МЕТОДИЧЕСКИЕ АСПЕКТЫ ИСПОЛЬЗОВАНИЯ СОЛЯНОКИСЛЫХ СПЕРМИНА И СПЕРМИДИНА ДЛЯ ИДЕНТИФИКАЦИИ УРОПАТОГЕННОЙ МИКРОФЛОРЫ

Бойко О.В.¹, Николаев А.А.², Бойко В.И.³_, Доценко Ю.И⁴

¹Доктор медицинских наук, ²доктор медицинских наук, профессор, ГБОУ ВПО «Астраханский государственный медицинский университет», ³доктор медицинских наук, профессор ФГБОУ ВПО «Астраханский государственный университет», ⁴кандидат медицинских наук, доцент ФБГОУ  ВПО «Российская академия народного хозяйства и государственной службы»

Работа выполнена при поддержке гранта РГНФ № 13-06-00008

МЕТОДИЧЕСКИЕ АСПЕКТЫ ИСПОЛЬЗОВАНИЯ СОЛЯНОКИСЛЫХ СПЕРМИНА И СПЕРМИДИНА ДЛЯ ИДЕНТИФИКАЦИИ УРОПАТОГЕННОЙ МИКРОФЛОРЫ

Аннотация

В статье предложен новый метод дифференциация этиологически значимых микроорганизмов от представителей нормальной микрофлоры или контаминантов на основании устойчивости выделенных из спермы бактерий к полиаминам. Метод отличается простотой исполнения и хорошей воспроизводимостью, а также высокой достоверностью получаемых результатов.

Ключевые слова: полиамины, спермин, спермидин, бактерионосительство, сперма

Boiko O.V.¹, Nikolaev A.A.², Boiko V.I.³ , Dotsenko Y.U.⁴

¹ MD, ² MD, full professor, State Budgetary Educational Institution “Astrakhan State Medical University”, ³ MD, full professor, Federal State Budgetary Educational Institution “Astrakhan State University”,  ⁴Ph D, Federal State Budgetary Educational Institution “Russian Academy of National Economy and Public Administration”

The investigation was carried out under supporting the grant of the Russian state scientific fund 13-06-00008. 

METHODICAL ASPECTS OF HYDROCHLORIC SPERMINE AND SPERMIDINE USING FOR THE PROCESS OF UROPATHOGENIC MICROFLORA IDENTIFICATION

Abstract

According to stability of bacteria extracted from sperm to polyamines we offer new method of differentiation of etiological importance microorganisms from normal microflora representatives or contaminants. The method is unique due to its simplicity and high trustworthiness of its results.

Key words: polyamines, spermine, spermidine, bacteria carrier, sperm

One of integral biochemical systems playing an important role in protective reactions of an organism and sperm, is collection of polyamines (spermine, spermidine, putrescine).

Levenguck began to investigate representatives of polyamines more than 300 years ago. In 1677 he described crystalline substance extracted from sperm which probably was sperminphosphate. Later many scientists observed crystals which were generated in  slow drying up of fresh human and animal sperm if it was in stable condition during an hour; these crystals were like acute-angled prisms, often curved with prominent surfaces, however the nature of these crystals was obscure. In 1878 Schreiner could extract the base of these crystals – spermine though not in pure condition. The name “spermine” was offered by Ladenburg and Abel in 1888.

Previously it was erroneously supposed that spermine could be found only in prostate ant this substance was peculiar to only a man organism. But the Russian scientist professor A. Poehl first discovered that spermine was not exceptional constitutive part of man’s prostate but in testicles, ovaries, pancreas, spleen, thyroid gland and in blood. He created preparation “Sperminum-Poehl” which was widely used at the end of the XIX^th – the beginning of the XX^th century for treatment of different diseases.  In the 50^th of the XX^th century it was ascertained that Poehl sperminm was not hormonal preparation and after that its production was stopped.

During many years after investigation of A. Poehl polyamines did not attract many scientists. Only descriptions of some polyamines periodically appeared; these polyamines were named according to the subjects they were extracted from: putrescine from products of putrefaction, kadaverine from ptomaine, spermidine from animal sperm. Chemical structure of spermine was interpreted by Rosenheim only in 1924-1926. Besides he synthesized spermine and on its base - spermidine.

