Development and N2-Fixing Activity of the Benthic Microbial Community in Transplanted Spartina alterniflora Marshes in North Carolina

Growth and maturation of transplanted salt marshes is often limited by the availability of nitrogen (N). We examined the role of N₂-fixing benthic microbial assemblages (microalgae and associated bacteria) in two restored marshes (1-year-old and 6-year-old marsh) and a natural salt marsh in the Newport River Estuary, North Carolina. Benthic N₂ fixation (nitrogenase activity, NA), chlorophyll a (Chl a ) concentration, Spartina alterniflora (smooth cordgrass) stem counts, and sediment organic matter content were determined in the three marshes. Significant differences were observed between sites for both Chl a and NA. The 1-year-old marsh always exhibited the highest levels of NA and Chl a . Sediment organic matter content was lowest in the 1-year-old marsh (∼2%), intermediate in the 6-year-old marsh (∼5%), and highest in the natural marsh (∼10%). Carbon and nitrogen analyses were also performed on the 1-year-old marsh sediments, which were depleted in N. A positive correlation was observed between surface sediment N and Chl a . Remineralized, microbially derived N may provide growth-limiting inorganic N to Spartina transplants. N₂-fixing microbial assemblages in the 1-year-old marsh may also be an important food source for marsh infauna. Benthic N₂-fixing microbial assemblages play a key role in the N economy of restored salt marshes.     

Growth of tissue culture–regenerated salt marsh monocots in a simulated marsh field plot: Implication for wetland creation and restoration

Phenotypically and genetically variable salt marsh plants are needed for wetland creation and restoration efforts. Selected tissue culture regenerants of five salt marsh monocots, Spartina patens, Spartina alterniflora, Juncus gerardi, Juncus roemerianus, and Scirpus robustus, were planted in a simulated marsh field plot that was flood-irrigated with 10 ppt salt water to compare their phenotypic variation for potential use in wetland projects. Plant growth was evaluated after one growing season. Phenotypic variation among regenerants was found in S. alterniflora, S. patens and J. gerardi, indicating the occurrence of somaclonal variation. In S. alterniflora, significant differences occurred among regenerants in stem density. In J. gerardi, significant differences occurred in height and clone circumference. In S. patens, two of the nine regenerants exhibited higher biomass and stem density than some of the other regenerants. By using the random amplified polymorphic DNA (RAPD) technique, genome DNA variation in S. patens regenerants was detected. Genetic variation not only occurred among phenotypically different regenerants, but also among those phenotypically similar for the characteristics measured. Tissue culture–regenerated plants often have desirable genetic characteristics and adaptability as a result of somaclonal variation and may enable a species to perform its ecological functions in created or restored wetlands where ideal environments cannot be achieved. Thus, some previously unrestorable sites may be restorable or marginal marshes made more productive.     

Dimethylsulphoniopropionate (DMSP) and related compounds in higher plants

Dimethylsulphoniopropionate (DMSP) is produced in high concentrations in many marine algae, but in higher plants only in a few salt marsh grasses of the genus Spartina, in sugar canes (Saccharum spp.), and in the Pacific strand plant Wollastonia biflora (L.) DC. The high concentrations found in higher plants (up to 250 micromol g(-1) dry weight) suggest an important role, but though many functions have been suggested (including methylating agent, detoxification of excess sulphur, salt tolerance, and herbivore deterrent), its actual functions remain unclear. The fact that the ability to produce DMSP in high concentrations is found in species that have no taxonomic or ecological relationship suggests that the compound evolved independently and serves different functions in different plants. This is supported by observations that DMSP in W. biflora behaves differently from that in Spartina species. While DMSP concentrations in W. biflora have been found to increase with increasing salinity, suggesting a role in osmotic control, such a relationship has not been found for DMSP in Spartina species. Recent observations on tissue culture showed that, while undifferentiated tissue of W. biflora produced DMSP, such material of Spartina alterniflora Loisel. did not. Ongoing studies with tissue culture of both species have opened up new avenues of research on DMSP in higher plants, ultimately to elucidate the functions of this enigmatic compound.     

