1. Streptococci M.O : Disease S.agalactia,S.dysagalactia : mastitis in cattle,sheep S.equi subsp. Equi : Strangles, Suppurative conditions (horse) S.zooepidemicus horse,cattle,pigs,lambs S.pyogens scarlet fever,septic sore throat, rhumatic fever (human) S.pneumoniae pneuomonia (human) Enterococcus faecalis mastitis, endocarditis, urinary tract Enterococcus faecium Fixed film Direct (milk deposits, By FAT (tissue for S.suis) after culture on B.A. Microscpe :- Gr+ve cocci, chains, facutative anaero. Catalase and oxidase negative,non motile,non spore forming

2. Streptococcus pyogenes Gram +ve cocci arranged in chains Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 8 In culture In tissue

3. Streptococcus pneumoniae Gram +ve lancet shape encapsulated diplococci arranged singly or in chains Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 9

4. Streptococcus pneumoniae Capsule swelling test after using type specific antisera Gram stain of sputum stain Capsule swelling

5. Culture Pus, CSF, Swab Blood agar (B.A.), no growth on nutrient agar(fastidious) milk Selective,diff. medium Edwards medium ( B.A. + crystal violet + sod. Azide + esculine) Medium for enterococci ( 6% Nacl + 40%bile, Ph. 9.6) Incubate aerobically 37C,24-48h. Or at 10-45C,24-48.(+ve growth for enterococci) Colony : dew drop like with or withot hemolysis B-hemol. S.Pyogens S.equi α -hemol. S.agalctia S.dysagal. Esculin –ve S.agalactia Escuin +ve Non pthog. sterpt

6. Partial and complete hemolysis Partial or alpha Complete or beta

7. Streptococcus pyogenes S.pyogenes colonies surrounded by zone of B-hemolysis B-hemolysis Dew drop like colonies Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 10

8. Streptococcus pneumoniae Mucoid colonies surrounded by alpha hemolysis(greening) Mucoid colonies Green colour Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 13

9. Differentiation of streptococci from other cocci 1-Micrococci : Catalase positive Oxidative reaction( OF test) bacitracin sensitive 2-Streptococci : Catalase negative Fermentative reaction (OF test) 3-Corynebacterium equi : Catalase positive OF( unreactive) Strept. Catalase -ve OF(fermentative )

10. Classification of streptococci Brown classifi. Acc. To hemolysis type Beta ( S.pyogens, S.equi) Alpha (S.pneumoniae, viridans) Gamma (s.lactis ) Sherman classifi. based on growth at 10-45C arginin hydrolysis growth at Ph9.6 Groups : pyogenic trept, enterococci, lactic strept. Viridans strept. Lacifield Classifi.(serological) Ag is unknown.(cell wall polsacchrid(hapten) of sterpt.) colony suspention +hot HCL centrifuge supernatant Ab. Is known( from market) test : ring test in capillary tube reading results: G: A S.pyogens (human) G:B S.agalactia (cattle) G:C S.equi (horse) S.zooepidemicus (man, other animals) S.dysagalactia (cattle, other animals) Groups: A to H K to T

11. Exotoxins 1-hemolysins : (alpha, beta, gamma- 2-streptolysin O : antigenic protein produced by most of group A and by many strains of group C and G. It stimulate antibody formation (Antistreptolysin O) formed in serum of human and animals (ASO ) 3-fibrinolysin (streptokinase): lysis of fibrin 4-erythrogenic toxin: formed by group A strept. 5-hyalourinidase : dissolve hyalouronic acid Biochemical tests: sugar ferment.of lactose,maltose, trehalose diff. group C strept group B(S.agalactiae) identified by CAMP test

12. Lab.diagnosis of mastitis streptococci A-Field tests : 1-Ph indicator test :milk + bromothymol blue(ALK or blue for abn. Milk and yellow or acid for normal milk) 2- white side test : NaoH +milk white precipitate B-Laboratory tests: M.B. stain of milk( direct staining) Hotis test : milk + bromocresol purble (incubate 37C/24h. Acid(yellow color) for S.agalactia CAMP test : +ve for S.agalactia Identification tests for streptococci Bile solubility Optochin susptibility Na hippurate CAMP Bacitraci suseplity ----+ Group A Strepto. --++- Group B Strepto. ++--- S. pneum- oniae

13. 13 Beta hemolytic Streptococci S.faecalisCAMP test Dew drop like colonies on NaCl agar 6.5 % S.aureus streak Streptococcus agalactia streak Arrow head hemolysis Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 11

16. S. agalactiae S. faecalis S. aureus CAMP Test Don’t let the streak lines touch

17. Streptococcus pneumoniae Culture on blood agar with a zone of inhibition around optochin disc Optochin disc Inhibition zone Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology

18. Streptococcus viridans Alpha hemolytic colonies with no inhibition by Optochin Optochin disc Alpha hemolytic colonies Zagazing University Fac. Of Vet. Med. Dep. Of Microbiology 15

19. Differentiation of bovine mastitis streptococci Serological typing : Latex aggl. Of lancifield groups A,B,C,D,F and G (Wellcome Lab. Slide aggl. Of groups A,B,C,F and G Growth on MacConkey medium Aesculin hydrolysis. On Edward medium CAMP test --+ S.agalactia --- S.dysagalactiae -+- S.ubris ++- E.faecalis

20. Beta haemolytic streptococci identification So that M.O belongs to Group A So this M.O does not belong to group A

21. Serum antistreptolysin O (ASO) Positive titre is the highest dilution of Ab which inhibit lysis of RBCs( Positive titre 1/300)

22. Identification of S. agalactiae Differentiation from other Strep. spp. relies on CAMP test Therefore, the CAMP test is only done on milk Principle: Group B streptococci (S. agalactiae is only Group B mastitis pathogen) produce CAMP factor, which acts synergistically with partial hemolysin of Staph. aureus Result is enhancement of S. aureus partial hemolysis

23. Case Milk sample from a cow, sample label shows alkaline millk, with precipitates, staining revealed blue chains of cocci 1-What are the suspected bacteria in the sample 2-What is the specific medium required for isolation 3-Confirmatory identification of the suspected isolates.

24. Streptococci are fastidious m.o. ensuring isolation on. a-Maconckey agar. b-nutrient agar. c-blood agar. d-specific agar agar Final identiffication of all sreptococci depends on. a-C.substance of cell wall. b-biochemical test. c-haemolysis on blood agar. d-CAMP test What would you do, when mastitic milk show long chains by methylen blue. a-culture on nutrient agar. b- culture on Edward’s media. c-biochemical tests. d-serologicol tests. I can differentiate enterococci from streptococci as they are. a-grow on Bl. agar. b-incubate for I week. c-grow at 45C, tolerate 40% bile. d-a and b. When you run for CAMP test for streptococci,you need. a-culture of s.aureus. b- culture of brucella. c-blood agre plate. d-two of them In bovine mastitis we could differentiate S.agalactia from S.dysagalactia through. a-Macfydean reaction. b-animal inoculation. c-CAMP test. d-all of them.

25. Identification of Streptococci **In some cases speciation is very important!** example: differentiation of S. equi from S. zooepidemicus Also, it’s important to specifically I.D. S. agalactiae, the cause of “contagious mastitis” in cattle transmitted from cow to cow, so the cow is the PRIMARY RESERVOIR proper treatment can eliminate the infection from the herd