Abstract：
Most microorganisms resist pure cultivation under conventional laboratory conditions. One of the primary issues for this un-culturability is the absence of biologically produced growth-promoting factors in traditionally defined growth media. However, whether cultivating microbes by providing spent culture supernatant of pivotal microbes in the growth medium can be an effective approach to overcome this limitation is still an under-explored area of research. Here, we used the spent culture medium (SCM) method to isolate previously uncultivated marine bacteria and compared the efficiency of this method with the traditional cultivation (TC) method. In the SCM method, Ca. Bathyarchaeia-enriched supernatant (10%) was used along with recalcitrant organic substrates such as lignin, humic acid, and organic carbon mixture. Ca. Bathyarchaeia, a ubiquitous class of archaea, have the capacity to produce metabolites, making their spent culture supernatant a key source to recover new bacterial stains. Both cultivation methods resulted in the recovery of bacterial species from the phyla Pseudomonadota, Bacteroidota, Actinomycetota, and Bacillota. However, our SCM approach also led to the recovery of species from rarely cultivated groups, such as Planctomycetota, Deinococcota, and Balneolota. In terms of the isolation of new taxa, the SCM method resulted in the cultivation of 80 potential new strains, including one at the family, 16 at the genus, and 63 at the species level, with a novelty ratio of ~ 35% (80/219). In contrast, the TC method allowed the isolation of ~ 10% (19/171) novel strains at species level only. These findings suggest that the SCM approach improved the cultivation of novel and diverse bacteria.
摘要：
大多数微生物在常规实验室条件下抵抗纯培养。这种不可培养性的主要问题之一是在传统定义的生长培养基中不存在生物产生的生长促进因子。然而,通过在生长培养基中提供关键微生物的废培养上清液来培养微生物是否可以是克服这一限制的有效方法仍然是一个未被探索的研究领域。这里,我们使用废培养基（SCM）方法分离了以前未培养的海洋细菌，并将该方法的效率与传统培养（TC）方法进行了比较。在SCM方法中，Ca.富含Bathyarchaeia的上清液（10％）与顽固的有机底物（如木质素）一起使用，腐殖酸,和有机碳混合物。Ca.Bathyarchaeia,一类无处不在的古细菌，有能力产生代谢物，使用过的培养上清液成为回收新细菌污渍的关键来源。两种培养方法均可从假单胞菌门中回收细菌，拟杆菌,放线菌,还有Bacillota.然而,我们的SCM方法还导致了从很少种植的群体中恢复物种，比如Planctomycetota，Deinococcota,和Balneolota.在新分类群的分离方面，SCM方法培养了80个潜在的新菌株，包括家庭中的一个，属16，物种水平为63，新颖性比率为~35%(80/219)。相比之下,TC方法仅允许在物种水平上分离〜10％（19/171）的新菌株。这些发现表明SCM方法改善了新型和多样化细菌的培养。
