Penka Moncheva

Biodiversity and Incidence of Burkholderia Species

ABSTRACT The members of genus Burkholderia are various microorganisms, which occupy surprisingly wide range of ecological niches—soil, water (including marine water), rhizosphere, animals and human. The genus includes over 30 species that are important causes of human, animal and plant diseases, as well as organisms useful in promoting plant growth and bioremediation. Extremely important is the question if the “good” and the “bad” strains are equivalent in their nature.

The composition of the microflora of boza, an original bulgarian beverage

ABSTRACT A total of 293 strains of lactic acid bacteria and 78 yeast strains from original Bulgarian boza were isolated. The rod-shaped strains were identified as Lactobacillus salivarius, Lactobacillus sakei, Lactobacillus maltaromicus, Lactobacillus fermentum, Lactobacillus parabuchneri, Lactobacillus paracasei subsp. paracasei and Weissella confusa. The strains of genus Leuconostoc were identified as Leuconostoc lactis, Leuconostoc amelibiosum, Leuconostoc mesenteroides subsp. dextranicum and Leuconostoc pseuclomesenteroides. The yeasts isolated belonged to the species Saccharomyces cerevisiae, Pichia membranifaciens, Dekkera bruxellensis and Nadsonici commutata. During the storage of boza the total number of the lactic acid bacteria increased slightly up to 48 h, while the n umber of yeasts increased considerably. The changes in pH and total acidity during the storage of boza were in accordance with the changes in viable count of lactic acid bacteria and their activity.

Differentiation of Xanthomonas spp. Causing Bacterial Spot in Bulgaria Based on Biolog System.

During the last 20 years, the causative agents of bacterial spot of tomato and pepper have been subjected to many studies and reclassifications. According to the current data, the species are four (X. euvesicatoria, X. vesicatoria, X. gardneri, and X. perforans) and cause similar symptoms in plants but possess different phenotypic properties. This work provides the full metabolic characteristics obtained by Biolog system of bacterial spots xanthomonads based on a large selection of strains from different vegetable-producing regions of Bulgaria with accent on their major differentiating properties which could be used for species differentiation by metabolic profiles. The results are compared to the data available in the literature in order to clarify the strong features of each species and distinguish the variable ones. Simple characteristics like amylase activity and utilization of cis-aconitate cannot serve alone for differentiation.

Evidence for Proteolytic Activity of Lactobacilli Isolated from Kefir Grains

ABSTRACT The proteolytic activity of 276 natural isolates from kefir grains was investigated. Evidence for extracellular proteolytic activity was demonstrated for 49 Lactobacillus strains. One strain, Lactobacillus kefir DR22x, was selected as a producer of proteinases. Lactobacillus kefir DR22x strain produces a cell-wall-bound proteinase. The proteinase was removed from the cell envelope by washing the cells with a Ca2+- free buffer. The crude proteinase extract showed the highest activity at pH 7.2 and 37 °C. The proteolytic activity was shown to be maximal in the late exponential growth. A study using several protease inhibitors suggested that this activity is associated with serine-type proteases. Considering the substrate specifity, the enzyme is similar to the lactococcal P1—type proteinases, since it completely hydrolysed only β-casein, showing very low activity towards α-casein.

Antibacterial potential of streptomycete strains from Antarctic soils.

The exploration of habitats with unusual environment and poorly explored areas such as Antarctica is one of the strategies for discovery of new biologically active substances and/or new producers. The aim of this study was to identify the actinomycetes isolated from the soils of the island Livingston – Antarctica and to investigate their potential to synthesize antibacterial agents against phytopathogens. Twenty-three actinomycete strains were the object of this study. Using PCR (polymerase chain reaction) amplification all strains were affiliated to genus Streptomyces. The sequencing of the 16S rRNA for three of the strains showed greatest similarity to Streptomyces tendae for one of them, and revealed that the other strains had closest relations to streptomycetes isolated from anthropogenically unaltered regions including Antarctica. The isolates were studied for production of antibacterial substances both by molecular and culture methods. PCR targeting specific biosynthetic genes involved in the production of some groups of antibiotics was performed. The screening showed that all strains possessed the gene for Type-II polyketide synthase, 11 strains – for non-ribosomal peptide synthetase; 6 strains – for polyene antibiotics; and 4 strains – for glycopeptide antibiotics. The production of antibacterial substances by the strains was tested in vitro against phytopathogenic bacteria. The strains differed in the number of inhibited test – bacteria and in their spectrum of action. Four strains showed a wide range of activity against Gram-positive and Gram-negative phytopathogens. The results obtained revealed that the Antarctic soils are potential source for isolation of streptomycetes producing antibiotics from different groups.

