Pyogenic cocci (Staph&Strep) Stijn van der Veen

Pyogenic cocci (Staph&Strep) Stijn van der Veen

Pyogenic cocci Spherical-shaped bacteria that are able to cause purulent inflammation of the mucous membranes of serous cavities (abdomen, pleura, pericardium) or in deep tissues (purulent infiltration, phlegmon). Staphylococcus sp. Streptococcus sp. Neisseria sp.

Cocci A coccus is a spherical bacteria Staphylococcus sp. tend to cluster in grape-like irregular bunches Streptococcus sp. tend to line up in chains Neisseria sp. tend to form a kidneyshaped diplococcus morphology

Major pathogenic species Staphylococcus sp. Staphylococcus aureus Staphylococcus epidermidis Staphylococcus saprophyticus Streptococcus sp. Streptococcus pyogenes (GAS) Streptococcus pneumoniae (pneumococcus) Streptococcus agalactiae (GBS) Neisseria sp. Neisseria meningitidis (meningococcus) Neisseria gonorrhoeae (gonococcus)

Overview of bacterial infections

Gram positive cocci

Pathogenic Staphylococcus infections Staphylococcus aureus Many types of infections Staphylococcus epidermidis Most common cause of infection in indwelling prosthetic devices (prosthetic joints, heart valves, etc.) Staphylococcus saprophyticus Leading cause of urinary tract infections in sexually active young women

Differentiation of Staphylococcus sp. The bacteria produce liposoluble pigments which make the colonies appear in certain color: Staphylococcus aureus golden yellow Staphylococcus epidermidis white Staphylococcus saprophyticus white or lemon yellow

Mannitol salt agar Combination of selective and differential medium. It contains high salt concentrations (<7.5% NaCl), which inhibits most bacteria except for Staphylococci (and few others). It also contains the carbohydrate mannitol and ph indicator phenol red to detect acid production from mannitol fermentation. Staphylococcus aureus produce yellow colonies, while other Staphylococci produce pink-red colonies.

Coagulase testing Staphylococcus aureus produces the enzyme coagulase, while other Staphyloccus sp. generally do not. Coagulase is an extracellular enzyme that clots fibrinogen proteins in human blood. The coagulase enzyme can be detected by the latex agglutination test, using fibrinogen coated latex beads.

Staphylococcus aureus A very versatile bacterium that is often found on the human skin and in the respiratory tract. It is asymptomatically carried by approx. 20% of the human population. Notorious for hospital acquired infections and for developing antibiotic resistance (MRSA).

Staphylococcus aureus infections Staphylococcus aureus causes a variety of suppurative (pus-forming) infections and toxinoses in humans.

Staphylococcus aureus infections Superficial skin lesions Boils, Styes Furunculosis Serious infections Pneumonia, Mastitis (inflammation of mammary gland) Phlebitis (inflammation of vein, usually in leg) Meningitis (inflammation of meninges) Urinary tract infections Septicemia

Staphylococcus aureus infections Deep-seated infections Osteomyelitis (inflammation of the bone) Endocarditis (inflammation of heart)

Staphylococcus aureus infections S. aureus is a major cause of hospital acquired (nosocomial) infection of surgical wounds and infections associated with indwelling medical devices.

Toxicoses Scalded Skin Syndrome Food poisoning A gastrointestinal illness from the release of enterotoxins into food. Toxic shock syndrome Release of superantigens into the blood stream. Commonly occurs in women.

Staphylococcus epidermidis Part of the normal human skin micro-flora Able to form biofilms on plastic devices Cause for infections on medical indwelling devices, e.g. catheters, medical prostheses, etc. Catheters are medical devices that can be inserted in the body to treat diseases or perform a surgical procedure.

Device related infections If devices are COLONIZED, the patient is INFECTED Bacteria can survive/multiply in biofilm. Biofilm provides reservoir for bacteria Microorganisms are detached from the biofilm and can infect a patient Antibiotics are largely ineffective in clearing biofilms, making treatment very difficult

What is a biofilm? Biofilms are structured communities of microorganisms, which are adhered to a surface and often surrounded by a self produced matrix of extracellular polymeric substances

Biofilm related diseases The Centers for Disease Control and the National Institutes of Health estimate that 65% to 80% of all chronic infections can be attributed to microbial biofilms. Device related infections Implant related infections Plaque; Gingivitis Cystic fibrosis Ear infection Bacterial endocarditis Chronic prostatitis

