RU2716593C1 - Method for rapid diagnosis of inflammatory pharyngeal diseases using fluorescent spectrometry and special algorithm for assessing morphometric, metabolic and functional changes in tissues in dynamics - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to otorhinolaryngology, and can be used for diagnosing chronic pharyngeal diseases, such as compensated and decompensated forms of chronic tonsillitis, and chronic pharyngitis. For this purpose, spectral curves are recorded using fluorescence spectrometry from upper pole surface, lower pole, lacunae of palatine tonsils, as well as from posterior pharyngeal wall. That is followed by calculating absolute fluorescence intensity, spectral half-width, normalized fluorescence with respect to the intact point. Thereafter, the obtained values are compared with spectral indices in healthy volunteers. Based on the obtained data, the patient is diagnosed with a compensated or decompensated form of chronic tonsillitis or chronic pharyngitis.

EFFECT: invention provides higher accuracy and sensitivity of non-invasive diagnosis of chronic pharyngeal diseases in a patient.

1 cl, 6 tbl, 30 dwg

Description

Technical field

The invention relates to medicine: to otorhinolaryngology, and can be used for express diagnostics of inflammatory diseases of the pharynx using fluorescence spectrometry and a special algorithm using index parameters to evaluate basic morphometrics. metabolic ”functional features of tissue in dynamics and in situ, both in vivo and in vitro.

The method of express diagnostics using fluorescence spectrometry and a special algorithm involves the use of laser radiation of various wavelengths (405 nm, 532 nm) to record the metabolic, morphometric and functional parameters of pharyngeal tissues in normal and inflammatory diseases in dynamics. The effect of fluorescence can be used to increase the efficiency of the diagnosis of inflammatory diseases of the pharynx; to conduct accelerated identification of the form of inflammatory diseases of the pharynx.

The method is aimed at conducting highly sensitive, non-invasive rapid diagnosis of the inflammatory process in the pharynx in situ in dynamics, both in vitro and in vivo.

The method according to the invention allows the dynamics to evaluate indices that illustrate the state of the tissue in a given period of time and allow with high accuracy and sensitivity (close to 100%) to characterize the physiological changes that occur in the tissue during inflammation and to register its structural features, as well as the presence of certain metabolites in it for the purpose of immediate (quick) diagnosis and early identification of the pathological process (for preventive measures, using this method as a screening and monitoring of treatment, the most rational choice of its form, the correction treatment in different stages of a pathological process).

To date, there are many ways to diagnose inflammatory diseases of the pharynx. Their main disadvantages are relatively low sensitivity, high duration, complexity in the technique, inadequacy at the time of treatment (inconsistency of the therapy with the stage of the inflammatory process), inability to use it in situ, in vivo and in vitro at the same time and evaluate what is happening in tissue processes in dynamics in express mode.

State of the art

A known method for the diagnosis of various forms of chronic tonsillitis (Brudastov Yu.A., Gonchar-Zaikin A.P., Dyukov L.A., Ryazanov V.D., Shulga I.A., 2009, patent No. 2429477). The essence of this method is that the patient determines a set of indicators of intestinal microbiocenosis: the content of bifidobacteria, lactobacilli, Escherichia coli with normal properties, Escherichia coli with hemolytic properties, Escherichia coli with lactose-negative properties, enterococci, saprophytic staphylococci, Staphylococcus aureus, Candida staphylococcus aureus, comparing them with indicators of the relative norm on a point scale. The dysbiosis index is calculated using a special formula. The dysbiosis index judges the form of chronic tonsillitis: compensated or decompensated.

The disadvantage of this method is that the form of chronic tonsillitis can be judged indirectly, based on bowel function. In addition, the method is quite time-consuming to perform and lengthy.

A known method for differential diagnosis of the clinical form of chronic tonsillitis, based on ultrasound examination of regional lymph nodes (Rychkova IV, RU 2638429), which is used for differential diagnosis of toxic-allergic and simple forms of chronic tonsillitis using ultrasound imaging of regional upper lateral cervical lymph nodes . The clinical form of chronic tonsillitis is determined by the size and number of lymph nodes, however, with all the advantages of the method, the main disadvantage is the possibility of error in the presence of other concomitant diseases that cause an increase in the size and number of lymph nodes, that is, additional examinations of the patient are required to clarify the diagnosis; in addition, pharyngeal tissue studies are not carried out directly in vivo with an assessment of all parameters at a morphometric, metabolic, and functional level.

Also known is a method for the diagnosis of chronic tonsillitis (patent 2004134651/14, Livenets V.P., 2006). The method consists in registering the active resistance and the electrical parameter from the surface of the tonsils, which are used to diagnose the form of chronic tonsillitis. The method allows to study the functional state of tonsil tissue. The main disadvantage is the difficulty in performing the technique; also the sensitivity and specificity of this technique are significantly inferior to those when using the method of fluorescence spectrometry.

A known method for the diagnosis of chronic tonsillitis, based on measuring changes in the impedance of the vascular system in the region of the tonsils (rheotonsillography) (RU 2014103 C1 from 06.15.94). In this case, the electrodes are applied to the skin in the maxillary region in the projection of the tonsils and to the hard palate along the midline, a rheogram is recorded and the severity of chronic tonsillitis and treatment tactics are determined by the value of the rheographic index. The disadvantages of this method is the indirect measurement of impedance, which can be influenced by pulsation from nearby large vessels. The use of one parameter characteristic of functional disorders in the tonsils with chronic tonsillitis, namely, a decrease in the blood supply to the tonsils, may indirectly indicate a violation of the function of the tonsils partially or completely.

The location of the electrodes directly on the tonsils during rheography was not widespread due to the complexity of the process and the need for their long fixation, necessary for recording rheograms.

A known method of measuring the electrical parameter directly of the mucous membrane of the tonsils using silver-plated electrodes. In this case, a study is carried out of the functional state of tonsil tissue with the goal of a qualitative assessment of it.

An essential property of this method is that a passive electrode is placed under the tongue along the midline, and the active one alternately on the surface of the right and left palatine tonsils and the electrical parameter is measured. With this technique, the active resistance is studied when the passive electrode is located under the tongue along the midline, and the active one alternately on the surface of the right, then left palatine tonsils. In this case, the influence of changes on the skin and in the neurovascular bundle, passing in the immediate vicinity of the tonsils, is eliminated. Also, the impedance is determined at frequencies of 1 kHz and 100 kHz. The resistance values are determined and displayed on a digital display. Electrodes made of silver-plated brass are used and a recording unit determining active biological resistance is used. The disadvantages described in the literature include the small sensitivity and specificity of the method, the impossibility of using it in patients with changes in blood rheology (O.Y. Plepis. Geography in otorhinolaryngology, - L .: Medicine, 1988. P. 50-62; manual on otorhinolaryngology. // Under the editorship of IB Soldatov, Moscow: Medicine, 1994. 574 p .; Patent RU 2014103 C1 dated 06.15.94. Method for the diagnosis of chronic tonsillitis).

An analogue of the proposed new method is also a method for diagnosing compensated and decompensated forms of chronic tonsillitis based on the method of infrared spectrometry of saliva (Portenko G.M. et al., 2005, patent No. 2261048). This method is based on the application of the probabilistic-logical training algorithm of the binary tree of CART decisions according to the indicators of the infrared spectrum of saliva and relates to the express methods of identification and differential diagnosis of forms of chronic tonsillitis, in combination with a probabilistic neural network according to the energy-information indicator of saliva samples. However, the method is indirect in the differential diagnosis of forms of chronic tonsillitis, since the study of biological saliva-fluid is carried out, the biochemical changes occurring in the body can thus be judged only indirectly, that is, in situ diagnosis of a biological object, in this case, tonsils and other structures of the pharynx, in this work it is not provided that it can not carry complete information about the studied object.

The closest prototype of the proposed new method is a method for monitoring the treatment of a disease, including fluorescence diagnosis of the disease, and a device for its implementation (Boriskova E.Yu., 2014, patent RU 2511262).

The invention relates to the field of fluorescence analysis, namely, modifications of optoelectronic equipment and methods for studying biological objects with its help, and can be used to identify multicomponent systems, in particular, to diagnose a disease and monitor its treatment based on changes in the fluorescence response of biological fluids.

