[Mycobacterium tuberculosis detection by polymerase chain reaction and identification of M. tuberculosis strain]. / Vyiavlenie mikobakterii tuberkuleznogo compleksa metodom polimeraznoi tsepnoi reaktsii s odnovremennoi identifikatsiei vida M. tuberculosis.

A rapid multiprimer PCR method for detection of Mycobacterium tuberculosis complex (MTC) and simultaneous identification of M. tuberculosis in clinical samples has been developed. The method is based on simultaneous amplification of two targets: a 401 bp region from the mtp40 species-specific gene sequence of M. tuberculosis and a 544 bp fragment from the RD1 genome region which is specific for MTC but absent in BCG strains. Polymerase inhibitors in this study were detected by internal control in each test. Detection sensitivity was 25 copies of M. tuberculosis genomic DNA. Seven methods for isolation of mycobacterial DNA were compared and the technique with chloroform extraction was selected as the most efficient. The proposed method was used for analysis of 37 clinical samples and the results were compared with the results of culturing, acid-fast bacilli staining, and clinical diagnosis. The method proved to be sufficiently sensitive and specific for detection of mycobacterial DNA. Moreover, in countries with only two main pathogenic species of MTC circulating (M. tuberculosis and M. bovis) this method can be used for differentiation of these two species.

[Antiviral effect of various oligonucleotide derivatives, complementary to tick-born encephalitis virus RNA]. / Protivovirusnyi éffekt razlichnykh proizvodnykh oligonukleotidov, komplementarnykh RNK virusa kleshchevogo éntsefalita.

Antiviral effect of two nucleotides complementary to tick-borne encephalitis (TBE) virus genome and their derivatives was compared to that of noncomplementary oligonucleotides. All the tested reagents influenced TBE multiplication in cell culture, this manifesting by various degrees of suppression of the cytopathic effect of the virus. Intact oligonucleotides, both complementary and noncomplementary to TBE, reduced virus titer by 2-4 orders, whatever the concentration of oligonucleotide. In some experiments a higher virus-inhibiting effect of complementary oligonucleotides (by 3-4 orders) was observed vs. noncomplementary (by 1-2 orders). Moreover, different oligonucleotide derivatives suppressed virus multiplication in porcine embryo kidney cell culture. In parallel with investigation of virus-inhibitory effect of oligonucleotides in cell culture, their effects on the synthesis of virus-specific and cellular proteins was studied. Screening of oligonucleotide derivatives by capacity to suppress biosynthesis and multiplication of virus in cell culture showed the highest efficacy of reaction-capable and cholesterol derivatives.

[Infection by influenza virus induces production of nucleases by fibroblasts]. / Infitsirovanie virusom grippa indutsiruet produktsiiu nukleaz fibroblastami.

The stability of oligodeoxyribonucleotides and activity of DNAses and RNAses in chicken fibroblasts and in the cells infected by influenza virus have been investigated. It was found that viral infection results in an increase of nucleolytic activity in cells. The fact should be taken into account when planning experiments with antisense oligonucleotides and virus infected cells.

[Preparation of hybrid proteins consisting of human interleukin-2 and the cytotoxic A-subunit of Shigella toxin]. / Poluchenie gibridnykh belkov, sostoiashchikh iz interleuikina-2 cheloveka i tsitotoksicheskoi A-sub''edinitsy toksina shigelly.

Recombinant plasmids providing the synthesis of chimeric proteins consisting of amino acid sequences of human interleukin-2 (IL-2) and Shiga toxin cytotoxic A-subunit (ILA and AIL chimeric toxins) were constructed. The ILA and AIL chimeric toxins were shown to inhibit protein synthesis in the rabbit reticulocytes cell-free system. These chimeric toxins displayed two opposite activities of the constituent parts of their molecules on T-lymphocytes from the peripheral blood of healthy volunteers. Hybrid protein AIL (approximately 10(-6) g/ml) has caused the most significant depression of T-lymphoblast proliferation.

[In vitro suppression of translation of tick-borne encephalitis virus RNA using antisense nucleotides]. / Podavlenie transliatsii in vitro fragmentov RNK virusa kleshchevogo éntsefalita s pomoshch'iu antismyslovykh nukleotidov.

Effect of antisense oligodeoxyribonucleotides on in vitro translation of RNAs corresponding to fragments of the tick-borne encephalitis virus genome has been investigated. Sequences optimal for oligonucleotide binding and translation arrest have been identified. The most efficient oligonucleotide (17-mer) at a concentration of 2.5 microM completely arrests translation of the RNA coding for the NS3 protein.

[Suppression of translation in vitro of the mRNA of the M1 protein of influenza virus using antisense oligonucleotides]. / Podavlenie transliatsii in vitro mRNK belka M1 virusa grippa s pomoshch'iu antismyslovykh oligonukleotidov.

Effect of antisense oligonucleotides on the in vitro translation of the influenza virus M1 protein mRNA was investigated. The most efficient arrest of mRNA translation was achieved by simultaneous action of two or three oligonucleotides (14-16-mers) complementary to the juxtaposed sequences in the 5'-terminus of the molecule around and upstream of the initiation codon.

[Inhibition of translation of immunoglobulin mRNA in vitro using an alkylating oligonucleotide derivative]. / Podavlenie transliatsii immunoglobulinovoi mRNK in vitro s pomoshch'iu alkiliruiushchego proizvodnogo oligonukleotida.

Effect of complementary oligonucleotides and their reactive derivatives on translation of mouse immunoglobulin G kappa light chain was investigated. It was found that oligonucleotide pTGCTCTGGTTT and shorter oligonucleotides complementary to the coding sequence of the mRNA (nucleotides 205-215) do not arrest translation of the mRNA in the rabbit reticulocyte cell-free translation system. Preincubation of the mRNA with the alkylating 4-(N-2-chloroethyl-N-methylamino)benzyl-5'-phosphamide derivative of the oligonucleotide completely suppresses the synthesis of the protein thus demonstrating higher efficiency of the reactive oligonucleotide derivatives as inhibitors of the mRNA function.