The causes of emergence and development of pneumonic plague outbreak in 2017 on Madagascar were analyzed. Summarized were the literature data, characterizing the spatial and biocoenotic structure of natural plague focus on Madagascar, assessed were the risk factors of plague infection among rural and urban population of the island, as well as the properties of Yersinia pestis strains isolated there in the XX century. It is substantiated that the distinctive type of plague outbreaks on Madagascar is a rural one. It is demonstrated that one of the main causes of wide spread of plague in 2017 is importation of the infection from central regions of the Republic of Madagascar into big cities with further formation of multiple epidemic foci. Confirmed was the circulation of Y. pestis strains that belong to the third branch of irradiation of oriental biovar (1.ORI3). Discussed are the issues of diagnostics and treatment of plague in epidemic foci in the territory of the Republic of Madagascar. The evidence is provided to the fact that the plague outbreak in 2017 on Madagascar is the consequence of a lower surveillance level, first and foremost undervaluation of specific and non-specific prophylaxis capacities regarding this infection.

Objective of the study is to generalize epidemiological data and assess the situation in the countries of East-Mediterranean region on infectious diseases that require the measures on sanitary protection of the territory of the Russian Federation, CIS member-states, and Eurasian Economic Union. Materials and methods. Epidemiological analysis was carried out using the data of official websites and periodical publications of WHO, WHO East-Mediterranean Office, National Health Ministries, Center for Disease Control and Prevention, other international organizations and open-access literature sources. Results and conclusions. Systematized and summarized have been the data on morbidity rates and spatial dissemination of diseases in each particular country from the point of view of possible risks to visiting them individuals. The data presented on the infectious diseases allow for addressing the issues associated with the risk of infection; identification of the factors and terms of high risk of infection; for forecasting the probability of disease importation into the Russian Federation. The information may prove to be useful for quantitative and qualitative evaluation of potential epidemic hazard in the course of sanitary-epidemiological welfare provision under international mass events.

Objective of the study is to assess the effectiveness of the methods for molecular diagnostics and identification of Yersinia pestis strains in the field material obtained from Sarydzhas high-mountain focus in Kyrgyz Republic. Materials and methods. Investigated were the samples of the field material, isolated in 2016 in Sarydzhas high-mountain plague focus, using conventional methods of laboratory diagnostics and PCR with hybridization-fluorescent and electrophoretic registration of results. Results and conclusions. It is demonstrated that in a number of cases molecular-genetic method has a higher resolution as compared to conventional methods of laboratory diagnostics of plague. It proves the necessity of wider usage of molecular-diagnostic methods in epizootiological monitoring of plague in natural foci in Kyrgyz Republic.

Objective of the study is to evaluate the serological activity of rabies virus antigens isolated from the brain tissue of mice by homogenization on FastPrep followed by ultracentrifugation. Materials and methods. Producer strain of the rabies virus “Ovechiy” GNKI. The rabies virus was isolated from the brain tissue of experimentally infected mice, followed by the study of the electrophoretic profile. The serological activity of the virus components was assessed by immunoblot and ELISA using specific anti-rabies sera.
Results and conclusions. In the course of comparing the methods of isolation and purification of the rabies virus antigen, it was found that most optimal one is to use a homogenization on FastPrep-24, followed by fractionation in a sucrose gradient. As a result of fractionation in a graded sucrose density gradient with a concentration of 15–50 % at 25000 g for 120 min, five fractions of the rabies virus components were obtained. The maximum purified protein fraction was from 15–20 % sucrose zone, which corresponded to a molecular weight of 67 kDa. The specific antigen activity of the fraction in ELISA reached up the titers of 1:1280 (Specificity coefficient 2.2).
Using immunoblot of antigens, obtained from the sucrose gradient in the range of 40–45 % and 20–35 % after ultracentrifugation, one major fraction of polypeptides (54 kDa) was detected, which showed the highest antigenic activity. The results obtained will be useful in the design of test systems for rabies screening and monitoring the effectiveness of anti-epizootic measures.

