A deep-sea bacterium related to coastal marine pathogens

Abstract

Evolution of virulence traits from adaptation to environmental niches other than the host is probably a common feature of marine microbial pathogens, whose knowledge might be crucial to understand their emergence and pathogenetic potential. Here, we report genome sequence analysis of a novel marine bacterial species, Vibrio bathopelagicus sp. nov., isolated from warm bathypelagic waters (3309 m depth) of the Mediterranean Sea. Interestingly, V. bathopelagicus sp. nov. is closely related to coastal Vibrio strains pathogenic to marine bivalves. V. bathopelagicus sp. nov. genome encodes genes involved in environmental adaptation to the deep-sea but also in virulence, such as the R5.7 element, MARTX toxin cluster, Type VI secretion system and zinc-metalloprotease, previously associated with Vibrio infections in farmed oysters. The results of functional in vitro assays on immunocytes (haemocytes) of the Mediterranean mussel Mytilus galloprovincialis and the Pacific oyster Crassostrea gigas, and of the early larval development assay in Mytilus support strong toxicity of V. bathopelagicus sp. nov. towards bivalves. V. bathopelagicus sp. nov., isolated from a remote Mediterranean bathypelagic site, is an example of a planktonic marine bacterium with genotypic and phenotypic traits associated with animal pathogenicity, which might have played an evolutionary role in the origin of coastal marine pathogens.

Fig. 1

Maximum‐Likelihood (GTR+G+I parameters) phylogenetic tree based on concatenated sequences of atpA, pyrH, recA, rpoA and rpoD genes. Only bootstrap values (1000 replications) above 50% are shown. Vibrio cholerae CECT 514^T = ATCC 14035^T was used as an outgroup. Black circles: Vibrio species found in association with oysters; white circles: Vibrio species found in association with other marine bivalves (Lemire et al., ; Destoumieux‐Garzón et al., 2020).

Fig. 2

Full genome map of Sal10^T indicating the adaptation features (orange colour), putative virulence factors and prophage (blue colour) position on each chromosome.

Fig. 3

In vitro effects of Vibrio bathopelagicus sp. nov. on haemocyte lysosomal membrane stability (LMS) and bactericidal activity in the Mytilus galloprovincialis (upper panel) and C. gigas (lower panel). Upper panel: A. LMS: mussel haemocytes were treated with V. bathopelagicus (V.b.) 10⁷ CFU ml⁻¹. For comparison, data obtained with Vibrio tasmaniensis LGP32 (V.t) at 10⁷ and 10⁸ CFU ml⁻¹ are reported; Controls haemocytes (C) were treated with artificial sea water (ASW). B. Bactericidal activity: haemocytes were incubated for different periods of time (60–90 min) with V. bathopelagicus sp. nov. at the same concentration utilized in the LMS assay, and the number of viable, cultivable bacteria (CFU) per monolayer was evaluated. Percentages of killing were determined in comparison to values obtained at zero time. Lower panel: oyster haemocytes treated with V. bathopelagicus (V.b.) 10⁷ CFU ml⁻¹. C. LMS; D. bactericidal activity. Data obtained with V. tasmaniensis LGP32 (V.t) at 10⁷ CFU ml⁻¹ are also reported. * = P < 0.05, Mann–Whitney U test.

Fig. 4

Effects of Vibrio bathopelagicus sp. nov. (V.b.) and Vibrio tasmaniensis LGP32 (V.t; 10⁶ CFU ml⁻¹) on Mytilus galloprovincialis larval development in the 48 h embryotoxicity assay. A. Percentage of normal D‐shaped larvae at 48 hpf. Data represent the mean ± SD of four experiments carried out in 96‐multiwell plates (six replicate wells for each sample). * = P < 0.01, Mann–Whitney U test. B–D. Representative images of control (B) and vibrio‐exposed larvae (C,D). Scale bars = 100 μm. B. normal D‐veligers, characterized by regular shells with straight hinge; C. V. bathopelagicus induced larval death, as shown by open shells and release of soft tissues. D. Vibrio tasmaniensis resulted in arrested development, as shown by the presence of larvae withheld at the trocophora stage.