University:
Kentucky State University
Course:
BIO 204 | Fund of Microbiology
Academic year:

2021
Views:

175

Pages:

47
Author:

Campbell Wagner

Medical Microbiology Gram Positive Cocci (Staphylococci &Streptococci ) Staphylococci-Introduction ⚫ Staphylococci are gram-positive cocci ⚫ In Greek; staphyle - Bunch of grapes Kokkus – Berry ⚫ Spherical cells arranged in irregular clusters (grape like clusters) ⚫ Common inhabitant of the skin and mucous membranes ⚫ Lack spores and flagella ⚫ May have capsules ⚫ Catalase-positive ⚫ About 40 species SHEET NOTE #1 • The spores found in some of gram positive rods or bacilli . • The flagella found in some of gram negative bacilli . • The bacteria that has capsules mean that it a virulence factor its be pathogens • There is 40 species but the most important for us (the medical importants are 3 types) Classification A) Based on coagulase production: 1. Coagulase-positive: Staphylococcus aureus 2. Coagulase-negative: Staphylococcus epidermidis Staphylococcus saprophyticus B) Based on pathogenicity: 1. Common pathogen: S. aureus 2. Opportunistic pathogens: S. epidermidis S. saprophyticus 3. Non pathogen: S. homonis SHEET NOTE #2 If we have a gram positive coccus the first step is : • We make a catalase test (if the result was positive that means it a staphylococcus ) Then we must make a test for the coagulase production • If it positive : S.aureus • If it negative : S. epidermidis or S. saprophyicus The S. epidermidis & S. saprophyicus areopportunistic don’t cause inflammation exept in some special cases Staphylococcus aureus General Characteristics ⚫Optimum temperature of 37oC ⚫Coagulase-positive ⚫Facultative anaerobe ⚫Withstands high salt, extremes in pH, and high temperatures ⚫Produces many virulence factors Epidemiology ⚫ Present in most environments frequented by humans ⚫ Carriage rate for healthy adults is 20-60%, mostly in anterior nares, skin, nasopharynx and intestine ⚫ Hospital infections caused by staphylococci are frequent & they are caused by strains resistant to various antibiotics ⚫ Staphylococci are a common cause of postoperative wound infection and other hospital cross infections ⚫ Source of infection: A) Exogenous: patients or carriers B) Endogenous: from colonized site (throat or skin ) ⚫ Mode of transmission: A) Contact: direct or indirect B) Inhalation of air borne droplets Virulence factors of S. aureus Cell associated factors: A) Cell associated polymers 1. Cell wall polysaccharide 2. Teichoic acid 3. Capsular polysaccharide B) Cell surface proteins 1. Protein A 2. Clumping factor (bound coagulase) Enzymes: ⚫Coagulase – coagulates plasma and blood, produced by 97% of human isolates, diagnostic . ⚫Hyaluronidase – digests connective tissue ⚫Staphylokinase – digests blood clots ⚫DNase – digests DNA ⚫Lipases – digest oils; enhances colonization on skin ⚫Penicillinase – inactivates penicillin Toxins: ⚫Hemolysins (α, β, γ, δ) – lyse red blood cells ⚫Leukocidin – lyses neutrophils and macrophages ⚫Enterotoxin – induce gastrointestinal distress ⚫Exfoliative toxin – separates the epidermis from the dermis ⚫Toxic shock syndrome toxin (TSST) – induces fever, vomiting, shock, systemic organ damage SHEET NOTE #3 • Leuko : white • Enterotoxin (very important) anther name {exotoxin} o It produce in food & sweetmeats . o It causes also food poisonic Clinical Presentations ⚫ Infections: 1) Skin and soft tissue: Folliculitis, furuncle (boil), carbuncle, styes, abscess, wound infections, impetigo 2) Musculoskeletal: Osteomyelitis, arthritis, bursitis 3) Respiratory: Tonsillitis, pharyngitis, sinusitis, otitis, bronchopneumonia, lung abscess, empyema 