Study of ultrafast processes in matter by means of time-resolved electron diffraction and microscopy

One of the most fundamental problems of modern natural science is the direct observation of atomic motions in the course of various processes. For this purpose, in the experiment it is necessary to provide high spatial-temporal resolution. The solution to this problem is achieved by using a pulsed electron beam of ultrashort duration to create a stroboscopic diffraction pattern in the method of time-resolved electron diffraction (TRED). Three types of experimental schemes have been developed at our lab. The experimental complex includes (i) 20-keV table-top apparatus for TRED, (ii) 75-keV ultrafast transmission electron microscope and (iii) lensless table-top device for femtosecond electron diffraction. The obtained experimental results are presented

Genetic variations among PRRSV strains isolated in Italy and in Russia

European and American isolates of porcine reproductive and respiratory syndrome virus (PRRSV) represent two distinct genotypes. An analysis of available data indicates that the American strains exhibit significant genetic diversity whereas the European strains are highly conserved and genetically related to each other. The PRRSV strains isolated in the Netherlands, Spain, France and Great Britain are closely related, with a homology of 96-100% for the nucleotide sequence of the ORF7 gene and 98-100% for the amino acid sequence of the nucleocapsid (N) protein. In this work, an attempt has been made to extend our knowledge of the European genotype of PRRSV by analysing the Italian and Russian virus groups. The ORF7 genes of eleven PRRSV strains isolated in Italy (1992-1998) and fourteen Russian isolates (1996-1999) were amplified by PCR and their nucleotide and deduced amino acid sequences were determined. A comparative analysis of corresponding nucleotide and amino acid sequences of the Italian group of virus isolates and other Western European strains revealed up to 8.5% nucleotide sequence diversity and up to 8.7% amino acid sequence diversity. Only one Italian strain, the 2156/92, isolated in Northern Italy in 1992, was closely related to the Lelystad strain with a homology of 95.9% for the nucleotide and 98.5% for amino acid sequences. The analysis revealed that the ORF7 of ten Italian isolates encode a 128 amino acid (a. a.) N protein while the ORF7 of the isolate 3152/97 encodes a 127 a. a. N protein. The level of diversity of the N protein sequence of the Russian isolate was much higher - up to 16% compared with the homologous sequence of the Lelystad strain. Alignments of corresponding sequences of both genotypes and the plotted phylogenetic tree showed that all the Russian isolates studies belong to the European genotype but represent two distinct genetic subgroups. Eight Russian isolates from one subgroup had 387 nucleotide long ORF7 encoding a 128 a. a. N Protein. But six isolates from the other subgroup had only 378 nucleotide long ORF7 encoding a 125 a. a. N protein with a stop-codon at the same site as isolates of the American genotype (the size of the N protein of American genotype isolates is 123 a. a.). It is interesting that the six isolates with 125 a. a. N protein had insertions of Asn between amino acid residues 46 and 47 (at the same positions as those of Lelystad strain N protein). The N protein of the American genotype isolates also had the same insertion at the same site. The analysis of the deduced amino acid sequences of the Russian isolates, demonstrated that the level of non-synonymous substitutions was up to 38% of total substitutions for different isolates, and part of them was observed at the same sites of the N protein. Some of these sites were identical for the isolates of both genotypes. The data obtained concerning the level of genetic diversity of the Italian and Russian isolates was suggestive of a more continuous history of PRRSV in Europe. To estimate the rate of PRRSV genome mutagenesis in field conditions, ORF7 genes of a number of isolates from the same farms were sequenced. The results were ambiguous. The isolates from different farms had mutation rates that differed very essentially. This could be partly explained by the heterogeneity of virus populations on large farms, but this problem requires further studying. Thus, the European genotype of PRRSV is not less genetically heterogeneous than the American one. Russia and Italy have probably been endemic for PRRSV for a long time. Regarding the problem of the emergence and spread of the virus, it is necessary to take into consideration the high mutation rate of the PRRSV genome and essential distinctions of this parameter. A large group of Russian PRRSV isolates has some structural features of the N protein characteristic of the American genotype isolates. The presence of these structural features can form the basis for speculations about a determinacy of the mutative process and possible directional genetic divergency of PRRSV