Genetic, morphological, and chemical divergence in the sponge genus Latrunculia (Porifera: Demospongiae) from New Zealand

We undertook a comprehensive study of Latrunculia in New Zealand to determine the relationship between taxonomic, environmental, and chemical variation within the genus. Sponges were collected from five locations around New Zealand: Three Kings Islands, Tutukaka, Wellington, Kaikoura, and Doubtful Sound. Allozyme electrophoresis at nine polymorphic loci indicated that sponges from each geographic location were genetically distinct, and that they displayed genetic differences of the magnitude usually associated with reproductively isolated species (Nei's D between locations =0.375-2.476). Additionally, the comparisons revealed that the green and brown colour morphs of Latrunculia that are sympatric at Three Kings Islands and Kaikoura are distinct from each other, and that there are two genetic groups within the green sponges in Doubtful Sound. On the basis of genetic data we conclude that there are at least eight species of Latrunculia in New Zealand waters, not one to four as had been previously thought. Morphological comparisons of the eight genetic species based on skeletal characters (i.e. skeletal organisation of the choanosome, spicule composition, size, and geometry) indicated that the eight Latrunculia species fell into only two morphological groups that could be easily diagnosed on the basis of discorhabd type. Within these two primary morphological groups, skeletal characteristics among the eight species largely overlap and are not diagnostic. These findings emphasise the limitations of traditional taxonomic methods based solely on skeletal characters for distinguishing species of Latrunculia. However, multivariate analysis (MANOVA and CDA) based on six measured skeletal variables did reveal significant morphological variation among the species (Pillai's Trace=3.28, F=6.90, P<0.0001), supporting the division of the genus into eight species. Comparisons of chemical extracts from Latrunculia also showed that the amounts of five different bioactive compounds (discorhabdins A, B, C, D, and J) varied predictably among the eight species. This finding suggests that discorhabdin variation within Latrunculia, previously thought to be associated with intra-specific environmental variability, is more likely to reflect differences among species rather than phenotypic plasticity. Our results also highlight the importance of thorough taxonomic studies associated with marine natural products research to understand fully the variation in bioactive properties among individuals. The potential processes underlying the unusually high speciation rates in New Zealand Latrunculia that are indicated in our study are discussed.     

Allozyme variation in European populations of the oyster Ostrea edulis

Variations at 22 enzyme coding loci were surveyed in 11 populations of the oyster Ostrea edulis L., which were sampled between 1988 and 1990 along the Atlantic and Mediterranean coasts of Europe. Atlantic oyster beds suffered a steady decline during the last century, and restocking of beds with oysters of foreign origin has probably resulted in a high degree of interbreeding of natural oyster stocks from all Atlantic Europe. Our study confirms the low levels of genetic variability previously reported for the oyster populations from the Atlantic coasts, and extends it to the Mediterranean coasts. The locus arginine-kinase (ARK ^*) exhibited a high degree of interpopulation differentiation (F _ST=0.289), resulting from extensive variation in gene frequencies along a geographical cline. However, the overall genetic differentiation between populations was slight, and similar to that reported for other local populations of bivalves (mean genetic distance between populations is 0.010, mean F _ST=0.062). A general pattern of increasing differentiation along the coastline in an Atlantic-mediterranean direction emerged; but genetic differentiation among the Atlantic populations was not significantly lower than that observed among the Mediterranean populations. This and other results suggest that the effects of extensive transplantation of oysters among various areas in Europe are detectable only in some particular localities. The geographical distribution of low-frequency alleles suggests a restriction to gene flow outwards from the Mediterranean Sea, across the Straits of Gibraltar.     

