[Problems of diagnosis of the hepatitis B virus].

The recent data on the genetic variability of the hepatitis B virus and problems of serological ELISA-mediated detection of this virus are discussed.

[Comparative evaluation of effectiveness of traditional serologic and modified methods of herpes zoster laboratory diagnostics].

AIM: Comparative evaluation of effectiveness of traditional serologic and modified diagnostic methods of disease arising due to varicella and herpes zoster virus (VZV) reactivation. MATERIALS AND METHODS: 2 groups of patients were examined. The main group consisted of 39 patients with manifest form of herpes zoster (HZ), control--20 healthy donors. Sex composition of the groups did not differ. Traditional method of serologic diagnostics included determination of anti-gE VZV IgG and anti-VZV IgG and anti-IgM in patient and donor blood sera by using EIA. Modified methods consisted of isolation in density gradient and cultivation for 48 hours of peripheral blood mononuclears (PBMC) in RPMI-1640 complete culture medium containing 10% of fetal bovine serum, 4 mM L-glutamin and gentamycin. Concentrations ofanti-VZV IgG and IgM were then determined in culture medium by using EIA. RESULTS: In all the examined HZ patients and healthy donors anti-VZV IgG were detected in blood. Only in 26 (67%) of 39 HZ patients anti-gE VZV IgG and anti-VZV IgM were determined in blood sera. Among donors false positive results for these markers were detected in 10% and 5% of cases, respectively. During simultaneous determination of anti-gE VZV IgG and anti-VZV IgM the specificity of the method increased to 100%, sensitivity of the diagnostic method based on simultaneous determination of anti-gE VZV IgG and anti-VZV IgM was 59%. During analysis of spontaneous production of anti-VZV antibodies by PBMC in 38 (97.4%) of 39 patients anti-VZV IgG were determined in PBMC culture, anti-VZV IgM production was observed only in 4 patients. In control group false positive results of anti-VZV IgG and IgM production by PBMC was not detected by the modified method (100% specificity). At equal specificity level sensitivity of the modified method based on determination of spontaneous anti-VZV IgG production by PBMC culture was significantly higher than effectiveness of the traditional serologic diagnostics (97.4% and 59%, p < 0.0001). CONCLUSION: The data obtained allow to recommend during diagnostics of manifest and atypical VZV infection forms arising due to endogenous virus reactivation the new modified method of laboratory diagnostics of the disease as having higher sensitivity compared with traditional serologic method.

[Heterogeneity of hepatitis B virus and diagnostic potential of modern test systems for the detection of HBsAg].

AIM: Study heterogeneity ofhepatitis B virus in adult patients with chronic hepatitis B and determination of diagnostic potential of modern test systems with the detection of HBsAg with amino acid substitutions in the main hydrophilic region (MHR). MATERIALS AND METHODS: In 27 hepatitis B virus samples isolated from patients with chronic hepatitis B virus infection living in Vladimir, nucleotide sequence ofgenome region corresponding to preS1/preS2/S genes was determined. RESULTS: In all of the 27 isolates genotype D virus presented by 3 subgenotypes D1, D2, D3 was detected in 18%, 26% and 56% respectively. Based on the distribution of nucleotide substitutions in the compared functional regions of hepatitis B virus (virus entry into the cell coding site (2875 - 2991 n.b.), pre-S2/S promoter region (2994 - 3171 n.b.), 5'-end pre-S2 and S-genes sequences (3172 - 154 n.b. and 155-455 n.b.), MHR (455 - 635 n.b.) and 3'-end S-gene sequence (636 - 835 n.b.), substitutions are mostly concentrated in the promoter region of the S2/S-genes (30.8%). HBsAg serotypes were determined in 24 of 27 cases by using the predicted amino acid sequence, and in 17 cases HBsAg belonged to ayw2 (71%) serotype and in 7 cases - to ayw3 serotype (29%). Amino acid substitutions G145A, M133I, S132T localized in the main hydrophilic region and P217L, S207N, V184A localized in the C-end of the protein C that are connected with diagnostic and vaccine escape were identified in 5 isolates. CONCLUSION: Diagnostic potential of test systems with the detection of HBsAg with known amino acid sequence of the MHR region were studied. Approximately equal potential of 6 test systems to detect HBsAg with amino acid substitutions G145A, M133I and S132T localized in the MHR region were shown.

