Physiological and proteomic analysis of salinity tolerance of the halotolerant cyanobacterium <Emphasis Type="Italic">Anabaena</Emphasis> sp

The halotolerant cyanobacterium Anabaena sp was grown under NaCl concentration of 0, 170 and 515 mM and physiological and proteomic analysis was performed. At 515 mM NaCl the cyanobacterium showed reduced photosynthetic activities and significant increase in soluble sugar content, proline and SOD activity. On the other hand Anabaena sp grown at 170 mM NaCl showed optimal growth, photosynthetic activities and comparatively low soluble sugar content, proline accumulation and SOD activity. The intracellular Na⁺ content of the cells increased both at 170 and 515 mM NaCl. In contrast, the K⁺ content of the cyanobacterium Anabaena sp remained stable in response to growth at identical concentration of NaCl. While cells grown at 170 mM NaCl showed highest intracellular K⁺/Na⁺ ratio, salinity level of 515 mM NaCl resulted in reduced ratio of K⁺/Na⁺. Proteomic analysis revealed 50 salt-responsive proteins in the cyanobacterium Anabaena sp under salt treatment compared with control. Ten protein spots were subjected to MALDI-TOF–MS/MS analysis and the identified proteins are involved in photosynthesis, protein folding, cell organization and energy metabolism. Differential expression of proteins related to photosynthesis, energy metabolism was observed in Anabaena sp grown at 170 mM NaCl. At 170 mM NaCl increased expression of photosynthesis related proteins and effective osmotic adjustment through increased antioxidant enzymes and modulation of intracellular ions contributed to better salinity tolerance and optimal growth. On the contrary, increased intracellular Na⁺ content coupled with down regulation of photosynthetic and energy related proteins resulted in reduced growth at 515 mM NaCl. Therefore reduced growth at 515 mM NaCl could be due to accumulation of Na⁺ ions and requirement to maintain higher organic osmolytes and antioxidants which is energy intensive. The results thus show that the basis of salt tolerance is different when the halotolerant cyanobacterium Anabaena sp is grown under low and high salinity levels.     

Differential Responses of Two Wheat Varieties Differing in Salt Tolerance to the Combined Stress of Mn and Salinity

The effect of Mn and NaCl on growth, mineral nutrients and antioxidative enzymes in two tetroploid wheat genotypes differing in salt tolerance was investigated in this study. 100 mM NaCl and Mn stress significantly inhibited plant growth, photosynthesis and Ca uptake, while stimulated ROS accumulation, MDA and proline content in wheat plants, Mn stress also increased SOD, APX, GR and DHAR activities. Durum wheat (AS780) was less affected by 100 mM NaCl and Mn stress than emmer wheat (AS847) due to more proline production, higher antioxidative enzymes activities and less-affected mineral nutrients. Application of 10 mM NaCl to Mn-stressed durum wheat alleviated Mn-induced damage by reducing Mn accumulation and translocation, while promoting proline accumulation and SOD, APX and GR activities. Irrespective of NaCl level, the combined stress of Mn and NaCl caused more severe oxidative stress, result in further reduction of photosynthetic rate and plant growth in emmer wheat as compared to Mn stress alone. The additively negative effects of NaCl and Mn stress on growth of emmer wheat results from reduced SOD and APX activities as well as Ca, Cu and Fe accumulation in both shoots and roots. These results suggest that salt-tolerant durum wheat is superior to emmer in adapting to Mn stress and the combined stress of salinity and Mn.

     

Effect of salt stress on physiological and antioxidative responses in two species of <Emphasis Type="Italic">Salicornia</Emphasis> (<Emphasis Type="Italic">S. persica</Emphasis> and <Emphasis Type="Italic">S. europaea</Emphasis>)

