Botanica Lithuanica, Supplementum 3

1999 m.

The samples of cell cultures were received from some Lithuanian institutions and laboratories. They were tested for mycoplasma infection using two methods: DNA staning with fluorescent dyes and broth-agar microbiological culturing. 66.3 and 93.3 per cent of cell lines, respectively werw contaminated by mycoplasma. We studied the effectiveness of immunological system for mycoplasma elimination from cell culture. MH-22A cell line was first cultivated in vitro and then in vivo – is cell protection from secondary mycoplasma infection.

Keywords: mycoplasma, cell culture, mice, decontamination.

The permeability characteristics of cell envelope of plant pathogen Pseudomonas syringae pv. Phaseolicola have been investigated. The method of distribution of the lipophilic cation tetraphenylphosphonium (TTP+) across the bacterial envelope was used to evaluate barrier properties of the cell envelope. Studies revealed the permeability changes of the cellular outer and inner membranes upon the interaction of Pseudomonas syringae cell with membranoactive chemical compounds (EDTA and polymyxin B) and bacteriophage ?6.

Keywords: Pseudomona syringae, cell envelope permeability, bacteriophage ?6, tetraphenylphosphonium.

As a result of recently changed keeping and feeding conditions of animals and fowl in farms the cases of various infection diseases occur more frequently. This is proved by the investigations carried out in 1992–1998. During this period in 27 district farms, pig-breeding farms, poultry farms, and individual farms 90 strains of bovine Salmonella agents, 192 pig and fowl Salmonella agents, 227 E. coli, 12 pig Erysipelas agents, 2 Bacillus subtilis, 11 Pasteurella multocida, and 2 Actinobacillus pleuropneumoniae strains were isolated. The agent of pig erysipelas was isolated in agricultural companies and individual farms. The cultures of infectious disease agents isolated in 1992–1998 that have stable immunogenic and antigenic properties are used in production of diagnostic preparations and inactivated vaccines of pig and bovine Collibacteria, Salmonella, and Pasteurellae.

Keywords: epizootology, salmonellosis, colibacillosis, pasteurellosis, pig erysipelas, identification of infectious diseases.

82 micromycetes species were isolated from the rhizosphere of the Ficus genus plants imported from the Netherlands. Most of the isolated species are typical for soils rich with organic mater. Together with saprophytic species, some rather pathogenic species were found in the rhizosphere of all plants. From the rhizosphere of some species (Ficus benjamina, F. elastica, and F. pumila) the following fungal species, able to parasitize plants of this genus were isolated: Pythium irregulare, Cladosporium herbarum, C. cladosporioides, Verticillium albo-atrum, and Fusarium oxysporum.

Keywords: micromycetes, the Ficus genus, rhizosphere, occerrence frequency.

Phytoindicators used for immunological investigations of apple-tree European canker were flax and garden pepper-grass. Germination indices of flax were rather distinctly inversely correlated with resistance of apple-tree varietes to European canker. The cortex supplement of sicf apple-tree compared to healthy ones resulted in 4.4-15 % reduction of germinability indices. The correlation between garden pepper-grass and resistance of apple-tree varietes against European canker was not sufficiently distinct.

Keywords: phytoindication, European canker, Nectria galligena, allelopathy.

The Bacillus subtilus BS 2924 suppressive volatile activity against broad range of test-cultures and its production at the presence of various carbohydrates were studied. IR-spectroscopy was done, however, more detailed study is necessary for identification of the nature of volatile antagonistic compound(s).

Keywords: Bacillus, suppressive volatile, spectrum of action, carbohydrates effect, IR-spectroscopy.

Echinacea extract, vermicompost, and chitosan applied as peat drech or plant spray significantly decreased the spread of some soil-borne pathogens, powdery mildew and rust of some ornamental plants. Chitosan used as bulb soak or spray probably induced plant resistance toward the tested pathogens.

Keywords: vermicompost, chitosan, concentration, Echinacea, Phytophtora, Pythium, Rhizoctonia, Fusarium, Sphaerothera, Puccinia, control.

Five viruses affecting irises have been isolated and identified: Iris mild mosaic potyvirus, Cucumber mosaic cucumovirus, Tobacco mosaic tobamovirus, Bean yellow mosaic potyvirus nad Tomato ringspot nepovirus (ToRSV). Viruses were identified technique by test-plants, electron microscopy, and serological methods. The optimum technique for obtaining a purified preparation for ToRSV was selected and the purified preparation was obtained.

