[Molecular and biological properties of the African swine fever virus (Asfarviridae: Asfivirus) isolate ASF/Tatarstan 20/WB-12276].

INTRODUCTION: Up-to-date data and full characterization of circulating ASFV isolates play a crucial role in virus eradication and control in endemic regions and countries. The aim of the study was to evaluate and characterize the molecular and biological properties of the ASFV isolate ASF/Tatarstan 20/WB-12276, conduct phylogenetic analysis, and compare the results with isolates circulating in Europe and Asia. MATERIALS AND METHODS: For bioassay, eight heads of the Large White pigs weighing 15-20 kg/head were used. Detection of specific anti-ASFV antibodies by ELISA and immunoperoxidase method. Detection of ASFV genome was performed by qPCR. Isolation of ASF/Tatarstan 20/WB-12276 and determination of titer were performed in pig spleen cell culture. Sequencing was carried out by the Sanger method. RESULTS: The virus was characterized as highly virulent and capable of causing acute to subacute forms of ASF. Phylogenetic analysis revealed substitutions in the genome of the ASF/Tatarstan 20/WB-12276 isolate (IGR/I73R-I329L and I267L markers) that supported the clustering of the studied variant with isolates prevalent in most of Europe and Asia. CONCLUSION: For the first time, the molecular and biological properties of the ASF/Tatarstan 20/WB-12276 virus isolate taken from a wild boar shot on the territory of the Republic of Tatarstan were studied and analyzed.

[Alternative approaches to the diagnosis of African swine fever in the Russian Federation in 2017-2021].

INTRODUCTION: Prevention and control of African swine fever (ASF) transmission on the territory of the Russian Federation requires monitoring based on testing of samples from pigs and wild boars. Specific anti-ASFV antibodies are rarely detected in samples during routine serological diagnostics. Although, ASF isolates with weakened virulence were confirmed in Russia and neighboring countries.The aim of this work was to determine the possibility of using alternative samples for ASF diagnosis and evaluate the effectiveness of the diagnostic methods used on the territory of Russia. MATERIALS AND METHODS: Biological materials obtained from experimentally infected animals and samples collected in the "field" conditions were used in this study. RESULTS: Complex testing (RT-PCR and ELISA) is a more effective approach to diagnose chronic and asymptomatic forms of ASF compared to the separate use of these techniques. The possibility and efficiency of using alternative samples in diagnostics are demonstrated. It was confirmed that IPT method overcomes ELISA by high diagnostic sensitivity and detection of antibodies on earlier stages in extended range of samples. Anti-ASFV antibodies were detected in domestic and wild pigs in five regions of Russia. Samples from infected pigs that are negative in RT-PCR can be positive for anti-ASFV antibodies. The detection of antibodies in samples from shot wild boars (negative or uncertain in RT-PCR test) suggests the existence of animals surviving ASF infection. CONCLUSION: The data obtained suggest a revision of the ASF surveillance strategy, by introducing complex diagnostic methods aimed at detection of both the virus genome and anti-ASFV antibodies simultaneously.

[Analysis of the whole-genome sequence of an ASF virus (Asfarviridae: Asfivirus: African swine fever virus) isolated from a wild boar (Sus scrofa) at the border between Russian Federation and Mongolia].

INTRODUCTION: The causative agent of African swine fever (Asfarviridae: Asfivirus: African swine fever virus) (ASF) is a double-stranded DNA virus of 175-215 nm. To date, 24 of its genotypes are known. Clustering of ASF genotype II isolates is carried out by examining a limited number of selected genome markers. Despite the relatively high rate of mutations in the genome of this infectious agent compared to other DNA viruses, the number of known genome molecular markers for genotype II isolates is still insufficient for detailed subclustering. The aims of this work were the comparative analysis of ASFV/Zabaykali/WB-5314/2020 virus isolate and determination of additional molecular markers which can be used for clustering of viral genotype II sequences. MATERIAL AND METHODS: ASF virus isolate ASFV/Zabaykali/WB-5314/2020 was used to extract genomic DNA (gDNA). Sequencing libraries were constructed using the Nextera XT DNA library prepare kit (Illumina, USA) using the methodology of the next generation sequencing (NGS). RESULTS: The genome length was 189,380 bp, and the number of open reading frames (ORFs) was 189. In comparison with the genome of reference isolate Georgia 2007/1, 33 single nucleotide polymorphisms (SNPs) were identified, of which 13 were localized in the intergenic region, 10 resulted to the changes in the amino acid sequences of the encoded proteins, and 10 affected the ORF of ASF virus genes. DISCUSSION: When analyzing intergenic regions, the ASFV/Zabaykali/WB-5314/2020 isolate is grouped separately from a number of isolates from Poland and three isolates from People's Republic of China (PRC), since it does not harbor additional tandem repeat sequence (TRS). At the same time, the construction of a phylogenetic tree based on DP60R gene sequencing relates ASFV/Zabaykali/WB-5314/2020 to isolates from PRC and Poland. Moreover, phylogenetic analysis of full-genome sequences confirmed previous studies on the grouping of viruses of genotype II, and as for the studied isolate, it was grouped with the variants from China. CONCLUSION: A new variable region was identified, the DP60R gene, clustering for which gave a result similar to the analysis of full-length genomes. Probably, further study of the distribution of ASF virus isolates by groups based on the analysis of this gene sequences will reveal its significance for studying the evolution of the virus and its spread.

