Keller-Costa Tina, Jousset Alexandre, van Overbeek Leo, van Elsas Jan Dirk, Costa Rodrigo
Centre of Marine Sciences (CCMAR), University of Algarve, Faro, Algarve, Portugal ; Department of Microbial Ecology, Centre for Ecological and Evolutionary Studies, University of Groningen, Groningen, The Netherlands.
Department of Ecology and Biodiversity, Utrecht University, Utrecht, The Netherlands.
PLoS One. 2014 Feb 12;9(2):e88429. doi: 10.1371/journal.pone.0088429. eCollection 2014.
Bacteria are believed to play an important role in the fitness and biochemistry of sponges (Porifera). Pseudomonas species (Gammaproteobacteria, Pseudomonadales) are capable of colonizing a broad range of eukaryotic hosts, but knowledge of their diversity and function in freshwater invertebrates is rudimentary. We assessed the diversity, structure and antimicrobial activities of Pseudomonas spp. in the freshwater sponge Ephydatia fluviatilis. Polymerase Chain Reaction--Denaturing Gradient Gel Electrophoresis (PCR-DGGE) fingerprints of the global regulator gene gacA revealed distinct structures between sponge-associated and free-living Pseudomonas communities, unveiling previously unsuspected diversity of these assemblages in freshwater. Community structures varied across E. fluviatilis specimens, yet specific gacA phylotypes could be detected by PCR-DGGE in almost all sponge individuals sampled over two consecutive years. By means of whole-genome fingerprinting, 39 distinct genotypes were found within 90 fluorescent Pseudomonas isolates retrieved from E. fluviatilis. High frequency of in vitro antibacterial (49%), antiprotozoan (35%) and anti-oomycetal (32%) activities was found among these isolates, contrasting less-pronounced basidiomycetal (17%) and ascomycetal (8%) antagonism. Culture extracts of highly predation-resistant isolates rapidly caused complete immobility or lysis of cells of the protozoan Colpoda steinii. Isolates tentatively identified as P. jessenii, P. protegens and P. oryzihabitans showed conspicuous inhibitory traits and correspondence with dominant sponge-associated phylotypes registered by cultivation-independent analysis. Our findings suggest that E. fluviatilis hosts both transient and persistent Pseudomonas symbionts displaying antimicrobial activities of potential ecological and biotechnological value.
细菌被认为在海绵动物（多孔动物门）的健康状况和生物化学过程中发挥着重要作用。假单胞菌属（γ-变形菌纲，假单胞菌目）能够定殖于多种真核宿主，但我们对其在淡水无脊椎动物中的多样性和功能的了解还很有限。我们评估了淡水海绵河流埃弗海绵中假单胞菌属的多样性、结构和抗菌活性。全局调控基因gacA的聚合酶链反应-变性梯度凝胶电泳（PCR-DGGE）指纹图谱揭示了与海绵相关的假单胞菌群落和自由生活的假单胞菌群落之间的不同结构，揭示了这些群落此前未被怀疑的在淡水中的多样性。不同的河流埃弗海绵标本中的群落结构各不相同，但通过PCR-DGGE可以在连续两年采集的几乎所有海绵个体中检测到特定的gacA系统型。通过全基因组指纹图谱分析，在从河流埃弗海绵中分离出的90株荧光假单胞菌中发现了39种不同的基因型。在这些分离株中发现了较高频率的体外抗菌活性（49%）、抗原生动物活性（35%）和抗卵菌活性（32%），相比之下，对担子菌（17%）和子囊菌（8%）的拮抗作用则不那么明显。高度抗捕食的分离株的培养提取物能迅速导致原生动物斯坦尼尔肾形虫的细胞完全不动或裂解。初步鉴定为耶氏假单胞菌、类产碱假单胞菌和栖稻假单胞菌的分离株表现出显著的抑制特性，并且与通过非培养分析确定的优势海绵相关系统型相对应。我们的研究结果表明，河流埃弗海绵宿主中既有具有潜在生态和生物技术价值的抗菌活性的短暂性假单胞菌共生体，也有持久性假单胞菌共生体。