Keywords: iris, virus, Iris mild mosaic, Cucumber mosaic, Tobacco mosaic, Bean yellow mosaic, Tomato ringspot.

The symptoms caused by viruses occuring on dahlia are described and virological state of dahlia is evaluated. Viruses were identified by test-plants, electron microscopy, and DAS-ELISA methods. Two viruses affecting dahlia have been identified: Dahlia mosaic caulimovirus and Tomato spotted wilt tospovirus. Viruses were in complex infection,

Keywords: dahlia, Dahlia mosaic caulimovirus, Tomato spotted wilt tospovirus.

8 potato virus M (PVM) isolates and 20 potato virus S (PVS) isolates have been separated and identified by serological, test-plants, and electron microscopy methods. The investigation of biological and morphological properties of the isolates revealed the prevalence of typical PVS strain in potato crops in Lithuania. The potato leaf rolling symptoms were caused not only by PVM K strain, but also by other strains of this virus.

Keywords: potato, viral diseases, carlaviruses, Potato virus M, Potato virus S, identification.

In Lithuania Tomato ringspot nepovirus (ToRSV) from vegetable crops (cucumber, tomato, and squash) was isolated and identified. This virus was identified by methods of electrn microscopy and test-plants. The purified preparation of ToRSV isolate from cucumber was prepared. The purified ToRSV preparation was used as an antigen for the preparation of diagnostic antiserum in rabbit.

Keywords: vegetable crops, viral diseases, Nepoviruses, Tomato ringspot virus, identification.

The ways in which antimicrobials (of microbial and chemical origin) get into milk are described. Modern requirements for the microbiological methods of determining the residues of antimicrobials in milk are presented. The sensitivily of three methods widely used in Europe for the detection of antimicrobial residues in milk was investigated. It has been established that Valio T-101 method was sensitive to a considerable number of antimicrobials such as antibiotics, detergents, and disinfectants.

Keywords: antimicrobials, sources, milk, methods, antibiotics, sensitivity.

Results of the investigations on environment influence on the distribution of the cercosporoid fungi in Lithuania in 1992–1998 are presented. Meanwhile 49 taxa ascertained on 23 families, 48 genera, 75 plant species have been identified in Lithuania. Most taxa – 11 (22 % of all indicated cercosporoid fungal taxa) were ascertained on the plants of the Fabaceae family. Some of these fungi – Mycovellosiella murina – parasitize the plants of even two families – Violaceae and Oleaceae. Passalora depressa was ascertained on host plants of 3 genera (Aegopodium, Anethum, Angelica), Cercospora beticola – on host plants of 2 genera (Beta and Chenopodium). Passalora microsora was ascertained on 9 plant species of the Tilia genus, Cercospora zerbina – on 8 plant species of the Trifolium, C. radiata – on 4 species of the Anthyllis genus, C. medicaginis and C. beticola – on 3 species of the Medicago and the Beta genera. The exception is C. nasturtii var. capsellae wicth adapted itself to parasitize only Capsella bursa-pastoris. Sometimes one host plant species is parasitized by two species of the cercosporoid fungud. Cercospora beticola and Passalora dubia parasitizes on Chenopodium album; Cercospora lathyri and C. lathyrina on Lathyrus pratensis; C. ribis and Cercostigmina ribicola on Ribes rubrum; Cercospora peckiana and Pseudocercospora acetosellae on Rumex acetosa. The humidity and active solar radiation influence the development of these plant pathogens more than temperature. They occurred only in the second half of summer when the amount of summed radiation decreased. During rainy and foggy summers they may appear earlier. Area pollution, industrial and health-resort cities and small towns have no influence to distribution of bronw leaf spot (Passalora microsora) of lime-tree. The spreading of fungus basically depends on meteorological conditions.

Keywords: cercosporoid fungi, environmental conditions, host plants, plant pathogens, Lithuania.

Activity of plant cell wall-degrading enzymes (two types of polygalacturonases: endo- (EC, exo- (EC and endo-1,4-?-glucanase (EC produced by the fungi pathogenic to weeds was ascertained. According to primary selection quantitative analysis was carried out. Strain 97-1 showed the highest pectic activity, 97-2, 98-2 and 98-3 –the highest cellulolytic activity.

Keywords: cellulase, pectinases, plant cell wall.