[Hepatitis virus genotype structure in patients with chronic hepatitis C].

AIM: Studies of hepatitis C virus (HCV) genotype and subtype structure in patients with chronic hepatitis C in 3 regions of the Central federal district of Russia. MATERIALS AND METHODS: Hepatitis C virus genotype and subtype structure was determined in patients with chronic HCV infection in Moscow (1993 - 1995 and 2005), Moscow region (2008) and Vladimir region (1993 -1995, 2005-2007). HCV genotype was determined by using A. Widell et al. (1994) technique, PCR (AmpliSens diagnostic kits), Genotype C test system. RESULTS: In all studied regions and during all the time periods the first position in rating belonged to HCV 1b subtype. In 1993 - 1995 and 2005 - 2007 period changes in HCV genotype and subtype structure were registered that consisted of relative weight of 1b subtype decrease and 3a subtype increase. Subtype 1b in females with chronic hepatitis C was registered more often than in males. In Vladimir region 3a subtype in males was detected more often than in females. In males older than 30 years the first rating position belongs to 1b subtype and in males younger than 30 years--subtype 3a. In females older than 30 years in Moscow region and Vladimir region, as well as in females younger than 30 years in Vladimir region subtype 1b was detected more often, while in Moscow region HCV subtypes 1b and 3a were detected with the same rate of 47.6%. CONCLUSION: Currently there is an urgent need to include mandatory monitoring of hepatitis C virus genetic variants into the system of hepatitis C epidemiologic control in Russia. This approach will allow for a significant increase in quality of hepatitis C serological diagnostics, and can be used in the prognosis of evolution of the epidemic process of this disease.

[Impact of changes in the amino acid sequence of the "a" determinant on the diagnostic capacities of test systems for the detection of HBsAg].

The diagnostic capacities of 4 commercial test systems were comparatively estimated for the detection of HBsAg, by applying a panel of samples with the established amino acid sequence of the "a" determinant of HBsAg. The Roche Elecsys HBsAg test system demonstrated the highest sensitivity - the maximum HBsAg concentrations were found in 19 of 31 cases. Escape mutations in the major hydrophilic region (MHR) of HBsAG (P120S, M133T) were responsible for differences in the sensitivity of 4 test systems by 10- to 40-fold. There were also samples that showed differences in the diagnostic capacities of the test systems to detect HBsAg, but without amino acid replacements in the area of the "a" determinant, which seems to be associated with amino acid replacements in other regions of HBsAg.

[Evaluation of the efficiency of external assessment of the quality of determining antibodies to hepatitis C virus in the network of screening laboratories].

A regional external quality assessment (EQA) system for determining anti-hepatitis C virus (HCV) was developed and introduced. For this, a control panel comprising 14 samples with anti-HCV and 6 samples without anti-HCV was tested in the reference laboratory of Moscow Infectious Hospital One. A total of 6 sessions were conducted with the participation of 8 laboratories. The regional EQA system covering a limited number of laboratories was shown to significantly improve the quality of detection of hepatitis C virus antibodies. The optimal multiplicity of sessions for the developed EQA system was ascertained to be once fortnight. It is possible to define the current rating of screening laboratories in the sensitivity criterion and to elaborate address organizational-and-methodic measures according to the results of control tests.

[Blood levels of HbsAg in patients with different variants of chronic HBV-infection].

