C hair of Medical Biology, M icrobiology, V irology, and I mmunology

1. Chair of Medical Biology, Microbiology, Virology, and Immunology Staphylococci. Streptococci. Meningococci and Gonococci Lecturer Prof. S.I. Klymnyuk

2. Classification. Staphylococci are included in the Firmicutes Bacteria, family Micrococcaceae, genus Staphylococcus. According to the contemporary classification, staphylococci are subdivided into more then 30 species. Among them: S. aureus, S. epidermidis, and S. saprophyticus, S. haemolyticus, S. capitis, S. hominis, S. warneri, S. xylosus etc.

3. Morphology. Staphylococci are spherical in shape, 0.8-1 mcm in diameter, and form irregular clusters resembling bunches of grapes. In smears from cultures and pus the organisms occur in short chains, in pairs, or as single cocci. Large spherical (L-forms) or very small (G-forms) and even filterable forms may be seen in cultures which have been subjected to various physical, chemical, and biological (antibiotics) factors.

9. Virulence factors Staphylococci express many cell surface-associated and extracellular proteins that are potential virulence factors. For the majority of diseases caused by this organism, pathogenesis is multifactorial. Thus it is difficult to determine precisely the role of any given factor. This also reflects the inadequacies of many animal models for staphylococcal diseases.

11. Other Extracellular Proteins. Coagulase is an extracellular protein which binds to prothrombin in the host to form a complex called staphylothrombin. The protease activity characteristic of thrombin is activated in the complex, resulting in the conversion of fibrinogen to fibrin. This is the basis of the tube coagulase test, in which a clot is formed in plasma after incubation with the S aureus broth-culture supernatant. Coagulase is a traditional marker for identifying S aureus in the clinical microbiology laboratory.

12. Enzymes. S aureus can express proteases, a lipase, a deoxyribonuclease (DNase) and a fatty acid modifying enzyme (FAME). The first three probably provide nutrients for the bacteria, and it is unlikely that they have anything but a minor role in pathogenesis. However, the FAME enzyme may be important in abscesses, where it could modify anti-bacterial lipids and prolong bacterial survival. The thermostable DNase is an important diagnostic test for identification of S aureus.

18. Laboratory diagnosis. Test material may be obtained from pus, mucous membrane discharge, sputum, urine, blood, foodstuffs (cheese, curds, milk, pastry, cakes, cream, etc.), vomit, lavage fluids, and faeces. The material is examined for the presence of pathogenic staphylococci. Special rules are observed when collecting the material since non-pathogenic strains are widespread in nature.

19. Treatment. Staphylococcal diseases are treated with antibiotics (penicillin, phenoxymethylpenicillin, tetracycline, gramicidin, etc.), sulphonamides (norsulphazol, sulphazol, etc.), and antistaphylococcal gamma-globulin.

20. Streptococci The streptococcus {Streptococcus pyogenes) was discovered by T. Billroth (1874) in tissues of patients with erysipelas and wound infections and by L. Pasteur and others (1880) in patients with sepsis. A. Ogston described the organisms in studies of suppurative lesions (1881). A pure culture of the organism was isolated by F. Fehleisen (1883) from a patient with erysipelas and by F. Rosenbach (1884) from pus. Streptococci belong to the family Streptococcaceae.

24.  Cultivation. Streptococci are facultatively aerobic, and there are also anaerobic species. The optimal temperature for growth is 37° C, and no growth occurs beyond the limits of 20-40° C for enterococci the limits are 10-45 C).

26. Fermentative properties. Streptococci are non-proteolytic, do not liquefy gelatin, and do not reduce nitrates to nitrites. They coagulate milk, dissolve fibrin, ferment glucose, maltose, lactose, saccharose, mannitol (not always constantly), and break down salicin and trehalose, with acid formation.

27. Toxin production. Streptococci produce exotoxins with various activities: (1) haemolysin (haemotoxin, 0- and S-streptolysm) which loses its activity after 30 minutes at a temperature of 55 C; disintegrates erythrocytes; produces haemoglobinaemia and haematuria in rabbits following intravenous injection; (2) leucocidin which is destructive to leucocytes; occurs in highly virulent strains and is rendered harmless by a temperature of 70 C (3) lethal (dialysable) toxin which produces necrosis in rabbits when injected intracutaneously; it also causes necrosis in other tissues, particularly in the hepatic cells; (4) erythrogenic toxin produces inflammation in humans who have no antitoxins in their blood; (5) Streptococcus pneumoniae produces alpha-hae molysin secreted into the culture fluid and beta-haemolysin which is released after lysis of the streptococci.

