Simultaneous TLR gene expression estimation (the prognosis of abortion and miscarriage), mucosal microbiocenoses types, mucosal immunity state estimation, presence or absence of clinical manifestation of miscarriage, pregnancy course (unfavorable prognosis-immunological rejection of the fetus or complicated pregnancy course and miscarriage, favorable prognosis-immunological irritation or normal fetal development, abortion-miscarriage), presence or absence of infection and/or clinical manifestation of infectious and inflammatory conditions can be objective additional criteria for estimation and prognosis of pregnancy course as well as to personalize the adequate treatment and prevention of abortion and miscarriage. Pathogenetic (concomitant increased reaction of mucosal epithelial cells receptors for developing fetus and infectious process leads to abortion and miscarriage) and TLR diagnostic role in pregnancy is established. In obstetric infections in pregnant women levels of gene expression of TLR-2 – 21.2 and above, TLR-4 – 23.0 and above, TLR-8 – 25.7 and above indicate a high degree of infection with clinical manifestations of urogenital infections, and in the levels of genes expression of TLR-8 - 26 and below verifies chronic urogenital infections, while levels of gene expression TLR-8 above 26 verifies acute course of urogenital infections TLR-8 level above 28 is a predictor of pregnancy termination and miscarriage.

Background and aims Innate protection recognising systems of human organism are important and perspective in respect of investigation autoimmune and infectious diseases. The aim was to evaluate lectin system (LS) of complement and its importance for innate interactome against infectious and autoimmune diseases. Methods Patient complement components (PCC) of sera were estimated by immunochemical methods in microplates (hybrid plate for C4A and C4B functionality testing) and on the blot (for PCC separated by isoelectrophoresis). Peroxidase activity was detected using TMB or chemiluminescent substrate (for blot, BioChemi System, UVP). Results Additional glycoconjugates(GC)-binding PCC were registered. C1-inh revealed affinity to heparin. Patient (SLE, antiphospholipid syndrome, rheumatoid artritis) subisotypes of C4B (up to 5) and C4A (up to 7) were observed as aggregated forms together with GC. Conclusions 1. Extended complement LS includes MBL, Factor H, C1-inh, CR1, CR2, CR3, C3, C4B, others. 2. Complement serve as a universal communicator among protective systems involving their LS—GC communications. 3. Complement (as mostly advanced innate protection system) possesses structure-function principles prognostic for any innate recognition systems. 4. There is superLS network in organism. Probiotic LS is important cofunctioning part of it. 5. The data support idea that any protection protein system partially functions involving LS—GC recognition (also for antibodies recognised by Fc-receptors as LS). 6. New prognostic-diagnostic possibilities in investigation of autoimmune and infectious diseases are opened using interactome LS—GC network.

Background and aims The aim was to summarise own data on the human complement system involving cofunctioning complement and other innate protective systems against infectious and autoimmunity diseases. Methods Components of the patient serum complement (CPSC) were registered by immunochemical methods in microplates (variants of functional analyses of isotypes C4A and C4B, and C1-inhibitor upon supramolecular assembling on well bottom) and on the blot (preliminary isoelectrophoresis of sera in the plate of polyacrylamide gel). Rabbit and goat polyclonal antibodies against purified CPSC were used. Activity of antibodies-peroxidase conjugates bound to CPSC was detected in the presence of TMB (microplate) or chemiluminescent substrate in a real time [BioChemi System (UVP)]. Results 1.Sera of patients having autoimmune diseases were characterised on the blot by appearance of aggregated C4B and C4A within pI 4,0–4,7 compared to less acidic free isotypes. Functional status of isotypes was confirmed in microplate. Absolute amounts of isotypes and their subisotypes as well as ratio of isotypes characterised prognostic-diagnostic patient groups of diseases (SLE, antiphospholipid syndrome, rheumatoid arthritis). Appearance and relative intensities of the system of aggregated isotypes and subisotypes of C4 indicated the presence of disease, its initiation, reached phase of disease and disease character. 2.Similar localization on the blot for the complex C4B and C1-inhibitor of patients was registered. Conclusions Results indicate possible cofunctioining C4B and C1-inhibitor in protection complement network upon development of autoimmune diseases. New mechanisms of cascade protection involving new combinations of CPSC may be revealed. Results open new practical possibilities in diagnostics of early, progressive and chronic autoimmune diseases.

