Streptococci

STREPTOCOCCI (from the Greek streptos-chain and coccus-coccus), spherical bacteria arranged in chains. One part of them is pathogenic and capable of causing local and general diseases in humans and animals, while the other is devoid of pathogenic properties. Streptococci were first described in tissues during erysipelas in 1881 by R. Koch and in pus by Ogston. They were isolated in pure culture and studied in detail by Fehleisen in erysipelas in 1883 and Rosenbach in phlegmon in 1884. To date, up to 60 species of streptococci have been described. The shape of individual spheres is most often round, sometimes oval, with a diameter of about 1 µm. The length of the chains varies both in different species and in the same culture depending on the properties of the nutrient medium. On solid nutrient media, streptococci most often form either very short chains or are arranged in uncharacteristic groups. In the tissues of the affected organ, streptococci mostly form short chains or sometimes lie in pairs [see separate table, figure 1]. Streptococci, with rare exceptions, are Gram-positive; streptococci do not possess active motility, and sometimes very lively molecular movement is observed. They do not form spores, and capsules are very rare. Usually, streptococci are facultative anaerobes, with the exception of S. putridus, which is an obligate anaerobe. On ordinary nutrient media, streptococci develop slowly, scantily, and die at 37° within 1-3 days. Streptococci require media rich in protein or carbohydrates (see Nutrient media), but even in such media, most streptococci die comparatively quickly (2-4 weeks). Streptococci cultures are preserved longest in serum-gelatin, as well as in semi-solid meat-peptone agar (0.1%) without the addition of sugar, under a layer of vaseline or paraffin oil, in an icebox. The rapid death of streptococci in culture is mainly due to the increasing action of acid, which is formed from the carbohydrates of the nutrient medium under the influence of streptococcal life activity. According to Foster, the optimal hydrogen ion concentration for streptococci is 7.4. When cultivating streptococci in sugar broth, the pH in a mature culture reaches 4.85-5.40. The addition of buffer substances to the medium (alkaline salts of the same acids, proteins) helps preserve streptococci. In broth, streptococci mostly grow in the form of a granular or flaky loose sediment, leaving the medium transparent. Turbidity of the broth or the formation of a film is observed in strains growing in the form of short chains. On agar, streptococci form small, about 0.5 mm in diameter, transparent colonies with an even or slightly serrated edge, and under low magnification, they are fine-grained. (Growth on blood media—see Microorganisms, table.) On gelatin, the appearance of the colonies is the same as on agar, with the exception of those rare varieties that liquefy gelatin. Streptococci tolerate drying well. When dried in a protein medium (sputum, pus, blood, etc.), streptococci survive for up to several months. At 60° they die in 1 hour, at 100° in a few seconds. A 1% sublimate solution, as well as a 5% phenol solution, kills streptococci in 15 minutes. The following biochemical properties can be considered characteristic of streptococci. Streptococci ferment various carbohydrates with the formation of only acid (mono-, di-, polysaccharides, alcohols, and glycosides). Gas formation (CO₂) is produced by only one variety—S. kefir. The acids formed are mainly lactic, followed by acetic, propionic, and traces of formic and butyric. Streptococci do not produce a proteolytic enzyme, with the exception of S. liquefaciens (Orla-Jensen), which liquefies gelatin. They do not form indole. They do not reduce nitrates. In litmus milk, they either form only acid, or form acid and coagulate, or grow without changing the medium. It should be noted that streptococci of various origins can be biochemically identical, and streptococci of the same origin can differ in a number of biochemical characteristics. Pathogenic to humans are mainly hemolytic streptococci, less often green (viridans), and very rarely other varieties. Mice, rabbits, to a lesser extent guinea pigs, and very little rats, cats, dogs, as well as cattle, which usually react only with a local reaction, are susceptible to streptococci. In general, hemolytic streptococci can be significantly more pathogenic to animals than green and non-hemolytic ones. By passages through animals, it is possible to increase the virulence of streptococci to high limits. Marmorek obtained a strain of hemolytic streptococcus in this way with a lethal dose for rabbits of 0.000001 cm³, and Aronson for white mice of 0.00000001 cm³. The virulence of streptococci usually increases in relation to the