{"id":2241,"date":"2018-10-30T15:15:25","date_gmt":"2018-10-30T12:15:25","guid":{"rendered":"http:\/\/microbio.bas.bg\/wordpress\/?page_id=2241"},"modified":"2018-10-30T15:25:54","modified_gmt":"2018-10-30T12:25:54","slug":"results","status":"publish","type":"page","link":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/en\/projects\/project-d03-100-05-06-2015\/results\/","title":{"rendered":"Results"},"content":{"rendered":"

Results:<\/strong><\/p>\n

Stefanova K., Tomova I.,\u00a0Tomova A.,\u00a0Radchenkova N., Atanassov I., Kambourova\u00a0<\/sup>M.\u00a0<\/strong>Archaeal and bacterial diversity in two hot springs from geothermal regions in Bulgaria as demostrated by 16S rRNA and GH-57 genes.\u00a0International Microbiology (in press), IF 1.326.<\/span><\/p>\n

Margarita Stoilova-Disheva, Dimitrina Lyutskanova, Ivanka Boyadzhieva, Nadja Radchenkova, Margarita Kambourova. Unusually high archaeal diversity in Vlasa hot spring, Velingrad, Bulgaria, revealed by phylogenetic analysis. Participation in Seminar of Ecology with International Participation \u2013 2016, Sofia, 21-22 April, 2016.<\/p>\n

 <\/p>\n

I N T E R M E D I A T E\u00a0\u00a0 R E P O R T\u00a0\u00a0 1<\/strong><\/p>\n

On the project D03-100 \/ 05.06.2015<\/p>\n

Programme of the Financial Mechanism of the European Economic Area (EEA grants), measure „Projects for inter-institutional cooperation“<\/p>\n

DIVERSITY AND BIOTECHNOLOGICAL POTENTIAL OF ARCHAEA FROM BULGARIAN HOT SPRINGS<\/span><\/p>\n

For the period 04.09.2015-31.12.2015<\/p>\n

1. Billboard<\/strong><\/p>\n

In implementation of the Plan for publicity a billboard indicating the financial support of EEA program was installed at the entrance of the Institute.<\/span><\/p>\n

2. Website<\/strong><\/p>\n

A page devoted to EEA program was created on the website of the Institute of Microbiology, BAS (www.microbio.bas.bg\/wordpress\/index.php\/en\/projects\/project-d03-100-05-06-2015\/, which includes information on:<\/span><\/p>\n

(A) the program operator<\/span><\/p>\n

(B) the beneficiaries of the grant and the contact person<\/span><\/p>\n

(C) the funding with clear indication of the mechanism of sponsorship<\/span><\/p>\n

(D) planned activities in the project by EEA program<\/span><\/p>\n

(E) cooperation with staff from the donor country, information on the work of young Bulgarian scientist at Norwegian team<\/span><\/p>\n

(F) the role of the donor country in project implementation<\/span><\/p>\n

3. Project Management<\/strong><\/p>\n

This activity involved the following activities:<\/span><\/p>\n

Corr. Memb. Hristo Naydenski, Director – Meeting with the EEA governing body, organization of procurement for supply of chemicals and consumables, a proposal for modification of the budget.<\/span><\/p>\n

Antoaneta Tsareva, Ch. Accounter – Processing of financial documents, preparation of lists procurement, preparation of financial statements and contacts with the governing body.<\/span><\/p>\n

4. Research activity<\/strong><\/p>\n

Activity 2,\u00a0Sample collection<\/strong><\/span><\/p>\n

Springs with different geothermal origin and different properties of water, suggesting divergence of their inhabitants in them were selected for analysis of the archaeal biodiversity, namely Mizinka and Vlasa, Velingrad, and Sapareva banja. The trip to Velingrad was carried out on 5-6.10.2015 and to Sapareva banja – 31.10-1.11.2015. One liter of each sample water was analyzed for the chemical composition. The analysis was carried out in a trade company Dial LTD.<\/span><\/p>\n

Activity 3,\u00a0Isolation of DNA<\/strong><\/span><\/p>\n

The successful detection and characterization of microbial 16S rDNA in natural samples require extraction of high molecular weight DNA and its proper disposal of pollutants such as certain organic acids and ions, which inhibit the next stage of amplification.<\/span><\/p>\n

The procedure for the isolation of total DNA included filtration of water, treatment of the filter with a lysis solution, RNase, protease, and DNA purification. As a result of the implementation of this phase high molecular total DNA was successfully isolated.<\/span><\/p>\n

Activity 4,\u00a0\u0410mpli<\/strong>fication of 16S rRNA gene and\u00a0initialization the\u00a0construction of\u00a0clone\u00a0libraries<\/strong><\/span><\/p>\n

