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TOM 17. N 1-2 2013 P.
20. The threal of multidrug-resistant tuberculosis: results of 1 yr of surveillance in the Lombardy region of Italy / G. Salamina, L. Sodano, F. Mezzetti, M. L. Moro // Mon-
aldi Arch Chest Dis. - 1999. - Aug., Vol. 54 (4). - P. 332336.
English version: VALUE MOLECULAR GENETIC METHODS OF DNA EXTRACTION IN THE TREATMENT OF NEWLY DIAGNOSED CHEMORESISTANCE TUBERCULOSIS
Filatova O.V.
Higher State educational if Ukraine "Ukrainian Medical Dental Academy", Poltava
The results of the genetic research of DNA from the sputum of patients with tuberculosis. Found that the detection of Mycobacterium tuberculosis resistance to antituberculosis drugs (isoniazid and rifampicin) and assigned the correct treatment, increases the effectiveness of the treatment of newly diagnosed tuberculosis chemoresistance. Performance criteria were: the timing of bacteriological and healing the destructive changes in the lungs. The results of this study allow us to recommend the molecular-genetic method of DNA extraction for the correction of the treatment of tuberculosis chemoresistance.
Key words: chemoresistant tuberculosis, molecular-genetic method of DNA.
which is associated with negative social factors, the increasing resistance of Mycobacterium tuberculosis to anti-TB drugs that are prone to rapid formation of drug resistance [1, 2, 3, 4]. In recent years among clinical isolates of Mycobacterium tuberculosis (MBT) has been increasingly meet both strains with resistance to almost all modern anti-TB drugs used in the clinic [5, 6, 7]. WHO estimates that about 50 million people on Earth are infected with multidrug resistant strains. The causative agent of tuberculosis has some unique features. We found a sufficient number of genes that can produce protein products responsible for the penetration of microbes into the host cells and its intracellular existence [8, 9, 10, 11, 12]. Due to the intracellular localization of M. tuberculosis and especially its membrane transport resistance genes seems difficult, in the case of resistant strains dominate the mutational changes of chromosomal genes [13, 14, 15, 16]. Thus, the probability of occurrence and selection of resistant strains of Mycobacterium large populations today is quite high [17, 18, 19, 20].
Objective: To increase the effectiveness of treatment of newly diagnosed himiorezystentny infiltrative and disseminated tuberculosis using molecular genetic methods DNA. Set the minimum time which of anti-TB drugs (isoniazidi or rifampicini) resistance is observed and immediately assign the correct treatment.
Materials and methods
60 patients were studied - 30 of 30 infiltrative and disseminated tuberculosis. Patients sick the first time, all had cavities and bacterial degradation. All patients were from Poltava and Poltava region. Among the patients studied was dominated men - 46 (76,6%) women were 14 (23,4%).
Analysis of resistance of Mycobacterium tuberculosis to anti-TB drugs (Table 1). In the group with infiltrative tuberculosis appeared to be the major indicators multyrezystant, it was found in 18 (60%) patients had monorezystant 12 (40%) patients. With disseminated
* To cite this English version: Filatova O.V. Value molecular genetic methods of dna extraction in the treatment of newly diagnosed chemoresistance tuberculosis / / Problemy ekologii ta medytsyny. - 2013. - Vol 17, Ns 1-2. - P. 92 -96.
Learning problems himiorezystentnyy TB incidence is determined in Ukraine and worldwide steady increase of patients. Detection of long-standing cases of tuberculosis with destruction, common process, improper treatment, violations of the sick, poor supply of medicines have increased cases of resistance that is the cause of increase in TB. Today is not sufficiently studied the problem of treatment of newly diagnosed destructive pulmonary tuberculosis with primary resistance of Mycobacterium tuberculosis to anti-TB drugs, the relevance of which is of particular importance given that the modern methods of primary resistance can be determined in 2-4 months after diagnosis. During this period, the patient should be assigned the correct anti-TB drugs but the best mode to use them at this time is not justified. Objective: To increase the effectiveness of treatment of newly diagnosed himiorezystentnoho infiltrative (n = 30) and disseminated (n = 30) TB lenen using molecular genetic methods DNA. Set the minimum time which of anti-TB drugs (isoniazid or rifampicin) resistance is observed and immediately assign the correct treatment. Among the patients studied were dominated by men - 46 (76,6%), 14 women (23,4%). Among men prevailed age 40-49, women 20-29, respectively. In the group with infiltrative tuberculosis appeared to be the major indicators multyrezystentnosti Office, it was found in 18 (60%) patients had monoresistance in 12 (40%) patients. With disseminated tuberculosis rates were almost identical: monorezystentnist in 10 (33,4%), multyrezystentnist in 20 (66,6%). The data obtained after treatment show the relevance of the application of molecular genetic methods in modern Tuberculosis. Increases the effectiveness of therapy (healing destructive changes in lungs,stopping bacteria).
Introduction
Recent years are characterized by a decrease the effectiveness of treatment of pulmonary tuberculosis,
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tuberculosis rates were almost identical: monorezystant in 10 (33,4%), multyrezystent in 20 (66,6%).