Background Isotypes C4A and C4B of human complement component C4 are innate immunity factors. We found that gel electrophoresis of patient sera in the new diagnostic area of C4 complexes/aggregates (pI 4-5) followed by immunoblotting allowed evaluation of patient C4 isotype deficiency grouping in accordance to our developed functional C4 isotype analyses in microplates [1]. Solid phase peroxidase kinetic chemiluminescence (CL) in the presence of CL substrate is sensitive process to detect serum glycoproteins distribution. Objectives The objective was further study of patient C4B and C4A subisotypes within C4-isotype-(non-deficient or deficient) sera groups of patients. Methods C4 subisotype patterns were obtained using separation of desialylated sera by isoelectric focusing in plate of polyacrylamide in gradient pH 3-5 followed by gel blotting on Immobillon P (Millipore). Blots were treated with goat anti-C4 polyclonal antibodies conjugated to horse radish peroxidase (Ab-HRP) or Ab-HRP against human IgG, IgM or IgA. Protein-A–HRP was also used. Substrate (BioWest, UVP; with increased sensitivity) was added and kinetics of the blot HRP CL was stepwise registered in live imagination regime using BioChemi System (UVP). Subisotypes were revealed by blot treatment with acetate buffer pH 4.5. Analysis was especially effective in optimized by us conditions (desialylation control, blot washings, substrate using, Cl registration regimes). Results Patient C4B or C4A subisotypes were presented as combinations of mainly up to 5 bands of different contribution. Contribution of subisotype(s) in C4A region was confirmed by the presence of increased levels of IgG in less acidic band(s) and/or IgM as additional early CL of mostly acidic 5th band in cases of SLE with or without antiphospholipid syndrome (group of C4B-deficiency). In cases of non-C4-deficiency groups (examples of rheumatoid arthritis etc.) preferential localization of non-early IgM, IgA and binding Protein A within C4B region indicated infectious nature of contribution to subisotype(s) bands. Ig Fc-fragments were masked in complexes (C1q binding was not observed). Relative resistance of subisotypes patterns upon sera storing indicated partial masking glycans in C4 subisotypes complexes possessing increased hydrophobicity. Algorithm of revealing very low levels of serum subisotypes (invisible in comparison to other sera) is proposed. Conclusions Results support both biomarker diagnostic and prognostic potential of patient sera C4B and C4A subisotypes and prospects of its further development of fine resolution CL patterns of C4 associates together with increased levels of classes and subclasses of Ig and lipoproteins, glyco- and lipid-containing antigens (also of microbial origin), others sera components using extended set of Ab. Few diagnostic kinetic CL pattern states (early and later) are possible to characterize disease(s) type. References Lakhtin M et al. Lakhtin M et al. Chemiluminescent visualization and quantification of the complement glycoprotein C4 containing systems in patient whole sera separated with isoelectric focusing in polyacrylamide gel and electroblotting: sialidase- and peroxidase-based new approaches. Glycoconjugate J 2011; 28: 296. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.3511

Бактериофаги пробиотические средства регуляции мик робиоценозов и деконтаминации микроорганизмами продуктов питания, животных и растений В обзоре рассматриваются вопросы безопасного и эффек тивного применения бактериофагов в качестве пищевых добавок средств деконтаминации пищевых продуктов (на всех стадиях производства), животных, растений и пробиотических биологически активных добавок к пище, служащих дополнительным источником бактериофагов для организма человека, присутствие которых в нормоф лоре позволяет снизить риск развития у индивидуума та ких высококонтагиозных пищевых

Background Probiotic bacterial lectins (PBL) isolated by us from industrial strains of human origin are important regulators of microbiocenoses. The Aim is to evaluate potential of PBL in medicine. Results The data obtained indicate that preparations of PBL imitate general functions of probiotics, increase or create new useful activities. In biotopes PBL act as intrinsic tool of microbial selection, factor of supporting probiotic compartment. PBL can be used for screening probiotic microbes, sensitive and resistant clinical strains, and also for monitoring balance of biocenosis in directions “Microbes - Microbes” and “Microbes - Organism”. Application of PBL (combinations of L of lactobacilli and bifidobacteria [LL and LB] as the model functional imitator of biotope probiotic compartment) is multidirected and use antipathogen cascade multisynergism of PBL in time and in space. Results indicate that PBL are able to destruct biofilms of pathogen combinations “Staphylococcus > or < Candida” (synergistic contribution: LL > or < LB). LL+LB possess advantages of probiotic action in cases of biotopes where bifidobacteria or lactobacilli are currently absent. It seems, being delivered into organism, PBL will complete cell system “Lactobacillus + Bifidobacterium + Other probiotic-like microbes” and increase synbiotic biotope ingredients. The data on synergistic action between PBL and antibiotics allow using PBL in cases of non-effectiveness of antibiotics or for decreasing antibiotic doses. Conclusions Results indicate prospects of PBL in supporting probiotic and synbiotic systems of organism for example during chemotherapy and radiotherapy of tumors, or as ingredients of anti-infectives against eukaryotic pathogens (microfungi and protozoans).