A, Amplification of 16S rRNA gene<\/span><\/p>\n

Using the 16S rRNA gene as a marker\u00a0gene\u00a0allows characterization\u00a0the structure of microbial communities in nature and reaching\u00a0the\u00a0real microbial\u00a0diversity. Archaeal DNAs were amplified with universal primers for archaea 21F and 958R.\u00a0PCR products \u00a0were\u00a0observed into two samples,Vlasa hot spring\u00a0and Sapareva banja.\u00a0A product was not observed\u00a0in the sample\u00a0from\u00a0Mizinka.<\/span><\/p>\n

B,\u00a0Ligation of\u00a0the archaeal\u00a016S rRNA into a plasmid and transformation\u00a0into E. coli cells<\/span><\/p>\n

\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0The amplified 16S rRNA genes were ligated into\u00a0a\u00a0vector and the plasmid was transformed into competent cells.\u00a0More than 200 clones were selected and tested for correctness of the insert.<\/span><\/p>\n

\u00a0\u00a0C,\u00a0Grouping\u00a0of\u00a0clones according to their restriction profiles<\/span><\/p>\n

The diversity\u00a0in the clone\u00a0library was\u00a0preliminary\u00a0assessed by ARDRA analysis (Amplified Ribosomal DNA Restriction Analysis). To establish the correct polymorphism between cloned 16S rRNA genes, they were subjected to restriction with two 4-basic enzyme – MspI and HaeIII. Based on the diversity in the profiles obtained after restriction of 16S rDNA, 28 correct\u00a0clones were allocated to 14 groups.<\/span><\/p>\n

D,\u00a0Sequencing<\/span><\/p>\n

Amplified PCR product of one representative from each restriction group was\u00a0sent for sequencing,\u00a0a total of 14 PCR product. Additionally, \u00a0six arhaeal\u00a0clones from\u00a0hot spring Levunovo were sent for sequencing. Sequencing was conducted in Macrogen,\u00a0South Korea. An\u00a0analysis of\u00a0the\u00a0results\u00a0and further work with the rest clones from the library is forthcoming.\u00a0\u00a0The genes of the remaining\u00a0clonesof\u00a0Vlasa\u00a0library will be amplified\u00a0in the\u00a0next stage, they\u00a0will be grouped according to their restriction profiles, sequenced and analysed.\u00a0Based on the results\u00a0received the structure of archaeal community will be revealed.<\/span><\/p>\n

\u00a0<\/span><\/p>\n

The following team members have participated in the implementation of activities 2, 3 and 4:<\/span><\/p>\n

1. Assoc. Prof. Margarita Kambourova<\/span><\/p>\n

2. Assis.Prof. Nadia Radchenkova<\/span><\/p>\n

3. Assis. Prof.\u00a0 Ivanka Boyadjieva<\/span><\/p>\n

4. Assis.Prof. Margarita Stoilova-Disheva<\/span><\/p>\n

5. Assis. Prof. Dimitrina Lyutskanova<\/span><\/p>\n

 <\/p>\n

13\/01\/2016 \u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0Project leader:<\/span><\/p>\n

Assoc. Prof. M.Kambourova<\/span><\/p>\n

Director\u00a0IMicB:<\/span><\/p>\n

Cor. Mem. H. Naydenski<\/span><\/p>\n

 <\/p>\n

 <\/p>\n

I N T E R M E D I A T E\u00a0\u00a0 R E P O R T\u00a0 2<\/strong><\/p>\n

For the project D03-100 \/ 05.06.2015<\/span><\/p>\n

Programme of the Financial Mechanism of the European Economic Area (EEA grants), measure „Projects for inter-institutional cooperation“<\/span><\/p>\n

 <\/p>\n

DIVERSITY AND BIOTECHNOLOGICAL POTENTIAL OF ARCHAEA FROM BULGARIAN HOT SPRINGS<\/span><\/p>\n

 <\/p>\n

For the period\u00a0<\/span>01.01.2016-30.04.2016<\/strong><\/p>\n

\u00a0<\/strong><\/p>\n

Website<\/strong><\/p>\n

The page on the website of the Institute of Microbiology, BAS, dedicated to the project has been updated (www.microbio.bas.bg),http:\/\/microbio.bas.bg\/IMB%20web%20page\/microbio\/Norwegia_Priekt_Kamburova_bg.html<\/a>, in which the next information was included:<\/span><\/p>\n

(A) The publication activity of the group and participation in a conference<\/span><\/p>\n

(B) First and second 4-month reports<\/span><\/p>\n

(C) Gallery of photos demonstrating the research activity of the team<\/span><\/p>\n

 <\/p>\n

Activity 1, Project Management<\/strong><\/p>\n

Corr. Mem. H. Naydenski, Director \u2013 Control of the project implementation.<\/span><\/p>\n

Antoaneta Tsareva, Ch. Accountant – Financial records, preparation of financial statements, modification of the budget.<\/span><\/p>\n

Assoc. Prof. DSc M. Kambourova, Project Manager:<\/span><\/p>\n

– Meetings of the commission for consideration, evaluation and ranking of participants in the procurement for delivery of materials, chemicals and consumables were organized.<\/span><\/p>\n