Table 1
Characterization of resistance to forms of the disease
Form Total TB Patients Monorezystant Patients Multyrezystant
Infiltrative n=30 30 12 18
Abs % 100 40 60
Disseminated n=30 30 10 20
Abs % 100 33,4 66,6
In patients with infiltrative tuberculosis dominated by complaints of cough 10 (33,4%), and weakness of 6 (20%). Some less worried about losing weight 2 (6,7%), fever 3 (10%) and stuffiness 2 (6,7%). In patients with disseminated process picture is different: complaints of cough 25 (83,4%), weight loss 21 (70%), fatigue 27 (90%). Some less worried fever 20 (6,67%) and stuffiness 10 (33,4%).
We studied the performance of ESR and WBC in both groups at the beginning of treatment. Shows that changes in the blood as accelerating SHOYE and leukocytosis, are identified more patients with disseminated process. SHOYE from 30 to 60 mm / h and above are also more common with this form. Acceleration SHOYE (30 mm /h) in 21 (70%) cases marked with infiltrative tuberculosis, disseminated - in 24 (80%), respectively.
Consider the prevalence of tuberculosis in the lungs. X-ray examination of patients showed that the prevalence of pathological process in the lungs was varied. The destruction was observed in all patients. When analyzing the table we can say that the infiltrative process was greatest prevalence of 1-2 cm in 25 (83,3%) with disseminated most <2cm 16 (53,4%).
Taken in the development of bacillary patients in whom bacterial necessarily confirmed by microscopic and culture results, followed by determination of susceptibility of Mycobacterium tuberculosis to anti-TB drugs. Comprehensive Assessment of massiveness bacteria according to dosage smear microscopy and by culture
with quantitative counting of colonies on nutrient dense medium was performed 60 patients. Microscopically poorly bacteria dominated in patients with infiltrative tuberculosis - 20 people (66,6%), somewhat less than it was observed with disseminated tuberculosis - 5 people (16,7%). Moderate in 9 (30%) and 14 (46,7%) patients, respectively, had a massive bacterial infiltration in patients with 1(3,4%) and dissemination 11 (36,6%), respectively. Poorly bacterial culture dominated in patients with infiltration - 17 (56,6%), somewhat less than it was observed with disseminated process - 4 (13,4%). Somewhat more moderate bacterial dissemination turned out - 14 (46,6%), with infiltration it was in 13 (43,4%). Massive bacteria were observed in patients with infiltration 0 (0%), and dissemination of 12 (40%), respectively.
In cultures of Mycobacterium tuberculosis isolated DNA were tested for susceptibility to isoniazidi and rifampicini molecular genetic methods. All patients studied specimens and isolated from her cult M. tuberculosis by rpoB locus and katG. According to the definition of mutations in genes responsible for sensitivity of Mycobacterium tuberculosis to the above mentioned anti-TB drugs intended regimens. Knowing the resistance of MBT, we initially treat increased dosage of fluoroquinolones (levofloxacini and Ethionamidi) including the time for oral medication. After 4 hours after taking the pill when bacteriostatic activity in the blood peaked, we introduced above mentioned drugs intravenously, thereby enhancing their properties.
Table 2
Chemotherapy regimen based on resistance to Mycobacterium tuberculosis TB drugs
Resistant Chemotherapy regimens
Intensive phase Maintenance phase
H R0,6+Z 2,0+E 1,2+S1,0+PAS 400,0 R 0,6+E 1,2+Ofx 0,8
R H 0,6+Z 2,0+ E 1,2 + PAS 400,0+Lfx 1,0 H 0,6+R 0,6+Z 2,0+ Et 1,0
H+R Z 2,0+E 1,2+K 1,0+PAS 400,0+Pt 1,0 Z 2,0+ E 1,2 +Ofx 0,8 + PAS 400,0+ Pt 1,0
The results are processed and analyzed by a computer program «MS Excel 2007". Also, analysis of the data was performed using software packages «Statistica 6.0». Data for the indicators are categorized in the text of the absolute values and for continuous parameters, standard methods of variational statistics: averages calculated RMS deviation and average values of arithmetic errors. Using the criterion Styudent estimated difference of averages in the two groups (p). Significant difference was considered one that was within the accuracy Styudent less than 0,05.
Results and Discussion
At the end of inpatient treatment were collected complaints in patients with both tuberculosis. With infiltrative form of complaints, none of the patients did not
show with disseminated complained of slight weakness only 1 (3,4%) patients. Zminy levels after treatment in the form of accelerated SHOYE and leukocytosis, were almost identical in both forms. SHOYE from 30 to 60 mm / h and was not measured. Acceleration SHOYE (15 mm / h) with infiltrative - in 28 (93,4%), 81,6% with disseminated ((p <0,05)). Slight leukocytosis was 1 (3,4%) patient with disseminated tuberculosis. The figure number 1 in the group with microscopically infiltrative process abatsylyuvannya achieved after 6 months of treatment in 15 (50%) with disseminated in 12 (40%) (p <0,05) patients. After 7-8 months abatsylyuvannya occurred in 24 (80%) of infiltration and in 20 (66,6%) (p<0,05) dissemination. At 9-10 months of treatment abatsylyuvannya stopped in all patients.