Objective of the study is to construct recombinant E. coli strain – producer of TcpA protein of cholera agent El Tor, carrying tcpACIRS gene in its genome, and use the strains for antigen production. Materials and methods. Utilized was non-toxigenic genovariant strain of Vibrio cholerae biovar El Tor from the “State Collection of Pathogenic Bacteria” at the premises of RusRAPI “Microbe”, as well as commercial E. coli strains and plasmids for cloning (Invitrogen, USA). Chromosomal DNA from V. cholerae cells was extracted using Charge Smitch gDNA Mini Bacteria Kit applying nucleo-sorption. To extract plasmid DNA from E. coli cells PureLink Quick Plasmid DNA MiniprepKits were used. The presence of tcpACIRS gene was assayed by PCR, using designed through our own efforts primers. DNA fragments were isolated from agarose gel with the help of PCR Clear-Up-System panel. SDS-PAGE was performed according to U.K.Laemmli method. Protein content of samples was measured by M.M.Bradford method. The panel for affinity chromatography was applied for recombinant TcpA protein purification. Results and conclusions. Constructed safe strain of E. coli is the producer of recombinant TcpA protein, basic subunit of toxin-coregulated pilus of adhesion of cholera agent biovar El Tor. The region of tcpA gene of Vibrio cholerae biovar El Tor was cloned as part of vector plasmid pET302 by the restriction sites XhoI-BamHI in E. coli strain BL21(DE3)Star. In the stated design protein biosynthesis is under transcriptional control of phage promoter T7 and induced by isopropyl-ß-thiogalactoside (IPTG). Tested were the conditions for optimum TcpA protein production and the layout of its purification using affinity chromatography. It was demonstrated that TcpA is present in cells of intestinal bacterium, both in native form and as inclusion bodies. Overall TcpA protein production amounted to 60 mcg/ml. Obtained purified TcpA protein can be used for studies of its immunogenic and physical-chemical properties, as well as development of immune-diagnostic preparations to evaluate the level of TcpA production in various V. cholerae strains, and identification of antigen composition of cholera vaccine preparations.

Objective of the study is experimental substantiation of possibility to produce new presentation form of cholera diagnostic sera – lyophilizate in bottles. Materials and methods. Cholera diagnostic sera. Liophylization was performed in Martin Christ Epsilon 2-6D. Residual moisture of dry sera was determined using humidity meter Sartorius MA 150. Solubility of experimental samples was assessed visually. pH was evaluated by potentiometry with the help of PK SevenExcellence-475 Mettler Toledo device, measuring pH/ORP/Ion content/conductivity/concentration. Specific activity and specificity was analyzed through expanded agglutination reaction, using relevant control strains of cholera vibrio. Results and conclusions. Experimentally justified is the possibility to lyophilize cholera diagnostic sera in flasks. Specified are the optimum parameters of lyophilization. It is determined that lyophilization does not result in deterioration of properties, as judging by the quality indicators they meet the requirements of regulatory documentation. Using accelerated aging test, it is demonstrated that the new presentation form of cholera diagnostic sera – lyophilizate in flasks – maintains their specific activity within five years term (the set out shelf life), which fully conform to normative standards.

Spatial indicators of epizooties in natural plague foci are used for justified planning and carrying out measures on epidemiological welfare provision for the population residing in enzootic territories. Plague affected regions are identified by real geographical coordinates of epizootic sites. Objective of the study is to develop numerical methods for determination of boundaries and area of epizootic regions according to the results of epizootiological monitoring. Materials and methods. Utilized are the digital maps of the natural plague foci, the data on epizootic points, and computer software. Results and conclusions. Buffer zones in the form of circumference of 5 km radius were established around each epizootic point with the help of computer application “ArcGIS 10.x” as a counterpart of circular extrapolation method. Resultant epizootic region over a certain season was generated by several overlapping circumferences.
Evaluated were the area and extension of the region, as well as the presence of epidemiologically significant objects within its bonds.
Actual positioning of the region, visualized on the focus map, is used for specified task assignment as regards carrying out of various preventive measures in this or that part of the area.