The effects of salt stress on growth parameters, free proline content, ion accumulation, lipid peroxidation, and several antioxidative enzymes activities were investigated in S. persica and S. europaea. The seedlings were grown for 2 months in half-strength Hoagland solution and treated with different concentrations of NaCl (0, 85, 170, 340, and 510 mM) for 21 days. The fresh and dry weights of both species increased significantly at 85 and 170 mM NaCl and decreased at higher concentrations. Salinity increased proline content in both the species as compared to that of control. Sodium (Na⁺) content in roots and shoots increased, whereas K⁺ and P_i content in both organs decreased. At all NaCl concentrations, the total amounts of Na⁺ and K⁺ were higher in shoots than in roots. Malondialdehyde (MDA) content declined at moderate NaCl concentrations (85 and 170 mM) and increased at higher levels. With increased salinity, superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (GPX) activities also increased gradually in both species. In addition, it seems that GPX, CAT, and SOD activities play an essential protective role in the scavenging reactive oxygen species (ROS) in both species. Native polyacrylamide gel electrophoresis (PAGE) indicated different isoform profiles between S. persica and S. europaea concerning antioxidant enzymes. These results showed that S. persica exhibits a better protection mechanism against oxidative damage and it is more salt-tolerant than S. europaea possibly by maintaining and/or increasing growth parameters, ion accumulation, and antioxidant enzyme activities.     

The effect of osmotic shock and subsequent adaptation on the thermotolerance and cell morphology of Listeria monocytogenes

The relationship between the time of exposure to different levels of NaCl and the corresponding changes in thermotolerance and cell morphology of Listeria monocytogenes was investigated. The kinetics of the increase in thermotolerance, after an osmotic upshift, showed a very rapid initial response (<2 min) followed by a more gradual increase whereby cells, after 4 h exposure at 30°C, became nearly as heat resistant as those grown for 48 h under the same conditions. Cells grown in media with 0.09 mol l⁻¹ NaCl subjected to a short osmotic up-shock in media containing 0.5, 1.0 or 1.5 mol l⁻¹ NaCl showed a 1.3, 2.5 and 8-fold increase in thermotolerance, respectively. Osmotic adaptation, signified by growth at the higher NaCl concentration, however, resulted in a 2- to 3-fold additional increase in thermotolerance. An osmotic down-shock caused a very rapid loss of thermotolerance (<5 min). Osmotic shock and adaptation experiments were also performed in minced beef where similar changes in thermotolerance were observed. Cell morphology was markedly affected by the osmolarity of the growth medium. Cells grown in media containing 1.5 mol l⁻¹ NaCl became up to 50 times longer than cells grown in media with 0.09 mol l⁻¹ NaCl, but no direct link to thermotolerance could be made.     

Carbohydrate and Proline Contents in Leaves, Roots, and Apices of Salt-Tolerant and Salt-Sensitive Wheat Cultivars1

Intra-specific variations in nonstructural carbohydrates and free proline were determined in leaves, apices, roots, and maturing seeds of two salt-tolerant cultivars (CR and Kharchia-65) and one salt-sensitive cv. Ghods of spring wheat (Triticum aestivum L.) grown in sand culture at various levels of salinity (0, 100, 200, and 300 mM NaCl and CaCl₂ at 5 : 1 molar ratio) under controlled environmental conditions. The levels of leaf, apex, and root ethanol-soluble carbohydrates, fructans, starch, and proline increased in line with elevating level of salinity in all three cultivars under investigation. The contents of proline, soluble and insoluble carbohydrates in the apex increased to levels exceeding those in the leaves and roots. Soluble carbohydrate content of salt-sensitive cv. Ghods was higher in the leaves, apices, and roots and lower in the maturing seeds than in the other cultivars at all levels of salinity except at 300 mM. The results show considerable variation in the amount of soluble, insoluble sugars, and proline among plant tissues and wheat genotypes in response to salinity. Higher soluble carbohydrates and fructan in leaves, roots and maturing seeds of stressed plants indicate that their accumulation may help plant to tolerate salinity. Salt-sensitive cv. Ghods accumulated less soluble sugars in the maturing seeds and higher soluble sugars in the apices, which might be used as an indicator in screening wheat genotypes for salinity tolerance.     

Salinity adaptation of plasma membrane H+-ATPase in the salt marsh plant Spartina patens: ATP hydrolysis and enzyme kinetics