Observations on shell form and its ecological significance in thaisid gastropods of the genus Lepsiella in New Zealand

This paper is concerned with the ecological significance of variation in shell form within the thaisid gastropod genus Lepsiella in New Zealand. Shell form has been investigated by measurement of shell height and breadth, aperture length and width, the diameters of consecutive whorls, apical angle, shell weight, and shell capacity, although in many cases shell height and shell breadth could not be measured because of erosion. L.albomarginata has been studied intensively at 4 stations in the South Island, and L. scobina less intensively at 6 stations in the North and South Islands. Comparisons of pairs of characters between stations have been tested by regression analysis and analysis of covariance where appropriate. Shells of L. albomarginata are relatively taller and narrower, and have a thicker wall, at a very sheltered station (Hakahaka Bay) that at more wave-exposed stations. L. scobina (sensu stricto), characterised by the presence of spiral ribs on the shell, exhibits less striking but comparable differences in shell shape. In laboratory tests in a tidal tank the thicker-shelled L. albomarginata from a sheltered station (Hakahaka Bay in Port Underwood) was much better able to resist attack by the shore crab Hemigrapsus edwardsi than was L. albomarginata from a nearby wave-exposed station (Whites Bay, near Cape Campbell, South Island). L. scobina from both stations was resistant to attack. H. edwardsi abounds at sheltered stations, but is missing from wave-exposed rock reefs such as those at Whites Bay, so that the ability to survive encounters with shore crabs is ecologically important to L. albomarginata inhabiting sheltered stations. L. scobina occupies a lower zone on the shore, where it is probably liable to encounter other more powerful predators. Its spiral ribs probably strengthen the shell. We do not know to what extent differences in shell form and thickness depend on environmental factors, and to what extent they originate genetically. Thin shells are associated with an abundance of mussels (Mytilus edulis ssp. aoteanus or Modiolus neozelanicus). There is an interesting possibility that a scarcity of mussels or other food caused by superior nonspecific predators might result in the production of better-protected Lepsiella.     

General and specific combining abilities of larval and juvenile growth and viability estimated from natural oyster populations

Diallel crosses of oysters from three geographically isolated natural populations were produced to evaluate the relative importance of genetic, maternal, and environmental effects on larval and juvenile growth and viability. Significant additive genetic effects were observed only in larval viability at Day 12 and larval shell length at Day 2. The presence of significant male and female mean square for larval viability (suggesting non-additive genetic variance) is consistent with fitness related characters. Important maternal effects were observed for the larval and juvenile shell length and viability characters. These female mean squares are probably affected by both real and spurious maternal effects and potential contributing influences are discussed. The performance of the crosses can be largely explained by two factors: parental performance and the heterotic gene effects. This is based on an apparent positive correlation of mean values between the parental populations and their crosses. The crosses' mean viability at the end of the larval phase was 14.0% lower than the pure matings and support a previous observation of lower heterozygote viability in the larval phase (Mallet and Haley, 1983b).     

Phaeomelanin- and carotenoid-based pigmentation reflect oxidative status in two populations of the yellow warbler (Setophaga petechia)

Carotenoid- and phaeomelanin-based sexual pigmentation may signal a capacity to maintain oxidative balance and viability. However, diverse empirical results leave the association between pigmentation and oxidative stress (OS) unclear. We assessed the hypothesis that population-specific levels of oxidative challenge, or strategies for managing OS, affect relationships between sexual pigmentation and OS. Specifically, intense oxidative challenge in migratory, temperate breeding birds might enhance correlations between pigmentation and OS relative to allied tropical breeders, since quality-based differences in OS may arise only under intense oxidative challenge. Alternatively, in temperate breeders with intense within-season reproductive effort, high-quality birds may invest in reproduction over oxidative balance, dampening negative correlations between pigmentation and OS. To assess these alternatives, we compared prenesting relationships between pigmentation and OS in a migratory, Californian population of yellow warblers (Setophaga petechia brewsteri) and in a resident, Mexican population (Setophaga petechia bryanti, “mangrove warblers”). Yellow warblers displayed higher OS than mangrove warblers. However, year of capture and sex had bigger influences on correlations between pigmentation and OS than population. Males with more intense melanin pigmentation had lower OS among mangrove warblers and yellow warblers captured in 2011, but not among yellow warblers captured in 2012. In females only, lower OS levels were associated with more colorful carotenoid pigmentation. Results suggest that both phaeomelanin- and carotenoid-based pigmentation have the potential to correlate with OS levels, but that the signaling potential of pigmentation may shift with inter-annual variation in environmental conditions and display sex-specific dynamics.     

