SU1761809A1 - Method for estimation of residual staphylococcal bacteriocarrierness - Google Patents

Abstract

Usage: medical microbiologists. The essence of the invention: resident staphylococcal bacteriocarrier is determined by seeding the studied material on nutrient media c. subsequent isolation of pure cultures, their identification and testing. The isolated pure cultures are tested for anti-immunoglobulin activity and the resident staphylococcal bacteriocarrier is determined by its value. With an anti-immunoglobulin activity value of 170 mg% and above, the resident bacterial carrier due to Staphylococcus aureus is determined and, at 280 mg% and above, Staphylococcus epidermidis. 2 tab.

Description

The invention relates to medical microbiology and can be used to detect the resident staphylococcal bacteriocarrier in surgical departments and hospitals, maternity hospitals, and in mass screening of the population in environmental monitoring.

Staphylococcus plays a large role in the etiological structure of infections. The main source of purulent staphylococcal infection is resident bacterial carriers. For the prevention of staphylococcal infection, it is very important to timely detect this type of bacterial carriers with subsequent reorganization. In addition, it became known that

the formation of a high level of resident staphylococcus bacteriocarrier occurs as a result of anthropogenic pollution of the environment. Therefore, this indicator can serve as a microbiological test of environmental monitoring.

Considering the above, the obvious need for an effective method for the diagnosis of resident staphylococcal bacteria carrier

A known bacteriological method for the detection of staphylococcal carriers. It requires a large expenditure of nutrient media, laboratory glassware, working time (72 hours), which limits its use in practice.

00

oh oh

ticking, especially when one-time examination of a large number of individuals is necessary. In addition, this method does not allow differentiation of resident (permanent) and transient (temporary) bacteria carriers, whereas the first type represents the greatest epidemiological significance. You need at least a triple bacteriological examination with a time interval in order to establish resident bacteria carriers. Those. the method has a low accuracy of diagnosis of resident staphylococcal bacteria.

The known method of detecting staphylococcal bacteria carriers consists in determining the level of antilysocyme activity in isolated staphylococci. In it, by the presence of antilyzozyme activity in Staphylococcus aureus, and in the epidermal one, when its level exceeds 6 µg, carrier strains are differentiated from contaminating microflora. The method is based on the presence in resident strains of a special property - antilysocyme activity, that is, the ability to inhibit cell lysozyme, which leads to long-term preservation of microorganisms in host cells. This method allows a single examination in the diagnosis of resident staphylococcal bacteriocarrier.

However, the known method has insufficient accuracy in diagnosing resident staphylococcal bacteriocarrier. This method does not reveal resident strains of Staphylococcus aureus, which have an antilysocyme activity of 1-2 µg. In addition, it is difficult to differentiate the strains into resident and transient epidermal staphylococci, which lack an antilysozyme activity of 4 and 5 µg. The consequence is that using this method, part of the resident bacteria carriers are not detected.

The aim of the invention is to improve the accuracy of the method for diagnosing resident staphylococcal bacteriocarrier.

This goal is achieved by the fact that in a known diagnostic method, which includes sowing the material under study on nutrient media with subsequent isolation and identification of pure cultures, anti-immunoglobulin activity is determined from isolated strains of staphylococci, by its presence a resident staphylococcal bacterium carrier is diagnosed. Resident bacteriocarrier is diagnosed if a strain of Staphylococcus aureus has

anti-immunoglobulin activity of 170 mg% and higher, and the epidermal staphylococcus strain 280 mg% and higher.

New features of the claimed method are:

anti-immunoglobulin determined

activity from isolated staphylococcal strains;

in terms of its value, Staphylococcus aureus 170 mg% and above are diagnosed with a resident staphylococcal bacteriocarrier;

according to its size, 280 mg% and more of epidermal staphylococcus are diagnosed with resident staphylococcal bacteriocarrier.

The presence of the first trait in the claimed method allows to reveal a new, more informative feature of the resident staphylococcal bacteriocarrier, which ultimately improves the accuracy of diagnostics of this bacilli carrier in the range of values of the defining indicator, expanding the sensitivity of the method at the lower indicator.

