Correspondence to cloescher@biology.sdu.dk Significance statementThis study addresses the important process of N 2 fixation, based on a whole-metagenome and -transcriptome screening, reports an increased diversity of N 2 fixing microbes in the sulfidic shelf water off Peru, as compared to previous target-gene based studies from the same waters. In addition to a generally higher diversity, genes encoding for alternative nitrogenases, which were previously not subject of any study on N 2 fixation in oxygen minimum zones, were detected. The ecological meaning and evolutionary history of those alternative nitrogenases is subject of ongoing debates, however, their presence in OMZ waters would allow for N 2 fixation at extreme anoxia, which may become important in a future ocean challenged by progressive deoxygenation. AbstractBiological dinitrogen (N 2 ) fixation is the pathway making the large pool of atmospheric N 2 available to marine life. Besides direct rate measurements, a common approach to explore the potential for N 2 fixation in the ocean is a mining based on molecular genetic methods targeting the key functional gene nifH, coding for a subunit of the nitrogenase reductase. As novel sequencing and single cell techniques improved, our knowledge on the diversity of marine N 2 fixers grew exponentially. However, to date one aspect of N 2 fixation in the ocean is commonly left aside. This is the existence of two alternative types of nitrogenases, which are besides the Mo-Fe nitrogenase Nif, the Fe-Fe nitrogenase Anf, and the V-Fe nitrogenase Vnf, which differ with regard to their metal co-enzymes, as well as regarding their operon structure and composition. nifH-based studies may thus be biased, and alternative nitrogenases could not be recovered. Here, we screened a set of 6 metagenomes and -transcriptomes from a sulfidic water patch from the oxygen minimum zone off Peru for genes involved in N 2 fixation. We identified genes related to all three nitrogenases, and generally increased diversity as compared to our previous nifH-based study from the same waters. While we could not confirm gene expression of the alternative nitrogenases from our transcriptomes, we detected additional diazotrophs involved in N 2 fixation. We suggest that alternative nitrogenases may not be used under conditions present in those waters, however, depending on trace metal limitation in the future they may become active.

<p>Both oxygenic and anoxygenic phototrophic bacteria (OPB and APB, respectively) are widely distributed in the ocean and play significant roles in carbon cycle and marine productivity. These organisms capture light as energy source via chlorophyll or bacteriochlorophylls-based photosystems. While OPB are relatively well studied, information on APB is rather scarce although they have been shown abundant in some ocean ecosystems and may play an important role in oxygen depleted environments. Here, we investigate the spatial profile of OPB and APB, gene abundance and expression of the key functional marker gene <em>pufM </em>(APB specific photosynthetic reaction center subunit M), in one fjord and three basins of the Baltic Sea using 16S rRNA amplicon sequencing and qPCR. Among the microbial community, abundances of OPB and APB were found to be similar thus emphasizing a potential importance of APB, with APB representing 1.6-17.5% and OPB representing 0.5-20%. Among APB, we identified eleven different orders, with <em>Rhodobacterales</em> being quantitatively dominant. The identified seven orders of OPB were dominated by <em>Synechococcales</em>. OPB were more abundant than APB in surface waters (<8m), while APB were comparably more abundant in deeper waters. Besides a depth-dependent distribution, we observed an impact of salinity on the distribution of APB and OPB, both of which being suggestive of distinct niches for those primary producer clades. <em>pufM</em> gene abundance ranged from 10<sup>4</sup> to 10<sup>9</sup> copies/L, with highest counts detectable in the mixed layer (<40m), however, even in deeper waters where gene abundances decreased APB <em>pufM</em> gene expression was high with up to 10<sup>4</sup> copies/L. These results indicate APB may play a more important role in marine primary productivity which has been underestimated before. </p>