Influence of the molybdenum cofactor biosynthesis on anaerobic 1 respiration , biofilm formation and motility in Burkholderia 2 thailandensis 3

20 Burkholderia thailandensis is closely related to B. pseudomallei, a bacterial pathogen 21 and the causative agent of melioidosis. B. pseudomallei can survive and persist 22 within a hypoxic environment for up to one year and has been shown to grow 23 anaerobically in the presence of nitrate. Currently little is known about the role of 24 anaerobic respiration in pathogenesis of melioidosis. Using B. thailandensis as a 25 model, a library of 1,344 transposon mutants was created to identify genes required 26 for anaerobic nitrate respiration. One transposon mutant (CA01) was identified with 27 an insertion in BTH_I1704 (moeA), a gene required for the molybdopterin 28 biosynthetic pathway. This pathway is involved in the synthesis a molybdopterin 29 cofactor required for a variety of molybdoenzymes, including nitrate reductase. 30 Disruption of molybdopterin biosynthesis prevented growth under anaerobic 31 conditions, when using nitrate as the sole terminal electron acceptor. Defects in 32 anaerobic respiration, nitrate reduction, motility and biofilm formation were observed 33 for CA01. Mutant complementation with pDA-17::BTH_I1704 was able to restore 34 anaerobic growth on nitrate, nitrate reductase activity and biofilm formation, but did 35 not restore motility. This study highlights the potential importance of molybdoenzyme 36 dependent anaerobic respiration in the survival and virulence of B. thailandensis. 37