Complete genomes of mutualistic bacterial co-symbionts “Candidatus Sulcia muelleri” and “Candidatus Nasuia deltocephalinicola” of the rice green leafhopper Nephotettix cincticeps

ABSTRACT The genomes of obligate bacterial co-symbionts of the green rice leafhopper Nephotettix cincticeps, which is notorious as an agricultural pest, were determined. The streamlined genomes of “Candidatus Sulcia muelleri” and “Candidatus Nasuia deltocephalinicola” exhibited complementary metabolic pathways for synthesizing essential nutrients that contribute to host adaptation.

M utualistic relationship with endocellular microbial symbionts is indispensable for growth and reproduction of diverse insect species, especially those that rely on nutritionally unbalanced diets like plant sap, vertebrate blood, and woody material (1-4). Many plant sap-feeding hemipteran insects belonging to the Auchenorrhyncha harbor bacteriome-associated co-primary symbionts: the ancient obligate endocellular bacterial symbiont "Candidatus Sulcia muelleri, " (hereafter Sulcia) and another microbial partner (5). Despite genome reduction, the co-symbionts retain the capability of synthesizing essential amino acids in a complementary manner, plausibly reflecting their role in nutritional supply to their plant-sucking host insects (6).
The complete circular genomes of Sulcia and Nasuia were reconstructed from 223,209 and 23,316 paired reads, corresponding to 580-fold and 103-fold coverage, respectively. The genome of Sulcia was 192,344 bp with 23.7% G + C content. It encoded 186 putative protein-coding sequences (CDS), 30 transfer RNAs, and a single copy of 16S/23S/5S ribosomal RNA operon. The genome of Nasuia was 113,316 bp with 15.5% G + C content. It encoded 133 CDSs, 29 transfer RNAs, and a single ribosomal RNA operon. In Nasuia, the UGA codon was reassigned to tryptophan instead of the stop codon, as seen in other extremely reduced symbiont genomes (6).
Judging from the gene repertoire, Sulcia synthesizes 8 of 10 essential amino acids (Thr, Val, Leu, Ile, Lys, Phe, Trp, and Arg), while Nasuia is responsible for the remaining two (Met and His). It is noteworthy that methionine could be derived from the direct sulfhydrylation pathway (17). These results are consistent with prior studies highlighting the complementary nutritional roles of Sulcia and Nasuia (10,(18)(19)(20)(21). Our new symbiont genome data will contribute to comparative symbiont genomic studies that provide insight into how sophisticated insect-microbe mutualisms have evolved and diversified.

ACKNOWLEDGMENTS
We thank Hiroaki Noda for his pioneer work on the symbiotic bacteria of N. cincticeps and his kind support. We also would like to thank the staff of Comparative Genomics Laboratory at NIG for supporting genome sequencing.
This study was supported by the JSPS KAKENHI Grants JP17K15399, JP19H02973, and JP16H06279 (PAGS) to M.M. and the JST ERATO Grant JPMJER1902 to T.F.

DATA AVAILABILITY
The sequence data have been deposited to DDBJ under the accession numbers AP028057-AP028058 for assemblies and DRR489300-DRR489301 for raw reads.