The main disadvantage of the method compared to the proposed one is the possibility of carrying out it only in vitro, as well as insufficient accuracy, due to the incomplete analysis of all informative parameters in numerical terms, allowing to evaluate all the constituent tissues of the pharynx (metabolic, morphometric, functional) at various stages of the inflammatory process and its various types.

All of the above methods have significant useful properties and advantages, however, the main disadvantage of these methods is the complexity of the execution, duration, relatively low sensitivity and specificity, as well as the inability to use the method simultaneously - in vivo, in vitro and in situ (inside the body, organ and “In place”), non-invasively and in express mode, in dynamics evaluating all the main parameters of the pharyngeal tissue (metabolic, morphometric, functional).

SUMMARY OF THE INVENTION

The proposed method of fluorescence spectrometry using a special algorithm and indices is a highly sensitive and accurate diagnostic method, since it allows you to record minimal changes in physiology, morphometric and metabolic parameters non-invasively and quickly (within a few seconds). The main difficulty is the need for a clinical interpretation of the results using this technology. In addition, this method is the fastest compared to other methods. The inventive method relates to methods of rapid diagnostics using the fluorescent component and a special diagnostic algorithm using indices to evaluate the morphometric, metabolic, functional changes that occur in the tissues of the pharynx normal and when they are involved in the inflammatory process with chronic pharyngitis and chronic tonsillitis, which allows you to register the main characteristics of the studied points for the purpose of differential diagnosis.

Thus, the method of fluorescence spectrometry in the diagnosis of forms of inflammatory diseases of the pharynx using a special algorithm and indices is accurate, non-invasive, highly sensitive, allowing in situ dynamics, both in vivo and in vitro, to expressly evaluate the nature of the inflammatory process in diseases pharynx, each of which is characterized by a certain set of metabolic, structural, as well as functional changes, which allows you to use the method to differentiate the type of pathological th (inflammatory) process.

The method of fluorescence spectrometry using a special algorithm and indices is implemented based on the use of hardware-software complexes with laser wavelengths of 405 nm and 532 nm and software that contains these indicators (indices) that allow differential diagnosis of inflammatory diseases of the pharynx in express mode. The calculation of indices is described in detail below in the description. In addition, using the in vitro technique, the spectral characteristics of microorganisms and / or their associates, their concentration in inflammatory diseases of the pharynx can be recorded, which is an additional factor indicating the possibility of using this method in the diagnosis of the inflammatory process in the pharynx, since it is known that with various forms of inflammatory diseases of the pharynx there is a different microflora. Thus, in addition to the available spectral characteristics of tissues in inflammatory diseases of the pharynx, one can determine the type of metabolism: aerobic or anaerobic; type of pathogen and its concentration for prescribing rational antibacterial therapy in the early stages.

A method for the rapid diagnosis of inflammatory diseases of the pharynx using fluorescence spectrometry and a special algorithm for assessing morphometric, metabolic, functional changes in tissues in dynamics

The problem solved by the invention is the possibility of establishing early inflammatory changes in the tissues of the pharynx during the inflammatory process and differentiating the type of inflammatory disease of the pharynx to prevent the disease at its initial stage, taking preventive measures, selecting rational therapy and monitoring it (evaluating the effectiveness of treatment) at all stages of the inflammatory process. Using the proposed method of fluorescence spectrometry in conjunction with the use of indices characterizing the metabolic, morphometric, functional features of pharyngeal tissues in normal and inflammatory diseases, it seems possible to accelerate the diagnosis of the form of inflammatory pharyngeal disease, since the method allows for minimal time with high sensitivity, non-invasively , in express mode, in situ, both in vivo and in vitro using a portable hardware-software computer Lex to identify the nature of the inflammatory process, as well as its variety.

EFFECT: implementation of a method for differential diagnosis of the form of an inflammatory pharyngeal disease using fluorescence spectrometry and a special algorithm and indices embedded in the software, together with objective examination data, as well as recording spectral characteristics of the tissue of the tonsils and posterior pharyngeal wall in express mode in situ, in vivo and in vitro using the highly sensitive and non-invasive rapid method.

The method is as follows: for recording spectral data, a device is used for the Raman-fluorescence diagnostics of the state of human tissues in normal and pathological conditions with wavelengths of laser radiation of 405 nm for in vivo measurements and 532 nm for in vitro measurements, which includes a laser with a laser filter, a system of mirrors and lenses; a system that collects a signal coming from the object under study and a cut-off filter; a spectrometer with a CCD camera, characterized in that the spectrometer additionally contains special nozzles and / or devices for express sample preparation of objects for their investigation in a liquid, loose, solid state and / or smear - an “imprint” of biological tissue. The spectrometer is connected to a computer on which special software is installed that implements algorithms for monitoring, diagnosing and correcting the condition of the subject and / or human biological tissues in normal and pathological conditions, using special software, calibration, normalization, background subtraction and scaling of the obtained data are performed and a diagnosis of the condition of the subject and / or biological tissues of a person is carried out, the spectrometer is connected to the feedback laser, the device also rzhit fiber optic cable for exposure by laser radiation, the microscope images for precision imaging and flash digital microscope with autofocus and illumination for micro-and macro-objects for the simultaneous imaging of fluorescence images and pattern of distribution, for in vitro studies and in vivo. When conducting differential diagnosis of forms of inflammatory diseases of the pharynx in vivo, laser radiation with a wavelength of 405 nm is applied using a special optical fiber to the tissue of the tonsils and posterior pharyngeal wall. The measurement is carried out by the contact method after preliminary processing of the fiber with an antiseptic solution.

Using laser radiation with a wavelength of 532 nm, it seems possible to analyze in vitro express mode the metabolic, morphometric and functional features of the tissues of the pharynx, as well as biological fluids of patients (blood, urine, saliva, etc.) and microbes that cause one or another type inflammatory diseases of the pharynx, which may be relevant not only in otorhinolaryngology, but also in other areas of clinical medicine.

The method of differential diagnosis of the form of inflammatory diseases of the pharynx is carried out using hardware and software systems operating on the basis of wavelengths of 532 nm (in vitro) and 405 nm (in vivo), which consists in recording measurements and spectral characteristics of the tissue in normal and inflammatory process in pharynx with chronic tonsillitis (toxic-allergic form 1 and toxic-allergic form 2) or chronic pharyngitis of hypertrophic form in a contact, stable way, without lighting, with preliminary treatment By operating the fiber. Measurements were taken at several points of the tonsils (with tonsillitis): at the upper pole, at the lower pole, in the gap, and also in the region of the posterior pharyngeal wall (with pharyngitis). In total, from 4 to 10 measurements were performed to obtain a more accurate result and reproducibility of spectral characteristics. Before each measurement, the fiber was processed with an antiseptic solution. Exposure time is 1-5 seconds. Then, video recording of the spectrum and calculation of absolute fluorescence intensity, spectral half-width, normalized fluorescence index (normalization is carried out with respect to the intact point), aerobicity index (ratio of a wavelength of 665 nm to a wavelength of 670 nm), structuredness and tissue consistency index, characteristic features are carried out peaks recorded at certain wavelengths and other indicators - informative parameters to identify the type of tissue: healthy tissue or tissue, affected inflammatory process, as well as identification of the form of the inflammatory process in chronic tonsillitis or pharyngitis. The calculation and interpretation of each of the above indicators is given below, including a complete diagnostic algorithm.

In the presented algorithm, indicators such as the generalized fluorescence signal, the characteristics of the peaks themselves are analyzed: their shape, amplitude, width, wavelength, their number is normal and for various forms of inflammatory diseases of the pharynx, tissue structure and consistency index, aerobicity index, individual metabolic index , as well as many additional parameters, which are calculated as follows.

In FIG. 1 shows the fluorescence maxima for various fluorochromes (Koltova N.A., Fluorescence in medicine. 2017).

It is known that each type of chemical, including various fluorochromes, has a maximum fluorescence at certain wavelengths, making it possible to identify it. This phenomenon allows you to register changes in metabolism in the tissue in dynamics using the proposed new method based on the registration of the qualitative and quantitative composition of a biological object. A feature of the proposed method is that not only the fluorescence phenomenon is used as a diagnostic criterion, but also a special diagnostic algorithm and indices are used, which allow us to characterize the type of inflammatory process in the pharyngeal tissues in dynamics.