29. Classification. By means of the precipitation reaction founded on the detection of group specific carbohydrates, streptococci are subdivided into groups which are designated by capital letters from A to H and from K to T. Five out of the 21 known Streptococcal species cannot be related to any antigenic group. Nine species are of interest for medical microbiology; The haemolytic streptococci, recovered from sick human beings, were subdivided by F. Griffith into 51 serovars. He attributed 47 serovars to group A, serovars 7, 20, and 21 to group C, and serovar 16 to group G.

30. The organisms grow at temperatures ranging from 10 to 45 C. They are resistant to high temperature (e. g. withstand exposure to 60 C for half an hour). Enterococci can be grown in broth containing 6.5 per cent common salt at pH 9.6 and on blood agar containing 40 per cent bile or an equivalent amount of bile salts. They ferment glucose, maltose, lactose, mannitol, trehalose, salicin, and inulin, with acid formation. They reduce and coagulate litmus milk in the presence of 0.1 per cent methylene blue. Enterococci differ from other streptococci in their ability to grow over a wide range of temperatures (10-45 C) and in a medium of pH 9.6, in their resistance to high concentrations of salt and to penicillin (a number of strainsshow growth in media containing 0.5-1 U of antibiotic per 1 ml of media). All enterococci decarboxylate tyrosine.

31. Enterococci inhabit the small and large intestine of man and warm-blooded animals. The organisms possess properties antagonistic to dysentery, enteric fever, and paratyphoid bacteria, and to the coli bacillus. In the child's intestine the enterococci are more numerous than the E. coli. In lesions of the duodenum, gall bladder, and urinary tract enterococci are found as a result of dysbacteriosis. Isolation of enterococci serves as a criterion of contamination of water, sewage, and foodstuffs with faeces.

32. Streptococcus pneumoniae

36. With an exogenous mode of infection streptococci invade the human body from without (from sick people, and animals, various contaminated objects and foodstuffs). They gain access through injured skin and mucous membranes or enter the intestine with the food. Streptococci are mainly spread by the air droplet route. When the natural body resistance is weakened, conditionally pathogenic streptococci normally present in the human body become pathogenic. Penetrating deep into the tissues they produce local pyogenic inflammations, such as streptoderma, abscesses, phlegmons, lymphadenitis, lymphangitis, cystitis, pyelitis, cholecystitis, and peritonitis. Erysipelas (inflammation of the superficial lymphatic vessels) and tonsillitis (inflammation of the pharyngeal and tonsillar mucosa) are among the diseases caused by streptococci. Invading the blood, streptococci produce a serious septic condition. They are more commonly the cause of puerperal sepsis than other bacteria.

44. Role of Streptococcus in the Aetiology of Scarlet Fever Scarlet fever has long been known as a widespread disease but at the present time its aetiology has not yet been ascertained. Four different theories were proposed: streptococcal, allergic, viral, and combined (viral-streptococcal). Most scientists and medical practitioners favoured the streptococcal theory. It is assumed that scarlet fever is caused by group A beta-haemolytic streptococci which possess M-antigen and produce erythrogenic exotoxin.

47. Laboratory diagnosis. Test material is obtained from the pus of wounds, inflammatory exudate, tonsillar swabs, blood, urine, and foodstuffs. Procedures are the same as for staphylococcal infections. Tests include microscopy of pus smears, inoculation of test material onto blood agar plates, isolation of the pure culture and its identification. Blood is sown on sugar broth if sepsis is suspected. Virulence is tested on rabbits by an intracutaneous injection of 200-400 million microbial cells. Toxicity is determined by injecting them intracutaneously with broth culture filtrate.

48. The group and type of the isolated streptococcus and its resistance to the medicaments used are also determined. In endocarditis there are very few organisms present in the blood in which they appear periodically. For this reason blood in large volumes (20-50 ml) is inoculated into vials containing sugar broth. If possible, the blood should be collected while the patient has a high temperature. In patients with chronic sepsis an examination of the centrifuged urine precipitate and isolation of the organism in pure culture are recommended. Besides, the group and type of the isolated streptococcus are identified by means of fluorescent antibodies. Serological methods are also applied to determine the increase in the titre of antibodies, namely streptolysins O and antihyaluronidase.

49. Meningococci The meningococcus (Neisseria meningitidis) was isolated from the cerebrospinal fluid of patients with meningitis and studied in detail in 1887 by A. Weichselbaum. At present the organism is classified in the genus Neisseria, family Neisseriaceae