Objective of the study is to investigate phenotypic and molecular-genetic features and perform whole genome sequencing of Y. pestis strains isolated in Vietnam. Materials and methods. Studied were phenotypic and genotypic peculiarities of 20 plague agent strains isolated in different prefectures of Vietnam. Carried out was SNP-analysis of the strains, sequenced were genomes of 8 Y. pestis strains. Results and conclusions. Based on the results of studies of differential biochemical characteristics all the investigated strains were attributed to oriental biovar of the main subspecies of plague agent, which was confirmed by the presence of marker indel mutation – deletion of 93 bps in glpD gene. Investigated was also the plasmid composition of the strains. On the basis of the conducted genome sequencing and SNP-analysis appurtenance of 19 out of 20 strains under examination was determined. They belong to 1.ORI2v phylogenetic branch, relative to the strains isolated in Yunnan Province, China, which points to their common origin. Identified was a marker SNP and developed the method of SNP-typing for 1.ORI2v strains from Vietnam.

Objective of the investigation was a comparative bioinformatics analysis of Vibrio cholerae О1 Cef (CHO cell elongating factor) genes and proteins. Materials and methods. 36 Vibrio cholerae О1 strains from the Rostov-on-Don Research Anti-Plague Institute collection have been utilized. DNA sequencing was conducted on the MiSeq platform (Illumina); gene identification and analysis was carried out by means of BioEdit 7.2.5, BLASTN 2.2.29, BLASTP, MEGA 7, Vector NTI Advance 11 software programs. Results and conclusions. The data obtained confirmed Cef to be rather conserved in choleragenic strains (carrying cholera toxin genes ctxAB as a part of genome-integrated CTX prophage): all of them shared closely related prototype alleles cefС or cefЕ1. The Е1 allele was also revealed in ctxAB– strains carrying the pre-CTX prophage and in a single strain lacking both prophages. In the rest of CTX–/pre-CTX– V. cholerae four novel cef variants, that were not previously described, have been identified, two of which (E2 and E3), belonging to the Russian isolates, appeared to be unique, while for the two others absolute homologues were found in NCBI. In this connection several strains which caused severe cholera-like diseases in humans were placed in the group of cefЕ4 host strains. Since Cef is one of pathogenicity/persistence factors of cholera vibrios, we presume that conservation of its altered variants in the course of natural selection embodies a certain biological sense in respect of possible acquisition of qualities, significant for realization of both pathogenic and persistence potential.

Objective of the study is to analyze the appurtenance, numbers, and dynamics of interaction of acanthamoeba from soils of GornoAltai plague focus with Yersinia pestis 367 strain, isolated in 2016 in enzootic territory of this focus. Materials and methods. Utilized were soil amoeba from Gorno-Altai high-mountain focus and the strain Y. pestis 367 of the main subspecies of antique biovar, isolated there in 2016. Determination of systematic relation of the isolated amoeba was carried out using PCR with genus specific primers and sequencing of the obtained PCR fragments followed by identification of nucleotide sequences against GenBank database. Localization of Y. pestis cells in acanthamoeba was performed using fluorescent antibody technique by means of Axio Imager Z2 (Carl Zeiss, Germany). Results and conclusions. For the first time ever established has been the presence of Acanthamoeba castellanii in soils of burrows of Marmota altaica in the numbers of up to 300000 cells/gr in Gorno-Altai high-mountain focus. Investigated has been the dynamics of interaction of these microorganisms. Preservation of the agent in vacuoles of endoplasmatic reticulum within 14 days has been revealed. It is an indicative of the possibility of Y. pestis persistence in amoeba of Acanthamoeba subspecies in soil biocoenosis of Gorno-Altai high-mountain plague focus.