The advantage of using these indices as diagnostic parameters is that they are normalized to an intact point within the studied individual, which makes it possible to level out the systematic error associated with the individual tissue parameters of each examined, which will be described in detail below, after statistical processing of the research data .

Due to the fact that the human body has a metameric structure, and symmetrical regions are innervated from the same ganglia, so there is a mutual influence of adjacent anatomical zones on blood flow and innervation (Alexandrov MT, 2008), as an intact point, an asymmetric point on the skin of the inner surface of the forearm is selected.

After processing the results, it turned out that the intact point, despite the fact that it is asymmetric, is closely interconnected with the studied points, which also confirms the possibility of using the proposed diagnostic method not only at the local, but also at the system level.

At the preliminary stage, we tried to choose as intact points those that were located in the head and neck, such as the skin of the buccal region, lips, etc., however, with appropriate measurements, we observed wave interference, and therefore as an intact we selected a point in the region of the skin of the inner surface of the forearm (asymmetric point).

In preliminary studies during the development of the technique, it was noted that the spectra taken from the surface of the tonsils on both sides had almost identical characteristics of the spectral curves, which additionally confirms not only the similar nature of the blood flow in this area, but also the systematic nature of the process involving symmetrical zones in pathological process. In addition to the intact point, we carried out measurements at 3 studied points within the same individual. Point 1 was located in the region of the upper pole of the palatine tonsil, point 2 - in the region of the lower pole of the palatine tonsil, point 3 - in the region of the lacuna of the palatine tonsil, point 3 in chronic pharyngitis - in the region of the posterior pharyngeal wall, and in healthy volunteers - point 3 - in the region mucous membrane of the posterior pharyngeal wall when compared with a group of patients with chronic pharyngitis. When comparing with a group of patients with chronic tonsillitis, we selected points 1, 2, 3 in the region of similar localization in healthy volunteers, namely, in the region of the upper pole, lower pole and lacuna of palatine tonsils. When compared with a group of patients with chronic pharyngitis, point 3 was a point on the back of the throat. Thus, the compared points in all groups were identical and depended on the localization of the pathological process. Also, in preliminary studies, it was proved that the relationship between the calculated indices at the intact point and the studied points, which made it possible to use the data obtained with respect to the intact point, also for diagnostic purposes.

The values of these indices at the studied points are calculated as follows:

if the spectrum is specified by a function of intensity versus wavelength: F = F (x),

then the indices presented can be designated as integral fluorescence indices (over the entire spectral range) for the Intact and Fundus functions.

Legend: Intact - intact point, in this case - the skin of the inner surface of the forearm, Fundus - point 1 - at the upper pole of the palatine tonsil, Middle - point 2 - on the mucous membrane of the palatine tonsil in the region of its lower pole, Internal - point 3 - at palatine tonsil lacunae (in case of chronic tonsillitis) and in the region of the posterior pharyngeal wall (in case of chronic pharyngitis).

The studies revealed that, given the difference in the morphometric structure of the pharynx tissue, as well as the nature of the metabolites for various types of inflammatory process in the pharynx, one can use an almost infinite number of relative indices characterizing the physiology of the processes in the pharyngeal tissues, but only a few of them are the most informative for diagnostics.

The formulas below describe the designations described. All functions are embedded in the software of the hardware-software systems used in the claimed method.

The 3 indices presented above are normalized (by the integrated fluorescence index at the intact point); these fluorescence indices are presented for points 1-3: Fundus, Middle, Internal and characterize the structure of the tissue, its viability, functional and metabolic changes in it normal and during the inflammatory process in the pharynx.

The 3 indices described above are indicated in the description as “normalized fluorescence index” (hereinafter referred to as “NPF” or “IF”).

The above four indices for the functions Intact, Fundus, Middle, Internal (intact points and points 1-3) are indicated in Tables 2, 4, 6 as the “aerobic index” (hereinafter referred to as “IA”), which allows to assess the degree of prevalence of the process - aerobic or anaerobic, the degree of tissue oxygenation, as well as the nature of the change in metabolism in intact tissue when it is involved in the inflammatory process.

As it turned out during the study, for pharyngeal tissues in normal and inflammatory diseases, there are countless such indices, based on which you can calculate and analyze spectral data, but the aerobicity indices described above, normalized fluorescence indices illustrating the degree of microbial contamination of the pathological focus are the most informative , the structure and consistency of the tissues of the pharynx, the stage of the inflammatory process, which will be illustrated in detail Based on the examples presented in the “Drawings and Drawings” section and described below.

The number of calculated indices varies depending on the individual tissue parameters recorded at specific wavelengths (wave numbers), however, when comparing the data obtained for healthy volunteers, patients with chronic tonsillitis and chronic pharyngitis, it turned out that some final number is the most informative indexes, which was confirmed in relevant studies. The results obtained are shown below, both on specific examples and as a result of statistical data processing .;

Also, normal, with chronic tonsillitis and chronic pharyngitis, various metabolites are recorded that can be identified by calculating the index values according to the formulas presented above.

It is important for the clinician not only which metabolites or microorganisms play a role in the development of inflammatory diseases of the pharynx, but also what changes in the tissue generally occur, and the latter aspect is more important when analyzing data in the studied groups, since it allows you to evaluate all the processes that occur in tissue at morphometric, metabolic, functional levels.

The calculation algorithm using the proposed method and the interpretation of each of the indices are as follows (in stages):

1. registration of the spectral curve at the intact point;

2. registration of the spectral curve at the studied point (points 1-3 in the study);

3. registration of all wavelength ranges in which changes are observed;

4. allocation of the most informative wavelength ranges at which changes are recorded;

5. calculation by formulas, examples of which are presented above, that is, calculation of the ratio of the integrals of the total fluorescent signal at certain most informative wavelengths with respect to each other and to the intact point;

6. comparison of the obtained values in automatic mode.

7. Conclusions: the corresponding values in the sample of healthy volunteers differ from those for chronic tonsillitis and chronic pharyngitis, which is confirmed by the relevant studies described in detail below.

All of the above indices allow us to judge the physiological, morphometric, metabolic changes in the tissue in the normal and inflammatory process with chronic tonsillitis and chronic pharyngitis. Assessing these indices using special software, it becomes possible to conduct an instant diagnosis of the type of pathological process in the tissues of the ENT organs in the inflammatory process, monitor treatment, evaluate its effectiveness, and differentiate tissues in the early stages of the disease.

When conducting relevant studies to prove the informativeness of the application of fluorescence spectrometry in combination with special indices as a diagnostic method for inflammatory diseases of the pharynx, a clinical algorithm for diagnosing inflammatory diseases of the pharynx was developed at the first stage.

At the second stage, the sensitivity and specificity of the method were determined, which were close to 100%. At the third stage, the most informative parameters were identified that characterize pharyngeal tissue normal and in case of chronic pharyngitis, chronic tonsillitis, which are special indices calculated according to the algorithm described earlier.

The developed algorithm for the diagnosis of inflammatory diseases of the pharynx, which allows one to evaluate the morphometric, metabolic, and functional changes in the intact tissue and the tissue involved in the inflammatory process in diseases of the pharynx, can be used to immediately identify the pathological process and then prescribe the most rational treatment and monitor it, and evaluate the effectiveness of throughout the course of therapy.

To assess the spectral characteristics of the pharynx tissue in healthy volunteers and patients with chronic tonsillitis and chronic pharyngitis, the following diagnostic algorithm was developed and a method for calculating and identifying its informative criteria for inflammatory diseases of the pharynx using fluorescence diagnostics using special indices for assessing morphometric, metabolic, functional changes in the tissues of the pharynx in normal and pathological conditions:

1. Assessment of the sensitivity and specificity of the LCD RFU technique.

2. Evaluation of fluorescence power indices at the studied points in normal and with VZG.

3. Evaluation of the aerobic index in normal and with VZG in the studied points.

4. Evaluation of the structured index of pharyngeal tissues in normal and with VZG in the studied points.

5. The analysis of the intensity of the main fluorescence signal in normal and with VZG in the studied points.

If the fluorescence intensity at all the studied points differs by no more than 5-10% from the reference values (in the absence of an inflammatory process in the pharynx), then we designate this result as + (table 3). If the fluorescence intensity at all the studied points differs by no more than 20-30% from the reference values (in the absence of an inflammatory process in the pharynx), then we designate this result as + - (table 3). If the fluorescence intensity at all the studied points differs by more than 31% from the reference values (in the absence of an inflammatory process in the pharynx), then we designate this result as - (table 3). The designation +++ is characteristic of reference values (table 3).