AIM: To estimate HbsAg in patients with different variants of chronic HBV infection. MATERIAL AND METHODS: Assay of HbsAg (IU/ml) in blood serum was made in 156 patients with chronic HBV infection (70 males and 86 females, age 19 to 78 years) using the test-system HbsAg Architect Lot 59665LF00 (Abbott) on the automatic analyzer Architect with construction of 4-parameter logistic curve. RESULTS: There are significant differences in the levels of HbsAg depending on the course of chronic HBV-infection: inactive carriers of HBV (12,884.14 +/- 5,512.26 IU/ml) had much lower blood levels of HbsAg than patients with HbeAg-negative (66,992.28 +/- 25,908.74 IU/ml) and HbeAg-positive chronic VHB (135,039.3 +/- 48,127.06 IU/ml) patients with chronic mixed hepatitis (82,783.12 +/- 21,001.34 IU/ml) and cirrhosis of HBV-etiology (67,477.86 +/- 24,081.9 IU/ml). No significant differences were found between two subgroups of pregnant women with or without viremia by HbsAg concentration in the blood. Maximal mean content of blood HbsAg was registered in patients with HbeAg-positive chronic hepatitis B. CONCLUSION: Significant differences in blood serum levels of HbsAg exist between patients with chronic viral hepatitis B and inactive carriers of HBV.

[The spread of hepatitis B and C viruses and the structure of HBsAg subtypes in Kabardino-Balkaria].

A seroepidemiological study of the spread of hepatitis B and C viruses (HBV and HCV) was conducted among some population groups in Kabardino-Balkaria. The structure of HBsAg subtypes was also studied in the residents of the republic. The presence of viral hepatitis B markers among the test groups of the healthy population corresponds to the parameters of moderate activity of an epidemic process. The analysis of the immunological structure of the population leads the author to assign Kabardino-Balkaria to highly HBC endemic areas. The occupational factor is demonstrated to be actively involved in the spread of HBC and it is practically of no significance for HCV. An examination could reveal the HBsAg subtype adrq+ that is uncharacteristic of this area.

[Prevalence of HBV and HCV markers in patients of specialized medical institutions].

240 medical histories of patients admitted to Moscow clinical narcological hospital No. 17 with "opium addiction" diagnosis in 2000 and 2003 (120 histories for each year) were analyzed. From 1997 to 2003 4220 patients registered in narcological dispensary of Cherepovets city (Vologda region) were tested for HBsAg and anti-HCV. It has been shown that in new injection opiate drug users young patients (15 - 19 y.o.) predominated (41.7%). Tendency to development of drug addiction in younger age was revealed - in 2003 compared to 2001 percent of new injection drug users aged 11 - 14 years admitted to hospital raised in 1.5 times. In 2000 and 2003 in Moscow markers of HIV and/or HBV/HCV infection were detected in 84.2 and 91.7% of injection drug users respectively. Maximal rates of HBV and HCV infection in injection drug users in Cherepovets city were detected in 2001 - 11.7 and 83.1% respectively. Leading role of injection drug users in maintenance of HBVand HCV epidemic process activity in Russia was established.

[Evaluation of the immunogenic and reactogenic properties of vaccine "Euvax B" in persons aged 16-18 years in different geographical zones of Russia].

The comparative evaluation of the reactogenic and immunogenic properties of vaccine "Euvax B" in adolescents aged 16-18 years was carried out in factory housing estate Lvovsky, the Podolsk district, the Moscow region, and in the Yamalo-Nenets Autonomous District (AD): in towns Salekhard, Noyabrsk, Muravlenko, in housing estate Tarko-Sale. The evaluation demonstrated that the use of this vaccine in a dose of 20 mg/ml produced no postvaccinal complications and the frequency of postvaccinal reactions was insignificant. Postvaccinal anti-HBs were detected, on the average, in protective concentrations in 98.4 +/- 1.8% of the vaccines (99.0 +/- 1.9% in the Moscow region, 97.7 +/- 3.2% in the Yamalo-Nenets AD). In most of the vaccinated adolescents high concentrations of postvacinal anti-HBs were detected: in 51.4 +/- 9.7% of the vaccinees in the Moscow region and in 89.7 +/- 6.4% in the Yamalo-Nenets AD they exceeded 1,000 I.U./lit. The conclusion was made that in the territories located in subarctic latitudes vaccinal prophylaxis against hepatitis B was safe and highly effective and have no limitations connected with the conditions of the Far North.