4) Central nervous system: Abscess, meningitis 5) Endovascular: Bacteremia, septicemia, endocarditis 6) Urinary: Urinary tract infection ⚫ Intoxications: 1) Food poisoning 2) Toxic shock syndrome 3) Staphylococcal scalded skin syndrome Folliculitis Furuncle (boil) Carbuncle • It maybe found in the neck , thigh , armpits • Carbuncle is a grouping of many furuncles. Laboratory Diagnosis 1. Specimens collected: Pus, sputum, blood, stool, and For the detection of carriers- Nasal swab 2. Gram Stain: Gram-positive cocci in grape like clusters 3. Culture: ⚫ Culture media: Non selective: Nutrient agar, Blood agar, MacConkey’s agar Selective media: Mannitol Salt Agar ⚫ Culture conditions: Ambient conditions, 37 °C, 18-24 h ⚫ Colonial morphology: Nutrient agar- golden yellow pigments MacConkey’s agar- small & pink in colour Blood agar- most strains produce β- haemolytic colonies 1. 2. 3. 4. SHEET NOTE #4 1. 2. 3. 4. first picture : gram stain Second picture : Mannitol Salt Agar Third picture : Nutrient agar Fourth picture : blood agar 4. Biochemical tests: ⚫Catalse -positive ‫مهم جدا‬ ⚫Coagulase-positive ‫يتم عمله من اجل‬ ⚫Ferments mannitol S. aureus ‫تمييز‬ • 5. Antibiotic sensitivity tests: very important Such as kerne power 6. Serological test: of limited value SHEET NOTE #5 .1 .2 .3 Catalse enzyme positive (Staphylococcus) negative .4 (streptococcus) .1 .2 Staphylocci S.auerus .3 S. epidermidis .4 Treatment ⚫ Drug resistance is common ⚫ Benzyl penicillin is not effective antibiotic, if the strain is sensitive ⚫ Cloxacillin or Methicillin is used against β-lactamase producing strains ⚫ Methicillin Resistant Staphylococcus aureus (MRSA) strains have become common ⚫ Vancomycin is used in treatment of infections with MRSA strains (Vancomycin used only for gram positive ) Staph. epidermidis ⚫Common habitat of skin ⚫It is a common cause of stitch abscesses ⚫It has predilection for growth on implanted foreign bodies such as artificial valves, shunts, intravascular catheters and prosthetic appliances leading to bacteraemia ⚫Endocarditis may be caused, particularly in drug addicts SHEET NOTE #6 • Stich abscesses • When someone perform artificial valve operation and infection occur its called endocarditis. • Cerebral shunts are commonly used to treat hydrocephalus, the swelling of the brain due to excess buildup of cerebrospinal fluid (CSF) and this tube can be connect between Sulcus and SVC. • If the S.epidermidis enter the shunts ( plastic tube ) as well as the intravascular catheters such as (central venus catheter that is placed in the neck or the cannula that placed in the hand ) and prosthetic appliances such as the artificial joints all of this will be dangerous for the human if the infection happens. • The drug addicts one of the most people that more vulnerable for infection because the use contaminated needles for drug abuse S. saprophyticus ⚫It causes urinary tract infections, mostly in sexually active young women ⚫The infection is symptomatic and may involve the upper urinary tract ⚫It is one of the few frequently isolated that is resistant to Novobiocin SHEET NOTE #7 • • For distinguish between the S.epidermidis and the S.saprophyticus we use the Novobiocin (antibiotic) • The steps are : 1. We put the two bacteria in a dish 2. We add the Novobiocin 3. If there inhibition that means the bacteria is S.epidermidis 4. If there nothing happen that means the bacteria is S.saprophyticus Distinguishing features of the major species of staphylococcus Characters Coagulase Novobiocin sensitivity Mannitol fermentation S.aureus + Sensitive + S.epidermididis - Sensitive - S.saprophyticus - Resistant - Streptococci General Characters ⚫Gram-positive cocci ⚫Chains or pairs ⚫Usually capsulated ⚫Non motile ⚫Non spore forming ⚫Facultative anaerobes ⚫Fastidious ⚫Catalase-negative (Staphylococci are catalase-positive) Classification Streptococci can be classified according to: ⚫Oxygen requirements 1. Anaerobic (Peptostreptococcus) 2. Aerobic or facultative anaerobic (Streptococcus) ⚫Serology (Lancefield Classification) ⚫Hemolysis on blood agar Serology: Lancefield Classification Streptococci Lanciefield classification Group A S. pyogenes Group B S. agalactiae Group C S. equisimitis Group D Enterococcus Other groups (E-U) ⚫ Streptococci classified into many groups from A-K & H-V ⚫ One or more species per group ⚫ Classification based on C- carbohydrate antigen of cell wall ⚫ Groupable streptococci ⚫ A, B and D (more frequent) ⚫ C, G and F (Less frequent) ⚫ Non-groupable streptococci ⚫ S. pneumoniae (pneumonia) ⚫ viridans streptococci ⚫ e.g. S. mutans ⚫ Causing dental carries SHEET NOTE #8 • The most medical important groups are group( A ,B &D) they are the most frequent to cause infection . • the Groupable streotococci cause beta Hemolysis • The Non- groupable streotococci cause ahpha hemolysis • Viridans are group not one Classification Based on Hemolysis on Bl. Agar 1. α-hemolysis (Partial : ⚫ Partial hemolysis with green discoloration around the colonies ⚫ e.g. non-groupable streptococci (S. pneumoniae & Viridans.S) 2. β-hemolysis ⚫ Complete hemolysis with clear zone of hemolysis around the colonies ⚫ e.g. Group A & B (S. pyogenes & S. agalactiae) 3. γ-hemolysis ⚫ No lysis ⚫ e.g. Group D (Enterococcus spp) Hemolysis on Blood agar β-hemolysis α-hemolysis γ-hemolysis Group A streptococci ⚫ Include only S. pyogenes ⚫ Group A streptococcal infections affect all ages peak incidence at 5-15 years of age ⚫ 90% of cases of pharyngitis ⚫ Infection can leads to sever complications inscluding rheumatic fever and glomerulonephritis SHEET NOTE #9 • Why the children between 5-15 vulnerable to infection by S.pyogenes? • Answer : because they more vulnerable to pharyngitis and tonsillitis . • #The commonest bacteria cause of upper respiratory track infection is (group A ) S.pyogenes. • #The commonest cause of upper respiratory track infections are the viral infections. • The indirect complications for the group A are rheumatic fever and glomerulonephritis . Pathogenesis and Virulence Factors ⚫ Structural components (cross reactivity) ⚫ M protein-interferes with lysis of the bacteria ⚫ Lipoteichoic acid & F protein-adhesion ⚫ Hyaluronic acid capsule ⚫ Enzymes-facilitate the spread of streptococci through tissues ⚫ Streptokinases ⚫ Deoxynucleases ⚫ C5a peptidase ⚫ Pyrogenic toxins mediates bacteremia and shock ⚫ Streptolysins (lyse red blood cells, white blood cells, and platelets) ⚫ Streptolysin O ⚫ Streptolysin S Disease caused by S. pyogenes ⚫ Suppurative ⚫ Non-Invasive 1. Pharyngitis “strep throat” 2. Skin infection: Impetigo ⚫ Invasive 1. Scarlet fever-rash that begins on the chest and spreads across the body 1. Pyoderma- pus-producing lesion that usually occurs on the face, arms, or legs 2. Necrotizing fascitis-toxin production destroys tissues and eventually muscle and fat tissue (flesh eating bacteria) ⚫ Non Suppurative ⚫ Rheumatic fever ⚫ Glomerulonephritits Group B Streptococci ⚫ Include mainly S. agalactiae ⚫ Infections: ⚫ Neonatal disease 1. Early onset in neonates who are less than 7 days old. Vertical transmission of the organism from the mother. Manifests in the form of pneumonia or meningitis with bacteremia. Associated with a high mortality rate 2. Late-onset infection between 1 week