6. When assessing the consistency of tissue, we use the same notation: “++” means the complete structure of the tissue (there is no inflammatory process, the tissue is intact), “+ -” means that the structure of the tissue is partial (the process is compensated), “-” is used to designations of a complete violation of the structure of the tissue (decompensated process or pronounced changes associated with the inflammatory process),

7. Based on the results of calculations of all indicators, we evaluated the tissue solvency index in normal conditions and in chronic tonsillitis (the classification of chronic tonsillitis of B.S. intact tissue, which was completely solvent, “++ -” and “+ -” - tissue partially solvent, to varying degrees, which is observed in chronic tonsillitis of toxic-allergic form 1 (XT TAF 1) and toxic-allergic form 2 ( XT TAF 2) and at various stages of rehabilitation depending on the form of the disease (chronic tonsillitis) and on the reactivity of the body; “---” - the tissue is completely unsound, marked changes are noted in its structure and metabolic; component, in comparison with reference values, in addition, with a particularly pronounced pathological process, especially when anaerobic microflora is attached, a shift of the fluorescence peak to the right is noted and, the more pronounced this process, the more intense its severity. The indicators presented by us are fixed in a software product and are calculated automatically on the principle of feedback in the online mode.

Using the new method, an accelerated differential diagnosis of the form of the inflammatory disease is carried out: chronic tonsillitis or chronic pharyngitis.

All indicators obtained in relation to inflammatory diseases of the pharynx were compared with indicators of intact tissues - tissues not involved in the inflammatory process.

It was found that, depending on the nature of the metabolic, morphometric, functional changes in the intact tissue during the inflammatory process, it is possible to conduct differential diagnosis of the form of the inflammatory disease of the pharynx using fluorescence spectrometry and special indicators calculated by the above method and embedded in the software with high accuracy and sensitivity non-invasively and in express mode, in situ, in vitro and in vivo.

Existing methods for diagnosing inflammatory diseases of the pharynx are either long-term or expensive, or give only an indirect idea of the processes occurring in the body, not allowing in-vivo in situ diagnostics, or difficult to perform. They are also not as sensitive methods as fluorescence spectrometry, which is not only highly sensitive, but also non-invasive and also fast. But the greatest emphasis to justify the use of the proposed method is carried out not on fluorescence spectrometry itself, which was previously used repeatedly in studies (Ryan K. Orosco, 2013, etc.), but on the diagnostic algorithm using special indices informative in relation to the rapid diagnosis of inflammatory diseases pharynx. The greatest advantage of this method is the ability to assess the physiological changes in the tissues of the pharynx that occur at earlier stages, compared with the morphometric ones, thus the method can be used for early diagnosis of inflammatory diseases of the pharynx. In addition, the method allows you to register numerous parameters in real time, that is, to carry out "diagnosis at the place of examination and treatment." To determine the morphological, functional and metabolic characteristics of pharyngeal tissues under normal and pathological conditions, including on the basis of recording autofluorescence characteristics, it is necessary to choose the most optimal method, which includes the method of fluorescence diagnostics. The development and implementation of the presented technology in wide clinical practice is promising and clinically feasible and will subsequently significantly improve the quality of diagnosis of inflammatory diseases of the pharynx and timely prescribe rational treatment.

The application of the proposed method also allows for reliable, highly sensitive (10 ⁴ -10 ⁵ CFU / ml) detection of the etiological factor in various forms of inflammatory diseases, as it helps to identify the type, concentration of the microorganism or microorganism associates.

The results of the rapid diagnosis of forms of inflammatory diseases of the pharynx are presented in the corresponding figures and tables located in the "Graphs and Drawings" section and described in detail below.

The choice of the method of fluorescence spectrometry using a special algorithm and indices and its software implementation in hardware-software complexes based on laser radiation with wavelengths of 405 nm and 532 nm for chronic tonsillitis and pharyngitis is justified by the need to increase sensitivity and specificity, speed up diagnostics with subsequent interpretation of informative criteria for individual spectral characteristics. To identify and confirm diagnostic informativeness of fluorescence spectrometry in combination with morphometric, metabolic, and functional characteristics of pharyngeal tissues, two groups were compared based on an analysis of the spectra of the tonsils and posterior pharyngeal wall of patients from the group of patients with chronic tonsillitis TAF 1, chronic tonsillitis TAF 2, chronic pharyngitis of hypertrophic form (HF GF).

The results are presented in the figures, the description of which is given below (see the appendix “Drawings and drawings”):

Differential diagnosis of forms of chronic tonsillitis and pharyngitis is one of the most important tasks facing otorhinolaryngologists and general practitioners. Using the method of fluorescence spectrometry in conjunction with the use of special indices, it becomes possible to analyze the spectral data of pharyngeal tissues in normal and inflammatory diseases of the pharynx in express mode, non-invasively and in dynamics, in the early stages. In the work used classification of chronic tonsillitis B.S. Preobrazhensky - V.T. Palm tree.

The separation between the studied classes was close to 100% with a sensitivity of about 100% and an accuracy of about 100% for tissue differentiation in chronic pharyngitis, chronic tonsillitis and normal in intact tissues that are not involved in the inflammatory process. When differentiating forms of chronic tonsillitis using the proposed method, high sensitivity and specificity were found only when comparing one point in the lacuna of the palatine tonsil: these points were different in all the studied groups after statistical processing of data, which is illustrated below.

However, it should be understood that the differential diagnosis in this case is more informative when analyzing the totality of data obtained in relation to the forms of inflammatory diseases of the pharynx, namely, objective research (pharyngoscopic picture and examination of other organs), registration of the corresponding metabolites in the blood plasma and other biological fluids of patients , assessing the microbial composition of the pathological focus, the concentration per gram of tissue of microbial bodies in a given period of time at the corresponding stage of the disease evanie, that is, a systematic approach is needed that can be implemented by a competent specialist.

A systematic approach with the analysis of the above indicators in dynamics at various stages of the disease will also make it possible to judge the physiological processes occurring in tissues, such as blood flow and microcirculation, proliferative activity of cells, difference in metabolites, features of morphometric data, etc. However, for the clinician it is not so important what metabolites, microbes, changes at the physiological and morphological levels, individually, occur in the tissues, how much all the parameters are important in the aggregate and in the comparative aspect: “norm-pathology”, which are presented in the proposed method using special diagnostic algorithm and are expressed in graphical and digital form, which is convenient and informative for the clinician. Thus, the doctor can use the method (in the future) without resorting to a huge variety of other diagnostic methods, which allows to reduce the time of diagnosis and treatment and provide rational therapy, however, it should be understood that the method is much more informative when using objective examination data.

Below are the clinical observations of various forms and stages of inflammatory diseases of the pharynx (VHI): studies using fluorescence spectrometry using a special algorithm and indices, using specific examples. The studies were conducted on the basis of clinical observations in dynamics using the method of fluorescence spectrometry. To confirm the results of the clinical objectivity of the method, the bacteriological method was used.

In FIG. 2 shows graphs of spectral characteristics of the tissue of the tonsils of healthy volunteers. I-spectrum at the intact point; 1, 2, 3 - spectra at the studied points on the surface of the tonsils - points 1, 2, 3, respectively. On the abscissa axis, the magnitudes of the wave numbers are the values inverse to the wavelengths, the unit of measurement is the inverse centimeters (1 / cm); along the ordinate, the fluorescence intensity (in relative units). The fluorescence intensity of the tissue of the tonsils, not involved in the inflammatory process, measured at different points, is almost the same and extremely low in intensity, compared with the amplitude-spectral indices of the comparison area (skin) - it is almost 1/10 of the fluorescence intensity in the intact region ; additional fluorescence peaks are not recorded; structured tissue and its viability are not violated; thus, the ratio of the fluorescence power integrals and other parameters (indices) is almost equal to unity within the spectra at the studied points, that is, they are almost the same.