Staphylococcus saprophyticus Part of the normal micro-flora of the female genital tract Common cause of community-acquired urinary tract infections Patients with urinary tract infections caused by S. saprophyticus usually present with symptomatic cystitis. Treatment Trimethoprim-sulfamethoxazole or norfloxacin

Gram positive cocci

Pathogenic Streptoccus infections Streptococcus pyogenes (GAS) Many types of infections Streptococcus pneumoniae (pneumococcus) Major cause of Streptococcal Pneumonia Streptococcus agalactiae (GBS) Major cause of bacterial septicemia in newborns

Hemolysins Hemolysins combine with the membranes of red blood cells, causing it to lyse. Streptococcus sp. can be distinguished by their hemolyses ability into α, β, and γ. α => Streptococcus pneumoniae β => Streptococcus agalactiae β => Streptococcus pyogenes γ => Most other Streptococcus sp.

Lancefield grouping Serological detection and identification of bacterial cell surface antigens. Binding of specific antibodies to the cell surface antigens. Agglutination assays (clumping of bacteria due to antibody binding.

Streptococcus pyogenes infections Streptococcus pyogenes (GAS) causes a variety of skin and soft tissue infections, pharyngitis (strep throat), and toxic shock syndrome (caused by toxin).

Streptococcus pyogenes infections Scarlet fever Pharyngitis (Strep throat) Rheumatic fever Skin infections Necrotizing fasciitis Streptococcal toxic shock syndrome Puerperal fever Glomerulonephritis (kidney)

Scarlet fever The causative agent Streptococcus pyogenes Sign and symptoms Pink-red rash on neck, chest, arms, strawberry-like tingue Transmission Respiratory droplets Treatment Penicillin, clarithromycin Prevention and control Practicing good hygiene

Streptococcal pharyngitis The causative agent Streptococcus pyogenes Sign and symptoms Sore throat, Fever, headache, swollen lymph nodes and tonsils Transmission Respiratory droplets Treatment Penicillin Prevention and control Practicing good hand hygiene

Streptococcus pneumoniae infections Streptococcus pneumoniae (pneumococcus) causes meningitis (infection of meninges) and a variety of diseases the respiratory tract

Streptococcus pneumoniae infections Pneumonia Bacterial meningitis Otitis media Sinusitis

Pneumococcal pneumonia The causative agent Streptococcus pneumoniae Sign and symptoms High fever, sharp chest pains, difficulty breathing, rust-colored sputum Transmission Respiratory droplets Treatment Penicillin, cefotaxime Prevention and control Vaccination, good hand hygiene

Acute bacterial meningitis The causative agent Streptococcus pneumonia, Neisseria meningitidis, Haemophilus influenzae type b Sign and symptoms Fever, stiff neck, severe headache, vomiting and nausea, sensitivity to light Transmission Respiratory droplets from prolonged contact Treatment Antibiotics Prevention and control Vaccination

Acute otitis media The causative agent Streptococcus pneumonia, Haemophilus influenzae Sign and symptoms Ear pain, red bulging eardrum Transmission Airborne contact Treatment Wait and see or antibiotics Prevention and control Limiting time in childcare

Streptococcus agalactiae Opportunistic bacterium that in some circumstances can cause severe invasive infections. Colonizes the gastrointestinal tract in up to 30% of healthy adults and the vaginal tract of up to 20-36% of sexually active women. Leading cause of bacterial neonatal infections in the baby during gestation and after delivery, causing septicemia, pneumonia, and meningitis which can lead to long-term sequelae or death.

Virulence factors Attachment to host tissues Production and delivery of various factors Replication and evasion of immunity Damage to host tissues

Capsule Capsule contributes to virulence of pathogens Mediates adherence Protects against phagocytosis Protects against antimicrobial compounds secreted by host cells, such as antimicrobial peptides and ROS. Examples include: Streptococcus pneumoniae Streptococcus mutans

Inverse binding of antibodies S. aureus produces protein A and S. pyogenes produces Protein G which bind the Fc portion of IgG Binding of IgG to the bacterial cell surface provides protection against phagocytosis.