[Morbidity in acute virus hepatitis B on the territory of Kabardin-Balkaria].

Morbidity in acute virus hepatitis B (AVHB) in Kabardin-Balkaria during the period of 1992 to 2003 was analyzed. The dynamics of changes in the age groups of AVHB patients, as well as in the structure of the transmission routes of the disease, was analyzed. The level of AVHB morbidity in the Kabardin-Balkar Republic was lower in recent years than the average level of such morbidity in the whole of Russia. The sharply defined irregularity in the territorial distribution of AVHB cases was established. The highest morbidity rates in AVHB were registered in Nalchik, as well as in Chegem and Prokhladnensk regions. The leading role in the formation of the morbidity in acute virus hepatitis B on the territory of Kabardin-Balkaria belongs to Nalchik, where 56.8% of AVHB cases were registered. In the structure of the transmission routes of AVHB the prevalence of artificial paths was noted; among them, the highest proportion belonged to parenteral medical manipulations in outpatient clinics (32.9%). The proportion of AVHB cases associated with the intravenous use of drugs was 6.9%. In the age structure of AVHB patients adults prevailed, and among them the highest number of cases was registered in the age groups of 20 - 29 years and 30 - 39 years. In 2002 the total proportion of AVHB cases among the patients of these age groups reached 68.3%.

[Immune status of patients with different outcomes of acute hepatitis C].

AIM: To ascertain parameters of immune response significant for prognosis of the disease outcome in patients with acute hepatitis C (AHC). MATERIAL AND METHODS: Seventy two examinees were divided into three groups: 30 patients of group 1 had AHC; 29 patients of group 2 had chronic hepatitis C (CHC) with the disease history 3-10 years; 13 AHC convalescents who had the disease 3-10 years ago. The control group consisted of 10 healthy subjects. The following parameters of immune status were studied: CD3+, CD4+, CD8+, CD16+, CD20+, HLA-DR+CD3+, CD95+, O-lymphocytes. RESULTS: By clinicobiochemical picture and the results of 12-month follow-up RT-PCR investigations of blood HCV RNA, AHC patients were divided into two groups: in 18 (72%) of 25 patients AHC became chronic, 7 (28%) patients achieved convalescence. The proportion and absolute number of subpopulations of lymphocytes CD8+, CD20+, CD16+ as well as CD4/CD8 in patients with AHC with different outcomes did not differ from the controls. AHC convalescents and patients with AHC transformation into chronic hepatitis had in the acute period of the disease much higher absolute number of subpopulations of the lymphocytes CD3+, CD4+, HLA-DR+CD3+ vs the controls. In patients with chronic hepatitis C with elevated levels of AlAT, 3-10 year follow-up registered significantly higher absolute amount of CD8+, CD3+, CD4+, HLA-DR+CD3+ vs the controls. Only patients with AHC which later became chronic had for 6 months of the disease significantly elevated absolute number of O-lymphocytes vs the controls. CONCLUSION: A high absolute count of O-lymphocytes in AHC patients may indicate probable development of chronic hepatitis C.

[A comparative evaluation of detection of antibodies to hepatitis C virus in different immunoassay test system and verification tests]. / Sravnitel'naia otsenka rezul'tatov opredeleniia antitel k virusu gepatita C pri ispol'zovanii razlichnykh immunofermentnykh test-sistem i podtverzhdaiushchikh testov.

Control tests and subsequent comparative analysis of their results related with the detection of anti-hepatitis C virus (anti-HCV) by different ELISA systems were made at a laboratory of the "Vector-Best" Company, Novosibirsk (A), a clinical diagnostic laboratory of Infection Clinical Hospital No. 1, No. 1 (B), and at a laboratory of chronic viral infections of Andjaparidze Institute of Viral Drugs of the Russian Academy of Medical Sciences (C). Two thousand three hundred and fifty blood sera of donors and 236 blood sera of patients with acute and chronic renal pathology were examined. The comparative research made at the above facilities by different ELISA systems produced diversified results. An analysis of 2350 blood samples from donors made in 3 ELISA systems at laboratory A showed divergence in 19 (0.8%) cases. At laboratories B and C, 5 ELISA systems (a set of 236 samples) denoted differences in 28 (11.9%) cases. Significant variances were registered in confirmation of disputable results of ELISA with immunoblot, which is apparently related with differing criteria applicable to confirming the positive results from using the confirmative tests. A possibility is discussed of using different ELISA systems in screening examination.