and 3 months after birth. Usually occurs in the meningitis form. Mortality rate is not as high as early-onset ⚫ Other: Systemic, Cutaneous, UTI's SHEET NOTE #10 Neonatal period : the first 4 weeks of life and especially the first week . The babies acquired the group B from vagina and during the delivery they might contract the bacteria and enter the blood cycle and cause septicemia then meningitis Group D Streptococci ⚫ Group D streptococci are divided into: 1. those that will grow in 6.5% saline (enterococci) 2. and those that will not (non-enterococci) ⚫ Enterococci are distantly related to other streptococci and have been moved into the genus Enterococcus which are members of the gut flora. ⚫ Associated infections ⚫ Bacteremia ⚫ Urinary tract infections ⚫ Wound infections ⚫ Endocarditis Streptococcus Pneumoniae ⚫ General characteristics ⚫ Inhabits the nasopharyngeal areas of healthy individuals ⚫ Typical opportunist ⚫ Gram-positive diplococci ⚫ Virulence factors ⚫ Polysaccharide capsule ⚫ Clinical infections ⚫ Pneumonia ⚫ Meningitis ⚫ Bacteremia ⚫ Sinusitis/otitis media Differentiation between β-hemolytic streptococci ⚫ The following tests can be used to differentiate between β-hemolytic streptococci ⚫ Lanciefield classification: serology with specific antibodies ⚫ Bacitracin susceptibility test: Specific for S. pyogenes (Group A) ⚫ CAMP test: Specific for S. agalactiae (Group B) Bacitracin sensitivity ⚫ Principle: ⚫ Bacitracin will inhibit the growth of gp A Strep. pyogenes giving zone of inhibition around the disk ⚫ Procedure: ⚫ Blood agar inoculated with heavy suspension of tested organism ⚫ Bacitracin disk (0.04 U) is applied to inoculated blood agar ⚫ After incubation, any zone of inhibition around the disk is considered as susceptible CAMP test ⚫ Principle: ⚫ Group B streptococci produce extracellular protein (CAMP factor) ⚫ CAMP acts synergistically with staph. β-lysin to cause lysis of RBCs ⚫ Procedure: ⚫ Single streak of Streptococcus to be tested and a Staph. aureus are made perpendicular to each other ⚫ 3-5 mm distance was left between two streaks ⚫ After incubation, a positive result appear as an arrowhead shaped zone of complete hemolysis ⚫ S. agalactiae is CAMP test positive while non gp B streptococci are negative Group B Streptococcus S. aureus (Spingomyelinase C) (CAMP Factor) Group A Streptococcus Enhanced Zone of Hemolysis Differentiation between α-hemolytic streptococci ⚫The following tests are used to differentiate between S. pneumoniae & viridans streptococci ⚫ Optochin Test ⚫ Bile Solubility Test ⚫ Inulin Fermentation SHEET NOTE #12 • Optochin test = antibiotic test • We add the bacteria to the blat if it sensitive it will be S.pneumonia and if it resistant it will be Viridans .S • When we add the bile to the blat direct the colones will appear directly • If the three tests are positive (sensitive) the Bacteria is S.pneumonia • if the three tests are negative (resistant ) the bacteria is Viridans .S Optochin Susceptibility Test ⚫ Principle: ⚫ S. pneumoniae is inhibited by Optochin reagent (<5 µg/ml) giving a inhibition zone ≥14 mm in diameter ⚫ Procedure: ⚫ Blood agar inoculated with organism ⚫ OP disk is placed on the center of inoculated blood agar ⚫ After incubation at 37°C for 18 hrs, accurately measure the diameter of the inhibition zone by the ruler ⚫ ≥14 mm zone of inhibition around the disk is considered as positive and ≤13 mm is considered negative ⚫ S. pneumoniae is positive (S) while S. viridans is negative (R) Optochin resistant S. viridans Optochin susceptible S. pneumoniae

Gram Positive Cocci

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