Isolation and characterisation of uncommon myxobacteria and other marine bacteria and their multilocus sequences analyses from Indonesia’s biodiversity Von der Fakultät für Lebenswissenschaften der Technischen Universität Carolo-Wilhelmina zu Braunschweig zur Erlangung des Grades einer Doktorin der Naturwissenschaften (Dr. rer. nat.) genehmigte D i s s e r t a t i o n von Rina Andriyani aus Bandung / Indonesia 1. Referentin oder Referent: Professor Dr. Michael Steinert 2. Referentin oder Referent: Privatdozent Dr. Joachim Wink eingereicht am: 01.03.2021 mündliche Prüfung (Disputation) am: 20.05.2021 Druckjahr 2021 The Entirely Merciful, the Especially Merciful This dissertation is dedicated to my beloved parents and our big family Vorveröffentlichungen der Dissertation Teilergebnisse aus dieser Arbeit wurden mit Genehmigung der Fakultät für Lebenswissenschaften, vertreten durch den Mentor der Arbeit, in folgenden Beiträgen vorab veröffentlicht: Posterbeiträge 1. R. Andriyani, K. I. Mohr, T. Mozef, M. Steinert, J. Wink, “Screening of Indonesian myxobacteria for new antibacterials and antifungals”, The 7th HIPS Symposium, Saarbrücken, 2017. 2. R. Andriyani, E. Sudarman, K. I. Mohr, T. Mozef, M. Steinert, J. Wink, “Compounds from Myxococcus sp. (MxGP53TM) a strain isolated from Indonesian soil”, The 10th International PhD Symposium, HZI, Braunschweig, 2017. 3. R. Andriyani, K. I. Mohr, T. Mozef, M. Steinert, J. Wink, et al., “A novel gliding bacterium of the family Flammeovirgaceae isolated from Bali Beach Indonesia”, The 11th International PhD Symposium, HZI Braunschweig, 2018. 4. R. Andriyani, K. I. Mohr, C. Risdian, T. Mozef, M. Steinert, J. Wink, “Drug discovery pipeline from a gliding bacterium to the isolation of antibacterial compounds by utilizing Indonesian biodiversity”, The 12th International PhD Symposium, HZI Braunschweig, 2019. 1 Preface and acknowledgements First of all, I would like to thank all of German-Indonesian Antiinfective Cooperation’s stakeholders in Germany and Indonesia. Thanks to the Federal Ministry of Education and Research, the Helmholtz Centre for Infection Research, the Graduate School of Helmholtz Centre for Infection Research and the Technische Universität Braunschweig in Germany. Thanks to the Ministry of Research and Technology of Indonesia, the Research Center of Chemistry and the Research Unit for Cleaning Technology in Indonesia. To Dr. Tjandrawati Mozef as the German-Indonesian Antiinfective Cooperation project’s lead collaborator from Indonesia who provided this opportunity and supported me. To the Head of Indonesian Institute of Sciences (LIPI), Dr. L.T. Handoko, the Head of Research Center of Chemistry at that time, Dr. Agus Haryono, the Head of Research Unit for Cleaning Technology (previous and present) Dr. Rike Yudianti and Dr. Ajeng Arum Sari, who gave me the eligibility for receiving this duty. Thanks to the staff as well. To the Director of Helmholtz Centre for Infection Research, the Director of the Graduate School of Helmholtz Centre for Infection Research, the Dean of The Faculty of Life Sciences of Technische Universität Braunschweig, the Head of Microbiology Department of Technische Universität Braunschweig and the staff as well who have assisted me for administration business during this study. Special thanks to my supervisors and thesis committee members, PD Dr. Joachim Wink and Prof. Dr. Michael Steinert. Without the Letter of Acceptance from you, it would have been impossible to receive the Riset-Pro scholarship from the Indonesian government to enrol in the PhD programme in the Technische Universität Braunschweig and the Graduate School of Helmholtz Centre for Infection Research. Also, to Dr. Kathrin I. Mohr and Dr. Raimo Franke as two of the thesis committee members. Thank you for your advice, support, keywords and guidance for my research. 2 To Prof. Marc Stadler as the Department of Microbial Drugs (MWIS) group leader and the staff for allowing me to work in the chemistry laboratory. To the team of Microbial Strain Collection (MISG): Stephanie Schulz, Klaus P. Conrad, Wera Collisi, Jolla Lulla, Birte Trunkwalter and Aileen Gollasch. To all the PhD students, the internship students and the postdoctoral researchers in the Helmholtz Centre for Infection Research, especially in the MISG and MWIS groups, also the PhD students from the Indonesian Institute of Sciences during this period. Thank you for the discussion moments and the togetherness on special occasions. To all teachers and lecturers who have shared their knowledge and broadened my perspective. It is not only about the science; thank you very much for your dedication and professionalism. To all friends everywhere, especially Christina Pape, Rani, Fitri, Okky, Yani, Rossy Chaerun Nisa, Rahma, Amelia, Mba Rere, Ulrike Tarajona, Mango, Uthay, Florian, Perda, Kikin, Astri, Khadija, Kwame, Tanya, Indonesian Moeslem Union of Braunschweig, and Indonesian Students Union of Braunschweig. Thank you for your support and friendship. Thanks to all authors in my references and Google, which made everything easier to find. I would like to thank Enago (www.enago.com) for the English language review. Last but not least, thanks to the Greatest who makes everything possible, has been guiding my soul to think clearly when I felt lost and doubtful. You always send answers, even in an unpredictable way. I have known You deeply in every chapter of my life. 3 CONTENTS Vorveröffentlichungen der Dissertation .............................................................................. 1 Posterbeiträge ...................................................................................................................... 1 Preface and Acknowledgements ......................................................................................... 2 Contents .............................................................................................................................. 4 Abbreviations ...................................................................................................................... 8 Chapter 1. Introduction ..................................................................................................... 11 1.1 Background ................................................................................................................. 11 1.2 Aims ............................................................................................................................ 12 1.3 Objectives ................................................................................................................... 12 1.4 Research location ........................................................................................................ 12 1.5 Research funding ........................................................................................................ 12 Chapter 2. Screening of culturable gliding bacteria isolated from some regions in Indonesia as sources for antiinfective substances .............................................................................. 13 2.1 Background ................................................................................................................ 13 2.2 Hypothesis................................................................................................................... 23 2.3 State of the art ............................................................................................................. 23 2.4 Materials and methods ................................................................................................ 24 2.4.1 Materials ................................................................................................................ 24 2.4.2 Methods.................................................................................................................. 24 2.4.2.1 Isolation of the bacteria .................................................................................... 24 2.4.2.2 Extraction, polymer chain reaction, and sequencing of DNA ......................... 26 2.4.2.2.a Extraction of Genomic DNA ...................................................................... 26 2.4.2.2.b Gel electrophoresis .................................................................................... 27 4 2.4.2.2.c Polymerase chain reaction .......................................................................... 27 2.4.2.2.d Strain identification .................................................................................... 27 2.4.2.3 Antiinfectives bioassay .................................................................................... 28 2.4.2.3.a Sample preparation ..................................................................................... 28 2.4.2.3.b Test strains preparation .............................................................................. 29 2.4.2.3.c Serial dilution test ....................................................................................... 29 2.4.3 Identification of the active compounds ................................................................. 29 2.5 Result and Discussion ................................................................................................. 30 2.5.1 Isolation of myxobacteria ...................................................................................... 30 2.5.2 Bioactivity screening and secondary metabolite analysis on gliding bacteria samples from Indonesia ................................................................................................