[Electron microscopy of the surfaces of bacillary spores].

The surface structures of the spores of Bacillus. cereus, B. thuringiensis, and Brevibacillus laterosporus were studied by transmission and scanning electron microscopy. Platinum deposition and negative staining with uranyl acetate revealed appendages and exosporium in B. thuringiensis and B. cereus. The exosporium structure was visualized by negative staining and ultrathin sectioning. For staining the exosporium polysaccharide, Alcian blue was used during fixation. The results obtained show the differences in structural organization of appendages and exosporium in different strains. Canoe-shaped inclusions were revealed in all Br. laterosporus strains, while strain IGM16-92 had a fibrillar capsule as well. Electron microscopy using a dual beam scanning electron microscope Quanta 200 3D provided the information of the spore surface relief without sample treatment (fixation and dehydration). The spores of Br. laterosporus strains had folded surface, unlike the smooth surface of B. cereus and B. thuringiensis spores. Diversity of external spore structures was shown within a species, which may be used for detection of bacteria at the strain level. Optimized procedures for visualization of spore surface by different electron microscopic techniques were discussed.

[Isolation, biological properties, and spatial structure of an antibiotic loloatin A]. / Vydelenie, biologicheskie svoistva i prostranstvennaia struktura antibiotika loloatina A.

Peptide antibiotic with cyanolytic activity was isolated from the IGM52 strain of the Brevibacillus laterosporus Gram-positive spore-forming bacteria. By 1H NMR spectroscopy, this antibiotic was identified as loloatin A, a cyclic decapeptide cyclo(-Asn-Asp-Tyr-Val-Orn-Leu-DTyr-Pro-Phe-DPhe-). The spatial structure of loloatin A in solution was determined. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2002, vol. 28, no. 4; see also http://www.maik.ru.

[The duration of action of Bacillus thuringiensis spp. israelensis and Bacillus sphaericus after encapsulation by infusoria Tetrahymena pyriformis]. / Prodolzhitel'nost' deistviia bakterii Bacillus thuringiensis spp. israelensis i Bacillus sphaericus posle inkapsuliatsii ikh infuzoriiami Tetrahymena pyriformis.

The study of the duration of the insecticidal activity of a mixture of infusoria with each of these bacteria revealed differences in the pattern of their relations during their long contact. T. pyriformis may be used for the prolongation of its insecticidal action if Bti is applied. An increase in the duration of this action depends on the viability of tetrahymena in the experimental flasks and on the presence of free spores and crystals of Bti. The prolongation of the action may be associated with the multiplication of this type of bacteria in the digestive vacuoles of infusoria. The use of Bsph in the mixture with T. pyriformis caused no increase in the insecticidal activity of bacteria. Tetrahymena increased the duration of Bsph and even reduced the period of its larvicidal activity. Tetrahymena used a sporocrystalline complex of Bsph as food and reduced its water concentration.

[Duration of the larvicidal effect of spore crystalline mass of bacteria Bacillus thuringiensis spp. israelensis and Bacillus sphaericus in the laboratory setting]. / Prodolzhitel'nost' larvitsidnogo deistviia sporokristallicheskoi massy bakterii Bacillus thuringiensis spp. israelensis i Bacillus sphaericus v laboratornykh usloviiakh.

The duration of action of sporocrystalline mass of Bacillus thuringiensis spp. Israelensis (Bti) and Bacillus sphaericus (Bsph) against three-age larvae of An. stephensi mosquitoes was studied in the laboratory setting. The duration of action depends on its initial concentration. The optimal concentration should be at least 10(5) spores/ml for An. stephensi larvae. Decreasing the concentration reduces the duration of larvicidal action. Increasing the number of the larvae placed also diminishes the duration of action shown by Bti. This has no significant influence on the duration of action by Bsph. The presence of dead larvae in the test vessel increases the duration of action by Bti only by 4 times whereas Bsph acts rather long. The follow-ups lasted 60 weeks with full death of the larvae placed again. A significant water resistance of Bti sporocrystalline mixture was found. The mixture retained its larvicidal activity 6 months following its presence in the dechlorinated water without larval placement. Bsph was less resistant and its complete inactivity was noted under the same conditions just following 2 months.

[Use of the recombinant bacterial strain to control blood-sucking mosquito larvae]. / Vozmozhnost' primeneniia rekombinantnogo shtamma bakterii dlia bor'by s lichinkami krovososushchikh komarov.

The recombinant strain of Methylobacillus flagellatum with the cloned synthesis gene Cry 4B of the toxic Bac. thuringiensis var. israelensis protein proved to be effective against larvae of the Anopheles stephensi, An. atroparvus, An. pulcherrimus, An. superpictus, and An. sacharovi cultured in the laboratory. The use of M. flagellatum in combination with T. pyriformis may greatly expand the scope of use of the recombinant strain to control malaria mosquito larvae. Their combined use shows a 6-fold increase in the rate of strain action and a 4-fold decrease in the concentration of the agent. The optimum effects are shown following 24-hour combined intubation of M. flagellatum and Tetrahymena pyriformis.