Spartina patens, an intertidal C4 grass, grows in the upper salt marsh and tolerates coastal seawater salinity. The regulation of ion movement across the plasma membrane (PM) for plant salt tolerance is thought to be achieved by an electrochemical gradient generated by plasma membrane H⁺-ATPase. In this study, the change of PM H⁺-ATPase in response to NaCl was characterized for S. patens callus. Callus was cultured for 10 weeks under salinity levels of 0 mM, 170 mM, 340 mM, and 510 mM NaCl. Plasma membrane was isolated from a Dextran/PEG aqueous polymer two-phase system and the purity was demonstrated with membrane enzyme markers. There was a significant increase (up to 2-3 fold) of PM H⁺-ATPase activity when callus was grown on media containing NaCl. The incremental activation of PM H⁺-ATPase activity would enable the cell to tolerate higher cytoplasmic NaCl concentrations. PM H⁺-ATPase appeared to have a higher Vmax and a lower substrate concentration (Km to reach Vmax. When growth medium salinity increased from 0 mM to 170 and 340 mM, the Vmax of H⁺-ATPase increased from 0.64 to 1.00 and 1.73, respectively, while the Km decreased from 3.58 to 2.07 and 2.44 mM, respectively. In vitro NaCl inhibition kinetic data revealed a pattern of non-competitive inhibition by NaCl on PM H⁺-ATPase. The response of PM H⁺-ATPase in S. patens callus suggests that this species has evolved mechanisms that can regulate this important enzyme when cells are exposed to NaCl.     

Interactive effects of Ca2+ and NaCl salinity on the ionic relations and proline accumulation in the primary root tip of Sorghum bicolor

The effects of supplemental Ca²⁺ supply and NaCl salinity on the ionic relations and levels of proline and other amino acids in the primary root of Sorghum bicolor (cv. Hegari) seedlings were investigated. Two days of exposure to 150 m M NaCl resulted in a 50-fold increase in the proline level in the 0–10 mm root tips of seedlings supplied with 5.0 m M Ca²⁺, but only a 4-fold increase in seedlings with 0.5 m M Ca²⁺. In contrast to the high levels of proline in the root tip, proline accumulation was only modest in the expanded tissues of the root. The enhancement of proline accumulation in the root tip of salinized seedlings with the Ca²⁺ supplement may be related to their more favorable tissue K to Na ratio. Thus, elevated Ca²⁺ may mitigate the NaCl-induced inhibition of S. bicolor root growth via the maintenance of net K to Na selectivity and the enhancement of proline accumulation in the root tip.     

Glutamine synthetase and glutamate dehydrogenase contribute differentially to proline accumulation in leaves of wheat (Triticum aestivum) seedlings exposed to different salinity

To investigate the roles of ammonium-assimilating enzymes in proline synthesis under salinity stress, the activities of glutamine synthetase (GS; EC 6.3.1.2) and NADH-dependent glutamate dehydrogenase (NADH-GDH; EC 1.4.1.2) were determined in leaves of wheat (Triticum aestivum) seedlings exposed to salt stress at 150 and 300 mM NaCl for 5d. At the lower salinity, only GS activity increased markedly. At 300 mM NaCl, however, NADH-GDH activity increased while GS activity decreased. A significant accumulation of proline was found only at high-salinity exposure while glutamate, a proline precursor, increased dramatically under both low and high salinity. These data suggests that GS-catalysis might be the main glutamate synthesis pathway under low salinity. At 300 mM NaCl, glutamate seems to be preferentially produced through the process catalyzed by NADH-GDH. The increase of ammonium in salinity-stressed wheat seedlings might have resulted from increased photorespiration, which is responsible for the higher NADH-GDH activity. The activity of Delta(1)-pyrroline-5-carboxylate reductase (P5CR; EC 1.5.1.2) was significantly enhanced at 300 mM NaCl but remained unchanged at 150 mM. Delta(1)-Pyrroline-5-carboxylate synthetase (P5CS) activity did not show a specific response, indicating that P5CR might be the limiting step in proline synthesis from glutamate at high salinity.     

Selection of callus cultures of sugarcane (Saccharum sp.) tolerant to NaCl and their response to salt stress

Stable callus cultures tolerant to NaCl (68 mM) were developed from salt-sensitive sugarcane cultivar CP65-357 by in vitro selection process. The accumulation of both inorganic (Na⁺, Cl⁻ and K⁺) and organic (proline and soluble sugars) solutes was determined in selected and non-selected calli after a NaCl shock in order to evaluate their implication in in vitro salt tolerance of the selected lines. Both salt-tolerant and non-selected calli showed similar relative fresh weight growth in the absence of NaCl. No growth reduction was observed in salt-tolerant calli while a significant reduction about 32% was observed in nonselected ones when both were cultivated on 68 mM NaCl. Accumulation of Na⁺ was similar in both salt-tolerant and non-selected calli in the presence of NaCl. Accumulation of Cl⁻ was lower in NaCl-tolerant than in non-selected calli while proline and soluble sugars were more accumulated in salt-tolerant than in non-selected calli when both were exposed to salt. K⁺ level decreased more severely in non-selected calli than in NaCl-tolerant ones after NaCl shock. The results indicated that K⁺ and Cl⁻ may play a key role in in vitro salt-tolerance in sugarcance cell lines obtained by in vitro selection and that organic solutes could contribute mainly to counteract the negative water potential of the outside medium.     