Seeds of lupine (Lupinus albus ‘Wat’ and L. angustifolius ‘Emir’) were treated with 3 different pesticides and then inoculation wuth Bradyrhizobium sp. After 0–1 h, 23–24 h and 47–48 h from inoculation the survival of root nodule bacteria on the seeds was examined. At the same time some seeds were sown into sand filled pots to assess the effect of chemical seed dressing on the symbiosis process. Of all the tested pesticides, mancozeb (fungicide) was most toxic to the survival of rhizobia on seeds and to the lupine root nodulation. On the other hand, carbosulphan (insecticide) and a mixture of carbendazim+thiuram (fungicides) were much less harmful to the bacteria, particularly when seeds had been sown within 0–1 after Bradyrhizobium sp. Inoculant application.

Keywords: lupine, Bradyrhizobium, seed dressing, pesticide, nodulation, biomass.

In three field experiments effects of different agricultural practices on root-nodule bacteria (rhizobia) populations in soil were assessed using the Most Probable Method. Populations of Rhizobium meliloti (Dangeard) nodulating lucerne were low in all studied soils, irrespective of location and agriculture practices. The highest number of these bacteria amounted only to 17 cells per g of soil in the experiment II located in Grabow. R. leguminosarum (Frank) biov. viciae and biov. trifolii populations ranged from less than 6 cells in the unlimed soils to 1.7 × 103 in the case of pea rhiozobia and up to 1.7 × 105 in the case of clover rhizobia in the manured soil without N fertisilation.

Keywords: root-nodule, bacteria, population, soil, agriculture practices.

Detached stems of soybean ‘Progress’ were dipped inyo Helriegel II nutreint medium and inoculated with freshly grown suspension of Bradyrhizobium japonicum Jordan strain 2490. It was determined that nodule bacteria might penetrate into soybean additioanl roots formed from detached stems. Roots of the detached stems, with were grown in not darkened nad not aerated Helriegel II medium without N, formed the greatest number of nodules – at an average 8–11 nodules per one plant. The most suitable time for the inoculation of soybean detached stems is 2–5 days after stem dipping.

Keywords: detached stems, soybean, nodule, nodule bacteria, inoculation, nutrient medium, pod, seed, root.

Two years monitoring of the impact of industrial emission of “Severstal” metallurgical works on soil micromycetes has been done. The data showed the double increase of quantity and species diversity of soil micromycetes in the zone of industrial pollution comparing with the unpolluted soil. Penicillium funiculosum, P. puberulum, aspergillus fumigatus, and A. terreus were confirmed as the species-indicators of heavy metals pollution in soil. The development of the phytopathogenic species of the Verticillium and the Fusarium genera in the polluted soild was revealed.

Keywords: soil-fungi, industrial pollution, micromycetes, dominant species, frequent species.

Both chemical and biological treatment of the substrate reduced the number of fungi propagules in cucumber rhizosphere and did not produce negative effect on cucumber growth. Biofungicides Mycostop (a.s. – Streptomyces griseoviridis) and Trichodermin (a.s. – Trichoderma spp.) are efficient against root rot in greenhouse cucumber and they did not differ in efficiency to compare with Previcur 607 SL. Mycostop and Trichoderma can be applied in greenhouse for plant protection against root rot agents in ecological production.

Keywords: cucumber, micromycetes, root rot, stem root, biofungicide, fungicide.

The inoculation efficiency of red clover, alfalfa, and vetch was investigated in the experiments established on slightly podzolized sod-podzolic loamy soil during 1993–1998. The effect of clover inoculation by Rhizobium strains was not very high: the yield of crude protein did not increase, but the amount of nitrogen was larger. For other legume plants the efficiency of inoculation was expressed by the increased yield of crude protein and amount of nitrogen. Meteorological conditions, especially humidity, had the greatest effect on the legumes inoculation by different Rhizobium strains. The link was determined between the number of nodules on the inoculated plants roots and percent of nitrogen: for clover r = 0.64, for alfalfa r = 0.44. A direct correlation was established between the yield of crude protein accumulated in legume plants and available contents of P2O5 and K2O in the soil. The coefficients of correlation were r = 0.70 and r = -0.60 for vetch, but the correlation for alfalfa was not determined.

Keywords: clover, alfalfa, vetch, rhizobium, inoculation, efficiency.

Thirty seven samples of calves nasal swabs were taken and examined for Mycoplasmas. Mycoplasmas have been isolated from seven samples (18.9 %). The isolates were identified as Mycoplasma bovis (8.1 %). Sixteen samples were taken from laboratory white mice lungs. Mycoplasma pulmonis was identified in 8 lung samples (50 %).

Keywords: calves, mice, mycoplasma, nasal swabs, lungs.