Background Probiotic human bacterial mixtures constructed are important for health [1]. The aim was to develop new perspective variants of the known Russian mixed probiotic “Acilact” (Lactobacillus strains NK1+K3III24+100ash; all strains of human origin) using approach of ranging new parameters of cultural fluids, their fractions and components. Methods The system of at least 32 parameters was ordered as decreased/increased sequences of 4 members (Acilact and its strains) depending on any selected parameter. The cases of leadership (inverted or not in the sequence) of Acilact or strain(s) are considered as technological advantages of consortium or strain(s). Results and discussion Among high molecular mass components (at least 30 kD), protein fractions, lectin systems, protease system, oxidase-reductase system, biosurfactants, intrinsic fluorofores containing protein and non-protein components, and activities against eukaryotic human pathogens were evaluated as perspective new features and criteria for construction of new system probiotics of differentially directed action. Non-linear proportional contributions of ingredient strains into final properties of Acilact were observed. Strains as dominant leaders in some features revealed in Acilact can be detected. Examples of significantly new advantages of ingredient strain(s) or Acilact are presented. The net of ordered ranges allows additional control of the results. Conclusion The approach developed can help in constructing qualitatively new effective and directed mixed extended probiotics based on traditionaliy used probiotic bacterial industrial strains. Reference [1]. V.V. Pospelova. Thesis (MD) [Microbial biological preparations for correction of intestinal bacteriocenoses, their designing and using (in Russian)]. Moscow. 1979.

Background: The urgency of the pertussis situation nowadays is caused by the remaining lethality and involving in epidemic process of primary school age children. Methods: Epidemiological, serological, microbiological, molecular-genetic. Results: The occurrence rate of pertussis in the RF between 2008-2011 stabilized at 2.5-3.8 per 100 000 people, including 1-4 lethal cases, mainly among children under age one. The severity of the illness depends on anatomic and physiological features of an organism, immunity of the individual, accompanying infectious diseases, as well as the agent's biological features. The distribution of the disease in Russia's 66 regions is sporadic. 9 regions show 5-10 cases per 100 000; in another 8 regions the rate is practically nil. In 2011 infection sources were established in 11,6% of cases. It is presumed that the morbidity rate is underestimated. Pertussis is registered in both non-vaccinated and vaccinated persons. The highest occurrence rate of vaccinated patients occurs in the age category of 6-10, which suggests a reduction of immunity to pertussis among children of this age, and leads us to question the appropriateness of the second revaccination at the age of 6. According to the WHO, many countries revaccinate a second and third time. Most of the modern-day circulating strains, as compared to the strains included in the current DPT vaccine, have a number of changes in their proteins, which determine the pathogenic and immunogenic features of the microbe. To construct a new pertussis vaccine, including an acellular vaccine, 3 candidate strains have been selected (together with I.I. Mechnikov's Research Institute for Vaccines and Serums of the Russian Academy of Medical Sciences). Being aware that laboratory diagnostics for pertussis require significant improvement, a rapid diagnosistic method based on loop-mediated isothermal amplification has been developed (LAMP). The LAMP does not require expensive equipment and the accuracy rate is 72.7% - 100%, depending on the terms and stage of the disease. Conclusion: It is necessary to develop a set of measures aimed to improve specific prophylaxis of pertussis, and the inclusion in healthcare practice of highly effective molecular - genetic methods of lab diagnostics.

Background Probiotic bacterial lectins are important factors in biotope. The aim was to study autoregulation properties of lectins of lactobacilli (LL) isolated by us [1]. Methods Clinical strains of lactobacilli were freshly isolated from urogenital tract samples of young and adolescent patients using growth on Lactobacillus MRS Agar (HiMedia) and variant of Shaedler medium. Additional differential indication media were used. Bacterial growth was studied in the presence of disc- or drop- applied of LL on solid media or LL within bacterial suspensions (0.5–5 McFarland units). Samples studied were characterized with lactobacilli at the level of 106–108 Cfu/ml, and did not contain a panel of standard bacterial and viral pathogens. Decreased contents of Staphylococcus ssp., Enterococcus ssp., E. coli, and Candida ssp. were registered. Results All methods used gave similar results (supporting each other) of action of LL on lactobacilli populations growth. The following groups of lactobacilli populations were identified: LL-resistant or sensitive, weakly suppressive or stimulated, highly or moderate suppressive, highly modulated, cell condensation depended or not. Conclusions Results indicate further evidences that probiotic LL imitate cellular probiotics. It seems LL are involved in signal events (QS, cross-talking) in biotope. LL may act as a biotope tool of microbiota selection and may be used for both supporting probiotic-like lactobacilli and simultaneous elimination of other lactobacilli possessing decreased survival (non-probiotic-like strains) in conditions of healthy or close to normal balance in biotope. Reference [1]. V. Lakhtin, M. Lakhtin, V. Pospelova, B. Shenderov (2006) Microb. Ecol. Health & Dis. 18: 55–60.

Successful implementation of the Russian Federation's national measles elimination program has been ensured by high vaccine coverage (>95%) of the target population (with 2 doses of measles-containing vaccine), case-based laboratory investigation of measles, and active surveillance of measles cases among patients with rashes and/or fever. As a result, the incidence of measles has decreased to <1 case per 1,000,000 population (2007-2009); no circulation of D6 genotype, which was endemic in the Russian Federation, has been observed since the second half of 2007; and the proportion of imported measles cases of different genotypes increased to 19.8% in 2007 from 1% in 2003. To confirm successful elimination of indigenous measles, a documentation system was initiated in the Russian Federation.