Cocci

Staphylococcus “staphyle” in Greek

Staphylococcus S.aureus: most virulent species. S.epidermidis: opportunistic pathogen. S.saprophyticus: rarely cause human diseases.

Biological characteristics Shape and structure Size: 0.5-1.0 m in diameter Shape: spherical Arrangement: grape-like Structure: capsule non-motile non-sporulating Staining: G+

Biological characteristics Cultivation Requirements media: all-purpose media gas: facultative anaerobes specially: tolerating 10-15% NaCl

Biological characteristics Growth properties on solid medium: small colony with various pigments on blood agar: zone of complete hemolysis (-hemolysis) S.aureus

Biological characteristics Biochemical reactions Catalase test “+” Staphylococci: “+” Streptococci: “-” Mannitol Fermentation “+” S. aureus: “+”; S.epidermidis & S.saprophyticus: “-”;

Biological characteristics Antigenic composition Capsules teichoic acid , peptidoglycan Protein antigen: SPA SPA capsule teichoic acid peptidoglycan

SPA (Staphylococcal Protein A ) Characteristics nonspecific bind to the Fc fragment of IgGs

SPA(staphylococcal protein A) Function: virulence factor Anti-phagocytosis damage platelet induce hypersensitivity and inflammation Phagocyte Fc receptor IgG SPA coagglutination test SPA-coated S.aureus is used as Ab carriers BACTERIUM

SPA (staphylococcal protein A ) S.aureus Ab

Biological characteristics---Classification Based on the pigments and chemical reactions Properties S. aureus S. epidermidis S. sarprophyticus Pigment Golden yellow White Citrine Coagulase + － － Mannitol + － － Thermostable nuclease + － － α-hemolysin + － － SPA + － － Pathogenicity strong Weak -

Biological characteristics---Classification Based on coagulase Coagulase” +”: e.g., S. aureus Coagulase” -”: e.g., S. epidermidis & S. saprophyticus Phage typing S. aureus: 3 phage groups, 26 phage types. group 1: TSST-1-producing strains group 2: exfoliative toxin-producing strains group 3: enterotoxin-producing strains ----is of epidemiological value

Biological characteristics Resistance: Drying Heat high concentration of salt (10-15% NaCl) Sensitivity: basic dyes (crystal violet) several antibiotics (penicillin, vancomycin) MRSA (methicillin resistance S.aureus)

Pathogenicity (S.aureus) Virulence factors Surface structure capsules, peptidoglycan, teichoic acid, SPA Enzyme Toxin

Pathogenicity (S.aureus) Virulence factors Enzymes: coagulase 血浆凝固酶 heat-stable nuclease 耐热核酸酶 hyaluronidase 透明质酸酶 lipase 脂酶 β – lactamase β-内酰胺酶 Exotoxins: Hemolysin 溶血素 Leukocidin 杀白细胞素 Staphylococcal enterotoxin 葡萄球菌肠毒素 TSST-1 毒性休克综合征毒素-1 Exfoliative toxin 表皮剥脱毒素

Enzyme Coagulase: An enzyme that converts fibrinogen into fibrin causing the coagulation of blood. Classification: Free coagulase 游离凝固酶 Bound coagulase 结合凝固酶

Coagulase Biological activity Significance Antiphagocytosis Inhibit the damage of bactericidal substances Formation of limited abscess Significance criterion for identification of S. aureus (pathogenic)

Coagulase test Tube test for free coagulase Slide test for bound coagulase

Hemolysins Roles: damage membrane permeability; cytotoxic effects on phagocytes and tissue cells Four kinds: -Lysin -Lysin -Lysin -Lysin S.Aureus in blood agar

Leukocidin Biological activity Impairment of membrane of WBC

Staphylococcal enterotoxin Source: 50% S. aureus Types: A、B、C1-3、D、E、G、H Chemical and physical characteristics Protein Heat stable (100℃, 30 min) Resistant to gut enzymes Mechanism: act on vomiting center, superantigen Disease: food poisoning