Investigations were carried out in order to evaluate the effects of under-usage of antimicrobials on bacteria. Field isolates of Enterobacteriaceae and quinolones, neomycin and amoxicillin were used. Five generations of bacteria were affected by media with antimicrobials. Development of resistance was determined. Results showed that effects achieved by using antimicrobials on bacteria may be various. Resistant strains of Enterobacteriaceae were cultivated together with a sensitive strain of Klebsiella pneumoniae. At the end of the experiment this strain did not acquire resistance.

Keywords: antimicrobial resistance, amoxicillin, bacteria, Enterobacteriaceae.

The most important bacterial diseases in Lithuanian pig breeding farms are salmonellosis and colibacillosis. Investigation was performed in 16 pig breeding farms. Examination was carried out using usual microbiological methods. The Salmonella was detected on the basis of international standard ISO 6579. Species were detected by biochemical properties. Serotypes were tested by agglutination with agglutination antiserum. The bacteriological tests revealed that pigs in the breading farms were mostly infected with the Salmonella and Escherichia coli strains. Salmonella choleraesuis and Escherichia coli O 108 K serotype were frequently isolated. Escherichia coli O 157 serotype was also detected. Other Enterobacteriaceae species are not important in causing swine diseases.

Keywords: enterobacteria, salmonellosis, colibacillosis, swine.

Cow‘s milk can be contaminated by microorganisms while in the udder. Further bacterial contamination depends on the conditions of milking, therefore, it can be reduced following the sanitary and hygiene requirements during milking and transportation. When milk samples are tested in a centralized way in an independent laboratory, it takes a certain period of time to collect and transport them, which affect the number of bacteria in them. To avoid this, the samples designated for microbiological analysis are preserved. The number of bacteria in them is determined by the bacterial analyser “Cobra”. The data of our experiments have demonstrated that the “Broad Spectrum Microtabs” preservative inhibits the growth of bacteria when milk is stored at various conditions within a three-day period, likewise similar effects can be achieved by acidiolli at 4 ?C only. At higher temperatures the growth of bacteria starts after 24 hours storage.

Keywords: raw milk, number of bacteria, preservative.

Response of natural and cultivated plants as well as microorganisms to air pollutions in Lithuania was estimated by methods of comparison. Influence of air pollutions upon the physiological processes of natural plants, cultivated plants and miroorganisms in different countries and in Lithuania was compared. The aim of the work was to study the influence of acid rain, sulphur dioxide, ozone, and carbon dioxide on physiological processes of plant and microorganisms: permeability of cuticle, stoma and membrane, on dark and light reactions of photosynthesis, photosyn-thesis intensity and transpiration, metabolism of organic matter, sensitivity and productivity of plants, external character of injurity.

Keywords: air pollutions, SO2, NO2, O3, pH, permeability, photosynthesis, respiration, transpiration, microorganisms.

In 1992–1997 laboratory, pot and field experiments, aiming to evaluate the resistance of different Rhizobium meliloti strains to soil acidity, were carried out. Possibility of artificial adaptation of strains to soil acidity was studied. For inoculation alfalfa was grown in derno-podzolic light loamy pHKCl – 4.0–5.3 (pot trials) and derno-podzolic gleyic light loamy pHKCl – 5.6–5.9 (field trials) soils. It was revealed that the minimal level of pH for Rhizobium meliloti was 4.6–5.0. Resistance of different strains to soil acidity makes from 57.8 to 81.2 % in comparison with the strains growing in neutral medium. Twenty selected strains of R. meliloti were being adapted to soil acidity, but only 7 strains acquired the resistance. Efficiency of plant inoculation depended on symbiotic activity of the strain and upon its resistance to soil activity. Application of the adapted strains A 425a and A 2M24 increased the yield of try matter up to 1.02–1.30 t/ha (28–35 % in pot trials) comparing with non-inoculated strains. The strains adapted to acid reaction increased the formation of nodules on alfalfa roots by 13 %.

Keywords: Rhizobium meliloti, strains, inoculation, adapted to soil acidity.

Eighteen species of powdery mildews belonging to 6 genera (Sphaerothera, Erysiphe, Microsphaera, Uncinula, Leveillula, Oidium) were identified in 1996–1998. Four powdery mildew species Microsphaera russellii, Leveillula taurica, Uncinula prunastri, and Oidium kalancho?ae were found for the first time. All the found fungi species were detected on 37 plants species that were ascertained for the first time in Lithuania as host-plants of powdery mildews.

Keywords: powdery mildews, host plants, anamorph, telemorph, Lithuania.