PHARMACEUTICAL PREPARATION FOR CONTROLLING PATHOGENIC INTESTINAL BACTERIA

Abstract: A pharmaceutical preparation for controlling pathogenic microorganisms causing diarrhoea and other gastrointestinal troubles in man and in animals contains Streptococcus lactis strain LIa in at least one pharmaceutically acceptable carrier medium in which the microorganism retains its viability. The use of the preparation for controlling pathogenic microorganisms causing diarrhoea and other gastrointestinal infections in man and in animals is also described.

Description: PHARMACEUTICAL PREPARATION FOR CONTROLLING PATHOGENIC INTESTINAL BACTERIA The present invention relates to a pharmaceutical preparation for controlling pathogenic intestinal bacteria in man and in animals, with a view to preventing and/or treating diarrhoea and other gastrointestinal ' troubles. Background of the Invention Diarrhoea and other gastrointestinal troubles caused by pathogenic microorganisms constitute a widespread health problem, which affects in particular tourists tra¬ velling abroad, people having received cancer therapy involving antibiotics or radiation, and people working in day nurseries. Such pathogenic intestinal bacteria causing diarrhoea include Salmonella, Shigella, Yersinia, E-coli, Pseudomonas, Clostridium dificile and sordelli, Stafylo- coccuε aureus and Campylobactus. These pathogenic micro¬ organisms adhere to the mucous membranes of the intestine and upset the intestinal function, thereby causing diar- rhoea. It is well-known that microorganisms affect each other positively or negatively by promoting or restraining each other's growth. This interference phenomenon has above all been studied in the flora of the skin, the pharynx and the intestine of human beings. Although the underlying causes are not fully known, it has been found that the normal bacterial flora is of considerable impor¬ tance in the defence against pathogenic bacteria. The bac¬ teria of the normal flora inhibit invading bacteria in many different ways, e.g. by producing antibiotic-like substances called bacteriocines. Unlike antibiotics, the bacteriocines mostly have a very selective effect on a specific group of bacteria without influencing the remain¬ ing bacterial flora. Treating infected patients with such harmless bacteria of specific effect has been not only discussed, but also tried to some extent. Naturally, the bacteria supplied must be able to settle in the intestinal area where they are to produce their effect. This is one of the ideas behind giving soured milk to patients whose intestinal flora is disturbed, and who suffer from severe diarrhoea after treatment with antibiotics. However, there has been no scientific follow-up of what actually happens with the intestinal flora, and the effect, if any, has mostly been uncertain, probably because no bacterial colo¬ nisation has taken place in the intestine or because the bacteria supplied lack the capacity to affect the diar- rhoea-inducing bacteria. Efforts have also been made to replace the existing bacterial flora in the nasal mucous membrane, the skin and the pharynx with harmless micro¬ organisms, and it has been shown that so-called recoloni- sation is possible and may be effective. As mentioned above, supplying bacteria to patients suffering from different illnesses induced by pathogenic microorganisms has mostly been inconclusive, obviously because of insufficient knowledge of what bacteria strains are best suited for bringing about colonisation and inter- ference. By working under given conditions, it has in recent years been possible to establish important principles for the mechanisms controlling the interaction between diffe¬ rent microorganisms, and also to develop methods for exa- mining this interaction. Tims, it has become possible to intervene in this interaction in a meaningful and repro¬ ducible fashion, which may be used therapeutically/prophy- lactically when treating infections. Summary of the Invention The present invention relates to a preparation for controlling pathogenic microorganisms in man and in ani¬ mals, with a view to preventing and/or treating diarrhoea and other gastrointestinal troubles, said preparation being characterised by containing a viable microorganism strain in the form of Streptococcus lactis strain Lla in at least one pharmaceutically acceptable carrier medium in which the microorganism retains its viability. Further characteristics of the invention are apparent from the following text