Objective of the study is to find areas characterized by different degree of potential epidemic hazard as regards tularemia in the territory of the Smolensk Region using GIS-technologies. Materials and methods. GIS-analysis of the archive data of the Center of Hygiene and Epidemiology in the Smolensk Region, obtained in the process of epizootiological and epidemiological monitoring (1941–2015) of enzootic as regards tularemia territories, as well as of Plague Control Center of the Rospotrebnadzor and literature sources was carried out. By means of Microsoft Excel created were the tables with the data on the sites of isolation (a total of 161) and residential areas (all in all 423), where patients with tularemia had been registered. Via Google Earth application, geographical coordinates of the base points (residential areas) were identified. Using GIS software package MapInfo Professional 10.5, a database was created, the layers on sites of human infections and agent isolation were obtained, and electronic map was generated. Results and discussion. Following GIS-analysis of epizootic and epidemic manifestations of natural tularemia foci in 1941–2015, differentiation of the areas by the degree of potential epidemic hazard was performed. It was established that the districts with high degree of potential epidemic hazard as regards tularemia and recurrence rate (more than 10–15 years) of its manifestations (culture isolation, human infection reports) occupy 30 % of the Smolensk Region. In addition, 42 areas of durable persistence (up to 60 years) of natural tularemia foci were spotted. The results revealed are the basis for the enhancement of epizootiological monitoring strategy and specifying the scope of prophylactic measures.

Objective of the study is to assess the effectiveness of anti-mosquito measures (against Aedes aegypti and Aedes albopiсtus mosquitoes) in the territory of Sochi and determine the ways to improve the tactics of insecticide, including larvicide, treatments.
Materials and methods. The monitoring data were analyzed in 156 points of mosquito numbers counting, 57 of which were located at sites that were subjected to treatment, including repeated ones. The count of mosquitoes was carried out by the “observer” method within 20 minutes. Statistical processing of the data was carried out using the “Past” package. Differences in mean values were calculated using the Mann-Wilcoxon-Whitney test. At the points of detection of Ae. aegypti or Ae. albopictus mosquitoes additional targeted anti-mosquito treatments were carried out. Results and conclusions. In 2016, an extremely low numbers of mosquitoes Ae. aegypti were observed. Abundance of mosquitoes Ae. albopictus is described as “high”: more than 11,000 specimens were collected in the territory stretching from the state border with the Republic of Abkhazia in the east to the city of Novorossiysk, inclusive, to the west.
The highest numbers of mosquitoes Ae. albopictus were registered in the territory of cemeteries. Performed anti-mosquito activities demonstrated their effectiveness in urbanized areas and some park zones with a high numbers of permanently residing population and tourists due to higher treatment density, small number of places favorable for the breeding of mosquitoes Ae. albopictus directly in these territories and nearby. To prevent the spread of mosquitoes Ae. aegypti and Ae. albopictus in the territory of epidemically significant objects from neighboring sites, monitoring of their numbers should be continued throughout the city and in all potential habitats. The continuation of monitoring activities across the entire Black Sea coast remains a topical issue due to the possibility of further expansion of the mosquitoes areal.

Objective is to study the role of small mammals, habitant in the Republic of Guinea, in Lyssavirus circulation. Materials and methods. Investigations were conducted using RT-PCR; nucleotide sequence of Lyssavirus cDNA fragments was identified with the help of sequencing with further phylogenetic analysis. Results and conclusions. Tested have been 356 brain samples from small mammals for the presence of Lyssavirus RNA using RT-PCR with genus-specific primers. The animals were caught in the suburbs of Kindia city in 2016. The samples were obtained from wild animals pertaining to Rodentia, Chiroptera, Eulipotyphla, and Carnivora orders.
Lyssavirus RNA was detected in 31 samples (8.7 %). For 14 PCR positive samples the appurtenance to Lyssavirus was confirmed through identification and analysis of nucleotide sequences of the collected short cDAN fragments of viral genome. The presence of rabies virus RNA in positive tests was excluded from PCR with the help of species specific primers. The pool of samples from black rats, Rattus rattus, positive for Lyssavirus RNA, contained RNA characteristic of Mokola lyssavirus species. Specified has been nucleotide sequence of matrix protein M gene fragment of Mokola virus. Genetic material of Mokola virus was detected in the Republic of Guinea for the first time ever.