Leukocidins Leukocidins are enzymes that destroy circulating white blood cells (WBC) such as neutrophils and macrophages, which are immune cells designed to phagocytize and destroy pathogens. Leukocidins destroy the cell membrane of WBC and trigger the release and rupture of lysosomes. Lysosomes contain powerful hydrolytic enzymes which then cause more tissue damage. Example: Staphylococcus aureus Streptococcus pyogenes Streptococcus pneumoniae

Research intermezzo Contribution of Panton-Valentine Leukocidin to S. aureus pathogenesis

Patient characteristics Variable PVL + (n=15) PVL - (n=90) P value Age (years), mean ±SD 63.7 ±11.3 59.1 ±24.9 0.91 a Age (years) 65, no. (%) 10 (67%) 47 (52%) 0.40 b Male sex, no. (%) 11 (73%) 60 (67%) 0.77 b Comorbidity score, mean±sd 2.47 ±1.55 2.60 ±1.85 0.79 b Diabetes, no. (%) 1 (7%) 8 (9%) 1.00 b Tumor, no. (%) 3 (20%) 14 (16%) 0.71 b Total of 105 patients with HAP or VAP due to MRSA No difference in patient baseline Chemotherapy, no. (%) 0 (0%) 2 (2%) 1.00 b COPD, no. (%) 0 (0%) 9 (10%) 0.35 b Sepsis, no. (%) 13 (87%) 62 (69%) 0.22 b Shock, no (%) 1 (7%) 4 (4%) 0.54 b Multiple organ damage, no. (%) 3 (20%) 6 (7%) 0.12 b VAP, no (%) 2 (13%) 3 (3%) 0.15 b Multilobar positive, no (%) 2 (13%) 26 (29%) 0.34 b a Mann-Whitney U test. b Fisher exact test

Severity of disease Variable PVL + (n=15) PVL - (n=90) P value APACHE II score, mean ±SD 14.3 ±7.8 10.1 ±4.7 0.005 a CRP >80 mg/l, no (%) 8 (53%) 16 (18%) 0.006 b WBC count >15 10 9 /L, no (%) 7 (47%) 12 (13%) 0.006 b Presence of PVL affects severity of disease in patients with HAP or VAP due to MRSA Platelet count <150 10 9 /L, no (%) 5 (33%) 17 (19%) 0.30 b Bilirubin >20 μmol/l, no (%) 5 (33%) 18 (20%) 0.31 b Urea-Creatinine ratio >100, no (%) 5 (33%) 28 (31%) 1.0 b APPACHE, Acute, Physiology and Chronic Health Evaluation; CRP, C-reactive protein; WBC, white blood cell. a Unpaired parametric two-tailed t-test. b Fisher exact test.

Clinical outcome Presence of PVL affects clinical outcome in patients with HAP or VAP due to MRSA Kaplan-Meier survival curves (log-rank test) Variable PVL + (n=15) PVL - (n=90) P value 30-day mortality, no (%) 4 (27%) 4 (4%) 0.01 a a Fisher exact test

Conclusion Presence of PVL genes in MRSA impact the severity of disease and survival of patients with HAP due to MRSA Leukocidins are an important virulence factor in S. aureus and might therefore be suitable targets for intervention

Coagulase Coagulase catalyses the formation of blood clots from Fibrinogen proteins in the human blood. Blood clots protect bacteria from phagocytosis by WBC s and other host defenses. Example: Staphylococcus aureus

Kinase Kinases have the ability to dissolve blood clots used as a defense by the body to restrict and isolate an infected area. Kinases help bacteria to spread and cause bacteremia Examples: Streptokinase - Streptococcus sp Staphylokinase - Staphylococcus aureus

Hyaluronidase Hyaluronidase breaks down hyaluronic acid, which is a polysaccharide that connects cells in tissues. It is called the spreading factor because it enhances penetration of bacteria into tissues. Examples: Streptococcus sp. Staphylococcus aureus

Hemolysins Hemolysins combine with the membranes of red blood cells, causing it to lyse. Lysing of red blood cells provide pathogens with the iron from hemoglobin, which is required for many metabolic processes. α, β, and γ hemolyses Example β-hemolyses: Streptococcus pyogenes Streptococcus agalactiae Staphylococcus aureus

Superantigens (sags) Antigens/toxins that cause non-specific activation of T- cells resulting in polyclonal T cell activation and massive cytokine release. Enterotoxin, 9 serotypes, responsible for Staphylococcal food poisoning. TSST-1, close to enterotoxin, cause toxic shock syndrome (TSS). Exfoliatin, 2 serotypes, cause Staphylococcal scalded skin syndrome (SSS). Examples: Streptococcus pyogenes Staphylococcus aureus

M protein The surface expressed M protein inhibits phagocytosis and complement mediated cell lyses. M protein binds fibrinogen in plasma which blocks complement deposition on the underlying peptidoglycan layer. Example: Streptococcus pyogenes

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