Salt tolerance and osmotic adjustment of Spartina alterniflora (Poaceae) and the invasive M haplotype of Phragmites australis (Poaceae) along a salinity gradient

An invasive variety of Phragmites australis (Poaceae, common reed), the M haplotype, has been implicated in the spread of this species into North American salt marshes that are normally dominated by the salt marsh grass Spartina alterniflora (Poaceae, smooth cordgrass). In some European marshes, on the other hand, Spartina spp. derived from S. alterniflora have spread into brackish P. australis marshes. In both cases, the non-native grass is thought to degrade the habitat value of the marsh for wildlife, and it is important to understand the physiological processes that lead to these species replacements. We compared the growth, salt tolerance, and osmotic adjustment of M haplotype P. australis and S. alterniflora along a salinity gradient in greenhouse experiments. Spartina alterniflora produced new biomass up to 0.6 M NaCl, whereas P. australis did not grow well above 0.2 M NaCl. The greater salt tolerance of S. alterniflora compared with P. australis was due to its ability to use Na(+) for osmotic adjustment in the shoots. On the other hand, at low salinities P. australis produced more shoots per gram of rhizome tissue than did S. alterniflora. This study illustrates how ecophysiological differences can shift the competitive advantage from one species to another along a stress gradient. Phragmites australis is spreading into North American coastal marshes that are experiencing reduced salinities, while Spartina spp. are spreading into northern European brackish marshes that are experiencing increased salinities as land use patterns change on the two continents.     

Ecological processes shaping Central Patagonian salt marsh landscapes

Plant zonation is one of the most conspicuous ecological features of salt marshes worldwide. In this work we used a combination of field transplant and greenhouse experiments to evaluate the importance of interspecific interactions and physical stress in the determination of the major plant zonation patterns in Central Patagonian salt marshes. There, Spartina alterniflora dominates the low marsh, and Sarcocornia perennis the high marsh. We addressed two questions: (i) What prevents Spartina alterniflora from colonizing the Sarcocornia perennis‐dominated high marsh zone? and (ii) What prevents Sarcocornia perennis from colonizing the Spartina alterniflora‐dominated low marsh zone? Our experimental transplants combined with neighbour exclusion treatments showed that the presence of Sarcocornia perennis negatively affects Spartina alterniflora, preventing it from surviving and/or spreading. Complementary field transplant and greenhouse experiments showed that Sarcocornia perennis did not survive the frequent tidal submersion by approximately 1.5 m of turbid seawater in the Spartina alterniflora zone, but its survival was independent of the presence of Spartina neighbours, and of the strong soil anoxia as well. Our results suggest that Spartina alterniflora is excluded by Sarcocornia perennis towards the low marsh, where frequent and prolonged submersion limit the survival of the latter. We provide and discuss key baseline information to facilitate the future design of ecophysiological experiments designed to accurately identify the exact mechanisms acting in every situation.     

Physiological Diversity of Rhizoplane Diazotrophs of the Saltmeadow Cordgrass,Spartina patens: Implications for Host Specific Ecotypes

Diazotrophic bacteria are important contributors to salt marsh productivity, but the biotic and abiotic factors that influence their distributions and function and the extent of their diversity cannot be understood in the absence of physiological information. Here we examine the physiological diversity and distribution patterns of diazotrophic bacteria associated with the rhizoplane of the saltmeadow cordgrass, Spartina patens, in comparison with diazotrophs from other intertidal grasses (tall and short form Spartina alterniflora and Juncus roemerianus) from the same salt marsh. S. patens plants were collected from two distinct habitats, and a total of 115 strains (111 Gram negative and 4 Gram positive strains) were isolated into pure culture by stab inoculating roots and rhizomes into combined nitrogen-free semisolid media. Most strains were microaerophilic and approximately one-half were motile. API test strips were used to eliminate redundancy within the culture collection, resulting in 21 physiologically different API groups (17 Gram negative and 4 Gram positive groups). A representative strain from each API group was selected for dot blot hybridization with a nifH specific probe and 16 strains (13 Gram negative and 3 Gram positive) were scored as positive. The nifH positive API group representative strains were characterized further using BIOLOG test plates. Substrate utilization potentials defined two S. patens strain clusters, and only one S. patens strain was physiologically similar to any other strain from a different host plant origin. No distinctions could be made based on the different S. patens habitats, suggesting that the host plant may have a greater impact than abiotic environmental conditions on the distributions of the rhizoplane diazotrophs recovered.     