[Structural and molecular-biological characteristics of a new transducing phage from Bacillus thuringiensis Tt 91]. / Strukturnaia i molekuliarno-biologicheskaia kharakteristika novogo transdutsiruiushchego faga Bacillus thuringiensis Tt 91.

A new bacteriophage Tt91 of larger size and broad lytical spectrum has been isolated from soil to widen the possibility to study new Bacillus thuringiensis strains. The Tt91 bacteriophage was shown to perform the intravariant transduction and, thus, can be used for genetic mapping. The ultrastructural analysis made it possible to refer bacteriophage Tt91 to morphotype A1 and showed similarity of Tt91 morphological structure to the one of CP-group bacteriophages. The performed comparative analysis of phage-specific proteins has confirmed a possible relation of the bacteriophages and revealed the differences between them. The data on the resistance of Tt91 DNA to majority of site-specific restriction endonucleases (17 out of 18 tested) and on DNA melting in the wide interval of temperatures suggests the presence of possible anomalies in the Tt91 DNA.

[Site-specific restrictases from Bacillus thuringiensis var. Kumantoensis]. / Saitspetsificheskie restriktazy iz shtamma Bacillus thuringiensis var.Kumantoensis.

The efficiency of bacteriophages CP-54 and CP-55 plating on Bacillus thuringiensis var. kumantoensis H18 (Kum) is decreased about 10-fold as compared with the efficiency of plating on Bacillus thuringiensis var. galleriae H5 (Gal). Bacteriophages having propagated for one cycle in Kum cells might be further grown in this strain without growth restriction. Two site-specific restriction enzymes isolated from Bacillus thuringiensis var. kumantoensis were designated BtkI and BtkII. The endonuclease BtkI recognises the same nucleotide sequence CGCG in DNA as recognised by the restriction endonuclease FnuDII; BtkII recognises the same nucleotide sequence GATC as the endonuclease Sau3A.

[Conjugation transfer of the pAMbeta1 plasmid to Bacillus anthracis]. / Kon''iugatsionnaia peredacha plazmidy pAMbeta1 Bacillus anthracis.

The possibility of conjugational transfer of the plasmid pAM beta 1 in the cells of different strains of Bacillus anthracis has been established. The efficiency of the plasmid replicon transfer in interspecies transfer B. thuringiensis X B. anthracis was n.10(-7), while in interspecies transfer it was n.10(-6). The capability of mobilization of extrachromosomal replicon pTG141 for conjugational transfer has been demonstrated. Bacillus anthracis transconjugants harbouring the pAM beta 1 plasmid have acquired the donor properties in conjugation.

[Site-specific restriction endonuclease BtcI from Bacillus thuringiensis var. canadensis]. / Saitspetsificheskaia éndonukleaza restriktsii PtcI iz Bacillus thuringiensis var. canadensis.

Efficiency of bacteriophage Tp4 plating on Bacillus thuringiensis var. canadensis H5 (Can) is decreased 10(7)-fold as compared with the efficiency of plating on Bacillus thuringiensis var. galleriae H5 (Gal). Bacteriophage Tp4 having propagated for one cycle in Can cells might be further grown in this strain without restriction. The sitespecific restriction endonuclease BtcI isolated from Bacillus thuringiensis var. canadensis recognises the same nucleotide sequence GATC in DNA as recognised by restriction endonuclease Sau3A.

[Plasmid transduction by Bacillus anthracis bacteriophage CP54]. / Transduktsiia plazmid bakteriofagom CP54 Bacisslus anthracis.

Possibility of plasmid transduction in Bacillus anthracis vaccine strains Sterne and STI-1 by bacteriophage CP54ant having an increased ability of adsorbtion and a shortened period of latent development in Bacillus anthracis cells has been isolated. The main parameters of plasmid transduction by the bacteriophage have been established for the plasmid pTG141 (TcR). They include the effect of multiplicity of infection, the level of UV-inactivation of bacteriophage, the presence of antiphage serum in the incubation medium. Plasmid transduction by the mutant phage CP54ant was found to be more efficient as compared with the one by the parent phage. The isolated transductants served as donors of the transduced plasmid for Bacillus anthracis and Bacillus thuringiensis strains.

[The role of plasmids in the regulation of delta-endotoxin synthesis in Bacillus thuringiensis H14]. / Rol' plazmid v reguliatsii sinteza delta-éndotoksina Bacillus thuringiensis H14.

The role of plasmids in regulation of delta-endotoxin synthesis by Bacillus thuringiensis H14 was studied. The derivatives of strain Is-1 H14 containing a 4Md plasmid integrated into the chromosome synthesize small crystals and are not toxic for the gnat larvae. The transceptional transfer into this strain of a plasmid coding for crystal synthesis from the strain 69-6 serotype H5 results in restoration of insecticidal activity to the level of the parental strain Is-1. Transcipients activity is increased 10-15 fold in case of 4Md plasmid excision from the chromosome and autonomous functioning. Evidently, 4Md plasmid from the strain Is-1 as well as a plasmid coding for crystal synthesis from the strain 69-6 contains the regulatory elements participating in the expression of crystalline protein genes localized on other plasmids. The existence of two cellular regulatory groups is supposed to result in the significant increase in crystalline protein synthesis.

