Search

Prokaryotic Argonautes (pAgos) are programmable nucleases involved in cell defense against invading DNA. In vitro, pAgos can bind small single-stranded guide DNAs to recognize and cleave complementary DNA. In vivo, pAgos preferentially target plasmids, phages and multicopy genetic elements. Here, we show that CbAgo nuclease from Clostridium butyricum can be used for genomic DNA engineering in bacteria. We demonstrate that CbAgo loaded with plasmid-derived guide DNAs can recognize and cleave homologous chromosomal loci, and define the minimal length of homology required for this targeting. Cleavage of plasmid DNA at an engineered site of the I-SceI meganuclease increases guide DNA loading into CbAgo and enhances processing of homologous chromosomal loci. Analysis of guide DNA loading into CbAgo also reveals off-target sites of I-SceI in the Escherichia coli genome, demonstrating that pAgos can be used for highly sensitive detection of double-stranded breaks in genomic DNA. Finally, we show that CbAgo-dependent targeting of genomic loci with plasmid-derived guide DNAs promotes homologous recombination between plasmid and chromosomal DNA, depending on the catalytic activity of CbAgo. Specific targeting of plasmids with Argonautes can be used to integrate plasmid-encoded sequences into the chromosome thus enabling genome editing.

Prokaryotic Argonaute proteins (pAgos) are homologs of eukaryotic Argonautes (eAgos) that were similarly proposed to play a role in cell defense against invaders. However, pAgos are much more diverse than eAgos and very little is known about their functional activity and target specificity in vivo. Here, we describe five pAgo proteins from mesophilic bacteria that act as DNA-guided DNA endonucleases and analyze their ability to target chromosomal and invader DNA. In vitro, the analyzed proteins use small guide DNAs for precise cleavage of single-stranded DNA at a wide range of temperatures. Upon their expression in Escherichia coli, all five pAgos are loaded with small DNAs preferentially produced from plasmid DNA and from chromosomal regions of replication termination. One of the tested pAgos, EmaAgo from Exiguobacterium marinum can induce DNA interference between multicopy sequences resulting in targeted processing of homologous plasmid and chromosomal loci. EmaAgo also protects bacteria from bacteriophage infection and is preferentially loaded with phage guide DNAs suggesting that the ability of pAgos to target multicopy elements may be crucial for their protective function. The wide spectrum of pAgo activities suggests that they may have diverse functions in vivo and paves the way for their use in biotechnology.

Parasitic liver zoonoses are endemic to some regions of Russia as well as to Mediterranean countries, Australia, New Zealand, South America, and the Indian subcontinent. However, the available data on the surgical treatment of patients with parasitic liver diseases are often contradictory, and such treatments remain a difficult task today. The effectiveness of surgical treatment was analyzed in 628 patients with echinococcosis and 58 patients with liver alveococcosis managed at the Republican Clinical Hospital during 1998-2018. The most commonly performed surgery for echinococcosis was closed echinococcectomy of different types, which was applied in 428 patients. Surgical outcomes were evaluated according to the type of surgery. Although there were no differences in outcomes in the immediate postoperative period, long-term postoperative outcomes differed with the type of surgery, compelling us to reconsider the surgical methods employed, with a preference for implementing cystectomy with omentoplasty of the residual cavity of the liver. In fact, the best outcomes in terms of recurrence and complications were obtained with minimal but adequate procedures, namely actual echinococcectomy without the resection of the residual cavity or liver. Thus procedures with low-traumatic access should be performed in cases with specific indications, such as the presence of easily accessible and well-visualized parasitic cysts.

Background : In Russia, cardiovascular mortality is among the highest in the world. Behaviours related to the development of cardiovascular disease are usually adopted in childhood and adolescence. Very little information exists on prevalence and trends of risk factors among Russian youth. This study aims to investigate changes in the prevalence of cardiovascular risk factors among adolescents in the Republic of Karelia, Russia, from 1995 to 2004. Methods : Cross-sectional surveys on chronic disease risk factors were carried out among 15-year-old adolescents in Pitkaranta region, in the Republic of Karelia. The surveys were conducted in all 10 secondary schools in the Pitkaranta region. All ninth-grade students in 1995 ( N = 385, response rate 95%) and in 2004 ( N = 395, response rate 85%) were included in the survey samples. Results : Systolic blood pressure decreased statistically significantly among boys (from 119 to 116 mmHg). Diastolic blood pressure decreased statistically significantly among both girls (from 64 to 59 mmHg) and boys (from 62 to 59 mmHg). Total cholesterol increased statistically significantly only among girls (from 3.9 to 4.1 mmol/l). Body mass index did not exhibit any significant changes in both genders. Daily smoking rate doubled statistically significantly among girls from 7% to 15%. Conclusion: The study results show changes in the prevalence of cardiovascular disease risk factors among adolescents in the Republic of Karelia occurring over the last decade. Active measures need to be taken to prevent the increase in smoking prevalence, especially among girls, and to avert the unfavourable development of other risk factors in the future.

Conference Proceedings of PHYSOR2006, Vancouver, Canada, 10-14 Sept. 2006; ISBN-0-89448-697-7 (2006) B104.