Synchronous effects of temperature, hydrostatic pressure, and salinity on growth, phospholipid profiles, and protein patterns of four Halomonas species isolated from deep-sea hydrothermal-vent and sea surface environments

Abstract

Four strains of euryhaline bacteria belonging to the genus Halomonas were tested for their response to a range of temperatures (2, 13, and 30 degrees C), hydrostatic pressures (0.1, 7.5, 15, 25, 35, 45, and 55 MPa), and salinities (4, 11, and 17% total salts). The isolates were psychrotolerant, halophilic to moderately halophilic, and piezotolerant, growing fastest at 30 degrees C, 0.1 MPa, and 4% total salts. Little or no growth occurred at the highest hydrostatic pressures tested, an effect that was more pronounced with decreasing temperatures. Growth curves suggested that the Halomonas strains tested would grow well in cool to warm hydrothermal-vent and associated subseafloor habitats, but poorly or not at all under cold deep-sea conditions. The intermediate salinity tested enhanced growth under certain high-hydrostatic-pressure and low-temperature conditions, highlighting a synergistic effect on growth for these combined stresses. Phospholipid profiles obtained at 30 degrees C indicated that hydrostatic pressure exerted the dominant control on the degree of lipid saturation, although elevated salinity slightly mitigated the increased degree of lipid unsaturation caused by increased hydrostatic pressure. Profiles of cytosolic and membrane proteins of Halomonas axialensis and H. hydrothermalis performed at 30 degrees C under various salinities and hydrostatic pressure conditions indicated several hydrostatic pressure and salinity effects, including proteins whose expression was induced by either an elevated salinity or hydrostatic pressure, but not by a combination of the two. The interplay between salinity and hydrostatic pressure on microbial growth and physiology suggests that adaptations to hydrostatic pressure and possibly other stresses may partially explain the euryhaline phenotype of members of the genus Halomonas living in deep-sea environments.

FIG. 1.

Growth of H. axialensis at 30°C (A), 13°C (B), and 2°C (C), with 4, 11, and 17% total salts, and under 0.1, 7.5, 15, 25, 35, 45, and 55 MPa of hydrostatic pressure. Warm colors (red) indicate faster growth. The 95% confidence intervals are approximated by the distance from the contoured plot surface to the black dots. Note the differences in the z-axis scale among Fig. 1 to 3.

FIG. 2.

Growth of H. meridiana strain Slthf1 at 30°C (A), 13°C (B), and 2°C (C). Other features are as described for Fig. 1.

FIG. 3.

Growth of H. hydrothermalis at 2°C (A) and growth of H. pacifica at 30°C (B) and 13°C (C). Other features are as described for Fig. 1.

FIG. 4.

Phospholipid fatty acid profiles of H. axialensis (columns 1 and 2) and H. hydrothermalis (columns 3 to 6) grown at 30°C under different conditions of hydrostatic pressure and salinity. C, growth under control conditions (4% total salts and 0.1 MPa); P, growth with elevated hydrostatic pressure (45 MPa and 4% total salts); S, growth with elevated salinity (11% total salts and 0.1 MPa); P+S, growth with both elevated hydrostatic pressure and salinity (45 MPa and 11% total salts). Only lipids that comprised ≥0.5% of the total are shown.

FIG. 5.

Cytosolic (A) and membrane (B) protein patterns of H. axialensis (H.a.; lanes 1 and 2) and H. hydrothermalis (H.h.; lanes 3 to 6) obtained at 30°C. Lanes M, molecular weight markers; other lanes are numbered identically to the columns in Fig. 4. Protein bands with changes in intensity are noted with letters.