THE GENETIC BASIS OF THE ECOLOGICAL AMPLITUDE OF SPARTINA PATENS. II. VARIANCE AND CORRELATION ANALYSIS

Reciprocal transplantations of Spartina patens genotypes from adjacent salt marsh, swale, and dune habitats provided evidence for genetic differentiation among subpopulations, due at least in part to contrasting selection regimes. Genet survival in the different habitats was related to the amount of genetic divergence. In the dune habitat, marsh ramets showed the lowest survival, swale ramets showed intermediate survival, and dune ramets showed the highest survival. This relationship was not reciprocal, however. The marsh habitat afforded an environment where survival was maximal for all genotypes. Thus, by comparison, the dune environment appeared to impose a more intense selection pressure, and the swale an intermediate selection pressure on Spartina patens. In each site resident genotypes tended to show greater relative fitness than aliens. This evidence for genetic divergence corroborates that previously reported on morphometric (Silander and Antonovics, 1979) and allozymic traits (Silander, 1984). High levels of phenotypic plasticity may permit greater adaptation to the spatially and temporally heterogeneous environment occupied by S. patens than would genetic variation alone. Dune and swale genets were more phenotypically plastic across traits examined than were marsh genotypes. The higher plasticity in these peripheral subpopulations may confer increased fitness among residents and compensate for observed declines in genetic variation. A slight decrease in genetic variability was evident from marsh to dune subpopulations. However, since the differences in genetic variation among subpopulations were small, and disparities did occur, it is unlikely that evolutionary divergence is retarded primarily by a lack of genetic variability in the characters considered. Evidence is presented to indicate that evolutionary divergence among subpopulations may be retarded by negative or unfavorable correlations among characters being selected simultaneously. These negative correlations may increase extinction probabilities in small peripheral populations, such as those represented by the dune or swale, and are likely to lower fitness. Based on these observations, I hypothesize that further microevolution may be retarded in peripheral dune and swale subpopulations, primarily by unfavorable genetic correlation structures among fitness components or characters under simultaneous selection. Contributing factors may include lowered genetic variance and higher levels of phenotypic plasticity.     

Molecular analysis of diazotroph diversity in the rhizosphere of the smooth cordgrass, Spartina alterniflora

N(2) fixation by diazotrophic bacteria associated with the roots of the smooth cordgrass, Spartina alterniflora, is an important source of new nitrogen in many salt marsh ecosystems. However, the diversity and phylogenetic affiliations of these rhizosphere diazotrophs are unknown. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified nifH sequence segments was used in previous studies to examine the stability and dynamics of the Spartina rhizosphere diazotroph assemblages in the North Inlet salt marsh, near Georgetown, S.C. In this study, plugs were taken from gel bands from representative DGGE gels, the nifH amplimers were recovered and cloned, and their sequences were determined. A total of 59 sequences were recovered, and the amino acid sequences predicted from them were aligned with sequences from known and unknown diazotrophs in order to determine the types of organisms present in the Spartina rhizosphere. We recovered numerous sequences from diazotrophs in the gamma subdivision of the division Proteobacteria (gamma-Proteobacteria) and from various anaerobic diazotrophs. Diazotrophs in the alpha-Proteobacteria were poorly represented. None of the Spartina rhizosphere DGGE band sequences were identical to any known or previously recovered environmental nifH sequences. The Spartina rhizosphere diazotroph assemblage is very diverse and apparently consists mainly of unknown organisms.     

A comparison of fungal communities from four salt marsh plants using automated ribosomal intergenic spacer analysis (ARISA)

Fungal decomposers are important contributors to the detritus-based food webs of salt marsh ecosystems. Knowing the composition of salt marsh fungal communities is essential in understanding how detritus processing is affected by changes in community dynamics. Automated ribosomal intergenic spacer analysis (ARISA) was used to examine the composition of fungal communities associated with four temperate salt marsh plants, Spartina alterniflora (short and tall forms), Juncus roemerianus, Distichlis spicata and Sarcocornia perennis. Plant tissues were homogenized and subjected to a particle-filtration protocol that yielded 106 microm particulate fractions, which were used as a source of fungal isolates and fungal DNA. Genera identified from sporulating cultures demonstrated that the 106 microm particles from each host plant were reliable sources of fungal DNA for ARISA. Analysis of ARISA data by principal component analysis (PCA), principal coordinate analysis (PCO) and species diversity comparisons indicated that the fungal communities from the two grasses, S. alterniflora and D. spicata were more similar to each other than they were to the distinct communities associated with J. roemerianus and S. perennis. Principal component analysis also showed no consistent, seasonal pattern in the composition of these fungal communities. Comparisons of ARISA fingerprints from the different fungal communities and those from pure cultures of selected Spartina ascomycetes supported the host/substrate specificity observed for the fungal communities.     

