Metagenomic bins and biosynthetic gene clusters in gut bacteria of turtle ants

Cephalotes are herbivorous ants (>115 species) feeding on low-nitrogen food sources and they rely on gut symbionts to supplement their diet in nutrients by recycling nitrogen food waste into amino acids. These conserved gut symbionts, composed of five bacterial orders, have been studied previously for their primary nitrogen metabolism, however little is known about their ability to biosynthesize specialized metabolites which can play a role in bacterial interactions between communities living in close proximity in the gut. We investigated the diversity of biosynthetic gene clusters (BGCs) producing specialized metabolites in the genomes and metagenomes of conserved gut symbionts by studying 17 Cephalotes species collected across several geographical areas. Our results reveal that (1) mining metagenomes and genomes show complementary results to retrieve BGCs especially when bacterial isolates are difficult to culture, (2) the conserved gut symbionts involved in the nutritional symbiosis have a large diversity of BGCs of different chemical families, (3) the phylogenetic analysis of BGCs encoding the production of arylpolyenes, non-ribosomal peptides (NRP), polyketides (PK), and siderophores shows high similarity between BGCs of a single symbiont across different ant host species, and between BGCs originated from different bacterial orders within a single host species. Additionally, the diversity of BGCs was found in four of the five conserved symbionts co-occurring in the hindgut except for one major player (Opitutales) localized alone in the midgut and lacking BGCs. This spatial isolation prevents direct interaction of Opitutales with other symbionts and suggesting that BGCs have an essential role for symbionts living in close proximity. These findings together pave the way for studying the mechanisms of BGCs conservation and evolution in gut symbionts genomes and the role of bacterial specialized metabolites involved in multipartite mutualism with Cephalotes turtle ants.

龟蚁属（Cephalotes）为植食性蚂蚁，目前已被描述的物种超过115个，它们以低氮食物为食，并依赖肠道共生菌将氮源食物废物循环转化为氨基酸，以此补充膳食营养缺口。这些由5个细菌目组成的保守肠道共生菌，其核心氮代谢功能此前已得到深入研究，但学界对其合成特殊代谢物的能力却知之甚少——这类代谢物可在肠道内近距离定植的菌群间发挥细菌互作功能。本研究针对采集自多个地理区域的17个龟蚁属物种，对其保守肠道共生菌的基因组与宏基因组中负责合成特殊代谢物的生物合成基因簇（biosynthetic gene clusters, BGCs）多样性展开了调查。研究结果显示：（1）宏基因组与基因组联合挖掘可互为补充，以获取生物合成基因簇，尤其在细菌分离培养难度较高的场景下优势显著；（2）参与营养共生的保守肠道共生菌拥有丰富多样的不同化学家族的生物合成基因簇；（3）对编码芳基多烯、非核糖体肽（non-ribosomal peptides, NRP）、聚酮（polyketides, PK）以及铁载体（siderophores）的生物合成基因簇进行系统发育分析后发现，同一共生菌在不同蚁宿主物种中的生物合成基因簇具有高度相似性，且同一宿主物种内不同细菌目来源的生物合成基因簇同样相似度极高。此外，除了单独定殖于中肠且缺乏生物合成基因簇的关键共生类群——奥氏杆菌目（Opitutales）外，在后肠中共存的5种保守共生菌中有4种均携带生物合成基因簇。这种空间隔离使得奥氏杆菌目无法与其他共生菌发生直接互作，这也暗示生物合成基因簇对近距离定植的共生菌具有至关重要的作用。上述研究结果共同为探索肠道共生菌基因组中生物合成基因簇的保守与演化机制，以及特殊代谢物在龟蚁属蚂蚁与菌群间多元共生关系中的功能奠定了研究基础。