and the appended claims. When studying the interaction between different bac¬ teria, we have concentrated on the ability of certain lactic-acid bacteria to prevent the growth of pathogenic bacteria. On a laboratory scale, cultures of Streptococcus lactis have been tested, and a large number of clones with different biological properties have been isolated there¬ from. Some of these have proved to have a pronounced inhi- biting effect on most of the infection-inducing bacteria in man, such as Stafylococcus aureus, various species of Salmonella, Shigella, Pseudomas, Klebsiella-Entero, and Campylobactus. An important reason for the effectiveness of the Streptococcus lactis strains is that some of these secrete bacteriocines which have a specific killing effect on some invading microorganisms. The inventive pharmaceutical preparation has a micro¬ organism strain content of 10 -10 , preferably 10 -10 , microorganisms/ml of final preparation ready for use. In a preferred embodiment, the carrier for the microorganism strain is an acidulated or fermented milk product, such as soured milk, ropy milk, yoghourt, kefir and the like. The microorganism strain may be used as starter culture when preparing such milk products, which results in acidulated or fermented milk products having an increased content, of the advantageous strain. Of course, it is also possible to add the bacteria to com¬ pleted milk products, thereby obtaining a higher content in the medium of the desired bacteria. This embodiment is of considerable importance, since the preparation obtain¬ ed is a wholesome and popular foodstuff. Such a prepara¬ tion may be administered for prophylactic or curative purposes against diarrhoea and other intestinal infec- tions often affecting infants. Further, it may be given to people who have received cancer therapy involving antibiotics or radiation, as well as to tourists travel- ling abroad where the bacterial flora is often different from that at home. The preparation may also be in dry form for oral administration, e»g. as capsules, tablets or powder. The carrier media used in this case are of conventional type and well-known to the expert in the field. The capsules may be resistant to gastric juice, so that the bacteria are protected therefrom and only released in the intes¬ tine. For the same reason, tablets may be provided with a known coating which is resistant to gastric juice. Fur¬ ther, powders may be stirred prior to use in an aqueous liquid, such as water, milk and fruit juice. The preparation has proved to be a highly effective prophylactic against various pathogenic intestinal bac- teria causing gastrointestinal troubles, such as "tourist diarrhoea". Advantageously, the preparation may also be used for treating Salmonella-infected people and animals. Further, the preparation may also be adapted for treating animals, in which case the carrier is a suitable animal feed. The microorganism should be kept in a freeze-dried state, preferably in skimmed milk in a dark and dry place, or frozen at a temperature of about -70°C. The requisite dose of the inventive preparation is determined depending on such factors as the age and gene¬ ral state of the patient, as well as the type and serious¬ ness of the illness. Anyone skilled in the art will have the competence necessary for determining the suitable dose. Another advantage of the invention is that a large overdose of the pharmaceutical preparation hardly involves any risks,at all to the patient. The Microorganism of the Invention The microorganism strain forming part of the inven- tive pharmaceutical preparation was isolated in the fol¬ lowing manner. A naturally-occurring Streptococcus lactis strain was isolated from a. steepgrass, and examined for its inhibitory effect on pathogenic intestinal bacteria. Seven clones were tested, and one called Lla gave the bes results. This microorganism strain, which is used in the inventive preparation, was deposited in 1989 at The National Collection of Industrial and Marine Bacteria (NCIMB), Torry Research Station, P.O. Box 31, 135 Abbey Road, Aberdeen AB9 8DG, Great Britain. Its NCIMB accessio number is 40157. The microorganism strain is typed or classified according to the API 20 System (API System, La Balme les Grottes, 38390 Montalieu, Vercieu, France). api 20 STREP TYPING TABLE RESULTS TESTS SUBSTRATES REACTIONS/ENZYMES NEGATIVE POSITIVE VP 1 + VP 2/Vait for 10 lin VP Pyruvate Formation of acetoin colourless rose/red NIN/wait for 10 min HI? Kippurate