We may say that polyamines were discovered in all alive organisms (in animals, plants, water plants, mushrooms, bacteria, viruses). In tissues and biological liquids of animals spermine and spermidine may be found, but these liquids contain only small amount of putrescine. Eeukaryotes tissues contain spermine and spermidine in millimolar concentrations and in much less amount they contain putrescine (nanomole). Concentration of putrescine of procariots is higher than of spermidine but most bacteria have not spermine at all.

There are not a lot of scientific papers about polyamines influence to the biochemistry of bacterial sell. Information of the most of them comes to the following: the growth of some kinds of bacteria is obligatory or facultative connected with polyamines presence. Higher concentrations of these biogenic poly kations depress blood pressure due to forming their active metabolite - aldehyd polyamines. Oxidized derivatives of polyamines (iminoaldehydes) take antibacterial, antivirus and antitumour effect oppressing the albumin and nucleic acids synthesis.

For polyamine antibacterial effect determination during our investigations we use the following kinds of microorganisms: strains of E. coli, K. pneumoniae, S. aureus and S. epidermidis extracted from sperm of patients with chronic prostate disease. As control group it was microbes from feces and substance extracted from umbilical wound of children and from pharynx mucous membrane of patients with chronic relapsing infection of higher respiratory ways.

This choice is explained by this fact that representatives of man autoflora are main pathogens of men urogenital canal infections influencing to the development of patosperm. Because of relatively pathogenic flora representatives dominate in the structure of microorganisms extracted from healthy men ejaculate and sterile men , it is ejaculate evident differentiation of etiology important flora and representatives of normal microflora or contaminants is based on revealing of qualitative features which are characterized for pathogenic microorganisms.

During the experiment it was ascertained that microorganisms had different level of survival after the contact with concentration 0,04 mg/ml of hydrochloric spermine solution or hydrochloric spermidine solution. First of all it was ascertained that spermine had higher antimicrobial activity than spermidine. For example number of atrains with more than 90,0% alive cells after the contact with solution of spermine was 40,0% and with solution of spermidine – 64,0% (p<0,05).

In 72 experiments of 100 during the test of surviving of the same strains the number of alive microorganisms was less after the contact with spermine and only in 12 tests after the contact with spermidine. And only 16 cultures had identical stability to the both polyamines.

Specific stability to spermine and spermidine are given in the table 1.

Tab. 1 – Specific stability of microorganisms to polyamines (M±m)

Information given in this table testifies to mainly bactericidal effect of spermine irrespective of extracted microorganism specific belonging. We did not reveal principal differences between different bacteria. However more convincing results were received during our attempt to reveal the level of bacteria sensitiveness to polyamines depending on their habitance: only 25% bacteria extracted from sperm were sensitive to polyamines (only 25% of them were sensitive only to spermidine, 25% - only to spermine and the rest were sensitive to the both polyamines). About 75% bacteria extracted from urogenital canal of men kept their viability but microorganisms extracted from the rest ecological niche were sensitive to polyamines (84,2% from them to spermidine and 89,9% - to spermine). So, we affirm (trustworthiness p<0,01) that different stability of bacteria stability to polyamines is to a considerable extent caused by their habitance (r=0,873).

Probably it is connected with the fact that during the process of evolution pathogenic microorganisms adapted to the growth in different tissues of a host. We suppose that high level of tissue specific features which is inherent in many microbes reflects difference between tissues in the respect of biochemical medium composition especially when there are not evident differences in protective mechanisms.   We suppose this direction of investigations is perspective because recommended criterion is correspond to two demands at least: to characterize an extracted microorganism pathogenic potential and to reflect the belonging of an extracted bacteria to the certain ecological niche. It is expediently to carry out such an investigation for deciding of vexed questions to ascertain the etiological agent when chronic difficult amenable treatment disease of urogenital system. According to information of the  Patent of Rassun Federation # 2251691 «The method of diagnosis of pathogenic of microorganisms Staphylococcus epidermidis extracted from sperm» was taken out.

References

1. Boiko O.V., Terentiev A.A., Nikolaev A.A. Molecular mechanisms of persisting infection. – 2006 - Astrakhan: Publishing house “Astrakhan university”. - P. 59-61