The second new feature allows reducing the sensitivity threshold of the diagnosis of a bacteriocarrier to 170 mg%, as a result of which the accuracy of diagnosis is improved

bacteriocarrier in the presence of Staphylococcus aureus strain, and, practically, resident bacteria carriers among the examined persons are detected.

The third new feature allows reducing the sensitivity threshold of diagnosing a bacterial carrier to 280 mg%, which ultimately improves the accuracy of diagnosing a bacterial carrier in the presence of an epidermal staphylococcal strain and

practically, identify all the carriers of this class from the examined persons.

These features have not been found in the prior art.

A statistically significant relationship was found between the ability of staphylococci to persist for a long time in a macroorganism and its property to degrade immunoglobulins in biological fluids.

Application of the claimed features

involves the determination of the anti-immunoglobulin activity of staphylococci, which is a test of the persistent characteristics of the strain and allows differentiation of the resident and transient bacteriocarrier.

Therefore, a technical solution meets the criterion of significant differences, and the combination of its known and distinctive features provides a new positive effect - an increase in the accuracy of the method. The proposal of this method became possible after the authors discovered using the experimentally developed anti-immunoglobulin activity of staphylococci, i.e. their ability to neutralize immunoglobulins of various classes in biological fluids. It has been suggested that this property may play a role in the persistent ability of a microorganism, where the anti-immunoglobulin activity is a factor that purposefully neutralizes the protective mechanisms of the microorganism. This was the basis for the determination of anti-immunoglobulin activity along with other characteristics, in particular, the anti-isozymic activity of staphylococcal strains isolated from 560 examined individuals in order to detect bacterial carriers. Analysis of the data obtained revealed a direct relationship between the values of the anti-lysozyme and anti-immunoglobulin G and M activities. Moreover, such dependence was most clearly traced in relation to the anti-immunoglobulin G activity of the microbe (Table 1).

You dreamed that staphylococcal strains with high antilysocyme activity, and, therefore, persistent properties, also have high anti-immunoglobulin activity. But at the same time, high anti-immunoglobulin activity was also found in some cases, when the epithelial staphylococcus strains had 4 μg or 5 μg of antimysozyme activity, and 0 μg or 2 μg in Staphylococcus aureus. It was decided to conduct a threefold bacteriological examination of a group of people who had epidermal staphylococci with an antilysocymic activity of 4.5.6 μg and golden staphylococci with an antilysocymic activity of 0, 2, 3 μg from the nasal mucosa.

A threefold examination showed that strains of resident bacteria carriers may have different antilysocyme activity - 7, 6, 5, less often 4 μg - epidermal staphylococcus, 3, 2, 0 μg - golden staphylococcus (Table 2). That is, strains of epidermal staphylococcus, having 4 and 5 kg, and Golden - 0; 2 mcg

can not be uniquely identified as transient.

The anti-immunoglobulin activity in this regard turned out to be a more informative indicator, since the strains of the epidermal staphylococcus isolated from bacteria carriers of the resident type had anti-immunoglobulin activity from 289.24 ± 5.7 mg% to 352.41 ± 11.4

mg%, golden - from 178.28 ± 6.4 to

190,31,7,5 mg%. Transient strains

epidermal staphylococcus had an anti-immunoglobulin activity of 112.86 ±

± 6.8-198.31 4.7 mg%, and transient

strains of Staphylococcus aureus - 98.64 ± 9.1-148.41 ± 5.8 mg%.

It is concluded that anti-immunoglobulin activity is a more informative indicator of persistent

microbial properties, rather than antilysocyme, and can be used as a diagnostic test for resident bacteria. This was the basis of the claimed method.