[Viral hepatitis in different population groups of Republic of Sakha (Iakutia)]. / Shirota rasprostraneniia virusnykh gepatitov sredi razlichnykh grupp naseleniia na territorii respubliki Sakha (Iakutia).

A survey for the presence of markers of viral hepatitis B, delta, C and E among different groups of the population of the arctic and agricultural zones of the Republic of Sakha (Yakutia), as well as in Yakutsk, was carried out. The survey revealed that viral hepatitides with the parenteral mechanism of the transmission of infective agents were widely spread among the population. In the arctic zone HBsAg was detected in persons aged 20 years and older in 10.8-17.1% of cases. On the average, in the arctic zone antibodies to virus delta were detected in 2.4% of cases. In different regions of the agricultural zone the detection rate of HBsAg was also high (10.4-23.8%). In the Namsk and Vilyui regions delta infection was registered among adolescents (15-19 years of age) in 8.7 and 22.4% of cases respectively. In the Mountainous region anti-delta antibodies were detected among persons aged 20 years and older in 31% of cases. The survey showed the presence of considerable differences in the detection rate of the markers of hepatitis B, depending on the ethnic origin of the examined patients. Similar tendencies were found to exist with respect to delta infection.

[Evaluation of morbidity and etiological structure of acute and chronic viral hepatitis in the Sakha Republic (Yakutiia)]. / Otsenka zabolevaemosti i étiologicheskoi struktury ostrykh i khronicheskikh virusnykh gepatitov na territorii Respubliki Sakha (Iakutiia).

The morbidity of viral hepatitis was epidemiologically analyzed and the etiological structure of severe and chronic viral hepatitis was studied in the Republic of Sakha (Yakutia). Acute viral hepatitis B, whose portion is 43.5%, was found to be of the key significance within the structure of acute viral hepatitis (AVH). Acute viral hepatitis C (with 4.7% in the AVH structure) is by far more rare. The latter is close to the value registered in other Russia's regions. Apart from the prevalence of viral hepatitis B (32.4%) an exceptionally high share of chronic hepatitis delta (24.5%) was established for the etiological structure of chronic viral hepatitis, which is a specificity of the discussed region. The epidemic situation in the Republic of Sakha (Yakutia) was stated to be linked with the mentioned extra dangerous diseases, which requires appropriate emergency measures with the prevention vaccination program against hepatitis B being a priority.

[Experience with vaccine prophylaxis of hepatitis B among different groups of children]. / Opyt vaktsinoprofilaktiki gepatita B sredi razlichnykh grupp detei.

The immunological activity of different vaccines against hepatitis B was evaluated in the Hepatological Center organized on the basis of the Infectious Hospital in Tula. Newborn infants were immunized with the use of the following vaccines: Engerix B (Belgium), Combiotech (Russia), Euvax B (Aventis Pastèur, South Korea). Altogether, after the full course of immunization anti-HBs were detected in 76 children out of 81 (in 93.8%). Vaccine Engerix B, when introduced according to the schedule 0-1-2-12 months, exhibited high immunogenic properties in a group of infants born of women with persistent HBs-antigenemia. Anti-HBs at a concentration exceeding 1000 I.U./I could be detected in 84.6%. In another group of children immunized according to the schedule 0-1-6 months first with vaccine Combiotech at the age of 0 and 1 month, then (at the age of 6 months) with vaccine Euvax, the presence of postvaccinal anti HBs at protective concentration was registered in all children. After immunization against hepatitis B with the use of all above-mentioned vaccines introduced according to both schedules high immunological activity and safety of immunization were noted.