Biochemical Alterations in Foliar Tissues of Citrus Genotypes Screened In vitro for Salinity Tolerance

Soil salinity is a prime impediment in the commercial production of citrus. In the present study two citrus rootstock genotypes viz. Citrus jambhiri and Citrus karna were cultured in vitro and exposed to NaCl salt stress. The previously standardized protocol was used for culture establishment and in vitro shoot and root regeneration. NaCl in different concentrations (25, 50, 75, 100 and 125 mM) was added in standardized regeneration and rooting media to note the biochemical changes due to salinity stress. Results revealed that salinity stress adversely affected the shoot and root differentiation and proved lethal above 100 mM NaCl. The hardening was also hampered due to salt stress. Among different biochemical parameters, proline, total soluble proteins and total sugars accumulation were enhanced however; total chlorophyll content was reduced under salinity stress. The revelation of some new protein polypeptides (21, 26 and 54 kDa) at different increasing salinity levels was attributed to their significance in stress alleviation.     

Changes on protein expression associated with salinity tolerance in Brassica cell cultures

The synthesis of proteins from salt-tolerant Brassica oleracea L. var. botrytis L. subvar. cauliflora (Gars.) DC. (cauliflower) cell cultures is modified in relation to controls in several features. There are nine newly induced polypeptides in tolerant cultures (absent in control conditions). Some of them are only present under low salt levels (85 mM NaCl). Another group seems to be representative of moderate and high salt levels (170 and 255 mM NaCl), and a third group is present in all the salt conditions tested. On the other hand, the synthesis of most of the polypeptides present in control conditions is modified in salt-tolerant cultures by increasing, decreasing or stopping their synthesis in any of the tested conditions. The relationship between these changes in Brassica and other plant systems is discussed.     

Heat shock and thermotolerance of Escherichia coli O157:H7 in a model beef gravy system and ground beef

Duplicate beef gravy or ground beef samples inoculated with a suspension of a four-strain cocktail of Escherichia coli O157:H7 were subjected to sublethal heating at 46 °C for 15–30 min, and then heated to a final internal temperature of 60 °C. Survivor curves were fitted using a linear model that incorporated a lag period (T_L), and D-values and 'time to a 4D inactivation' (T_4D) were calculated. Heat-shocking allowed the organism to survive longer than non-heat-shocked cells; the T_4D values at 60 °C increased 1·56- and 1·50-fold in beef gravy and ground beef, respectively. In ground beef stored at 4 °C, thermotolerance was lost after storage for 14 h. However, heat-shocked cells appeared to maintain their thermotolerance for at least 24 h in ground beef held at 15 or 28 °C. A 25 min heat shock at 46 °C in beef gravy resulted in an increase in the levels of two proteins with apparent molecular masses of 60 and 69 kDa. These two proteins were shown to be immunologically related to GroEL and DnaK, respectively. Increased heat resistance due to heat shock must be considered while designing thermal processes to assure the microbiological safety of thermally processed foods.     

Differential Solute Regulation in Leaf Blades of Various Ages in Salt-Sensitive Wheat and a Salt-Tolerant Wheat x Lophopyrum elongatum (Host) A. Love Amphiploid

