Uropygial gland secretions of birds consist of host and bacteria derived

compounds and play a major sanitary and feather-protective role. Here we

report on ourmicrobiome studies of theNew Guinean toxic bird Pachycephala

schlegelii and the isolation of a member of the Amycolatopsis genus from the

uropygial gland secretions. Bioactivity studies in combination with co-cultures,

MALDI imaging and HR-MS/MS-based network analyses unveil the basis

of its activity against keratinolytic bacteria and fungal skin pathogens.We trace

the protective antimicrobial activity of Amycolatopsis sp. PS_44_ISF1 to the

production of rifamycin congeners, ciromicin A and of two yet unreported

compound families.We performNMR and HR-MS/MS studies to determine the

relative structures of six members belonging to a yet unreported lipopeptide

family of pachycephalamides and of one representative of the demiguisins, a

new hexapeptide family. We then use a combination of phylogenomic, transcriptomic

and knock-out studies to identify the underlying biosynthetic gene

clusters responsible for the production of pachycephalamides and demiguisins.

Our metabolomics data allow us to map molecular ion features of the

identified metabolites in extracts of P. schlegelii feathers, verifying their presence

in the ecological settingwhere they exert their presumed active role for

hosts. Our study shows thatmembers of the Actinomycetotamay play a role in

avian feather protection.