A Novel Method for Coral Explant Culture and Micropropagation

We describe here a method for the micropropagation of coral that creates progeny from tissue explants derived from a single polyp or colonial corals. Coral tissue explants of various sizes (0.5?C2.5?mm in diameter) were manually microdissected from the solitary coral Fungia granulosa. Explants could be maintained in an undeveloped state or induced to develop into polyps by manipulating environmental parameters such as light and temperature regimes, as well as substrate type. Fully developed polyps were able to be maintained for a long-term in a closed sea water system. Further, we demonstrate that mature explants are also amenable to this technique with the micropropagation of second-generation explants and their development into mature polyps. We thereby experimentally have established coral clonal lines that maintain their ability to differentiate without the need for chemical induction or genetic manipulation. The versatility of this method is also demonstrated through its application to two other coral species, the colonial corals Oculina patigonica and Favia favus.     

The Contribution of Mangrove Expansion to Salt Marsh Loss on the Texas Gulf Coast

Landscape-level shifts in plant species distribution and abundance can fundamentally change the ecology of an ecosystem. Such shifts are occurring within mangrove-marsh ecotones, where over the last few decades, relatively mild winters have led to mangrove expansion into areas previously occupied by salt marsh plants. On the Texas (USA) coast of the western Gulf of Mexico, most cases of mangrove expansion have been documented within specific bays or watersheds. Based on this body of relatively small-scale work and broader global patterns of mangrove expansion, we hypothesized that there has been a recent regional-level displacement of salt marshes by mangroves. We classified Landsat-5 Thematic Mapper images using artificial neural networks to quantify black mangrove (Avicennia germinans) expansion and salt marsh (Spartina alterniflora and other grass and forb species) loss over 20 years across the entire Texas coast. Between 1990 and 2010, mangrove area grew by 16.1 km², a 74% increase. Concurrently, salt marsh area decreased by 77.8 km², a 24% net loss. Only 6% of that loss was attributable to mangrove expansion; most salt marsh was lost due to conversion to tidal flats or water, likely a result of relative sea level rise. Our research confirmed that mangroves are expanding and, in some instances, displacing salt marshes at certain locations. However, this shift is not widespread when analyzed at a larger, regional level. Rather, local, relative sea level rise was indirectly implicated as another important driver causing regional-level salt marsh loss. Climate change is expected to accelerate both sea level rise and mangrove expansion; these mechanisms are likely to interact synergistically and contribute to salt marsh loss.     

Screening for bacterial-fungal associations in a south-eastern US salt marsh using pre-established fungal monocultures

Both bacteria and fungi play critical roles in decomposition processes in many natural environments, yet only rarely have they been studied as an integrated community. We examined whether physical associations exist between individual bacterial and fungal species that co-occur on decaying smooth cordgrass, Spartina alterniflora, in a south-eastern US salt marsh. Fungal-pervaded decaying Spartina was used as "bait" for potential bacterial associates. The bundles (infiltrated with one of three dominant fungal members of the decomposer assemblage, or an autoclaved control) were placed in a salt marsh and collected biweekly for 6 weeks during the first experiment (late summer 2002), and weekly for 3 weeks during the second experiment (early summer 2003). Terminal-restriction fragment length polymorphism (T-RFLP) analysis of 16S rRNA genes was used to track colonization by bacterial taxa in association with the established fungal species. T-RFLP analysis of 18S-to-28S internal transcribed spacer (ITS) regions was used to monitor changes in fungal communities once bundles had been placed in the field. Results from both years were nearly identical, and showed that invasion by fungi other than the bait species was slow, resulting in a virtual fungal monoculture for several weeks into the experiments. Surprisingly, bacterial communities were unaffected by the identity of the fungal bait. Regardless of the fungal species, and even in the absence of prior fungal colonization, bacterial 16S rRNA profiles were remarkably similar. These results suggest that few species-specific associations, either positive or negative, exist between bacterial and fungal members of the Spartina decomposer community during initial colonization.     