Hydrolysis colourless/pale blue dark blue/violet 4 h 24 h 4 h 24 h colourless colourless grey black ESC Esculin β-glycosidaεe pale yellow pale yellow black light grey ZYM A + ZYK B/10 Ein (1) decolour with intense liαht if need be 4 h 24 h 4 h 24 h RIB Ribose Acidulation orange/yellow yellow m L-arabinose Acidulation orange/yellow yellow M Mannitol Acidulation orange/yellow yellow m Sorbitol Acidulation orange/yellow yellow ESC Lactoεe Acidulation orange/yellow yellow Tit Trehalose Acidulation orange/yellow yellow W Inuliπ Acidulation orange/yellow yellow KEF Raffinose Acidulation orange/yellow yellow Starch (2) Acidulation orange/yellow yellow GLYG Glycogen Acidulation light yellow (1) At a second reading after 24 h of incubation, a pre¬ cipitate may be noted in the tubes to which the rea¬ gents ZYM A and ZYM B have been added. This is a nor¬ mal phenomenon, which can be ignored. (2) The acidulation of starch is often weaker than that of other sugars. TEST WITH API 20 STREP VP + HIP + ESC + PYRA tGAL PGUR J3GAL PAL LAP + ADH + RIB + ARA MAN + SOR LAC + TRE + INV RAF AMD GLYG HEM The strain has a weak c-hemolysiε. It grows well at 20-37°C, and comparatively well at 39°C. A suitable bouillon is M17, see recipe. The strain belongs to the Lancefield N group. Making Soured Milk In one embodiment of the invention, the pharmaceu¬ tically acceptable carrier medium for the preparation is soured milk. When implementing this embodiment, the microorganism Lla according to the invention is allowed to grow in milk for 2-20 h, preferably 4 h. Then, the remaining soured- milk culture is added, which results in further growth, but at a reduced temperature of 21°C, if the temperature was higher at the beginning. When the remaining soured-milk culture is added, the content of Lla should be 10 -10 cfu/ml, preferably 8 9 10 -10 cfu/ml. Then, the soured milk is left for 20-24 h, preferably 20 h, whereupon it is cooled and filled into containers. After taking 260 ml of this soured milk in the morn¬ ing and in the evening, the Streptocuccus lactis strain of the invention can be found in the intestine even after 3-5 days.

Claims: CLAIMS 1. A pharmaceutical preparation for controlling pathogenic microorganisms in man and in animals, with a view to preventing and/or treating diarrhoea and other gastrointestinal troubles, c h a r a c t e r i s e d by containing a viable microorganism strain in the form of Streptococcus lactis strain Lla with NCIMB accession num- ber 40157 in at least one pharmaceutically acceptable car¬ rier medium in which the microorganism retains its viabi¬ lity. 2. The pharmaceutical preparation of claim 1, c h a ¬ r a c t e r i s e d in that the microorganism strain con- tent is 10 -10 microorganisms/ml of final preparation ready for use. 3. The pharmaceutical preparation of claim 2, c h a ¬ r a c t e r i s e d in that the microorganism strain con- tent is 10 8-109 microorganisms/ml of final preparation ready for use. 4. The pharmaceutical preparation of claim 1, c h a ¬ r a c t e r i s e d in that the pharmaceutically accept¬ able carrier medium is an acidulated or fermented milk product, such as soured milk, ropy milk, yoghourt, kefir and the like. 5. The pharmaceutical preparation of any one of the preceding claims, c h a r a c t e r i s e d in that the pharmaceutically acceptable carrier medium is a suitable animal feed when the preparation is to be administered to animals. 6. The pharmaceutical preparation of claim 1, c h a ¬ r a c t e r i s e d in that the microorganism strain has been added to the carrier medium in the form of a starter culture, and/or has been added to the final product. 7. The pharmaceutical preparation of any one of the preceding claims, c h a r a c t e r i s e d by being in dry form, e.g. capsules, tablets and powder, together with a pharmaceutically acceptable carrier medium. 8. The use of a pharmaceutical preparation containing a viable Streptococcus lactis strain Lla with NCIMB acces¬ sion number 40157 in at least one pharmaceutically accept¬ able carrier medium in which the microorganism retains its viability, for controlling pathogenic microorganisms caus¬ ing diarrhoea and other gastrointestinal troubles in man and in animals.

Inventors: GRAHN EVA ELISABETH, HOLM STIG EDVIN FOLKE, GRAHN, EVA, ELISABETH, HOLM, STIG, EDVIN, FOLKE
Applicants: GRAHN EVA E, HOLM STIG EDVIN FOLKE, GRAHN, EVA, ELISABETH, HOLM, STIG, EDVIN, FOLKE
Application year: 1990
Main classification: A23K3
IPC codes: A23C9, A23K1, A61K35, A61P1