In FIG. 3 shows the spectral characteristics of the tissue of the posterior pharyngeal wall of healthy volunteers. To the abscissa axis, the magnitudes of the wave numbers are the values inverse to the wavelengths, the unit of measurement is inverse centimeters (1 / cm); along the ordinate, the fluorescence intensity (in relative units).

The legend is embedded in the English software, the decoding of the indices is as follows: I_Intact - fluorescence intensity in the intact tissue, Air Intact - aerobicity index at the intact point, I_А, I_В, I_С - normalized fluorescence indices at points 1, 2, 3, respectively (NPF1, NPF2, NPF3, respectively); Air_A, Air_B, Air_C-aerobic indices (IA1, IA2, IA3) - at points 1, 2, 3, respectively; Infl. 1, Infl. 2, Infl. 3 - indices of microbial dissemination of tissue at points 1, 2, 3, respectively, calculated as follows:

The wavelength ranges of 665-675 nm and 713-723 nm are taken, since they are, in this case, the most informative with respect to recording the spectra of microflora and determining its concentration. In the figure along the abscissa, the reciprocal of the wavelengths is the wave numbers, the unit of measure is also embedded in the software.

From FIG. Figure 3 shows that normal (in the absence of an inflammatory disease of the pharynx), the fluorescence intensity in the region of the posterior pharyngeal wall is low (in this case, no more than 500 rel. Units), however, in the inflammatory process, chronic pharyngitis (hypertrophic form), a significant increase in intensity fluorescence, up to 5000 rel., as illustrated in FIG. 4. Moreover, the fluorescence intensity of the tonsils tissue is higher than the fluorescence intensity of the tissue of the posterior pharyngeal wall (Fig. 2, Fig 3), which indicates a more pronounced microbial contamination of the tonsils per gram of tissue. The latter position was proved using the bacteriological method in preliminary studies.

In FIG. Figure 4 illustrates the spectral characteristics of the tissue of the posterior pharyngeal wall with hypertrophic pharyngitis, where the I-spectrum is at the intact point, the OOS is the back-reflected signal, 1, 2, 3 are the spectra at the studied points - 1 (upper palatine tonsil), 2 (lower palatine pole tonsils), 3 (palatine tonsil lacuna), respectively.

As can be seen from FIG. 4, with hypertrophic pharyngitis there is a violation of the structure of the tissue, a significant increase in the fluorescence intensity over the entire frequency range; an additional peak appears at the value of the wave number 5750 / cm, as well as an increase in the width of the back-reflected signal, which indicates violations of the morphometric, metabolic, functional parameters of tissues, which is not observed normally (Fig. 3).

We give examples of calculating the indices of structured and tissue consistency.

FIG. 5 Spectral characteristics of the tissue of the tonsils with chronic tonsillitis, compensated form. The abscissa axis is the magnitude of the wave numbers, the ordinate axis is the fluorescence intensity in relative units.

I-spectrum at the intact point, 3 - spectrum in the lacuna of the palatine tonsil. When registering the spectrum at point 3, there is a significant increase in fluorescence intensity, which is not observed at point 3 in a healthy volunteer (Fig. 2).

According to the developed diagnostic algorithm using the method of fluorescence spectrometry in conjunction with informative indexes for assessing the morphometric, metabolic and functional parameters of pharyngeal tissues in normal and inflammatory diseases of the pharynx, we can conclude that:

- fluorescence intensity and its power in chronic tonsillitis (indicated by 3 in the figure - the spectrum recorded in the region of the 3rd point - in the region of the lacuna of the palatine tonsil, 2 - in the region of the lower pole of the tonsil, 1 - in the region of the upper pole tonsils), the fluorescence intensity at point 3 exceeds that at the intact point "And." In addition, there is an additional peak at the wave number of 6350 / cm, which is not observed normally (Fig. 2). The aerobic index at point 1 is, in this case, 0.993, at point 2-2.04, at point 3-1.79, which is illustrated in the table under FIG. 5.

The II fluorescence intensity at the intact point is 14,000 relative units (OE), the fluorescence intensity (IF) in the tissue of the tonsils not involved in the inflammatory process is 1700 OE (Fig. 2) - 100%, while the fluorescence intensity at point 3 - the lacuna of the palatine tonsil, - is equal to 19000 ОЕ-х%, х = 1118, increase of IF (fluorescence intensity) by 11 times. Thus, it is possible to calculate the indices of tissue structure and consistency in relation to all points:

At point 2, the IF is equal to 4200 rel. units - 100%, while at point 3 - 19000-x%, hence x = 452%, that is, 4.5 times more than at point 2, which indicates completely the disunity and violation of the structure of the spectra (as with reference values in the norm, and at the points of the palatine tonsil with respect to each other) and the consistency of pharyngeal tissues in chronic tonsillitis (---).

The aerobic index at the intact point is 1.85, at the second point is 2.04 (the table in Fig. 5 is embedded in the software), respectively,

1.85-100%,

2.04-x%, hence x = 110, that is, with chronic tonsillitis of the compensated form, an increase in IA at point 2 by 10% (+) is noted, which indicates the compensation of the inflammatory process in the tissue of the tonsils (relatively small physiological changes).

Thus, from FIG. 5 it follows that the intensity of fluorescence during the inflammatory process increases and correlates with microbial contamination of the pathological focus (tissue of the tonsils) per gram of tissue, since the greatest microbial contamination is observed in the region of the gaps of the tonsils.

Also at a value of 6350 / cm an additional peak is detected, which is not detected normally (in the absence of XT).

In FIG. 6 shows the spectral characteristics of the tissue of the posterior pharyngeal wall with hypertrophic pharyngitis. The main fluorescence signal is recorded at a wave number of 5000 / cm; there is a violation of tissue structure, an additional peak at the wave number of 5750 / cm. Symbols in the figure: I-intact point, 1, 2, 3 - spectra at points 1, 2, 3, respectively.

From the figure it follows that with hypertrophic pharyngitis, an increase in the fluorescence intensity is noted at all the studied points, the structure of the tissue is completely impaired, which can be confirmed by the corresponding calculations:

so, at point 1, the fluorescence intensity of 500 OE-100%, and at point 2-3000 OE-x% (600%), the changes are 500% (3000: 500 = 6), which indicates a violation of the structure of the tissue and its consistency, which is not observed normally (Fig. 2, Fig. 3).

In FIG. 7 illustrates the spectral characteristics of the tissue of the tonsils before washing the gaps of the tonsils with an antiseptic solution (miramistine 0.01%), patient K., 26 years old, with chronic tonsillitis, toxic-allergic form 2 (XT TAF 2) and chronic hypertrophic pharyngitis (HF GF). The number 3 in FIG. 7, 8 indicate the spectra taken at point 3 in the gaps of the palatine tonsils.

From FIG. 7 it follows that with chronic tonsillitis of TAF 2, an additional peak is observed at the wave number of 6370 cm – 1, which is recorded in chronic tissue hypoxia, according to preliminary studies and not observed normally (Fig. 2). There is also a violation of tissue structure. IF at point 3-1500 OE, at point 2-4000 OE, thus

1500-100%, 4000s, x = 267%, that is, with chronic tonsillitis, toxic-allergic form 2, there is a violation of the structure and consistency of pharyngeal tissues by 167%, which indicates a complete violation of the structure of the tissue (---) .

In FIG. Figure 8 shows the spectral characteristics of the tissue of the tonsils of a patient K., aged 28 with chronic tonsillitis, TAF 2 and chronic hypertrophic pharyngitis after washing the lacunae of the tonsils with miramistin solution, an additional peak at the wave number of 6350 / cm remains, but its amplitude increases, which indicates the death of microbes and the exit of porphyrins (which are endogenous fluorophores) of their cell wall; after exposure to miramistin 0.1% as a detergent, the amplitude of the peak recorded at a wave number of 6350 / cm increases; the obtained data are confirmed by studies conducted by domestic scientists (Popov S.N., 2012; Cherkesov I.V., 2005) .

In FIG. Figure 9 illustrates the spectral characteristics of tissue in the region of the amygdala niches of a patient K., 28 years old with chronic tonsillitis, TAF 2 and chronic hypertrophic pharyngitis 2 weeks after bilateral tonsillectomy.