Comparative effects of two antifouling paints on the oyster Crassostrea gigas

The effects of two antifouling paints, one containing organotin compounds, the other copper oxide, were studied from September 1983 to October 1984 on Crassostrea gigas (Thunberg) grown under natural field conditions in the Bay of Arcachon, France. The organotin paint reduced growth rate expressed as weight, length and width, but did not affect shell height; it drastically decreased the dry condition factor and shell density, but did not affect the viability of embryos and larvae from exposed oysters. However, some decrease in larval growth rate was observed. The copper paint had no effect on oyster growth, but lowered the condition factor compared to controls. Neither viability of embryos or larvae nor larval growth were affected by this paint.     

Contrast in the importance of arrow squid as prey of male New Zealand sea lions and New Zealand fur seals at The Snares,subantarctic New Zealand

New Zealand sea lions (Phocarctos hookeri) are threatened by incidental bycatch in the trawl fishery for southern arrow squid (Nototodarus sloanii). An overlap between the fishery and foraging sea lions has previously been interpreted as one piece of evidence supporting resource competition for squid. However, there is currently no consensus about the importance of squid in the diet of New Zealand sea lions. Therefore, we investigated this importance independently of spatial and temporal differences in squid availability through a simultaneous study with sympatric New Zealand fur seals (Arctocephalus forsteri), a species known to target arrow squid. Diet sampling at The Snares (48°01′S 166°32′E), subantarctic New Zealand, in February 2012 coincided with peak annual catch in the nearby squid fishery. Diets were deduced by analyses of diagnostic prey remains from scats (faeces) and casts (regurgitations). The contribution of each prey species to the diet was quantified using the per cent index of relative importance (% IRI) that combined frequency of occurrence, mass and number of prey items. Arrow squid was a minor component in sea lion scats (2 % IRI), and none was found in their casts. In contrast, arrow squid was the major component in fur seal scats and casts (93 and 99 % IRI, respectively). This study found that New Zealand sea lions ate minimal squid at a time when squid was clearly available as evidenced by the diet of New Zealand fur seals; hence, there was no indication of resource competition between sea lions and the squid fishery.     

Genetic evidence for two species of sprat (Sprattus) in New Zealand waters

The New Zealand sprat Sprattus antipodum (Hector) is phenotypically variable, with two body forms (deep and slender) being recognised in the Canterbury Bight of New Zealand. Electrophoretic analysis of tissue samples from the two body types showed them to be fixed for different alleles at two loci and to have very different allele frequencies at a further three loci. Heterozygosities of 0.088 and 0.104 were found for the deep type and 0.056 for the slender type. Genetic distances of 0.421 and 0.403 were found between samples of deep and slender types and 0.014 between samples of deep types. It is concluded that the two body types are separate species.     

Development and phenology of Ecklonia radiata at two depths in Goat Island Bay,New Zealand

Ecklonia radiata (C. Ag.) J. Ag. growing in Goat Island Bay, New Zealand produced up to 6 kg dry weight of tissue m^-2 yr^-1 in dense populations at 7-m depth. Half this tissue and an unknown amount of exudates were sloughed or torn off. Tissue production at 15 m was 0.3 to 0.5 kg m^-2 yr^-1. At this depth the plants grew only half as fast as in shallow water and the population density was decreased. Approximately 20% by weight of tissue produced at 7 m and 10% of that at 15 m was reproductive tissue. At both depths the elongation rate and percentage of water in stipes varied with stipe size. Initial sorus formation was related to plant size and not to age. Seasonal fluctuations in growth rate, haptera formation, frond size and sporulation were delayed in plants in deeper water.     