Objective of the study is to conduct epidemiological analysis of official statistical data on brucellosis morbidity rates over the period of 2005–2014 in different constituent entities of the Russian Federation, using Wald method. Materials and methods. Utilized were recording and reporting documents of the Federal Service for Surveillance in the Sphere of Consumers Rights Protection and Human Welfare, FBHI “Federal Center of Hygiene and Epidemiology” of the Rospotrebnadzor, and WHO information resources.
Results and conclusions. Studies of peculiarities of epidemic process development over the long-term period have allowed for identification of entities that are the most affected by the diseases. The results obtained on the morbidity rates in the Russian Federation over the period of 2005–2014 testify to the fact that first comes North Caucasian Federal District (NCFD) (62 %), next go Siberian (SbFD) (16 %) and Southern (SFD) (13 %) Federal Districts, second and third lines of the list respectively. Other regions account for 9 % of the load. The largest share of morbidity in NCFD entities belongs to the Republic of Dagestan – 62 %. Thereat, annual increment rate is 5.54 cases, which points to stabilization and some downward trend. Application of this morbidity rate prediction tool provides for in-time planning of clinical-diagnostic, prophylactic, and anti-epidemic measures in brucellosis foci. Wald method for forecasting of morbidity can be used for other infectious diseases too.

Objective of the study is to detect the circulation of the agents of ixodic tick-borne borreliosis (ITB), human monocytic ehrlichiosis (HME), and human granulocytic anaplasmosis (HGA) in samples collected from carriers and vectors of transmissible tick-borne infections using PCR-assay. Materials and methods. Field material was studied in PCR for evaluating the rate of infection by causative agents of tick-borne infections (tick-borne borrelioses, ehrlichioses, anaplasmosis) in combined natural foci on the territory of the Rostov Region. Results and conclusions. The data obtained during epizootiological investigations carried out in 21 administrative districts and 10 cities of the Rostov Region between 2014 and 2016 were used in the study. The results were indicative of spontaneous infection in ixodic ticks by ITB causative agent on the territory of the Rostov Region, as well as of presence of corresponding natural focus. Since 2013 the activity of different components of ITB natural focus was confirmed by the detection and official registration of patients. Moreover, in the Rostov Region for the first time ever the circulation of HME and HGA causative agents was recorded and territorially combined foci of HME and HGA with ITB were identified. The results obtained allow assuming the genesis of a focus of ixodic tick-borne borreliosis and monocytic and granulocytic ehrlichioses owing to annual expansion of the areal of ticks, infected by these pathogens, and animals on which the ticks are feeding. PCR-analysis is of significant value for assessment of spread of causative agents of the “tick-borne” infections in the Rostov Region, forecasting of epidemiological situation, as well as implementation of preventive measures in the region.

Objective of the work is to perform the sanitary and quarantine control (SQC) in the under emergency situation of on the neighboring with the Russian Federation territory. Materials and methods. Utilized were information materials and report data of the Rospotrebnadzor Administration in the Rostov Region on the activities of automobile border-crossing points in high alert regime (2013–2016). Considered were some peculiarities of the sanitary and quarantine control (SQC) in the Rostov Region under emergency situations of social nature in the neighboring with the Russian Federation territories. Results and conclusions. Performance efficiency of the sanitary-quarantine facilities (SQF) in terms of inspection of vehicles and population testify to the consistently high traffic and flow of migration through multilateral automobile points of entry (APE) (2014–2016), followed by their redistribution between the APE in the context of the unfolding situation. SQC at state border checkpoints was carried out in the high alert regime to identify sick persons suspected of infectious diseases of various etiologies – the multi-etiological risk factors. Demonstrated is the expediency of “symptomatic” approach for identification of persons with suspected communicable diseases using the maximum of capacities of other government control agencies. The main peculiarities of the high alert regime under social conflict in the neighboring areas are the strengthening of interdepartmental cooperation with the Ministry of Emergency Situations (MES) and the Federal Medical-Biological Agency of Russia.