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□ Infiltrative tuberculosis
□Disseminated tuberculosis
20 10
6 m
7-8 m
9-10 m
Figure 1. Dynamics terms abacilates by microscopy in both groups at the end of treatment Culture abacilates stopped after 6 months of (90%) with infiltrative tuberculosis and in 22 (73,4%) (p treatment in the first 25 (83,4%) in the second 20 (66,7%) <0,05) with disseminated. At 9-10 months of treatment (p<0,05). After 7-8 months abacilates occurred in 27 abacilates achieved in all patients in both groups (Fig. 2).
L00n
90 80 70 60 50 40 30 20 10
□Infiltrative tuberculosis
□Disseminated tuberculosis
7-8 m
-10 m
Figure 2. Dynamics terms abacilates culture method in both groups at the end of treatment
In the first and second group after 4 - 6 months of treatment, abnormal shadows resorption destruction and healed in 20 (66,7%) and in 14 (46,7%) (p <0,05),
respectively. After 8-9 months of healing degradation was noted in all patients (Fig. 3).
0
npoSAeMH eKOAorii Ta MCJHUHHH
100n 90 80 70 60 50 40 30 20 10 0
□ Infiltrative tuberculosos
□ Disseminated tuberculosis
-r
4-6 m
8-9 m
Figure 3. Dynamics timing resolution of lesions and scarring destruction in both groups at the end of treatment
Conclusions
1. These figures indicate that the use of immunosuppressive therapy in newly diagnosed patients with destructive pulmonary tuberculosis after receiving data on resistance of Mycobacterium tuberculosis to anti-TB drugs can increase the effectiveness of treatment in terms of both abacilates phlegm and healing to destructive changes in the lungs, compared with treatment of patients with only minutes.
2. At the end of hospitalization in patients with complaints of both groups were as follows: in the study group complained of cough 2 patients (3,4%) in the control - 4 (6,6%) (p<0,05). Complained of weakness 1 (1,6%) patients of the group, and 2 (3,4%) (p<0,05) from the control group. Acceleration SHOYE observed in the study group in 7 (11,6%) in the control group in 14 (23,4%) (p<0,05). Leukocytosis occurred in 1 (1,6%) patients of the main group and in 3 (5%) (p<0,05) in the control. In the analysis of resorption lesions and scarring destruction after 6 months and at discharge, we received the following information. In the study and control group after 6 months of treatment resorption abnormal shadow in 56 (93,3%) and 52 (86,6%), respectively (p <0,05). After 9 months of destruction was scarring in 60 (100%) of the patients in both groups. Through 6 months abacilates by microscopy occurred in 50 (83,3%) in the intervention group and 46 (76,6%) (p <0,05 ) in the control. After 7-8 months of suspension of bacteria in 56 (93.4%) in the intervention group and in 51 (85%) in the control group (p <0,05). At 9-10 months of treatment abacilates achieved in all patients in both groups. Abacilates culture method in both groups are presented below. After 6 months abacilates occurred in 52 (86,6%) in the intervention group and 47 (78,4%) in the control group (p <0,05). After 7-8 months in the intervention group stopped microscopic bacteria in 57 (95%) patients in the control group in 54 (90%) (p <0,05). At 9-10 months of treatment stopped abacilates all patients in both groups.
3. Molecular genetic analysis of M. tuberculosis showed that all patients had resistance to isoniazidi and rifampicini. Patients who had resistance to isoniazidi was
more of disseminated tuberculosis in both groups, 7 (23,4%) and 4 (13,4%), respectively (p<0,05).
4. With infiltrative form of complaints nobody showed, with disseminated complained of slight weakness only 1 (3,4%) patients. Changes in the blood after treatment in the form of accelerated SHOYE and leukocytosis, were almost identical in both forms. Significant increase SHOYE not measured. Slight leukocytosis was 1 (3,4%) patient with disseminated tuberculosis. In the analysis of resorption lesions and scarring destruction, we obtained the following data. In the first and second group after 4 -6 months of treatment resorption abnormal shadow in 20 (66,7%) and in 14 (46,7%), respectively (p<0,05). After 89 months of healing degradation was noted in all patients. In the group with microscopically infiltrative process abacilates achieved after 6 months of treatment in 15 (50%), and disseminated in 12 (40%) patients (p <0,05). After 7-8 months abacilates occurred in 24 (80%) of infiltration and in 20 (66,6%), dissemination (p <0,05). At 9-10 months of treatment abacilates stopped in all patients. Culture abacilates stopped after 6 months of treatment in the first 25 (83.4%) in the second 20 (66,7%) (p <0,05). After 7-8 months abacilates occurred in 27 (90%) with infiltrative tuberculosis and in 22 (73.4%) with disseminated (p <0,05). At 9-10 months of treatment abacilates achieved in all patients in both groups.
5. Thus, the data above indicate that the proposed addition of immunosuppressive therapy and the use of molecular genetic method, after which you can assign the correct treatment, increase the effectiveness of treatment of newly diagnosed destructive pulmonary tuberculosis himiorezystentny.
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