Leaf blades of different ages from a salt-tolerant wheat x Lophopyrum elongatum (Host) A. Love (syn. Agropyron elongatum Host) amphiploid and its salt-sensitive wheat parent (Triticum aestivum L.cv Chinese Spring) were compared for their ionic relations, organic solute accumulation, and sap osmotic potential ([pi]sap). The plants were grown for 18 d in nonsaline (1.25 mM Na+) and salinized (200 mM NaCl) nutrient solutions. The response of leaf blades to NaCl salinity depended greatly on their age or position on the main stem. Na and proline levels were highest in the oldest leaf blade and progressively lower in younger ones. Glycine betaine and asparagine levels were highest in the youngest blade. The [pi]sap was similar for corresponding leaf blades of both genotypes, but contributions of various solutes to the difference in [pi]sap between blades from control and 200 mM NaCl treatments differed greatly. The NaCl-induced decline in [pi]sap of the youngest leaf blade of Chinese Spring was predominately due to the accumulation of Na and to a lesser extent asparagine; in the amphiploid, it was due to a combination of glycine betaine, K, Na, and asparagine. Proline contributed little in the youngest blade of either genotype. In the older blades Na was the major solute contributing to the decline in [pi]sap. Thus, the maintenance of low Na and high K levels and the accumulation of glycine betaine in the young leaf tissues contributed to the NaCl tolerance of the amphiploid. No such role was evident for proline.     

Effects of abscisic acid and proline on adaptation of tobacco callus cultures to salinity and osmotic shock

Tobacco callus ( Nicotina tabacum cv. Badischer Geudertheimer) took up sorbitol rapidly and without a lag period from media with up to 0.7 M of the polyol. Accumulation of proline was greatly enhanced under these conditions and was proportional to the absorbed sorbitol, while the viability of the callus cultures was quite low after a few hours of incubation. Under moderate conditions (0.1 M sorbitol) as well as under severe osmotic shock (0.7 M sorbitol), the cells adapted by adjusting the sorbitol/proline ratio to ca 3. NaCl (0.1 M ) had the same effect as sorbitol (0.7 M ) on the survival rate, but only slightly affected proline synthesis in the first hours of incubation. Addition of 10⁷ or 10 ⁵ M abscisic acid (ABA) did not increase the proline content, but 10 ⁷ M ABA delayed the deleterious effect of NaCl and improved the state of the cells. No influence of abscisic acid during the incubation with sorbitol could be detected. Two different strategies for the adjustment of tobacco callus to salinity and sorbitol are suggested: Non-ionic stress is controlled by the accumulation of proline, whereas ABA could be involved in the adaptation to ionic stress.     

Correlated changes in thermotolerance traits and body color phenotypes in montane populations of Drosophila melanogaster: analysis of within- and between-population variations

Thermotolerance traits vary across geographical gradients but there is a lack of clinal variation in some Drosophila species. Thus, it is not clear whether thermotolerance or other correlated traits are the target of natural selection. In order to test selection responses, we investigated body melanization and thermotolerance traits in six altitudinal populations of Drosophila melanogaster . Based on rearing different geographical populations under uniform growth conditions at 21 °C (common garden experiments), clinal variations for cold resistance are in the direction opposite to heat resistance along an altitudinal gradient, that is darker flies from highland populations evidenced higher levels of cold resistance while lowland populations showed higher heat resistance. Phenotypic plastic responses for body melanization at 17–28 °C showed significant correlations with thermotolerance traits. At 17 °C, regression coefficients as a function of altitude are highly significant and positive for cold resistance but negative for heat knockdown. However, for flies reared at 28 °C, there is no elevational change in melanization as well as thermotolerance traits. Thus, both genetic and plastic changes of body melanization and thermotolerance traits suggest a correlated selection response. Further, within-population analyses of body melanization (based on dark, intermediate and light color phenotypes) showed significant associations with thermotolerance traits. Correlated variations in body melanization and thermal tolerances are associated with climatic thermal variability ( T _cv) but not with T _min. or T _max. along an altitudinal gradient.     

Signal regulation of proline metabolism in callus of the halophyte Nitraria tangutorum Bobr. grown under salinity stress

Callus of the halophyte Nitraria tangutorum Bobr. was used to investigate proline metabolism and its signal regulation under salinity stress. Enhanced levels of proline and hydrogen peroxide (H₂O₂) were observed in calli exposed to salinity stress, and elevated levels of calcium (Ca) were detected in early responses to 75?mM NaCl treatment. Additionally, NaCl treatment induced significant elevation of ornithine-??-aminotransferase (OAT) activity, but notable decreases occurred in the activities of glutamyl kinase (GK) and proline dehydrogenase (PDH). H₂O₂ scavenger dimethylthiourea and pyruvate inhibited the accumulation of proline and the stimulation of OAT in salinity-stressed calli. Moreover, the utilization of Ca chelator EGTA and Ca channel blocker verapamil abolished the enhancement of proline level induced by 75?mM NaCl treatment for 3?days. These results suggest that the accumulation of proline is correlated to the increase of OAT activity and the decrease of PDH activity in response to salinity, and that elevated Ca signal during the early stage of NaCl treatment and the excitation of OAT activity resulting from the increase of H₂O₂ generation are essential for proline accumulation in salinity-stressed calli.     