The spectral signs of chronic tonsillitis persist: tissue structure is partially impaired, the fluorescence intensity is higher than the values recorded in the norm, but there is no longer an additional peak in fluorescence at a wave number of 6350 / cm, which indicates the severity of microbial contamination per gram of tissue. After removal of the tonsils, there is no chronic pathological focus, gradually the microflora of the pharynx changes and approaches that of healthy volunteers, which was illustrated in previous studies.

In FIG. 10 shows the distribution of aerobicity index values in the studied groups. N-IA is normal; 1 - IA in chronic tonsillitis TAF 1, 2 - IA in chronic tonsillitis TAF 2.

The concentration of the average IA values on a certain area in each of the presented groups is noted. The highest aerobicity index is observed in the case of normal, but in the case of XT TAF 2 there are single IA values that are superior to IA in the group of healthy volunteers, which can be explained from the standpoint of attempts to restore homeostasis in the body, including through redistribution of blood flow in chronic tonsillitis, TAF 2 , at the maximum maximum permissible load on the body (XT decompensation).

In FIG. 11, the ratio of NPFs is normal in case of chronic tonsillitis, TAF 2. The XT distribution of NPFs in the group of patients with chronic tonsillitis TAF 2. The designation "CT" in FIG. 11 - chronic tonsillitis, toxic-allergic form 2.

Analyzing NPFs in dynamics, one can draw a conclusion about what stage the inflammatory process is in the pharynx, monitor and evaluate indicators in a comparative aspect to prescribe appropriate adequate therapy for a given period of time, which will be described in detail below, after the statistical processing of the data.

In FIG. 12 presents a comparison of the fluorescence intensity of the tissue of the tonsils (I max N) in normal and chronic tonsillitis, toxic-allergic form 2 (I max, decomp.), Where N is normal, 2-XT TAF 2.

In FIG. Figure 12 shows a comparison of the fluorescence intensity in normal and chronic tonsillitis: in the inflammatory process, a significant increase in IF is observed (7 times higher than normal), which is associated with both an increase in microbial dissemination of tissue and a more intensive proliferative process in the tissue of the tonsils at XT. In particular, with chronic tonsillitis, a decompensated form, not only tissue destructuring of the palatine tonsils, impaired blood flow and decreased oxygenation are noted, but also an intensive scar process, tissue sclerosis and impaired function.

In FIG. Fig. 13 shows an increase in IF in chronic tonsillitis, TAF 1 compared with tissue not involved in the inflammatory process, but this increase is less pronounced than in the normal series - “CT TAF 2”, which can be explained from the perspective of restoration of homeostasis and mobilization of body resources to fight the inflammatory process. In this case, the inflammatory process is relatively controlled and compensated. Legend in FIG. 13: H — norm, I max N — maximum fluorescence intensity in norm, 1-HT TAF1, I max comp. - maximum fluorescence intensity with CT TAF 1.

In FIG. 14 illustrates the spectral characteristics of the tissue of the tonsils with chronic tonsillitis, TAF 2. 3 - spectrum at point 3 - gap of the tonsils.

In addition to a violation of the structure of the tissue of the tonsils, a peak is recorded at the wave number of 6350-6400 / cm, as well as an increase in the fluorescence intensity of the tissue of the tonsils compared to the reference tissue values of the NM of healthy volunteers, as described in detail above.

FIG. 15 Spectral characteristics of the tonsils tissue of a patient with chronic tonsillitis, decompensated form, 1 day after opening of the paratonsillar abscess.

In FIG. 15 we see that during the decompensation of the process - with the toxic-allergic form 2 of chronic tonsillitis, the structure of the tissue and its viability are violated in comparison with the reference values, however, no additional peaks are recorded, due to the fact that the cavity is rinsed after opening the paratonsillar process with antibacterial and antiseptic drugs , which contributes to the death of microbes.

FIG. 6 Spectral characteristics of the tissue of the palatine tonsils of a patient with chronic tonsillitis, decompensated form, 6th day after opening of the paratonsillar abscess.

In FIG. 16 we see that after a certain period of time (6th day after the operation), the structure of the tissue is gradually restored.

FIG. 17 shows the spectral characteristics of the tonsils tissue of a patient with chronic tonsillitis, TAF 1 before washing the tonsils lacunae: there is an increase in the fluorescence intensity of the tonsils tissue, the appearance of additional peaks at the wave numbers of 6350-6400 / cm, violation of the structure of the tissue NM compared with normal values. In FIG. 17, figure 3 denotes a spectrum taken in the region of the lacuna of the palatine tonsil - at point 3.

In FIG. 18 (3 - spectrum at point 3- in the lacuna of the palatine tonsil) the spectral characteristics of the tissue of the palatine tonsils of a patient with chronic tonsillitis, TAF 1, immediately after washing the lacunae of the palatine tonsils (1 minute) with a miramistin solution of 0.01%: a significant increase fluorescence intensity of tissue NM, the appearance of an additional peak at wave numbers of 6350-6400 / cm, violation of the structured structure of the tissue of the tonsils, compared with normal values, as well as an increase in the amplitude of the recorded supplement Nogo peak by increasing the concentration of porphyrin, which is achieved miramistina influence on the cell wall of the bacteria, resulting in increased fluorescence intensity has been shown in previous studies.

In FIG. 19 (the numbers indicate the spectra taken at points 1, 2, 3, respectively); spectral characteristics of the tonsils tissue of a patient with chronic tonsillitis, TAF 1, immediately after washing the tonsils lacunae with miramistin solution (10 minutes) are illustrated: a significant increase in the fluorescence intensity of NM tissue, the appearance of additional peaks at wave numbers of 6400 / cm, violation of the structure of the palatine tissue tonsils compared with normal values, as well as an increase in the amplitude of the recorded additional peak due to an increase in the concentration of porphyry new, which is achieved by the action of miramistin on the bacterial cell wall, as a result of which the fluorescence intensity increases, as was shown in previous studies, then, after some time, the fluorescence intensity decreases.

FIG. 20 Spectral characteristics of the tissue of the palatine tonsils of a patient with chronic tonsillitis, TAF 1, after a full course of washing of the gaps of the palatine tonsils with miramistin solution (for 9 days): a significant decrease in the fluorescence intensity of the NM tissue was noted, no additional peaks at the wave numbers of 6400 / cm the structure of the tissue of the tonsils is restored, approaching normal values, which indicates the effectiveness of the conservative treatment.

After processing all the data obtained regarding the effectiveness of conservative treatment for chronic tonsillitis of TAF 1 (toxic-allergic form 1), the values were obtained, which are presented in table 1, where NPF 1, NPF 2, NPF 3 are normalized fluorescence indices, at points 1, 2 , 3, respectively, IA1, IA2, IA3 - aerobic indices at points 1, 2, 3, respectively. The calculation of the presented indices was described earlier.

Table 1 presents these indicators before treatment and after treatment.

According to this table 1, it is noted that aerobic indices after treatment (washing the lacunae of the tonsils with antiseptic solutions, treating the pharyngeal mucosa with anti-inflammatory drugs, additional physiotherapeutic procedures, etc.) increase, which indicates the normalization of oxygenation and microcirculation in tissues, the restoration of its structure and physiological properties, normalized fluorescence indices, in this case, on the contrary, decrease, which indicates a decrease in activity m krobnogo (inflammatory) processes in the tissues of the pharynx on the background of the therapy.

With inflammatory changes in the tonsils with CT, additional peaks appear on the graphs at values of 6350-6400, 7050 / cm, which indicates both the registration of pathogenic flora and an increase in the concentration of conditionally pathogenic flora, which can lead to disease.

In preliminary studies, it was shown that the more pronounced the anaerobic process, the more the peak shifts to the right along the abscissa. The more intense the additional peak of fluorescence on the abscissa axis, the more pronounced the inflammatory process in the tissues of the pharynx.

On the graphs (Fig. 20) we see that the tissue structure, oxygenation, microcirculation are normalized, the aerobicity index increased, spectral data in favor of the pathogenic flora are not visualized, the maximum fluorescence peak lies in the lower third of the curve recorded from the surface of the intact point, which indicates clinical improvement and morphological restoration of the tissue of the tonsils after conservative treatment, which is not observed when analyzing the tissue spectra of the tonsils with CT TAF 1 before treatment niya (Fig. 18, Fig. 19).