Analysis of internal transcribed spacer (ITS) regions of rRNA genes in fungal communities in a southeastern U.S. salt marsh

The ascomycete community colonizing decaying Spartina alterniflora blades in a southeastern U.S. salt marsh was characterized by analysis of internal transcribed spacer (ITS) regions of fungal rRNA genes. ITS sequences were amplified with ascomycete-specific primers from DNA extracted from S. alterniflora blades at two stages of decay (early and late) and were identified based on sequence analysis of a companion ascomycete culture collection. The S. alterniflora ITS libraries were dominated by clones from three species of ascomycetes: Mycosphaerella sp. 2, Phaeosphaeria spartinicola, and Phaeosphaeria halima. ITS sequences from five other less abundant ascomycete species were also found in the clone libraries, only two of which could be identified based on the culture collection, Hydropisphaera erubescens and a new species nicknamed '4clt'. Ascospore expulsion assays indicated dominance by the same three species as the ITS analysis, although this non-molecular approach differed from the molecular method in relative ranking of the dominant species and in characterization of minor species. Analysis of ITS amplicons from three replicate plots by terminal restriction fragment length polymorphism (T-RFLP) analysis showed significant spatial homogeneity in ascomycete community composition for both early- and late-stage decay. ITS sequence analysis identified morphologically cryptic subgroups for two of the three dominant salt marsh ascomycetes.     

Diversity of the ring-cleaving dioxygenase gene pcaH in a salt marsh bacterial community.

Degradation of lignin-related aromatic compounds is an important ecological process in the highly productive salt marshes of the southeastern United States, yet little is known about the mediating organisms or their catabolic pathways. Here we report the diversity of a gene encoding a key ring-cleaving enzyme of the beta-ketoadipate pathway, pcaH, amplified from bacterial communities associated with decaying Spartina alterniflora, the salt marsh grass that dominates these coastal systems, as well as from enrichment cultures with aromatic substrates (p-hydroxybenzoate, anthranilate, vanillate, and dehydroabietate). Sequence analysis of 149 pcaH clones revealed 85 unique sequences. Thirteen of the 53 amino acid residues compared were invariant in the PcaH proteins, suggesting that these residues have a required catalytic or structural function. Fifty-eight percent of the clones matched sequences amplified from a collection of 36 bacterial isolates obtained from seawater, marine sediments, or senescent Spartina. Fifty-two percent of the pcaH clones could be assigned to the roseobacter group, a marine lineage of the class alpha-Proteobacteria abundant in coastal ecosystems. Another 6% of the clones matched genes retrieved from isolates belonging to the genera Acinetobacter, Bacillus, and Stappia, and 42% of the clones could not be assigned to a cultured bacterium based on sequence identity. These results suggest that the diversity of the genes encoding a single step in aromatic compound degradation in the coastal marsh examined is high.     

不同发育时间的互花米草盐沼对大型底栖动物群落的影响

2004-2006年对长江口崇明东滩湿地芦苇(Phragmites australis)盐沼和不同发育时间的互花米草(Spartina alterniflora)盐沼的大型底栖动物群落特征进行分析研究.结果表明:互花米草盐沼发育初期,大型底栖动物群落以腹足类为主,物种丰富度(D=2.18)和多样性(H′=2.19)均低于芦苇盐沼(D=2.61, H′=2.29);随着时间的推移,互花米草与本地生物逐渐形成互动和稳定的格局,大型底栖动物群落组成中多毛类的种类逐渐上升(由3种变为6种),物种数和物种丰富度也上升,从而逐步形成新的大型底栖动物群落,物种丰富度(D=2.70)和多样性(H′=2.48)逐渐上升并高于芦苇盐沼(D=2.19, H′=2.09);从大型底栖动物群落的重新形成到稳定阶段,需要若干年的时间.     