[Changes in the genome of African swine fever virus (Asfarviridae: Asfivirus: African swine fever virus) associated with adaptation to reproduction in continuous cell culture].

INTRODUCTION: African swine fever virus (ASFV) is a large, double-stranded DNA virus in the Asfarviridae family. It is the causative agent of African swine fever (ASF). Only the genome of BA71V strain, adapted to Vero cell culture, was fully analyzed.The aim of this study was analyzing the complete genome sequence of two strains of adapted to the growth in CV-1 cell culture (CC) ASFV obtained after 30 and 50 passages, in comparison to the parental virus. MATERIAL AND METHODS: ASFV isolate Odintsovo 02/14 (parental), ASFV adapted variants ASFV/ARRIAH/CV-1/30 and ASFV/ARRIAH/CV-1/50 were all used to extract genomic DNA (gDNA). Sequencing library was constructed using the «Nextera XT DNA library preparation kit¼ («Illumina¼, USA). RESULTS: Genomes of ASFV/ARRIAH/CV-1/30 and ASFV/ARRIAH/CV-1/50 consisted of 186 529 bp and 186 525 bp, respectively. Total 78 single nucleotide polymorphisms (SNPs) were identified between the parental Odintsovo 02/14 and the two high passaged strains, as well as a 2947 bp large-size deletion in the 3' variable region of adapted viruses was detected. DISCUSSION: ASFV as a DNA-containing virus may not have a very high level of mutation, but this is the second study showing that adaptation to growth in continuous CC leads to large deletions in the genome of the virus. CONCLUSION: Mutations in the protein-coding regions of the genome can be synonymous and non-synonymous, i.e. leading to amino acid substitution. Additional research is needed to understand the influence of the mutations described in the adaptation process on the reproduction of the virus and its virulence.

A web-based survey of reproductive awareness and choices in women with endometriosis.

OBJECTIVE: Our aim was to study fertility issues, attitudes towards reproductive techniques and fertility preservation options in women of reproductive age with endometriosis. STUDY DESIGN: In 2018 we conducted a web-based survey on fertility issues in women aged 18-40 years with endometriosis. Participants were recruited via advertisements on social media and local endometriosis support groups. Participants completed a self-developed online questionnaire evaluating the following dimensions: sociodemographic, medical data, parental project, knowledge and attitudes toward endometriosis and fertility, means used to access information, and reproductive choices. RESULTS: The majority of women (96 %) worried about the impact of endometriosis on their fertility. Approximately half of them (52 %) reported having received sufficient information concerning the effect of endometriosis on fertility from their doctor, whereas 31 % had discussed fertility issues with their doctor but desired further information. In contrast, only a minority (27 %) of women considered themselves well-informed on fertility preservation options. Information given by specialists on endometriosis and reproduction was considered most useful. Information mediated through patient support groups was also highly rated, whereas information given by the general gynecologist was less highly rated. The majority of women would consider assisted reproductive techniques (74 %) or adoption (70 %) in case of infertility. Interestingly, 72 % of women would undergo oocyte vitrification for fertility preservation, whereas only 37 % would resort to oocyte donation. CONCLUSION: This is the first survey to address the topic of fertility issues from the patient's perspective in women with endometriosis. The vast majority of women attach great importance to a discussion about fertility possibilities and only a minority of women consider themselves well-informed. Our results highlight the importance of addressing the issue of fertility in women with endometriosis. Special attention should be given to information and counselling about fertility preservation options since most women consider their knowledge on the topic insufficient. Knowledge and attitudes to counsel endometriosis patients on fertility issues and fertility preservation options should be included in the training curricula of gynecologists. Adequate information on reproductive aging, risk factors for infertility, and reproductive choices, including oocyte vitrification, should be incorporated into follow-up visits for endometriosis patients.

Managing anxiety in eating disorders with knitting.