Superantigens and the non-specific stimulation of T cells

Toxic Shock Syndrome Toxin-1(TSST-1) Source: phage group Ⅰ of S. aureus Biological activity fever Increase the sensitivity to endotoxin superantigen Disease: TSS

Exfoliative toxin Source Biological activity protease activity phage group Ⅱ of S. aureus Biological activity protease activity Disease: SSSS Staphylococcal scalded skin syndrome

Pathogenicity Disease Invasive infection/pyogenic infection local infection: folliculitis; boil ; carbuncle ; impetigo organ infection: pneumonia; meningitis Systemic infection: Septicemia; pyemia Toxin-associated diseases

hair folliculitis boil

Pathogenicity Toxin-associated diseases Food poisoning (enterotoxin) TSS (Toxic shock syndrome) SSSS (staphylococcal scalded skin syndrome) Staphylococcal enterocolitis -dysbacteriosis

Staphylococcal scalded skin syndrome (SSSS) Most often occurs in infants and young children

CNS Coagulase-Negative Staphylococci Virulence factor: slime Antibiotic-resistance Opportunistic infection S. epidermidis, S.saprophyticus

Diseases caused by coagulase-negative staphylococci urinary tract infection Bacterial endocarditis Septicemia Infections associated with indwelling devices

Laboratory diagnosis Specimen Direct smear and Gram stain Isolation and identification Primary criterions: coagulase test, thermostable nuclease gold yellow pigmentation -hemolysis mannitol fermentation Enterotoxin test (animal test) Antibiotic susceptibility tests

Control Nosocomial infection Aseptic measures Prevention Drug resistance hygiene Treatment Autovaccino-therapy Antibiotic susceptibility tests

Streptococcus

Shape: G+ cocci structure: capsule in chains cell membrane cell wall pilus-like structure structure: capsule pilus-like (LTA-M protein) carbohydrate antigen protein antigen: M, R, T carbohydrate antigen and protein antigen

Classification Hemolytic activity Incomplete hemolysis -hemolytic strep. Incomplete hemolysis Opportunistic pathogens e.g., S.pneumoniae, S.viridans -hemolytic strep. Complete hemolysis() Major human pathogens e.g., S. pyogenes -streptococcus No hemolysis, No pathogenicity e.g., enterococci.

a b c

Classification Antigenic structure ( Lancefield 血清学分群 ) Streptococcus C carbohydrate antigen group-specific antigen 20 groups (A~H, K~V) Group A- main human pathogens Protein antigen type-specific antigen M, R, T protein Group A >100 types Cell wall Streptococcus capsule protein Polysaccharide Peptidoglycan

Classification Biochemical reaction 生化反应 血清学分类 溶血反应 化脓性链球菌 咽峡炎链球菌 无乳链球菌 停乳链球菌 牛链球菌 草绿色链球菌 肺炎链球菌 A A C F G 无相关性 B C G D 无相关性 溶血 溶血，偶见溶血或不溶血 溶血，偶见不溶血 溶血或不溶血，偶见 溶血 溶血或不溶血 溶血

S. pyogenes

Biological characteristics shape and size: spherical, 0.6～1.0 μm in diameter structures: capsule (hyaluronic acid) nonmotile nonsporeforming highly nutritive -hemolysis catalase negative

Pathogenicity Virulence factors Surface structures Enzymes Exotoxins

Pathogenicity Virulence factors Surface structures adhesin: LTA F protein M protein: anti-phagocytosis cross-reactive antigen lipoteichoic acid F-protein M protein fibronectin epithelial cells

Pathogenicity Virulence factors Enzymes Hyaluronidase Streptokinase (SK) Streptodornase (SD) (DNase, DNaseB-Ab)

Pathogenicity Virulence factors Exotoxins Streptolysin (hemolysin) Erythrogenic toxin

toxic to a variety of cell types Streptolysins streptococci group A, C, and G; Classification: Properties Streptolysin O (SLO) Streptolysin S (SLS) O2 oxygen-labile oxygen-stable Antigenicity Strong (ASO test) Weak Chemical Protein Polypeptide Activity toxic to a variety of cell types

