Phylogeny and phylogeography of the genus Patella based on mitochondrial DNA sequence data

Although some molecular phylogenies of Patella species have been published in recent years, unresolved questions concerning the phylogeny and taxonomy of this genus still remain. We sequenced the mitochondrial genes cytochrome c oxidase subunit I, 12S rRNA and 16S rRNA for all Patella species (Patella vulgata, Patella depressa, Patella candei, Patella caerulea, Patella lugubris, Patella ferruginea, Patella pellucida and the continental and Macaronesian forms of Patella ulyssiponensis and Patella rustica) and Cymbula safiana. Our results revealed the occurrence of five strongly supported clades, although relationships between these are not well supported. According to our data P. vulgata is a genetically distinct lineage and the close phylogenetic relationship between this species and P. depressa, found in previous mitochondrial DNA studies, is not supported. The Mediterranean Sea and the Macaronesian Islands seem to have played an important role in the speciation and diversification of this genus, although different clades show different phylogeographic patterns. Our dataset point to the necessity of a taxonomic revision, as P. candei is paraphyletic relative to P. lugubris.     

COUPLING MOTOGRAPHIC AND SONOGRAPHIC RECORDING TO ASSESS FORAGING BEHAVIOUR OF PATELLA VULGATA

Motographic recording of locomotor activity and sonographicrecording of rasping were simultaneously obtained in the fieldfrom a group of Patella vulgata L. inhabiting a sheltered shore(Menai Bridge, North Wales). Each limpet was equipped with apiezoelectric transducer, a small amplifier, and a micro-lighterencased in dental acrylic and glued to the shell. A remote filter-amplifier-recorderdevice continuously recorded rasping noise through the activityperiods (nocturnal low tide), when individual movements werealso recorded using an automatic camera. Movements of experimentallimpets did not differ from those of specimens equipped withthe micro-lighter only. Locomotor activity was definitely concentratedduring the first (outgoing) and last (homing) parts of eachactivity phase. Rasping started when limpets remained exposedto air at ebb tide, 1–2 hours before leaving home, andoccurred during the whole period spent away from home. Raspingwas fairly variable in rate, consisting in non-rhythmic boutsseparated by recovery phases. No significant correlation wasobserved between rasping rate and speed variation during differentparts of each excursion. However, the number of rasps performedin each spatial unit of the path was strongly correlated withthe time spent by the limpet in that unit. The data suggestthat in Patella vulgata the spatial organisation of foragingwithin each excursion is based on changes in travel speed relatedto local food density but not on modulation of rasping ratein time. (Received 4 July 1993; accepted 2 August 1993)     

Sewage pollution and extinction risk: an endangered limpet as a bioindicator?

The mollusc Patella ferruginea, endemic to the Mediterranean, is the most endangered marine species on the list of the European Council Directive 92/43/EEC and it is under serious risk of extinction. In spite of the low abundances and restricted distribution of this limpet, important populations have been found in the harbour of Ceuta, north Africa. The main objective of the present study was to characterise, for the first time, the effects of sewage pollution on P. ferruginea and related limpet species, and to evaluate the potential value of these limpet assemblages as bioindicators, using univariate and multivariate analyses. Physicochemical parameters and limpets were sampled in nine stations located at 0, 1, 2, 4, 8, 16, 32, 64 and 128 m away from the discharge point of a sewage effluent in Ceuta harbour. The stations closer to the outfall (0, 1, 2, 4 and 8) were characterised by higher values of turbidity, phosphate and ammonia in the water column, and organic matter, faecal coliforms and faecal Streptococci in sediments. A total of six limpet species were found and studied (Patella ferruginea, P. caerulea, P. nigra, P. rustica, P. ulyssiponensis and Siphonaria pectinata); the number of limpet species increased with increasing distance from the outfall, while diversity and evenness reached the highest values at intermediate sites. Siphonaria pectinata and P. caerulea were the most resistant and abundant species, while P. ferruginea was the most sensitive species to sewage pollution, only found at stations from 32 to 128 m. The distribution of this endangered limpet seems mainly affected by the pollution gradient, and not by the competition with the remaining limpets. The results of this study should be taken into account in future programmes of management and conservation of P. ferruginea.     