[Homologous and heterologous transcription of the Cry+-plasmid in Bacillus thuringiensis]. / Gomologichnaia i geterologichnaia transtseptsiia Cry+-plazmid Bacillus thuringiensis.

The possibility of homologous and heterologous transception of Cry+ plasmids in Bacillus thuringiensis is demonstrated. Cry+ plasmids from crystal bearing strain of Bacillus thuringiensis were transferred into acrystalline strain belonging to H5 serotype by mutual incubation. The donor strain was previously marked by the transmissive plasmid pAM beta 1 coding for erythromycin and lincomycin resistance. The transcipients having acquired the ability to synthesize delta-endotoxin were referred to H5 serotype due to their phenotype. By analogous method Cry+ plasmid was transferred from Bacillus thuringiensis to Bacillus cereus. Bacillus cereus strain GP7 was used as a recipient strain resistant to tetracycline. The presence of delta-endotoxin in transcipients was confirmed by bioprobes and immunoenzyme assay. To prove the transfer of Cry+ plasmid the plasmid profiles of the parent strains and transcipients have been analyzed. The formation of cellular contacts during mutual incubation of Bacillus thuringiensis and Bacillus cereus strains was demonstrated by electron microscopic study of ultrafine cuts.

[Characteristics of Bacillus thuringiensis phages with circular permutation in the DNA molecule]. / Kharakteristika fagov Bacillus thuringiensis s tsirkuliarnoi permutatsiei molekul DNK.

Bacteriophages Tm2 and Tg27 of different origins but identical in biological properties have been compared. Physicochemical characteristics of bacteriophages have revealed the existence of end repeats and circular permutation of phage DNA. Phages Tm2 and Tg27 share the same dimensions of incapsulated DNA, differing in the sizes of phage genome and end repeats. Bacteriophage Tm2 genome is 45.2 kb. long with the end repeats containing 5.7%. The genome of Tg27 is 42.53 kb and 11.8% of end repeat. The bacteriophages relation has been confirmed by heteroduplex and restriction analysis. Tm2 and Tg27 share 84% of homology. Two regions of nonhomology are found representing a single-stranded loop and equishouldered vesicle with the sizes 2.19 kb and 5.03 kb, respectively.

[Effect of the pC194 plasmid on the development of Bacillus thuringiensis phages]. / Vliianie plazmidy pC194 na rezvitie fagov Bacillus thuringiensis.

Bacillus thuringiensis var. galleriae strains were transformed by plasmid pC194, coding for chloramphenicol resistance (CmR). Efficiency of plating and the yields of bacteriophages Tg13 and Tg27 maturating in CmR transformant cells were decreased for 2-3 orders as compared with the ones in parental strains. The CmR transformants are characterized by the increased level of spontaneous induction of bacteriophage Tg22.

[Ultrastructure of the bacterial spores and crystals of various Bacillus thuringiensis serotypes]. / Ul'trastruktura spor i kristallov bakterii razlichnykh serotipov Bacillus thuringiensis.

Spores and crystals of various Bacillus thuringiensis serotypes were studied by electron microscopy. The serotypes were shown to differ in the fine structure of the exosporium and crystals. The packing parameters of morphological subunits were determined by analysing electron photomicrographs by the technique of optical diffraction and filtration. Some of the strains are characterised by hypersynthesis of surface spore structures. The individual morphological properties of crystals can be used for their differentiation.

[Intervariant transduction in Bacillus thuringiensis]. / Mezhvariantnaia transdultsiia u Bacillus thuringiensis.

This report deals with intervariant transduction in strains of Bacillus thuringiensis subsp, dendrolimus and galleriae with bacteriophage Tg13. This bacteriophage has a broad host range and is of a large DNA molecular mass (40.3 mD). Transducing properties of this bacteriophage were studied in Bacillus thuringiensis var. galleriae using auxotrophic mutants requiring purine and pyrimidine bases or tryptophan as recipients. The frequency of transduction was 10(-5)-10(-6). In order to prove the possibility of intervariant transduction, two series of experiments have been conducted. In the first case, bacteriophage tg13 has been propagated on the prototrophic strain 49-18 (var. dendrolimus), 48gua (var. galleriae) being used as a recipient. In the second case, the prototrophic strain 69-6 (var. galleriae) served as a donor and 49-18-1 trp (var. dendrolimus) - as a recipient. The frequency of heterologous transduction was 10(-7). The positive result obtained in cross-transduction experiments may be explained by common metabolic pathways of purine and pyrimidine bases and amino acids in strains of different serotypes.