Temperature and salinity: two climate change stressors affecting early development of the New Zealand sea urchin Evechinus chloroticus

Temperature and salinity are important environmental factors affecting the normal functioning of marine animals, particularly animals such as sea urchins living in shallow waters and tide pools. Here, we evaluated the effect of different combinations of temperature and salinity on early embryos of the endemic New Zealand sea urchin Evechinus chloroticus. Animals were collected at Matheson’s Bay (36º18′17′′S; 174º47′51′′E) in north-eastern New Zealand in February 2013. Embryos were exposed to five salinities (29, 31, 34, 35 and 37 ppt) and two temperatures (18 and 21 °C) during the first 24 h of development. Low salinity (29 ppt) affected all parameters (fertilization, development rate, gastrulation and normal development), with ca. 50 % of embryos surviving at 29 ppt, whereas seawater temperature only affected development rate and gastrulation. An increase in temperature from 18 to 21 °C minimized the negative effect of low salinity (≤31 ppt) on development rate and gastrulation of E. chloroticus. Overall, the results of this study suggest that early embryos of E. chloroticus have developmental plasticity to withstand reductions in salinity up to 29 ppt; however, it is still unknown whether the surviving embryos will be able to complete larval development at low salinities, particularly whether the embryos and larvae are carried into extreme environments such as estuaries where salinity is even lower. Multistressor studies are very important for climate change research as multiple environmental factors will act together in the wild, having major consequences for development and recruitment of marine invertebrates.     

Abundance of the New Zealand subantarctic southern right whale population estimated from photo-identification and genotype mark-recapture

The abundance of New Zealand subantarctic southern right whales (Eubalaena australis) was estimated for the first time using mark-recapture methods based on photo-identification and microsatellite genotyping (13 loci). Individual identification photographs of 383 whales and microsatellite genotypes of 235 whales were collected during annual austral winter field surveys from 1995 to 1998. Given the 4-year survey period and lack of geographic and demographic closure, we estimated super-population abundance using the POPAN Jolly-Seber model implemented in the software programme MARK. Models with constant survivorship but time-varying capture probability and probability of entry into the population were the most suitable due to the survey design. This provided estimates of abundance in 1998 of 908 non-calf whales (95% C.L. = 755, 1,123) for the photo-identification and 910 non-calf whales (95% C.L. = 641, 1,354) for the microsatellite genotype data sets. The current estimate of 900 whales may represent less than 5% of the pre-whaling abundance in New Zealand waters.     

Genetic structure of wild European populations of the invasive Pacific oyster Crassostrea gigas due to aquaculture practices

As a result of aquaculture activities, Pacific oysters Crassostrea gigas (Thunberg, 1793) have invaded European coasts. Using seven microsatellites, we found virtually no genetic differentiation between natural populations throughout the European range (from the south of the Wadden Sea (the Netherlands) to the south of France) and French cultivated oysters. The genetic homogeneity of Pacific oyster samples appears to be the result of repeated transfers from same seed stocks made for aquaculture and, to a lesser extent, widespread dispersal due to specific biological traits of this species. The only genetic differentiation of Sylt population in the north of the Wadden Sea (Germany) suggests a stronger, persistent impact of ongoing supply of new genetic material from hatchery production, corresponding to seeds selection made by breeders. All of our genetic data highlighted the importance of aquaculture practices on the genetic structure of the keystone invader C. gigas in Europe.     

Reproductive cycle of two populations of Ophionereis schayeri (Ophiuroidea) in New South Wales

Reproduction of two populations of Ophionereis schayeri (Müller and Troschel) in New South Wales was investigated from November 1991 through January 1993. The reproductive cycle was documented by histological examination of the gonads and by image analysis to determine the oocyte size-frequency distributions. Both populations of O. schayeri were mature in December and January and had a major summer spawning period between January and February. Thereafter, the condition of the gonads exhibited interindividual variability, with asynchronous low-intensity gamete release through August. O. schayeri has the potential to spawn for up to 8 mo of the year. This breeding pattern with synchronous spawning in summer and asynchronous gamete release through winter was similar in both populations. During autumn and winter, the gonads contained developing gametes and advanced gametes ready for spawning. The eggs spawned by O. schayeri during these seasons started their growth during the previous spring vitellogenic period, and continued to grow through summer. Upon reaching full size, they were stored for spawning outside the main breeding period. Spring is marked by increased spermatogenic and vitellogenic activity followed by maturation and spawning. Throughout their development, the oocytes of O. schayeri are surrounded by a follicle consisting of periodic acid-Schiff-positive (PAS+) haemal fluid and PAS+ yolk by the oocytes. In the testes, the haemal sinus projects into the centre of the spermatocyte columns, thus appearing to play a nutritive role in support of both oogenesis and spermatogenesis. Late vitellogenic oocytes of O. schayeri are firmly anchored to the germinal epithelium by an attachment complex consisting of specialised attachment cells and basophilic strands that radiate between the oolemma and the nucleus. Spawning is associated with rupture of the follicles, which remain as prominent, empty U-shaped profiles in the ovaries. O. schayeri produces copious numbers of 200 m-diam oocytes, suggesting that this species is a broadcast spawner and that it has a modified ophiopluteus or vitellaria larva.     