Erythrogenic toxin Pyrogenic toxin /scarlet fever toxin Protein Antigenicity Superantigen Scarlet fever

Abscess with surrounding cellulitis Pathogenicity Pyogenic infection: Local purulent infections pharyngitis, tonsillitis, puerperal fever 产褥热 erysipelas 丹毒, cellulitis 蜂窝织炎 impetigo Systemic infection: septicemia Abscess with surrounding cellulitis

Erysipelas Erysipelas on the cheek

Toxin-associated diseases scarlet fever Hypersensitive disease Acute glomerulonephritis Rheumatic fever Hypersensitivity type II & III

Laboratory diagnosis specimen S.aureus Isolation and cultivation Direct smear S.pneumoniae PYR test -hemolytic strep. S. Pyogenes 阳性

Laboratory diagnosis Serologic diagnosis ASOT (anti-SLO test) a neutralization test between the toxin (SLO) and its specific anti-toxin (ASO) helping to diagnose rheumatic fever and acute glomerulonephritis positive standard：ASO titer >1:400

Control Treat the pharyngitis and tonsillitis in time Antibiotics: penicillin G for the first choice

S. pneumoniae (Pneumococcus )

General properties G+, arranged in pairs, lancet-shape Capsule Polysaccharide Type-specific antigen Fastidious blood agar -hemolysis 有氧条件下可以 形成 -hemolysis，胆汁溶菌试验，菌液中加入10% 去氧胆酸钠或2%牛磺胆酸钠，或牛、猪、兔等新鲜胆汁，置室温或37，在5－10min内出现溶解

General properties Autolysis Bile solubility test: “+” (-) (+) (-) (+) (Bile solubility test)

Pathogenesis Virulence factor Main disease Capsule Pneumonia Neuraminidase Pneumolysin LTA, peptidoglycan Main disease Pneumonia Others: middle ear infections (otitis media), sinusitis, meningitis, septicemia

Non-typical pneumonia

Laboratory diagnosis Identification of pneumococci from S.viridans bile solubility test optochin sensitivity test quellung reaction (capsular swelling test) positive negative

Control Treatment: penicillin G Prevention: polysaccharide vaccine (for children and elderly)

Viridans streptococci S.mutans S.anginosus S.salivarius S.mitis S.sanguis dental caries subacute bacterial endocarditis The viridans group of streptococcus are a heterogeneous collection of alpha-hemolytic and nonhemolytic streptococci. Their group name is derived from viridis(Latin for “green’) because many of these bacteria produce a green pigment on blood agar media. The taxnomic nomenclature for these species is confusing because European and American microbiologist have not reached a consensus on it. Thus different species name are often used interchangeably in the literature. At least 15 species? Have been identified in the United States, and they ar bile solubility test （－） optochin sensitivity test（－）

S.agalactiae Group B streptococcus, GBS Transmission Disease vaginal flora Disease neonatal meningitis neonatal pneumonia septicemia

S. suis

Neisseria

Neisseria N. gonorrhoeae N. meningitidis N. sicca N. subflava N. mucosa N. lactamica

General properties Shape and structure G- diplococci Coffee bean-shaped kidney-shaped Lipooligosaccharide (LOS) Pili Closely related DNA homology 70%

General properties Cultivation Fastidious: chocolate agar plate Obligate aerobes, 5～10%CO2 Autolysis (N.meningitidis) Oxidase positive Fermentation certain sugars N. gonorrhoeae : Glucose N. meningitidis : Maltose & Glucose

Fragile - don’t survive long outside the host General properties Very weak: cool, heat, drying, disinfectants Fragile - don’t survive long outside the host

Pathogenesis LOS PILI N. gonorrhoeae N. meningitidis Virulence Factors Similar, but – Differences in utilization LOS LOS IgA1 protease Capsule iron-binding proteins PILI Outer Membrane Proteins (Por,Opa, Rmp proteins) IgA1 protease iron-binding proteins PILI Outer Membrane Proteins

N. gonorrhoeae /gonococcus PILI Nonpiliated gonococci not virulent Phase variation turns pilus production on/off Antigenic variation