The method is carried out as follows. In cultures examined from the nasal mucosa, pure cultures of epidermal golden staphylococci were isolated by culture.

by the method with the determination of common properties - plasma coagulase, hemolysin, lecitelase, etc. At the same time, anti-immunoglobulin activity is determined in these strains. For this take daily

a suspension of staphylococci, standardized on FEC-M (D-0.50, cuvette M 2, green optical filter). It was this suspension density that was experimentally selected as revealing the maximum manifestation of the anti-immunoglobulin activity of the microbe. 0.1 mg of this suspension is introduced into a test tube, to which 0.1 ml of serum is added with a known amount of Ig, determined according to the Mancini method. Test tube with mixture

put in a thermostat for incubation at 37 ° C for 30 minutes. After the time has elapsed, the mixture is centrifuged at 6000 rpm for 10 minutes and the supernatant is used to re-determine the level of immunoglobulins taking into account preliminary dilution 2 times. Then, the difference between the levels of immunoglobulins in the control and experimental serum samples is calculated and judged by it.

the anti-immunoglobulin activity of the microbe. If the level of the anti-immunoglobulin activity of Staphylococcus aureus is 170 mg% and higher, and that of the epidermal staphylococcus is 280 mg% and higher,

they consider these strains to be resident and diagnose resident staphylococcal bacteriocarrier,

PRI me R 1. During the examination for staphylococcal bacteriocarrier in a patient P.AD. Epidermal staphylococcus was isolated, having ALA - 5 µg, A IgG 283.4 mg%. It was assumed resident bacterium. Multiple repeated inoculation of identical staphylococcus confirmed the resident nature of the bacteriocarrier. At the same time, the titer of bacterial contamination increased.

Example 2. Patient M.S.K. isolated epidermal staphylococcus had the following characteristics of ALA - 5 µg, A IgG

-156.2 mg%.

Re-sowing did not give a similar result.

Example 3. The patient D.I.I. Staphylococcus aureus was isolated - ALA - 0 µg, A IgG

-171.2 mg%.

The assumption of the resident bacterium was confirmed by triple seeding of identical staphylococci (phage type 3 C). The titer of bacterial contamination increased to 10 -10 CFU / tampon,

Example 4. The patient K.AND.O. The isolated strain of Staphylococcus aureus possessed ALA-2 µg, A IgG - 121.3 mg%. When repeated seeding, identical staphylococci were not detected, the denial of transient bacteriocarrier, as was suggested by the test A IgG.

Comparing the accuracy of diagnostics of the proposed method and the prototype we take for 100% the number of resident bacteria carriers detected by a threefold bacteriological method upon examination of 560 people.

A threefold bacteriological examination revealed 198 resident bacteria carriers — 100%. Using the prototype method, 124 bacteria carriers were detected - 63.2%, and the claimed method was 195 people. or 98.4%, which is 35.2% more than in the way of the prototype.

Consequently, the claimed method allows to increase the accuracy of the method for diagnosing resident bacterial carriers by 35.2%, which allows timely and targeted rehabilitation of bacterial carriers. In addition, the inventive method has additional advantages — it is simpler than the prototype method. Simplification is manifested in the fact that

anti-immunoglobulin activity of different levels can be recorded on a single Petri dish, i.e. simultaneously investigate up to 45 strains with minimal material and labor costs. The determination of antilysocyme activity according to the method of the prototype is much more cumbersome, since it involves the use of several Petri dishes with nutrient egar, including different concentrations of lysozyme, in order to detect various levels of antilyzozyme activity of the strains studied.

Thus, the use of the method will improve the accuracy of diagnosis

resident staphylococcal bacteria carrier, and, consequently, to achieve a positive effect.

Claims (2)

1. The method of determining the resident staphylococcal bacteriocarrier by sowing the test material on nutrient media, followed by isolation of pure cultures, their identification and testing, characterized in that, in order to increase the accuracy of the method, the isolated pure cultures are tested for anti-immunoglobulin activity and their resident staphylococcal bacteriocarrier is determined by its value. 2. The method according to claim 1, characterized in that with an anti-immunoglobulin activity value of 170 mg% and above, the resident bacterial carrier is determined, caused by Staphylococcus aureus, and at 280 mg% and above - Staphylococcus epidermidis. Table 1 table 2