Shell shape variation in populations of Mytilus chilensis (Hupe 1854) from southern Chile: a geometric morphometric approach

The pattern of shell shape variation in populations of the mussel, Mytilus chilensis (Hupe 1854) from Southern Chile was analyzed as a function of sample origin (cultivated vs. wild) and latitude, using standard tools of geometric morphometrics for landmark data. Additionally, posterior adductor muscle index (PAMI), Freeman condition index and shell thickness were measured in each sample. Highly significant differences in shell shape components were found among mussel populations. These differences are related to the origin of samples (expansion of the posterior adductor muscle scar, elongation of the lateral ligament and of the ventral umbo position in non-cultivated samples) and to latitude (more elongated shells and more extended posterior adductor muscle scar in most southern samples when compared with the northernmost ones). PAMI and shell thickness were statistically higher in wild population, and Freeman condition index was higher in cultivated shells. It is suggested that in wild populations of M. chilensis, the mussels may face higher predator pressures and other environmental stress factors. Consequently, individuals may be using higher energy fraction to reinforce shells and to promote adductor muscle growth at the expense of somatic growth. In contrast, individuals found in calm aquaculture environments are relatively protected from predators and use most of their assimilated energy in somatic growth. In turn, this growth depends on changes that covariate with shell morphology.     

Effects of habitat fragmentation on the genetic structure and connectivity of the black-lipped pearl oyster Pinctada margaritifera populations in French Polynesia

Habitat destruction leading to increased fragmentation is detrimental to species by reducing population size and genetic diversity and by restraining population connectivity. However, little is known about the effects of naturally fragmented habitats on wild populations, especially when it comes to marine benthic invertebrates with long pelagic larval duration. In this framework, we investigated the connectivity and genetic diversity variation among nine wild populations of the black-lipped pearl oyster, Pinctada margaritifera, throughout French Polynesia using ten microsatellite DNA markers. Despite the naturally fragmented habitat (South Pacific oceanic islands), we found high values of genetic diversity and population admixture, indicating connectivity at small and large spatial scales within sampled sites of the Tuamotu, and between the Society and Tuamotu Archipelagos. In the meantime, habitat geomorphology increased genetic drift in populations occurring in small, closed lagoons. Significant genetic structure not correlated to geographic distance was detected mainly between closed and open lagoons. The Marquesas Islands hosted the most divergent populations, likely a result of vicariance. Our results also highlight that migration patterns among lagoons are not symmetrical. Altogether, the general pattern of gene flow, nonsymmetrical migration rates among populations, absence of isolation by distance and absence of recent extinction events found in our study strongly suggest that P. margaritifera populations of French Polynesia follow an asymmetrical island model of dispersal.     

Elevated CO2 alters larval proteome and its phosphorylation status in the commercial oyster, Crassostrea hongkongensis

