WO2016083990A1

WO2016083990A1 - Method of sanitizing microbiological environment

Info

Publication number: WO2016083990A1
Application number: PCT/IB2015/059070
Authority: WO
Inventors: Marek JAMROZY
Original assignee: Jamrozy Marek
Priority date: 2014-11-25
Filing date: 2015-11-24
Publication date: 2016-06-02
Also published as: PL410247A1

Abstract

The invention presents method of sanitization of microbiological environment surrounding people and animals. It relies on modification of environment by identification one or more pathogenes, choosing the way of biocontrol through previously selected bacteria strains, fungi or other microorganisms, able to produce own metabolites. Their action is checked and then, they are introduced to environment.

Description

Method of sanitization of microbiological environment

The main idea of invention is a method of sanitization of microbiological environment, which can be accomplished by modification of microbiologic environment. This aim is achieved by application of specially selected bacteria, fungi or other microorganisms, which are able to produce their own metabolites. This way enables to reduce or completely eliminates diseases caused by microorganisms found in human and animal habitats.

It is common fact that disease - causing organisms like bacteria, fungi, viruses and other are occurred in microbiological environment of people and animals. They are very important in livestock production and influence on farmers' profit. It is estimated that losses connected with single treatment of mastitis amount 1000 - 1200 PLN. This sum could be multiplied in case of repeated action. Moreover, sick cows might be scrapped of herds. Other diseases, mostly found among young animals, do not allow them to reach the highest possible potential of production.

There are some available methods, which are intended to decrease level of microorganisms in microbiological environment. Each of them has advantages and drawbacks depended on different factors. However, the best way of prevention is contamination avoidance. Therefore, sanitization is defined as the reduction of pathogenic organisms number to the level at which they do not pose a disease threat to their hosts and without any negative impact on products produced for customer. It is very crucial that sanitization has to be neutral for macro - environment.

As standard, two main methods of sanitization are used: chemical (detergents, chlorine, hypochlorite, ozone) and physical (rinsing, temperature, high pressure, electric and magnetic field, ultrasounds or UV radiation). Both methods give short-term and selective effect to proper modification of microbiological environment. Frequently, after their application in microenvironment, appears other adverse pathogenic microorganisms like fungi, causing additional health problems. Furthermore, wide range of antibiotics used in treatment results in increasing resistance of disease- causing organisms, and finally, encourage researchers to find new ways of reaction.

In sanitation process, some means of biocontrol are applied. Presented invention is concerned in these categories. Currently, bacteriophages are used as a mean of biocontrol in sanitization. American patent description US2004191224 reveals sanitization method which relies on microbiological modification by spraying previously cleaned surface with large droplet washing containing one or more bacteriophages, as a mean of biocontrol. This way of action is expensive and trouble-some due to high costs of bacteriophage's multiplication. Much more effective and easier in usage in industrial scale is presented invention, which assumes using bacteria or their metabolites as a method of biocontrol. Bacteria are multiplied easily at and at little cost.

During sanitization process, the natural abilities of bacteriaphages are used. They infect bacteria and cause cell lysis. It is essential to identify an etiologic factor of infection before induction of bacteriophage and choose the suitable strain of bacteriophage characterized with the highest lysis potential towards specified bacterium strain. Typically, phages affect on one kind of bacterium, so they are multiplied only within one species of bacterium or its strain. However, sanitizers containing bacteria and fungi are able to act on wider spectrum disease-causing bacteria and fungi. Treatment with highly specified phages can be troublesome owing to the fact of necessity of extraction infectious bacterium. On the contrary, antibiotic microorganisms, presented in invention, possess wide range of activity towards harmful microbes. Phages application also requires taking into account that phages encode genes of toxicity, as a consequence, transform harmless bacteria into pathogens. Despite of the fact that the newest research enable to isolate toxic genes, still all the virulence factors are not well known. The solution is lack of using lysogenic phages or precisely known phages with vector ones incapable to replication.