Molecular characterization of sulfate-reducing bacteria in a New England salt marsh

Sulfate reduction, mediated by sulfate-reducing bacteria (SRB), is the dominant remineralization pathway in sediments of New England salt marshes. High sulfate reduction rates are associated with the rhizosphere of Spartina alterniflora when plants elongate aboveground. The growth process concurrently produces significant amounts of new rhizome material belowground and the plants leak dissolved organic compounds. This study investigated the diversity of SRB in a salt marsh over an annual growth cycle of S. alterniflora by exploring the diversity of a functional gene, dissimilatory sulfite reductase (dsrAB). Because the dsrAB gene is a key gene in the anaerobic sulfate-respiration pathway, it allows the identification of microorganisms responsible for sulfate reduction. Conserved dsrAB primers in polymerase chain reaction (PCR) generated full-length dsrAB amplicons for cloning and DNA sequence analysis. Nearly 80% of 380 clone sequences were similar to genes from Desulfosarcina and Desulfobacterium species within Desulfobacteraceae. This reinforces the hypothesis that complete oxidizers with high substrate versatility dominate the marsh. However, the phylotypes formed several clades that were distinct from cultured representatives, indicating a greater diversity of SRB than previously appreciated. Several dsrAB sequences were related to homologues from gram-positive, thermophilic and non-thermophilic Desulfotomaculum species. One dsrAB lineage formed a sister group to cultured members of the delta-proteobacterial group Syntrophobacteraceae. A deeply branching dsrAB lineage was not affiliated with genes from any cultured SRB. The sequence data from this study will allow for the design of probes or primers that can quantitatively assess the diverse range of sulfate reducers present in the environment.     

Restoring Salt Marshes Using Small Cordgrass, Spartina maritima

The use of exotic cordgrasses in salt marsh restoration projects has caused important negative environmental impacts and little is known about the possibilities of applying the endangered cordgrass Spartina maritima as a biotool at many European estuaries where it is the only native cordgrass. This paper discusses the planning and the development of an innovative restoration project based on S. maritima plantations in Odiel marshes (S.W. Iberian Peninsula). Our ecological restoration project had four specific goals: (1) to recover native vegetation, restoring the degraded landscape; (2) to phytostabilize oil-polluted sediments; (3) to prevent erosion and stabilize banks; and (4) to promote the conservation of S. maritima . Spartina maritima was planted at two physiographical locations: slightly sloping channel banks and flat interior marshes. Nonsuccessional stands of S. maritima develop at the channel banks where the marsh surface was stabilized. In contrast, successional stands of S. maritima grown in flat interior marshes are being replaced naturally by Sarcocornia perennis .     

The role of salt marshes in the Mira estuary (Portugal)

The Mira estuary is a narrow entrenched pristine estuary of the Ria type, about 30 km long. It comprises an area of 285 ha of salt marsh, of which250 ha have been proposed for reclamation for aquaculture. Dredging, village and recreation development menace the yet undisturbed estuarine ecosystem. To assess the biological importance of this wetland, a multidisciplinary study was conducted in apart of the salt marsh, considered as being representative of the whole area. Halophytic vegetation covering 75% of the total salt marsh site is dominated by Spartina maritima (28% of total vegetation area). Total primary production attains63,766 kg/yr (dw). A net export of 1541 kg/yr of COM to the relatively oligotrophic adjacent waters was also found. Insects and birds are described for the first time in the saltmarsh. Macrobenthic communities are dominated by Hediste diversicolor, Nepthys caeca and Scrobicularia plana. The fiddler crabUca tangeri attains here its north distribution limit. The mud flats and creeks associated with the salt marsh act as a nursery for 40.8% of the fish species present. The food web is dominated by detritivorous species like the grey mullets. The results obtained in this study support the need for an effective conservation of this area. This revised version was published online in July 2006 with corrections to the Cover Date.     

SEASONAL PATTERNS IN THE BELOWGROUND BIOMASS OF SPARTINA ALTERNIFLORA (GRAMINEAE) ACROSS A TIDAL GRADIENT

Belowground biomass of two ht forms of the salt marsh cordgrass, Spartina alterniflora, in a New England salt marsh exhibited a seasonal cycle. Biomass was highest in midsummer with no secondary peak in biomass corresponding with the autumn dieback of aboveground parts. Total annual biomass production and the maximum depth that living tissue penetrated into the substrate decreased with increasing tidal ht. Substrate characteristics (soil aeration, pH, nutrient levels) known to affect aboveground biomass of S. alterniflora also decreased with increasing tidal ht and may similarly affect belowground biomass across the same tidal gradient.