonfermentative.
•  H2S production.
•  Classification of species and varieties is significant (2500 serotypes).
• Can cause enteritis, typhoid.
•  Some serotypes are more commonly associated with gastroenteritis from the consumption of contaminated meals.

Shigella

• Gram-negative, non-motile rods.
•  Lactose nonfermenters
• Does not produce H2S.
•  Humans are the only reservoir, commonly transmitted via the fecal-oral route.
•  Mainly a disease of infants and young children/school aged children.
• Key virulence factors for invasion are the types of toxin and their associated mechanism of action; specifically Shiga toxin and its mechanism of action.

Yersinia enterocolitica

• Gram-negative facultative rods.
•  Motile at 25°C, non-motile at 37°C
•  Urease +, but does not ferment lactose.
•  Usually found in colder climates or contaminated meat, milk.
• Can cause enterocolitis and post-infectious arthritis.

Yersinia pestis

• Gram-negative coccobacilli.
•  Non motile
• Facultative intracellular.
• Important zoonotic pathogen (transmitted via fleas.
• Plague is a disease that can occur in bubonic, pneumonic, and septicemic forms.
• Different cycles of infection are detailed, in both wild and domestic situations. 

Enterobacteriaceae Identification

• Key tests for identifying members of the Enterobacteriacea family are lactose fermentation, H2S production, motility, and Voges-Proskauer.

Test your knowledge on clinical manifestations and treatments related to various bacteria, including Helicobacter pylori and Enterobacteriaceae. This quiz covers identification tests, infections, characteristics, and therapies for specific pathogens. Perfect for students in microbiology courses or healthcare professionals looking to refresh their knowledge.