animal species through which the streptococcus is passaged. Thus, Koch and Petruschky infected humans without any effect with large doses of streptococci virulent for rabbits. Thus, the virulence for animals of a given streptococcus strain cannot serve as a criterion of its virulence for humans. According to Ruge and Philipp, streptococci virulent for humans, in contrast to avirulent ones, possess the ability to multiply in human blood. Based on this, for prognostic purposes, especially in postpartum diseases, Philipp proposed the following method for determining streptococcal virulence. One loopful of a daily broth culture of the test streptococcus is inoculated into 0.5-1.5 cm³ of freshly obtained, defibrinated human blood. An increase in the number of colonies after 3 hours of growth indicates virulence. With mixed or strictly localized infection, Philipp's test fails. Upon subcutaneous administration to animals, depending on the dose, strain virulence, and organism susceptibility, a various course of infection can be observed—from an infiltrate at the injection site to fatal septicemia. Upon intravenous administration of streptococci, animals also react variously. In this case, rapid death from septicemia or damage to individual organs can be observed. When administered by mouth, according to the experiments of Tonarelli and Bail, streptococci can also cause disease in animals, especially young ones. In this case, acute enteritis develops. Subsequently, the infection becomes generalized. Toxic products of streptococcus life activity. Hemolysins, streptolysins. Streptococcal hemolysins were discovered in 1893 by Knorr and obtained in pure form by Bezredka (1901), who named them streptolysin. The method of obtaining streptolysin, according to Bezredka, is as follows: a rabbit is inoculated subcutaneously with a daily virulent broth culture of streptococci. One day after the death of the animal, 2-3 drops of hemolyzed blood are taken from the heart of the carcass and inoculated into sterile rabbit serum or serum-broth. The 24-hour culture is filtered through a Chamberland candle or centrifuged. The resulting filtrate contains streptolysin. It lyses the erythrocytes of humans, cattle, guinea pigs, and rabbits. Streptococcal hemolysin differs from other bacterial hemolysins by a number of properties. In artificial cultures of streptococci, the amount of hemolysin reaches a maximum within 1-2 days of culture growth. In this connection, the zone of clearing around the streptococcal colony on blood agar, formed during the first day of growth, does not increase further in the future. Streptococci can cause the dissolution of erythrocytes in the living animal body, which is a specific feature of streptolysins only. Streptolysin is destroyed at 70° in 2 hours, and at room temperature in 20 days. The optimum action of streptolysin is 37°. Streptolysin is not toxic. Streptolysin does not possess antigenic properties. Sometimes normal sera possess antihemolytic action. There is no constant parallelism between the ability to hemolyze and the pathogenicity of streptococci. Although hemolytic streptococcus is most frequently isolated in human diseases, it can also be found in healthy people (saprophytes of the mouth, nose, vagina). Along with hemolysin, virulent streptococci produce streptoleukocidin. Leukocidin (see), a substance that kills white blood cells, can be determined using the reductase test with methylene blue (leukocytes + streptococci + methylene blue). Living leukocytes reduce methylene blue, while those subjected to the action of streptoleukocidin lose this ability. The question of the ability of streptococci to produce a true toxin has not been resolved. To date, toxigenic properties have been most studied in scarlatinal streptococcus (Savchenko, Dick). Besides hemolysin and leukocidin, streptococcus secretes a substance of the endoenzyme character—hemoglobinotoxin, which possesses the ability to convert oxyhemoglobin into methemoglobin. The formation of methemoglobin was first noted by Gilbert and Fournier in 1896 and then established spectroscopically. This property is persistent and is especially well expressed in green streptococci. On blood agar, the colonies of green streptococci have a characteristic brownish-green color. The medium around the colony takes on the same shade. Hemolytic streptococci can also possess this ability. Tunnicliff proposes to differentiate streptococcus of scarlatinal origin from other hemolytic streptococci on the basis of pigment formation on chocolate agar [see sep. table (art. 855-856), fig. 2]. The substance causing this ability of streptococci is destroyed at 65° in 30 minutes, and at 100° in 5 minutes. It can be isolated from living streptococci and while doing so retains its activity.