Phospholipases C and D modulate proline accumulation in Thellungiella halophila/salsuginea differently according to the severity of salt or hyperosmotic stress

Proline accumulation is one of the most common responses of plants to environmental constraints. Thellungiella halophila/salsuginea, a model halophyte, accumulates high levels of proline in response to abiotic stress and in the absence of stress. Recently, lipid signaling pathways have been shown to be involved in the regulation of proline metabolism in Arabidopsis thaliana. Here we investigated the relationship between lipid signaling enzymes and the level of proline in T. salsuginea. Inhibition of phospholipase C (PLC) enzymes by the specific inhibitor U73122 demonstrated that proline accumulation is negatively controlled by PLCs in the absence of stress and under moderate salt stress (200 mM NaCl). The use of 1-butanol to divert some of the phospholipase D (PLD)-derived phosphatidic acid by transphosphatidylation revealed that PLDs exert a positive control on proline accumulation under severe stress (400 mM NaCl or 400 mM mannitol) but have no effect on its accumulation in non-stress conditions. This experimental evidence shows that positive and negative lipid regulatory components are involved in the fine regulation of proline metabolism. These signaling pathways in T. salsuginea are regulated in the opposite sense to those previously described in A. thaliana, revealing that common signaling components affect the physiology of closely related glycophyte and salt-tolerant plants differently.     

Effect of ABA on the contents of proline, polyamines, and cytokinins in the common ice plants under salt stress

The effects of ABA treatment on the contents of proline, polyamines (PA), and cytokinins (CK) in the facultative halophyte the common ice plant (Mesembryanthemum crystallinum L.) subjected to salt stress were studied. Plants grown in the phytotron chamber on Jonson nutrient medium for 6 weeks were subjected to 6-day-long salinity by a single NaCl adding to medium. During first three days of salinity, half plants of each treatment were placed for 30 min on nutrient medium containing 0, 100, or 300 mM NaCl plus ABA in the final concentration of 1 μM. Salinity reduced biomass accumulation and water and chlorophyll contents in plants. This was accompanied by the increase in the levels of MDA, proline, and sodium ions. ABA treatment of salt-stressed plants favored biomass accumulation and photosynthetic pigment protection, reduced the intensity of oxidative stress and the level of NaCl-induced proline accumulation. ABA treatment increased the contents of putrescine (Put) and spermidine (Spd) in the leaves and roots of control plants (not subjected to salt stress), reduced the losses of Put in the leaves and roots and Spd in the roots in the presence of 100 mM NaCl, and suppressed cadaverine (Cad) accumulation in the roots in the presence of 300 mM NaCl. In the presence of NaCl, ABA reduced the contents of zeatin and zeatin riboside and increased the level of zeatin-O-glucoside in the roots and isopentenyladenosine and isopentenyladenine in the leaves. Thus, ABA protective action under salinity can be realized through the weakening of oxidative stress (a decrease in MDA content) and the regulation of PA, proline, and CK metabolism, which has a great significance in plant adaptation to injurious factors.     

Effects of salinity on egg development and hatching in grey mullet Mugil cephalus L.

Grey mullet, Mugil cephalus , collected from ponds were induced to spawn with carp pituitary homogenate and human chorionic gonadotropin (HCG) at 30%_o salinity and 26° C. The spontaneously spawned and fertilized eggs were transferred to sea water of different salinities in the range of 5–70%_o, either at the 2-blastomere or the gastrula stage. Those eggs transferred at the gastrula stage were more tolerant to the salinity change than were those transferred at the 2-blastomere stage. The eggs did develop to the embryonic stage within the salinity range from 5 to 60%_o. Hatching occurred in all salinities between 10 and 55%_o. However, no larvae survived at 10 or at 55%_o. The optimal salinity range for eggs incubating at 22.0–25.5° C was from 30 to 40%_o salinity, with the peak at 35%_o.