Deep-sea hydrothermal vents and cold seeps are chemosynthetic ecosystems characterized by high pressure, low oxygen, absence of light, and rich in reducing gases. In these completely lightless ecosystems, primary producers are not photosynthetic organisms commonly found on land, but chemoautotrophic bacteria that can utilize hydrogen sulfide, hydrogen, methane, etc. released from deep-sea hydrothermal vents or cold seeps. Shrimps of the family Alvinocarididae are endemic species of deep-sea chemosynthetic ecosystems. Like many other invertebrates in these ecosystems, alvinocaridid shrimps harbor important chemosynthetic bacterial communities in the gill chamber and gut. Different alvinocaridid species occupy different ecological niches in the Okinawa Through hydrothermal region, with Shinkaicaris leurokolos inhabiting the chimney walls of hydrothermal vents and Alvinocaris longirostris inhabiting the mussel beds far from vents. Moreover, the A. longirostris is the only alvinocaridid shrimps found co-distributed in hydrothermal vent and cold seep environment. Therefore, alvinocaridid shrimps have become an excellent model to study the adaptive evolution mechanism of extreme deep-sea chemosynthesis environment.
In this study, the full-length transcriptomes of S. leurokolos and A. longirostris in the Okinawa Through hydrothermal vent were sequenced and compared by using a combination of single-molecule real-time (SMRT) and Illumina RNA-seq technology technology. In total, 16,037 and 13,666 unigenes were obtained from S. leurokolos and A. longirostris, respectively, of which 11,718 (73.07%) and 10,243 (74.95%) unigenes were annotated with NR, NT, KOG, KEGG, KO, and GO databases. The GC contents of the transcriptomes were 46.5% and 45.8% in S. leurokolos and A. longirostris. Further analysis revealed that genes related to sulfide and heavy metal detoxification metabolism (SQR, ETHE1, TST, and FRI) were highly expressed in S. leurokolos, while more immune-related genes (BGBP, CHIT, CHIA and ALPS) were highly expressed in A. longirostris. In addition to this, genes related to oxidative and heat stress (SODM, GST, GPX, HSP70, and HSP7D) were significantly expanded in S. leurokolos, while genes related to immune defense genes (CHIA and CHIT) were significantly expanded in A. longirostris. Eventually, six genes involved in DNA damage repair-related genes (HIPK2, INT6, HEAT3, RUVB2, SMC1A, and SIR1), five genes involved in oxidative stress (CCD51, RT05, DHC24, IMP2L, and VA0E) and five genes involved in temperature stresses (PPID, EXL1, L2EFL, NU155, and TCPQ) were subjected to positive selection in S. leurokolos. These features may be related to the adaptability of S. leurokolos and A. longirostris to their microenvironments.
The symbionts of alvinocaridid shrimps also play an important role in helping hosts adapt to extreme environments. The third-generation full-length 16S amplicon sequencing technology was used to analyze the community composition and diversity of bacteria associated with the gills of the A. longirostris and S. leurokolos from the Okinawa Through hydrothermal vent and the A. longirostris from cold seeps in the South China Sea. In the end, a total of 568 OTUs were obtained by clustering, the number of OTUs on the gills of the cold seep A. longirostris, was the largest (average 208 per sample), and the number of OTUs in the hydrothermal vent S. leurokolos was the least (average 92 per sample). The analysis of diversity showed that the diversity of the gill-associated bacteria of the cold seep A. longirostris were the highest. In contrast, the lowest diversity was found in the hydrothermal vent S. leurokolos, and the gill-associated bacterial communities of cold seep A. longirostris and hydrothermal vent A. longirostris were more similar in composition. The difference in diversity may be due to the milder and more chemically stable environment of cold seeps than those of hydrothermal vents, and active selection by the host may also play an important role.
Further annotation found that the three groups of shrimp gill-associated bacteria were mainly composed of bacteria from the classes Campylobacteria, Gammaproteobacteria, and Alphaproteobacteria. However, at the family level, hydrothermal vent S. leurokolos gill-associated bacteria consisted almost entirely of sulfur oxidation-related Sulfurovaceae family (94.06%), and excluding Sulfurovaceae family (69.21%) in hydrothermal vent A. longirostris gill bacteria, Gammaproteobacteria un-group（14.37%）and Thiotrichaceae family (6.77%) involved in sulfur oxidation also accounted for a relatively high abundance. In comparison, the relative abundance of Methylomonadaceae family with methane oxidation capacity on the gills of cold seep A. longirostris was significantly more than that of the two hydrothermal vent A. longirostris and S. leurokolos (19.38% vs. 0.32% vs. 0.01%). In conclusion, we suggest that physicochemical factors of the habitat environments and the specific stable recognition of bacteria by the hosts may co-contribute to the composition of the gill-associated bacteria in alvinocaridid shrimps.
In summary, this study illustrates the molecular basis of adaptive evolution in alvinocaridid shrimps with different ecological niches and their gill-associated microbial diveristy through comparative full-length transcriptome analysis and full-length 16S amplicon sequencing. The results have greatly enriched the genetic resources of deep-sea life, and provided basis for further studies of adaptive evolution mechanisms of crustaceans from deep-sea chemosynthetic ecosystems.