N. gonorrhoeae /gonococcus Outer Membrane Proteins Porin proteins (Por) = prevent phagolysosome fusion & allow intracellular survival [protein I] Opacity proteins (Opa) = binding of organisms to epithelium [protein II] Reduction-modifiable proteins (Rmp) = protection against bactericidal antibodies [protein III]

N. meningitidis/menigococcus Capsule 13 serotype A, B, C, Y, W-135 LOS (lipooligosaccharide) vascular damage skin rash renal failure shock disseminated intravascular coagulation (DIC)

Pathogenesis X LOS PILI N. gonorrhoeae N. meningitidis Virulence Factors Similar, but – Differences in utilization LOS LOS IgA1 protease iron-binding proteins Capsule PILI Outer Membrane Proteins (Por,Opa, Rmp proteins) IgA1 protease iron-binding proteins PILI Outer Membrane Proteins X NO capsule

Disease Gonorrhea Adults Transmission: STD (sexually transmitted disease) Clinical disease: Genitourinary tract infection Male-Urethritis Female-Urethritis, cervix inflammation PID (Pelvic Inflammatory Disease) Sterility

Gonorrhea Symptomatic infections are notably PURULENT Urethritis

Disease Gonorrhea Newborns Ophthalmia neonatorum

Gonorrhea Purulent conjunctivitis Ophthalmia neonatorum Infection in newborns during vaginal delivery

Disease Source: patients and carriers Transmission: respiratory tract nasopharynx→local infection→septicemia→meningitis Disease: epidemic cerebrospinal meningitis

Laboratory diagnosis Specimens Smears Culture Thayer-Martin VCN Oxidase test Maltose fermentation Antigen Detection and Enzyme Immunoassays Radioimmunoassay ELISA SPA coagglutination test Others DNA Probe Hybridization

Treatment ＆Prevention N. gonorrhoeae Antibiotic 1%Silver nitrate-- ophthalmia neonatorum No effective vaccine

Antibiotic Resistance Increase in penicillin resistance since 1976 PPNG (penicillinase-producing N. gonorrhoeae ) plasmid mediated due to beta lactamase production Tetracycline resistance recognized in 1985 TRNG (Tetracycline resistance N. gonorrhoeae ) due to chromosomal mutation Sensitive to quinolones, cephalosporins Penicillin used to be the drug of choice for gonococcal infections because they were essentially all sensitive to it. However, penicillin resistant strains were initially reported in Southeast Asia and were brought to the U.S. by military men returning from Viet Nam. The first ones were reported in this country in 1976. Now the recommended dose of penicillin has risen from 200,000 units in 1945 to 4.8 million units (24 times higher) today. This resistance is due to beta lactamase production that is encoded for on a plasmid (like it is in Staphs), so it can be transmitted from one strain to another. Then in 1985, a chromosomal mutation encoding for tetracycline resistance was recognized in N. gonorrhoeae. So far, however, we can still treat these organisms with a number of our current antibiotics, including the quinolones and cephalosporins you will be hearing about.

Treatment ＆Prevention N. gonorrhoeae Antibiotic 1%Silver nitrate-- ophthalmia neonatorum No effective vaccine

Treatment ＆Prevention N. meningitidis Treatment penicillin: the first of choice sulfasulfonamide Prevention immunization: capsule polysaccharide group B meningitis

Pathogenic cocci/Pyogenic cocci Staphylococcus aureu Streptococcus pyogenes Streptococcus pneumoniae G+ cocci Neisseria meningitidis Neisseria gonorrhoeae G- cocci

Neisseriae of medical importance Properties N. meningitidis N. gonorrhoeae Gram stain Gram-negative Capsule Yes No Potal of entry Respiratory tract Genital tract Virulence factors Pili Outer membrane protein IgA1-protease Iron-binding protein LOS, Capsule LOS Disease Epidemic cerebrospinal meningitis Gonorrhea (adults) Ophthalmia neonatorum (newborns) Prevention Polysaccharide vaccine Tetracycline, erythromycin ointment or AgNO3 for ophthalmia No vaccine for gonorrhea