– Modification of the project budget<\/span><\/p>\n

– Procedure for granting a loan from BAS to cover expenses until the last installment of the contract.<\/span><\/p>\n

– Participation in the organized on 30.03.2016 meeting with the management of the EEA Program operator BG09, in which discussion on the progress on the projects implementation was performed.<\/span><\/p>\n

 <\/p>\n

Activity 2, Experimental work<\/strong><\/p>\n

– Members of the team together with Norwegian partner Prof. Birkeland visited hot springs in southwestern Bulgaria and in the region of Velingrad. Samples for isolation of culturable archaea were taken.<\/span><\/p>\n

– PhD student Nicoleta Boteva (team member) has achieved growth of mixed cultures from 9 samples in Norwegian laboratory. Their ability to synthesize biotechnologically valuable enzymes (xylanase, cellulase, gellan lyase) was established. She is working on the isolation of pure strains that will be sent to the Bulgarian laboratory for characterization.<\/span><\/p>\n

– The work on the construction of clone library with DNA isolated from a hot spring Vlasa, Velingrad continued in Bulgarian laboratory. The selected clones were splited into 57 groups according to the identity of their restriction profiles. One representative from each group was sent for sequencing in a Korean company Macrogen. The team successfully works with chemicals and consumables borrowed from colleagues from the institute and other scientific organizations.<\/span><\/p>\n

– A contract for suppling a system for anaerobic cultivation of archaeal strains has been signed. Incoming system components are assembled.<\/span><\/p>\n

\u00a0<\/span><\/p>\n

Activity 3, Exchange of visits<\/strong><\/p>\n

– The Norwegian partner Nils-Kare Birkeland visited Bulgaria in the period 3-7.01.2016. He participated in trips for collecting samples from hor springs in southwestern Bulgaria and the region of Velingrad. He met the working team and an interesting discussion on the specifics in archaeal cultivation took place.<\/span><\/p>\n

– The team member PhD student Nicoleta Boteva left on 10.01.2016 for 6 months work on the project in the laboratory of Prof. Nils-Kare Birkeland, Faculty of Biology, centered on Geobiology, University of Bergen, Bergen, Norway.<\/span><\/p>\n

\u00a0<\/strong><\/p>\n

Activity 4, Information and Publicity<\/strong><\/p>\n

– Results obtained from clone libraries of 16S rRNA gene and amylase gene from Levunovo and Vetren dol were summarized in a publication submitted and accepted for print in the journal International Microbiology.<\/span><\/p>\n

Stefanova K., Tomova I., Tomova A., Radchenkova N., Atanassov I., Kambourova M. Archaeal and bacterial diversity in two hot springs from geothermal regions in Bulgaria as demostrated by 16S rRNA and GH-57 genes. International Microbiology (in press), IF 1.326.<\/span><\/p>\n

– The team participated in the „Seminar on Environment – 2016“ with international participation with poster:<\/span><\/p>\n

Margarita Stoilova-Disheva, Dimitrina Lyutskanova, Ivanka Boyadzhieva, Nadja Radchenkova, Nicoleta Boteva, Margarita Kambourova. Unusually high archaeal diversity in Vlasa hot spring, Velingrad, Bulgaria, revealed by phylogenetic analysis. Seminar on ecology with international participation – 2016, Sofia, 21-22.04.2016.<\/span><\/p>\n

 <\/p>\n

30\/04\/2016 \u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0\u00a0Project leader:<\/span><\/p>\n

Assoc. Prof. M. Kambourova<\/span><\/p>\n

Director IMicB:<\/span><\/p>\n

Cor. Mem. H. Naydenski<\/span><\/p>\n

 <\/p>\n","protected":false},"excerpt":{"rendered":"

Results: Stefanova K., Tomova I.,\u00a0Tomova A.,\u00a0Radchenkova N., Atanassov I., Kambourova\u00a0M.\u00a0Archaeal and bacterial diversity in two hot springs from geothermal regions in Bulgaria as demostrated by 16S rRNA and GH-57 genes.\u00a0International Microbiology (in press), IF 1.326. Margarita Stoilova-Disheva, Dimitrina Lyutskanova, Ivanka Boyadzhieva, Nadja Radchenkova, Margarita Kambourova. Unusually high archaeal diversity in Vlasa hot spring, Velingrad, Bulgaria, […]<\/p>\n","protected":false},"author":1,"featured_media":0,"parent":2229,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"","meta":[],"_links":{"self":[{"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/pages\/2241"}],"collection":[{"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/comments?post=2241"}],"version-history":[{"count":4,"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/pages\/2241\/revisions"}],"predecessor-version":[{"id":2245,"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/pages\/2241\/revisions\/2245"}],"up":[{"embeddable":true,"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/pages\/2229"}],"wp:attachment":[{"href":"https:\/\/microbio.bas.bg\/wordpress\/index.php\/wp-json\/wp\/v2\/media?parent=2241"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}