Ocean acidification (OA) is beginning to have noticeable negative impact on calcification rate, shell structure and physiological energy budgeting of several marine organisms; these alter the growth of many economically important shellfish including oysters. Early life stages of oysters may be particularly vulnerable to OA-driven low pH conditions because their shell is made up of the highly soluble form of calcium carbonate (CaCO₃) mineral, aragonite. Our long-term CO₂ perturbation experiment showed that larval shell growth rate of the oyster species Crassostrea hongkongensis was significantly reduced at pH < 7.9 compared to the control (8.2). To gain new insights into the underlying mechanisms of low-pH-induced delays in larval growth, we have examined the effect of pH on the protein expression pattern, including protein phosphorylation status at the pediveliger larval stage. Using two-dimensional electrophoresis and mass spectrometry, we demonstrated that the larval proteome was significantly altered by the two low pH treatments (7.9 and 7.6) compared to the control pH (8.2). Generally, the number of expressed proteins and their phosphorylation level decreased with low pH. Proteins involved in larval energy metabolism and calcification appeared to be down-regulated in response to low pH, whereas cell motility and production of cytoskeletal proteins were increased. This study on larval growth coupled with proteome change is the first step toward the search for novel Protein Expression Signatures indicative of low pH, which may help in understanding the mechanisms involved in low pH tolerance.     

Purification and immunolocalization of a vitellin-like protein from the Pacific oyster Crassostrea gigas

A female-specific protein from the hemolymph, and a related ovarian protein were identified in the Pacific oyster Crassostrea gigas using immunological procedures. The latter protein was isolated from ovarian extract by a combination of precipitation in distilled water, ammonium sulfate fractionation, hydroxylapatite chromatography and gel filtration on a Sepharose 6B column, and was identified as a macromolecular glycolipoprotein of 500 kD; sodium dodecylsulfate polyacrylamide gel electrophoresis, under reducing conditions, revealed seven heterogeneous polypeptides. Using a specific antiserum against purified protein, we also examined the immunohistochemical distribution of the protein at light- and electron-microscopic levels. Light microscopy revealed its presence in the oocytes, and electron microscopy identified it in yolk granules and the vitelline coat of the oocytes. These results indicate that the purified protein is a vitellin-like protein. Since no organ other than the ovary reacted with the antiserum, it is probable that the protein is produced inside the ovary, probably autosynthetically by oocytes.     

Field evidence of the impact of two acanthocephalan parasites on the mortality of three species of New Zealand shore crabs (Brachyura)

The effects of the acanthocephalan parasites Profilicollis antarcticus and P. novaezelandensis on the fecundity and mortality of three species of shore crab (Macrophthalmus hirtipes, Hemigrapsus edwardsi and H. crenulatus) are examined. The number of eggs produced by female crabs was strongly correlated (all P<0.05) with carapace width; parasite load was not a significant determinant of female fecundity. Mortality was inferred from reduction in the mean number of parasites per crab in the largest crab size-classes, indicating that heavily infected individuals are removed from the population. Mortality attributable to the parasites was observed for all three species of crabs, although the effect of parasites varied in both time and space; significant curvilinear regressions between parasite load and crab size-classes were not found in all samples. Crab mortality appears to be influenced by more than the pathological influences exerted by the parasites. Parasite-induced behavioural alterations may cause crab hosts to be more susceptible to predation by definitive hosts. We support this suggestion with three lines of evidence: the lack of parasite effects on fecundity, the weakening of the parasite effect on mortality during the time of year when birds are absent, and previous indications of parasite-mediated alterations in crab burrowing behaviour.     

Are there two species of the polychaete genus Marenzelleria in Europe?

North Sea and Baltic Sea populations of the introduced polychaete Marenzelleria viridis (Verrill, 1873) reproduce at different times (spring and autumn, respectively). Enzyme separation by starch gel electrophoresis revealed major differences between specimens from the Baltic Sea and those from the North Sea (collected in 1992 and 1993) but a high degree of homogeneity among populations from the same sea. Three enzyme loci, glucose-6-phosphate isomerase (GPI-A, GPI-B) and malate dehydrogenase (MDH), were fixed to 100% by different alleles in the North and Baltic Sea populations, respectively. Different alleles are dominant for mitochondrial aspartate aminotransferase (mAAT) with allele frequencies of ca. 0.97 in all sampled populations from the North Sea and Baltic Sea, respectively, but heterozygotes were found in all populations. These genetic differences could be due to environmentally induced selection or genetically different origins of the populations, suggesting that populations of the genus Marenzelleria in the North and Baltic Seas may be two different species.