[Evaluation of the system for controlling the quality of the HBsAg detection in the system of screening laboratories]. / Otsenka sistemy kontrolia kachestva detektsii HBsAg v seti skriningovykh laboratorii.

The two-stage control system, ensuring the high quality of serological investigations in the network of screening laboratories in Moscow was developed and introduced into practice. At the first stage the entry control of the quality of the test system for the detection of HbsAg, coming to the screening laboratories, is made in the reference laboratory with the use of specially developed "representative" panels, as well as the test systems for comparison ("reference" test systems). Then "minipanels" are formed from specimens included into the "representative" panels, which are used for the evaluation of the quality of laboratory investigations made in the screening laboratories. The high quality of such system for controlling the quality of the detection of HBsAg in the screening laboratories of Moscow is shown.

[Propagation of hepatitis E virus in an endemic and nonendemic regions]. / Osobennosti rasprostraneniia virusa gepatita E v éndemichnom i neéndemichnom regionakh.

The role of hepatitis E (HE) in sporadic morbidity at an endemic territory (Southern Uzbekistan) and the incidence of anti-HEV in different populations of a nonendemic region (Russia) were evaluated. Antibodies to HEV were detected in 22.1% of patients with acute HE, including mixed infections (+ HA or HB), in the Dekhkanabad district of Kashkadarya region in 1993. The estimated incidence of acute HE was 51.7 per 100,000 population. Analysis of monthly incidence of acute HE demonstrated a seasonal pattern of the morbidity: more than 80% of total recorded cases occurred in August-September. These data indicate the presence of group HEV infections and an important role of this infection in the structure of acute viral hepatitis at the endemic territory. Anti-HEV were found in some population groups at a nonendemic territory: in free-of-charge blood donors in Surgut (4%), in patients with HIV infection (1.6%), and in medical workers in Moscow (1.1%).

[Spread of hepatitis C and separate genotypes of hepatitis C virus in a region with a moderately active epidemic process]. / Rasprostanenie gepatita C i otdel'nykh genotipov virusa gepatita C v regione s umerennoi aktivnost'iu épidemicheskogo protsessa.

The prevalence of hepatitis C virus (HCV) is studied in different age and risk groups in the city of Vladimir and the genotypes of the virus circulating at this territory are determined. Anti-HCV are detected in 2.4% donors and 1.4% pregnant women. Population immunity to HCV gradually increases with age. AntiHCV are detected in 0.9% children aged 3-6 years, in 1.8% in the group aged 20-29 years, and in 2.9% in the group aged 30-39 years. Medical workers, mental inpatients, hematological patients, patients hospitalized at hemoperfusion departments, and imprisoned subjects were referred to groups at risk of HCV infection: antiHCV were detected in 3.9, 5.4, 13.8, 53.6, and 24.5% of these groups, respectively. Genotype 1b predominates in the structure of HCV genotypes circulating in Vladimir: 72.9% in all examined population groups. In addition to this genotype, genotypes 2a, 2b, and 3a were detected (2.9, 2.9, and 15.7%, respectively). Genotype 3a was the most incident in children aged 0-14 years.

Evaluation of carcinogenic risk in friction product workers.

Mortality rates of two cohorts of asbestos friction product (FP) workers were studied in comparison with the population of the adjoining towns over periods of 20 and 40 years respectively. The second cohort was subdivided into 3 subcohorts exposed to chrysotile asbestos (CA), vulcanization and/or polymerization vapours and gases (VPGV) and asbestos bakelite (AB) or asbestos rubber (AR) dusts. In the first cohort no deaths from lung cancer were recorded, even though the total tumour mortality was higher than in the general population. In the second cohort an excess tumour mortality was observed in the first subcohort for stomach cancer only. In the other two subcohorts the expected rates were higher for all the tumour sites. Following intraperitoneal injection of CA, AB and AR dusts, malignant tumours developed in 31.5% of the rats which received CA, and in about 10% of the rats which were injected with AB and AR dusts. Also, the mean longevity of CA-treated animals was significantly lower than in the other two groups. Thus, the carcinogenic risk was real in the group of FP production workers with significant CA exposure only.