New host plants of Erwinia amylovora in Bulgaria

Nine strains of Erwinia amylovora were isolated from new host plants in Bulgaria D chokeberry and strawberry. The strains were characterized morphologically and biochemically using the API 20E and BIOLOG system. It was established that they showed three different API 20E metabolic profiles, not found by previous studies of E. amylovora. All strains were identified as E. amylovora due to their metabolic fingerprint patterns obtained by the BIOLOG system. The identification was confirmed by PCR amplification of a specific region of plasmid pEA29 and genome ams-region. This study is the first characterization and identification of E. amylovora strains isolated from chokeberry and strawberry by the API 20E and BIOLOG system and by polymerase chain reaction.

Proteolytic and antimicrobial activity of lactic acid bacteria grown in goat milk

We examined 62 strains and 21 trade starter cultures from the collection of LB Bulgaricum PLC for proteolytic and antimicrobial activity of lactic acid bacteria (LAB) grown in goat milk. The aim of this study was to investigate the fermentation of caseins, α-lactalbumin and β-lactoglobulin by LAB, using the o-phthaldialdehyde (OPA) spectrophotometric assay and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The proteolysis targeted mainly caseins, especially β-casein. Whey proteins were proteolyzed, essentially β-lactoglobulin. The proteolytic activity of Lactococcus lactis l598, Streptococcus thermophilus t3D1, Dt1, Lactobacillus lactis 1043 and L. delbrueckii subsp. bulgaricus b38, b122 and b24 was notably high. The proteolysis process gave rise to medium-sized peptide populations. Most of the examined strains showed antimicrobial activity against some food pathogens, such as Escherichia coli, Staphylococcus aureus, Salmonella cholere enteridis, Listeria monocytogenes, Listeria innocua and Enterobacter aerogenes. The most active producers of antimicrobial-active peptides were strains of L. delbrueckii subsp. bulgaricus and S. thermophilus, which are of practical importance. The starter cultures containing the examined species showed high proteolytic and antimicrobial activity in skimmed goat milk. The greatest antimicrobial activity of the cultures was detected against E. aerogenes. The obtained results demonstrated the significant proteolytic potential of the examined strains in goat milk and their potential for application in the production of dairy products from goats milk. The present results could be considered as the first data on the proteolytic capacity of strains and starter cultures in goat milk for the purposes of trade interest of LB Bulgaricum PLC.

Physiological diversity of bacterial communities from different soil locations on Livingston Island, South Shetland archipelago, Antarctica

Terrestrial food webs of Antarctica are simple and dominated by microorganisms. Soil bacteria play an important role in nutrient cycling, yet little is known about their capacity to utilize different carbon sources and to participate in site nutrient turnover. Biolog EcoPlate™ was applied to study the catabolic activity and physiological diversity of bacteria inhabiting the soil of moss, vascular plants, and fell field habitats from Livingston Island, Antarctica. Additionally, the number of oligotrophic and copiotrophic bacteria was counted by the agar plate method. Results indicated a lack of site-specific distribution of bacterial abundance, in contrast to bacterial catabolic activity and community level physiological profiles. Community level physiological profiles revealed a common capacity of soil bacteria to intensively utilize polyols, which are cryoprotectants widely produced by Antarctic organisms, as well as site-specific phenolic compounds (vegetated habitats), amino acids/amines (moss habitats), carbohydrates and carboxylic acids (fell field habitat). It was concluded that the physiology of soil bacteria is habitat specific concerning both the rate of catabolic activity and pattern of carbon source utilization.

Phenotypic diversity of Erwinia amylovora in Bulgaria.

Fifty-one strains of Erwinia amylovora isolated from nine host plants in Bulgaria were characterized phenotypically and identified by the API 20E and BIOLOG system. The identification was confirmed by PCR amplification of a specific region of the plasmid pEA29 and the genome ams region. The phenotypic diversity of the strains was studied on the basis of their API 20E and BIOLOG metabolic profiles, as well as of their SDS-PAGE protein profile. Metabolic diversity among the strains was established, but no connection with the origin of the strains was revealed. The Bulgarian strains showed API 20E metabolic profiles not found in previous studies of E. amylovora. The strains formed a homogenous group on the basis of their protein profiles. All the strains were sensitive to the antibiotics streptomycin, tetracycline and oxytetracycline. This study was an initial step towards an investigation of the diversity and evolution in the Bulgarian population of E. amylovora, and it was the first characterization of E. amylovora strains isolated from different host plants in the period 1995-2005 in Bulgaria.

DIVERSITY OF XANTHOMONAS SPP. CAUSAL AGENTS OF BACTERIAL SPOT ON PEPPER AND TOMATO PLANTS IN BULGARIA

ABSTRACT Seventy-one Xanthomonas strains isolated from infected pepper and tomato plants with bacterial spot from Bulgaria and Macedonia were examined in this study. The pathotypes and races of the strains were determined. The pepper-tomato pathotype predominated among Bulgarian strains, whereas the Macedonian strains belonged to the pepper pathotype. On the basis of their phenotypic characteristics the causal agents of bacterial spot on tomato and pepper possessed properties of at least three species (X. euvesicatoria, X. vesicatoria/X. perforans and X. gardneri). The identification of thirty strains to species Xanthomonas euvesicatoria was confirmed using species specific primers in PCR amplification.