There is no correlation between the hemometamorphosing property and pathogenicity of streptococci. There is not a single known trait of streptococci that has not been attempted to be used as a criterion for their classification. Kocher and Tavel, just like Lingelsheim, Baer, Knorr, and Kurth, attempted to classify streptococci by the length of their chains. Kurth, Behring, and others classified them by the character of growth of streptococci in broth, and Hiss by biochemical properties, particularly in relation to carbohydrates. However, all these attempts turned out to be untenable in view of the inconstancy of the indicated traits. The classification of streptococci according to their relation to red blood cells, which enjoys great popularity at present, also cannot be considered entirely satisfactory due to the variability of the hemolytic properties of streptococci and the proven transformation of hemolytic streptococci into green and non-hemolytic ones (Morgenroth and others). The classification of streptococci according to their agglutination properties (Meyer) has not gained widespread acceptance in view of frequent individual deviations in the receptor apparatus of individual strains of streptococci. Finally, the criterion of pathogenicity proposed by Neisser and Gins also turned out to be untenable, since not a single species of streptococci is either absolutely pathogenic or absolutely saprophytic. It also proved impossible to classify streptococci according to the disease they cause. Petrushka's experiments on animals showed that one and the same species of streptococcus under various experimental conditions can cause erysipelas, phlegmon, or sepsis. A natural classification of streptococci, just like that of other bacteria, does not exist, since the genetic relationship between various groups of streptococci is unknown, and even the question of whether all streptococci are a single species of bacteria or whether there are many different species of streptococci remains controversial. Practically, the most satisfactory classification should be recognized as that of Schottmüller, which is constructed on the basis of taking into account the most characteristic and stable traits of streptococci. Schottmüller divides streptococci into two groups: 1) S. longus, s. haemolyticus, exhibiting hemolytic properties on blood agar; 2) S. mitior, s. viridans, producing green colonies on blood agar without hemolysis of the surrounding medium. By American authors, the first type of streptococci is designated as β, and the second as α. It is advisable to add to Schottmüller's classification a third type designated by American authors as γ, to which belong streptococci that are neither hemolytic nor form a green pigment. All streptococci belonging to this type are saprophytes. In recent years, the classification of the Society of American Bacteriologists (S. A. B.) has attracted attention. This classification (Bergey, 1926) divides streptococci into 25 types on the basis of biochemical traits and agglutinin absorption. The S. A. B. classification undoubtedly has its defects: a large number of types, sometimes differing only in non-essential and labile traits, the dubious value of the agglutinin absorption reaction as a clear method for differentiating individual types of streptococci, etc. The role of streptococci in human pathology. In the healthy human organism, streptococci are very frequently normal inhabitants of the mucous membranes. In the oral cavity, the green streptococcus is most frequently encountered, but in a certain percentage, sometimes quite significant, hemolytic and non-hemolytic streptococci are found. Streptococci discovered in the intestines of healthy people and named S. faecalis are at present isolated from the group of streptococci as an independent species—the enterococcus (see). Both hemolytic and non-hemolytic streptococci are frequently found on the vaginal mucosa of healthy women, as well as in the anterior part of the urethra, in the region of the anus and perineum. However, the question of the role of these streptococci in the occurrence of postpartum complications is unclear. In any case, healthy, undamaged mucosa and skin serve as a sufficient barrier for the penetration of streptococci into the organism. In the presence of appropriate conditions (decrease in the body's resistance, disturbance of the permeability of mucous membranes, etc.), streptococci can penetrate the organism and affect any organ. Apparently, there is no organ or tissue that is not sensitive to the streptococcus. Streptococci are the causative agents of various suppurative processes, erysipelas (see), and sepsis (see); their role in rheumatism (see) and scarlet fever (see) has not been definitively established. Spontaneous streptococcal infections do not leave behind any solid and lasting immunity. By means of active immunization with increasing doses of weakened or killed cultures, it is possible to immunize laboratory animals, rabbits, and mice, against lethal doses of a streptococcal culture. The main factor of immunity in experimental animals is the appearance in the blood of various antibodies and especially specific bacteriotropins, which promote the phagocytic activity of leukocytes (Denys, Bordet, Neufeld). Antitoxins appear upon immunization of animals with broth cultures of streptococci or their filtrates. The question of the specificity of these antitoxins and their significance in immunity has not been resolved. Attempts to use specific serum therapy in streptococcal infections were made a very long time ago. Therapeutic serum was obtained by immunizing horses with one or many species of streptococci. Marmorek used strains of streptococci passaged through mice to immunize horses. The serum proved to be very effective in experiments on mice, but very weak with respect to streptococci freshly isolated from humans (Aronson). Therefore, in the future, the preparation of anti-streptococcal sera was carried out using streptococcal cultures freshly isolated from humans (Gabrichevsky, Tavel, Moser). Anti-streptococcal sera are used in many streptococcal infections (erysipelas, puerperal diseases, sepsis, etc.). The most widely used is anti-scarlet fever serum (see Scarlet fever). In addition to serum therapy in the treatment of streptococcal infections, mainly local ones, vaccine therapy is used, most often autovaccine therapy, as well as anti-virus therapy (see Vaccination, vaccines, and Anti-virus). Preventive immunization in streptococcal diseases has found application only in scarlet fever (see). Diagnostics of streptococcal infections. In blood examination, sterilely obtained blood from the cubital vein in an amount of 8–10 cm³ is seeded into 100–200 cm³ of sugar broth. After a 24–48-hour stay in the incubator, if streptococci are discovered in smears from the culture, a seeding of 1–2 loops onto blood agar in Petri dishes is made, from which a pure culture of streptococcus is isolated a day later. It is very convenient in difficult cases for practical purposes to use the following scheme for the differentiation of streptococci from pneumococci and enterococci: (stain by Ziehl). The site of the most frequent primary localization of streptothrix is the lungs. The disease usually proceeds extremely protractedly, presenting a picture of chronic bronchitis with the development of peribronchial granulation nodules and bronchopneumonic foci, which frequently undergo suppuration. The latter serves as the cause of the emergence of bronchiectases and cavities, which appears especially typical for streptothrix. The process for the most part long bears a local focal character, not being accompanied by the development of extensive destructive granulations, as in typical actinomycosis, and almost always is treated initially as tuberculous, until finally a bacteriological examination of the sputum clarifies the matter. In favorable cases, Microorganisms / Shape of cocci / Capsule formation / Growth on ascitic agar / Hemolytic properties / Dissolution by bile / Growth in bile / Aesculin-splitting ability in cultures / Pathogenicity for white mice / Str. haemolyticus / round / — / — / + / — / — / — / weak / Str. viridans / round / — / — / — / — / — / — / usually mildly pathogenic / Str. anhaemolyt. / round / — / — / — / — / — / — / non-pathogenic / Pneumococcus / lancet-shaped / + / + / — / + / + / — / pathogenic / Pneum. mucosus / lancet-shaped / + / mucous / — / + / + / — / pathogenic / Enterococcus / round / — / + / — / — / + / + / non-pathogenic. Examination of pleural and joint fluid is carried out in the same way as blood examination. In the examination of cerebrospinal fluid, seedings are made onto serum agar, ascitic agar, or Bailey's agar, since both streptococci and meningococci grow on these media. In the examination of pus, seeding is done onto blood agar in Petri dishes and simultaneously microscopy of the pus is performed. Mucus from the pharynx, taken with a sterile swab, is examined in the same way. It is recommended to simultaneously seed the material into sugar broth, from which, after a 24-hour stay in the incubator, subcultures are made onto Petri dishes with blood agar. This method increases the percentage of positive results. Lactic acid streptococci — see Lactic acid bacteria.