OBJECTIVE: Recovery from anorexia nervosa (AN) is often confounded by intrusive, anxious preoccupations with control of eating, weight and shape. These are distressing and represent a potential barrier to psychological change. Theoretical and empirical evidence suggests that performing a concurrent visuospatial task reduces the emotional intensity of distressing images. We assessed whether the visuospatial task of knitting influences the anxious preoccupation experienced by inpatients with AN. METHOD: Prospective interventional cohort. SUBJECTS: Thirty-eight women with AN admitted to a specialized eating disorder unit. INTERVENTION: All subjects were given knitting lessons and free access to supplies. MEASURE: Subjects were asked to report the qualitative effects of knitting on their psychological state using a self-report questionnaire. RESULTS: Patients reported a subjective reduction in anxious preoccupation when knitting. In particular, 28/38 (74%) reported it lessened the intensity of their fears and thoughts and cleared their minds of eating disorder preoccupations, 28/38 (74%) reported it had a calming and therapeutic effect and 20/38 (53%) reported it provided satisfaction, pride and a sense of accomplishment. DISCUSSION: This preliminary data suggests that knitting may benefit inpatients with eating disorders by reducing their anxious preoccupations about eating, weight and shape control. The specificity of this effect is yet to be determined. This preliminary outcome requires further controlled study in AN subjects. From a clinical perspective, knitting is inexpensive, easily learned, can continue during social interaction, and can provide a sense of accomplishment. The theoretical and empirical rationale for this observation, and implications for deriving alternative strategies to augment treatment in AN, are discussed.

[ASF virus replication features in the presence of recombinant proteins CD2v, pX69R and pE248R.]

INTRODUCTION: African swine fever (ASF), sever hemorrhagic disease of swine caused by a large DNA virus of the Asfaviridae family. Since there are no effective and safe vaccines against ASF yet, it is urgent to study the functions of its proteins, which is applicable by analyzing the features of ASF virus replication in the presence of recombinant proteins in vitro. PURPOSE: To study the effect of ASFV recombinant proteins CD2v, pE248R and pX69R on the speed and level of reproduction of ASF virus in vitro. Thus, obtain the necessary knowledge to develop approaches for creating a vaccine against ASF. MATERIALS AND METHODS: ASFV isolate Krasnodar 07/17 and strain ASF/ARRIAH/CV-1 were used. Cloning of X69R, EP402R, and E248R genes was performed in the pJET1.2 / blunt vector and pCI-neo in E. coli JM-109 cells, according to the manufacturer's manual. Localization of recombinant proteins in CV-1 cell line carried out by direct immunofluorescence reaction (DIF) using polyclonal antibodies conjugated to FITC. The ASF virus reproduction level was assessed by hemadsorption reaction and qPCR kit (Central Research Institute of Epidemiology). RESULTS: Recombinant plasmids pCI-neo / E248R, pCI-neo / EP402R and pCI-neo / X69R were constructed. The localization and the specificity of the obtained recombinant proteins CD2v, pE248R and pX69R was confirmed. It was established that these recombinant proteins induce the level of ASF virus reproduction on days 3-5 of the experiment by ~ 1.2-1.5 lgHADU50/cm3 in comparison with the negative control. DISCUSSION: The data obtained demonstrate the important role of CD2v, pX69R and pE248R proteins in the reproduction of the virus, since they significantly affect its level. The exact function of pX69R protein was not determined, however, in the experiments its positive effect on ASF virus reproduction was established, manifested in an increase in its reproduction level. CONCLUSION: This methodology allows us to study the nature of the effect of proteins with unknown function on ASF virus replication.

The interrater reliability of rating non-exercise activity of inpatients with eating disorders using a visual analogue scale.

BACKGROUND: Non-exercise activity thermogenesis (NEAT) is the energy expended by body movement, other than sleeping, eating or sports-like activities. The obese have been reported to have a lower NEAT (walking, standing, and fidgeting) than controls. We hypothesize that an elevated NEAT could explain why some patients with anorexia nervosa are resistant to weight gain. OBJECTIVE: To evaluate the interrater reliability of a rating of non-exercise activity of inpatients with eating disorders (ED) using a visual analogue scale (VAS). METHOD: Health care providers were asked to rate the non-exercise activity of inpatients by marking a VAS. RESULTS: Eight patients were individually rated by 10 clinicians. Results were analyzed using the intraclass correlation coefficient (ICC) and Cohen's multi-rater kappa statistic (kappa). The ICC(3,k) was 0.257 (p<0.01) and 0.708 (p<0.01) for average measures. DISCUSSION: The ratings of NEAT using a VAS were not reliable between clinicians. This indicates that the ward staff, even on a specialized ED unit, cannot reliably estimate non-exercise activity and physiological measurements should be used.