Usage of phages also needs consideration of negative lizogenia process, relies on bacteria immunization on phages. In case of acquired resistance, lysogenic cycle in bacterium is not proceeded, and virus is halted to pre-phage stadium. This process is frequently connected with bacterium adjustment and leads to higher virulence. Thus, lizogenia is undoubtedly dangerous side reaction and current scientific knowledge does not present the way of prevention. Phages are also characterized by expanding only in the presence of targeted species. When targeted species disappear from environment, their phages as well die out. Reappearance in environment pathogenic microbes requires introducing specific bacteriophages. It is very difficult to control and catch a moment when specified bacteriophages have to be necessarily introduced. If it is failed, and the infection symptoms occur again, the results for humans and animals could be dangerous. The other point is some number of disease asymptomatic carriers. For instance, it has been found that amount of carriers Staphylococcus aureus in human population is estimated from 10 to 40%. Thus, for protective existence of phage, living in fought bacterium environment is needed. Therefore, in case of host disappearing, bacteriophage does not have successful effectiveness and ensure reduction of microbial threat.

The method according to invention refers to other kind of microorganisms: bacteria, fungi and other microorganisms, which are able to form their own metabolites. Bacteria have wider spectrum of antibiotic properties towards other pathogenic species. As a pattern, strains of bacteria used in presented invention, successfully fight listed species: Pseudomonas aeruginosa, Salmonella typhimurium, Staphylococcus aureus, Staphylococcus epidermidis, Klebsiella pneumoniae, Sarcina lutea, Streptococcus pneumonia, Streptococcus agalactiae, Streptococcus faecalis, Streptococcus mutant, Escherichia coli, Candida albicans, what was proved in research which was checked towards bacteria strains mentioned before. In case of some pathogenic microbes remarked before, efficacy equals 100% and it is followed by completely elimination of pathogens. Such effectiveness is gained towards for example: E.coli, S. aureus, Candidia sp., S. agalactie and A. baumann. The range of efficient activity, presented in invention, is wider than in commonly known action of phages. It is worth emphasizing, the preventing action of existing in environment bacteria, even in spite of lack of pathogens. Antibiotic bacteria still remain in environment, ready to action in case of renew appearing pathogens. The invention gives an opportunity to enter into environment bacteria metabolites working against pathogens, instead of pure bacteria. In that moment, the protection is active only when there is sufficient amount of antibiotic compounds.

Presented invention assumes, sanitization of microbiological environment with selected and specially prepared microorganisms or their metabolites by modification of environment, using antibiotic properties of them. As a result, pathogens are reduced or completely removed from environment through application of single or mixture strains of bacteria or their metabolites.

During developing of invention and examining various microorganisms: bacteria, fungi, actinomycetes and other extracted from natural environment, their mutual reactions were analyzed and their influence on commonly found pathogens among humans and animals. In this research, influence on such a common pathogens: Escherichia coli, Staphylococcus aureus, streptococcus and salmonella were tested. Studies were made on referencial pathogenic species. They proved that possessed strains are highly effective towards widespread pathogens, encountered in animals and humans.

At the same time, previously mentioned strains are tended to show tolerance for common microorganisms found in environment and remain neutral to health of people and animals. This fact means that bacteria strains work against harmful microbes and simultaneously modify microbial environment into health desired direction. Thus, the sanitation effect is gained by reduction or complete elimination of disease causing pathogenes. As a result of this elimination, the number of diseases caused by microbes living in environment is reduced.

In each environment, surrounding people and animals, has very specific microflora and different from other. In some cases, the source of infections are well known and kinds of microorganisms responsible for certain diseases. Such a situation occurs in previously remarked mastitis, found in dairy cattle. Other diseases commonly happened among livestock are also examined and their reasons, too. In animal surround, still lots of health neutral microorganisms exist and do not let other harmful ones to multiply: for instance fungi. Excluding pathogens, other microbial organisms maintain in stable balance. So far, the methods of complete environment modification in farm buildings, relied on replacing pathogenic organisms with their natural foes (neutral for animals), have not been known.