When assessing the basic spectral characteristics of tissue in normal and inflammatory diseases of the pharynx (VHI), the fluorescence intensity at the intact point (II), the fluorescence intensity at the points on the surface of the tonsils in healthy volunteers and patients with chronic tonsillitis, as well as on the surface of the posterior wall, were analyzed pharynx in patients with and without chronic pharyngitis.

An important aspect of the diagnosis is not only an assessment of the functional state of the tissue during the inflammatory process, but also a qualitative and quantitative analysis of microflora for the selection of rational antibacterial therapy. In addition, it is necessary to evaluate the patient's rehabilitation after an infectious process, as well as prevent the widespread occurrence of epidemiologically significant diseases, such as inflammatory diseases of the pharynx.

In the experiment on microbes, it was also found that each microbe, depending on its type, has a certain set of spectral data that changes with an increase or decrease in their concentration per gram of tissue, it was also determined that additional peaks are recorded in tissues during the inflammatory process of the pharynx. shifted to the right along the ordinate axis, which indicates a microbial process, this effect is not observed when recording spectra of the pharyngeal tissue is normal. For the otorhinolaryngologist, as a clinician, not only the type of inflammatory pathogen is important, but also the concentration of microbes per gram of tissue, which can be registered using the presented method.

To assess the contribution of the microbial factor to changes in tissues that occur during inflammatory diseases of the pharynx, spectra of the tissues of the pharynx (tissue of the tonsils with chronic tonsillitis; tissue of the posterior pharyngeal wall with chronic pharyngitis) and some types of pathogens of inflammatory diseases of the pharynx were studied. To assess the information content of the use of fluorescent medical technologies in a series of in vitro experiments and in vivo studies, the main spectral features of pharyngeal tissues and some microbes colonizing it were demonstrated.

In FIG. 21 illustrates the spectral characteristics of the microbe Candida albicans (pure monoculture on non-fluorescent substrates).

FIG. 22 and FIG. 23 show the spectral characteristics of the microbe Candida dubliniensis (pure monoculture on non-fluorescent substrates) and the spectral characteristics of the microbe E. faecalis (pure monoculture on non-fluorescent substrates, respectively.

In FIG. 24 shows the spectral characteristics of the microbe Klebsiella pneumoniae - pure monoculture on non-fluorescent substrates.

In FIG. 25 shows the spectral characteristics of the microbe Proteus mirabilis (pure monoculture on non-fluorescent substrates).

FIG. 26 shows the spectral characteristics of the microbe Staphylococcus aureus (pure monoculture on non-fluorescent substrates).

In FIG. 27 shows the spectral characteristics of the microbe Streptococcus pneumoniae - pure monoculture on non-fluorescent substrates.

In FIG. 28 shows the spectral characteristics of the microbe Streptococcus pyogenes (pure monoculture on non-fluorescent substrates).

From the presented FIG. 21-28 it follows that each type of microbe is characterized by a set of specific spectral features that allow it to be calculated among many others. These spectral data can be presented both in digital and in graphical form, as can be seen in the corresponding figures. Also, in most microbes, an additional peak is detected at a wave number of 6700 / cm, which indicates the microbial component of the inflammatory process, namely, it can be noted that a similar peak is detected during the inflammatory process in the pharynx: especially in chronic tonsillitis (Fig. 5, 7, 8, 14, 17, 18, 19), which is not noted in the norm (Fig. 2). Thus, it can be concluded that the characteristic features of the spectral curves of the pharyngeal tissue in chronic tonsillitis and chronic pharyngitis are associated, in part, with the microbial factor, as shown in FIG. 21-28.

When the concentration of microbes per gram of tissue is exceeded, these changes become more obvious, namely, the fluorescence intensity increases, additional peaks appear at the wave numbers of 5350-6700 / cm, the more this peak is shifted to the right along the abscissa, the more pronounced the anaerobic process and the the inflammatory process in the pharynx as a whole proceeds more aggressively, which is confirmed by clinical and microbiological data.

As a result, after processing the obtained data by statistical methods, we obtained the following results.

As it turned out, the values of AI (fluorescence intensity at the intact point) and IAI (aerobic index at the intact point) are positively correlated, Spearman's rank correlation coefficient is 0.231 (p <0.001), which means that the absolute fluorescence intensity at the intact point is interconnected and dependent on aerobic index at this point in a particular individual.

In FIG. 29 this relationship is depicted more clearly. Legend: HF-chronic pharyngitis, AI - along the ordinate axis - fluorescence intensity at the intact point, in absolute units; IAI is the aerobic index at the intact point, in relative units.

Reliable information on the values of AI and IAI with gender and age has not been identified, that is, these values do not depend on gender and age.

From FIG. 29 it follows that the strongest relationship between the studied parameters is observed in the group of patients with chronic pharyngitis, in other groups it is less strong, so there is a correlation between the studied indices.

The study of the normalized fluorescence index and aaerobic index in the compared groups (0, 1, 2, 3) on the surface of the tonsils at the upper and lower poles (at points 1 and 2).

The considered indicators are distributed with a noticeable asymmetry and non-compactly, in connection with which, for the analysis of these indicators we will use mainly non-parametric statistics methods.

Table 2 presents a comparison of the statistical parameters of the studied quantities in the subgroups, where NPF1, NPF2 are normalized fluorescence indices at points 1 and 2, IA1, IA2 are aerobic indices at points 1, 2; M is the average value, σ is the standard deviation, Min and Max are the minimum and maximum values; Me is the median, m is the error of the mean.

After analyzing the aerobic indices and normalized fluorescence indices in the studied groups, we obtained significant differences between the NPF 1 and NPF 2 indices in all the studied groups, however, the IA indices in all the groups were unreliable (table 2). Thus, after verifying the reliability of the results by Mann-Whitney and Kolmogorov-Smirnov, it was noted that in the groups of “Healthy volunteers” and people with “Chronic tonsillitis, TAF 1” there are differences in the values of normalized fluorescence indices, the differences are significant, and in the absence inflammation (in healthy volunteers) these indicators are 1.7 (NPF 1) - 1.9 (NPF 2) times less than with inflammatory changes in the tonsils with chronic tonsillitis, TAF 1.

In chronic tonsillitis, TAF 2, the values of NPF 1 and NPF 2 exceed those in the normal range of 2.28-2.59; differences are significant.

In the group "Chronic pharyngitis" there are significant differences in the rates of NPF 1, NPF 2, which in the presence of the disease are 8.25-11.69 times higher than the corresponding values in the norm.

The forms “Chronic tonsillitis, TAF1” and “Chronic tonsillitis, TAF 2” do not have significant differences at the corresponding points IA1, IA2, NPF1, NPF2.

There are significant differences between the corresponding indices for chronic tonsillitis, TAF1 and chronic pharyngitis at all points: IA1, IA2, NPF1, NPF2 and NPF1 for HF 4.81 more than for XT 1, IA 1 1.33 times less for HF, NPF 2 is 6.25 times greater with HF than with XT 1, and IA2 is 1.279 times greater with XT 1 than with HF.

There are also significant differences at the corresponding points in chronic tonsillitis, toxic-allergic form 2 and chronic pharyngitis, so that NPF 1 is 3.623 times greater with CP than with CT TAF 2, IA 1 is 1.37 times more with CT TAF 2 than with HF, NPF 2, 5 times more with HF than with CT TAF 2, IA 2 1.31 times more with CT TAF 2 than with HF. All differences in the groups are significant.

Table 3 presents the normalized fluorescence indices, the indices of the structure and viability of the pharyngeal tissue in healthy volunteers, patients with chronic tonsillitis TAF 1 and TAF 2 and chronic pharyngitis of hypertrophic form. IP is the tissue structure index, IST is the tissue consistency index, NPF is the normalized fluorescence index, the numbers indicate the points at which the spectra were recorded.

Legend: +++ - the tissue is completely structured and consistent (typical for the tissues of the pharynx of healthy volunteers), ++ - the tissue is partially structured, + - the tissue is practically unstructured; ---- The fabric is not completely structured. ZD - healthy volunteers, CT TAF 1 - toxic-allergic form; CT TAF2 - chronic tonsillitis, toxic-allergic form 2; HF GF - chronic pharyngitis, hypertrophic form.