Objective. Comparative study of the protein profiles of typical virulent strains and their isogenic avirulent LPS-defective mutants of Francisella tularensis using mass spectrometric analysis. Materials and methods. Six typical virulent strains of F. tularensis of three main subspecies and eight isogenic avirulent variants which have different lesions in LPS structure were used in this investigation.
Protein spectra were obtained by MALDI-TOF MS using mass spectrometer Autoflex speed III Bruker Daltonics and Flex Control software of Biotyper. Earlier we expanded the database by introduction of the mass spectra of 40 typical F. tularensis strains of the three main subspecies. Results and conclusions. It was shown that the high quality spectrograms were obtained by means of the extraction of bacteria with ethanol supplemented with formic acid, as compared with two other methods (extraction matrix or TFA).
According to the data received, the typical virulent strains of tularemia agent of the three main subspecies have significant differences in the major peaks of protein profiles. This suggests that MALDI-TOF MS can be used for the subspecies differentiation of F. tularensis.
However, a comparative study of the protein spectra of virulent and isogenic avirulent F. tularensis strains of the same subspecies did not detect any essential differences. We were able to identify only two individual peaks (m/z 4673, 6929) which were characteristic of all virulent strains and one peak (m/z 7256) – of all avirulent strains. Thus, MALDI-TOF MS allows for reliable determination of the subspecies affiliation of F. tularensis strains but does not allow for evaluation of the strains` virulence. The results indicate that MALDI-TOF MS is a useful method for identification of typical and antigen-modified LPS-defective F. tularensis strains.

Objective of the study is to obtain the data on the state of Severokerulensky plague focus in Mongolia, adjacent to the Trans-Baikal steppe focus in the south. A reconnaissance investigation on plague in the territory of Dornod aimak was carried out by efforts of the joint Russian-Mongolian group of specialists. Material samples were tested under extended scheme using bacteriological, biological, serological, and molecular-genetic methods. Negative test results on plague allow for reporting the absence of risks of intensive and extensive epizooty development and agent importation into the territory of Russia. Though it is necessary to continue the survey in the territory of Khentii aimak, where the presence of active plague foci is registered.

Objective of the work was to compare the results of monitoring of Vibrio cholerae strains isolated in the territories of federal districts, their constituent entities and from certain water bodies between 2006–2016. Based on the methodology, using updated geoinformation system “Cholera 1989–2014”, a comparative study of the dynamics of isolation and biological properties of 586 Vibrio cholerae O1, O139 strains and R-variants, collected from the surface water bodies in the administrative territories of Russia, was carried out. Eventually it is established that Vibrio cholerae strains of various serogroups are found in the territory of all federal districts, but not in all the entities included in them. It is shown that the greatest number of isolated strains was registered in the Southern Federal District. The overwhelming number of Vibrio cholerae strains, isolated in the country, were non-toxigenic. It was noted that against the background of epidemiological welfare, the detection of single epidemiologically significant strains continued (Rostov Region).
ctxA–tcpA+ strains were found in the Southern, Far Eastern and North-Western Federal Districts. It is established that until 2013 in five constituent entities strains of Vibrio cholerae O1 and O139 in water bodies were not detected. Non-toxigenic strains of Vibrio cholerae O1 El Tor Ogawa serovar prevailed in the territories of the Southern and Ural, and Inaba serovar – in the remaining Federal Districts.
The most frequently isolated were non-toxigenic Vibrio cholerae O1 strains with phagotype 15, while isolates with phagotypes 4, 5, 10, 14 and 17 were traced only in the Southern Federal District. The data obtained formed the basis of the approach that contributes to a comprehensive assessment of the epidemiological situation on cholera in the territory of Russia and highlighted the prospects for using GIS to improve the effectiveness of cholera vibrio monitoring in surface water bodies.