The subject of invention is method of sanitization of microbiological environment of people and animals. It is achieved by modification of microbiological environment, in which at first, one or more pathogenic factors are identified, then suitable mean of biocontrol is adjusted by using specially selected microorganisms that are capable to produce their own metabolites, and finally selected microorganisms are introduced into environment. So, the primary feature of invention is opportunity to introduce metabolites instead of microorganisms. In that moment, protection acts, only if there is sufficient amount of antibiotic compounds in environment. Preferable and additional advantage of presented invention is an opportunity of introducing mean of biocontrol in different forms. It is implemented by: large or small droplet sprays, cold misting or powder and it covers surfaces giving the best contact with fought strains.

Identification of pathogenic factor is primarily based on well known pathogenic agents. If standard treatment does not give positive effect, more accurate analysis and identification of infectious microbes may be necessary and investigation of antibiotic effectiveness strains which fight with disease in particular environment.

In other cases, when infectious factors are still unknown, further research is needed. In order to find the source of infection, matching of antibiotic strains with identified pathogens is gradually checked.

The number of antibiotic strains (density of vaccine) proceeds by checking of antibiotic effectiveness bacteria used for sanitization of environment. The number of cells which are fought, and the number of antibiotic cells, are checked on petrie dishes. As a result, needful measure of antibiotic bacteria is determined and introduced to environment.

The other invention quality is possibility to apply it in whole farm buildings and their side area. The moment of application the mean of biocontrol is adjusted to successful effect of modification. Microorganisms used to fight against pathogens belong to low-risk microorganisms. Action of invention is submitted in pattern of mastitis among milking cows. In most important cases, infectious problems caused by environmental microflora are examined and pathogenic organisms are known. However, in every even well-maintained herd, some difficulties may occur. The pathogenic microbes, mostly responsible (90%) for mastitis, are: Ataphylococcus aureus, Streptococcus agalactiae, Streptococcus dysgalactiae and Staphylococcus uberis. The other reasons of mastitis are: Staphylococcus epidermis, Clostriduym perfmgens, Corynebacterium pygogenes, Escherichia coli, Klebsiella spp, Candida albicans or Mycoplasma spp.

Knowing this knowledge, some selected species of microorganisms are matched i.e. Ochrobacterum, Paenibacillus, Streptomyces, Humicola and others in single strain form or mixture of strains. Then, the action of selected bacterium strains are checked on petrie dishes. That kind of verification also tests the lack of negative influence on humans and animals health, which is vital condition of introducing to environment. This verification is gradually conducted, beginning from simple organisms to vertebratas. Checking antibiotic activity of chosen strains is proceeded not only towards pathogens, occurred in contaminated buildings, but also on referential strains. This kind of procedure guarantees high effectiveness of pathogens overpowering.

Knowing the number of fought pathogens and fighting against them antibiotic bacteria (checked in petrie dishes), the amount of vaccine is calculated and introduced to microbiological environment through small droplets spray.

If applied treatment does not give satisfactory effect more detailed identification of pathogenic microbes existing in certain environment is needed.

The presented invention is also concerned to other diseases and their pathogens: infectious atrophic rhinitis Pasteurella multocida an d Bordetella bronchiseptica), streptococci s,

Glasser's disease {Haemophilus parasuis), diseases caused by anaerobic bacteria(C I os iridium perfringes) , c o c ci di o s i s , exudative dermatitis {Staphylococcus hyicus), colibacilosis, salmonellosis, hyperplasia (Lawsonia (L) intracellular is) , dysentery Serpulina), MMA, paratuberculosis, and for poultry diseases like salmonellosis, streptococci s, colibacilosis or diseases caused by anaerobic bacteria.

Claims

Method of sanitization of microbiological environment surrounding people or animals which relies on modification of environment by identification of one or more pathogenes, choosing the way of biocontrol mean, which action is checked and then, the mean of biocontrol is introduced to environment characterized in that the mean of biocontrol, introduced in certain quantity, consists of selected strains of bacteria, fungi and other microorganisms which are able to produce their own metabolites.

Method according to claim 1, characterized in that introduction of biocontrol mean relies on small or large droplet, cold misting or introduction by powder.

Method according to claim 1 or 2, characterized in that identification of pathogenic factor is based on identification pathogens in definite environment.

4. Method according to claim 1 or 2, characterized in that after identification of pathogenic factor, antibiotic effectiveness is checked.