When analyzing the data of Table 3, it follows that with chronic pharyngitis and chronic tonsillitis, there is a violation of the structure and viability of the pharyngeal tissues, expressed to various degrees, in contrast to the corresponding results in the norm, in the absence of the inflammatory process in the pharynx.

Table 4 presents the correlation between the indicators of fluorescence intensity at intact points and at the upper pole of the palatine tonsil (point 1).

From table 4 it follows that there is an interdependence between the fluorescence intensity at the intact point and the aerobic index and the normalized fluorescence index. A similar interdependence can be traced in all the studied points. Thus, the consistency of the pathological process in inflammatory diseases of the pharynx is confirmed.

There is also an interdependence between aerobic indices and normalized fluorescence indices: when one group of indicators changes, others synchronously change.

Based on the presented modeling (Fig. 29, tables 2-4), we can finally formulate the basic laws of the interdependence of the most informative parameters from each other:

1. Fluorescence intensity, normalized fluorescence index, aerobic index at the studied points within the patient groups are interconnected;

2. In each of the studied groups there is an interdependence of these parameters of different strengths. The most severe dependence is observed in the group of patients with chronic pharyngitis;

3. In groups of healthy volunteers, as well as patients with chronic pharyngitis, there is the smallest variation in the values of IF, NPF, and IA indicators, both at the intact point and at the studied points in the palatine tonsils and posterior pharyngeal wall;

4. differences in the aerobic indices of IA 1 and IA 2 in the study groups were unreliable;

5. normalized fluorescence indices in the studied groups of NPF 1 and NPF 2 in all studied groups were different and the difference was significant;

6. in healthy volunteers, NPF indicators are 1.7 (NPF 1) - 1.9 (NPF 2) times less than with inflammatory changes in the tonsils with chronic tonsillitis, TAF 1.

7. with chronic tonsillitis, TAF 2, the values of NPF 1 and NPF 2 exceed those in the norm in 2.28-2.59; differences are significant.

8. In the group of patients with chronic pharyngitis there are significant differences in the rates of NPF 1, NPF 2, which in the presence of the disease are 8.25-11.69 times higher than the corresponding values in normal.

9. The forms “Chronic tonsillitis, TAF 1” and “Chronic tonsillitis, TAF 2” do not have significant differences at the corresponding points IA1, IA2, NPF1, NPF2.

10. between the corresponding indicators for chronic tonsillitis, TAF1 and chronic pharyngitis, there are significant differences at all points: IA1, IA2, NPF1, NPF2 and the rate of NPF1 with CP is 4.81 more than with XT 1, IA 1 is 1.33 times less with HF, NPF 2 6.25 times more with HF than with XT 1, and IA2 1.279 times more with XT 1 than with HF.

11. There are significant differences at the corresponding points in chronic tonsillitis, toxic-allergic form 2 and chronic pharyngitis, so that NPF 1 is 3.623 times greater with CP than with XT TAF 2, IA 1 is 1.37 times more with CT TAF 2 than with HF, NPF 2 is 5 times greater with HF than with CT TAF 2, IA 2 1.31 times more with HT TAF 2 than with HF.

Measurement of the normalized fluorescence index and aerobic index at point 3 (palatine tonsil lacunae in chronic tonsillitis and in healthy volunteers)

Table 5 presents the statistical parameters of the value of NPF 3 in the compared groups

The differences in the mean are significant with p = 0.004

According to table 5, there are significant differences between the average values of NPF 3 in groups of healthy volunteers and patients with chronic tonsillitis, TAF 1: NPF 3 in the group of healthy volunteers is 1.7 times less than in the group of patients with CT TAF 1, as well as in 3.6 times less than in the CT TAF 2 group, while NPF 3 in the group of patients with CT TAF 1 is 2.2 times less than in the group of patients with CT TAF 2, which can be used for the differential diagnosis of forms chronic tonsillitis.

Study of the normalized fluorescence index and aerobic index at point 3 (posterior pharyngeal wall in patients with chronic pharyngitis),

Of 51 patients in this group, NPF 3 values were observed in the range from 0.219 to 8.38 with an average of 2.66 ± 0.27 and σ = 1.91.

In FIG. 30 shows the frequency distribution of NPF 3 in patients with chronic pharyngitis, where Mean is the average value, Std. Dev. - standard deviation, N - number of subjects, Frequency - frequency of occurrence, along the ordinate axis, in relative units, NPF - normalized fluorescence index.

From FIG. 30 it follows that the variation of NPF 3 within the considered group is close enough to the normal distribution. In most cases, NPF 3 in this group is in the range from 0 to 4.5, however, it can reach 8.5 rel. units Normalization to the intact point in this case leads to leveling the difference between the dynamic parameters of the tissue, characterizing its metabolic, morphometric and functional changes.

Table 6 presents the correlation between the values of NPF 3 in patients with chronic pharyngitis with the aerobic index at the intact point, the aerobic index at points 1 and 2, as well as the normalized fluorescence indices at points 1 and 2 and the fluorescence intensity index c. intact point in patients from the same group.

From table 6 it follows that there is an interdependence between NPF 3 in chronic pharyngitis with other indicators, such as II, IAI, NPF1, IA1, NPF2, IA2, so when changing some indices, one can judge the nature of changes in others, as well as register the data obtained in express mode and in Dynamics using the proposed diagnostic method.

The proposed method and algorithm developed during the study allows differential diagnosis between the norm and pathology in inflammatory diseases of the pharynx, as well as to evaluate the main differences and characteristic features of the pharyngeal tissue in case of chronic pharyngitis and chronic tonsillitis of various forms.

Thus, the method of fluorescence spectrometry, together with informative criteria, has established itself as a non-invasive, highly sensitive, highly specific express method that allows us to evaluate the set of morphometric, metabolic, functional features of tissues in normal and inflammatory diseases of the pharynx.

Thus, in accordance with the data presented, it can be noted that the method of rapid diagnosis of inflammatory diseases of the pharynx using fluorescence spectrometry and a special algorithm for assessing morphometric, metabolic, functional changes in tissues in dynamics can be used in situ (at the "location" of the examination) , both in vivo and in vitro, to identify inflammatory diseases of the pharynx, as well as to assess the microbial factor (as an etiological) in the pathogenesis of the inflammatory process. Using the proposed method, it is possible in express mode, with high sensitivity and specificity, in dynamics to evaluate the morphometric, metabolic, functional features of the pharyngeal tissues in normal and inflammatory diseases of the pharynx, as well as to analyze the qualitative and quantitative composition of the microflora of the pathological focus. Thus, the development of an algorithm for the diagnosis of inflammatory diseases of the pharynx using this method can be used in the future for immediate identification of inflammatory diseases of the pharynx and, as a result, for the implementation of the most rational treatment and preventive measures.

Claims (1)

A method for the diagnosis of chronic diseases of the pharynx, selected from a compensated form of chronic tonsillitis, a decompensated form of chronic tonsillitis and chronic pharyngitis, using fluorescence spectrometry, which consists in the fact that the surface of the upper pole (point 1), lower pole (point 2), lacuna (point 3) palatine tonsils, as well as from the region of the posterior pharyngeal wall, spectral curves are recorded and absolute fluorescence intensity, spectral half-width, normalized fluorescence of the indicator relative to the intact point located on the inner surface of the skin of the forearm; comparing the obtained values with spectral indices in healthy volunteers; with an increase in the normalized fluorescence index at points 1 and 2 by 1.7-2.59 times compared with the norm, the appearance of additional peaks at the wave numbers of 5350-6700 / cm and a shift of these peaks to the right along the abscissa axis, chronic tonsillitis is diagnosed in the patient, and with an increase in the main fluorescence signal at a wave number of 5000 / cm, with an increase in the normalized fluorescence index at points 1 and 2, 8.25-11.69 times higher than the norm and the appearance of an additional peak at a wave number of 5750 / cm in the spectrum obtained from the back pharyngeal walls; chronic pharyngitis is diagnosed in a patient; the normalized fluorescence at point 3 in the group of healthy volunteers is 1.7 times less than in the group of patients with a compensated form of chronic tonsillitis and 3.6 times less than in the group with a decompensated form of chronic tonsillitis, and the normalized fluorescence at point 3 in patients with a compensated form of chronic tonsillitis is 2.2 times less than in the group of patients with a decompensated form of chronic tonsillitis.

Images