TEMPORAL VARIATION IN FORAGING BEHAVIOUR OF PATELLA GRANULARIS (PATELLOGASTROPODA) AND SIPHONARIA CONCINNA (BASOMMATOPHORA) ON A SOUTH AFRICAN SHORE

Foraging activity of two mid- to low- shore species of limpet,Patella granulans (Prosobranchia) and Siphonaria concinna (Pulmonata)from an exposed shore on the Eastern Cape coast of South Africawas monitored. In both species, activity was compared duringspring and neap tides and, in P. granularis between summer andwinter. Rhythms of activity of the two species were similar,with foraging excursions being mainly associated with nocturnallow tide times, although some P. granularis foraged during daytimelow tides. It is suggested that foraging excursions in P. granularisare triggered by wave action. Both species foraged further onspring tides than on neap tides and this is suggested to bea result of the limited time limpets have to forage. P. granulariswas also found to forage further during summer when comparedto winter and the possibility that seasonal micro-algal productivityinfluences foraging distances in limpets is discussed. The foraging activity of both species could be divided into3 distinct phases, a relatively rapid outward phase, a muchslower foraging phase and a rapid homeward phase. Whether ornot these limpets graze throughout an excursion is not known.S. concinna was found to home to a fixed scar, although duringthe experiment some scar-swapping occurred. P. granularis didnot home to a fixed scar but possessed a ‘home range’(approx. 5 cm²) to which it returned after each excursion. Patella granularis was found to move randomly during foraging,whilst S. concinna foraged in a non-random direction -whichtook individuals upshore. No tidal-influence is thought to bepresent in this case and the possibility of a learning componentin the foraging behaviour of certain limpet species in relationto the return to optimal feeding patches is discussed. (Received 18 May 1996; accepted 2 September 1996)     

AN ULTRASTRUCTURAL STUDY OF GAMETES AND FERTILIZATION IN PATELLA VULGATA L.

Gametes of Patella vulgata L. have been studied using electronmicroscopy and their ultrastructure described. Changes in ultrastructureoccurring during and after fertilization have been characterized,especially those occurring in the cortical region. These changesinclude appearance of Golgi bodies, mitochondria and agglomerationsof microtubular material. Cortical granules disappear and glycogengranules begin to be degraded. (Received 13 February 1984;     

Development of microsatellite loci for the black-footed limpet, <Emphasis Type="Italic">Patella depressa</Emphasis>, and cross-amplification in two other <Emphasis Type="Italic">Patella</Emphasis> species

Limpets (Patella spp.) are suitable organisms to investigate the effects of climate change in marine systems. They are widespread over NE Atlantic intertidal rocky shores and have been extensively studied in terms of population dynamics and ecology. Within genus Patella, microsatellites have only been developed for Patella caerulea and cross-species tests are unknown. In this work, we describe 11 primer pairs for Patella depressa and the results of cross-species testing on Patella candei and Patella rustica.     

THE ULTRASTRUCTURE AND FUNCTION OF THE OESOPHAGUS OF PATELLA VULGATA LINNAEUS

The oesophagus of Patella vulgata Linnaeus is divisible intotwo histologically and functionally distinct regions, a centralfood channel and a series of lateral pouches forming the oesophagealglands: both are lined by a glandular ciliated columnar epithelium.Subepithelial mucous cells are restricted to the epitheliumlining the food channel and posterior section of oesophagus,whilst amylase-secreting cells occur only in the oesophagealglands. The activity of the ciliated cells lining the food channelmixes ingested food with saliva and mucus to form a food stringthat is carried towards the stomach. Amylase-secreting cellsproduce an extracellular amylase in the form of proteinaceoussecretory granules, which are released by apocrine secretionwithin cytoplasmic blebs. Release is correlated with periodsof immersion when feeding is possible. The free blebs are carriedout of the pouches and into the food channel where the amylaseinitiates carbohydrate digestion. The oesophageal epitheliumabsorbs tritiated D-glucose by a Na⁺-dependent mechanism inhibitedby 2,4-DNP and phloridzin and the amylase-secreting cells pinocytoseferritin. It is suggested that the oesophagus absorbs smallsolute molecules derived from ruptured algal cells and theirpartially digested polysaccharide storage products. There alsoappears to be pinocytosis of dissolved organic matter. ^*Current address: The R L. Weston Institute for NeurologicalStudiet, University College & Middlesex Medcal School, MortimerSt., London Wl. (Received 8 March 1988; accepted 29 April 1988)     

MUCUS PRODUCTION AND PHYSIOLOGICAL ENERGETICS IN PATELLA VULGATA L.

Most energy budgets for marine gastropods lack a measured mucusterm. This is a major omission since mucus is widespread inmolluscan physiological processes. In this paper we outlinemethods for estimating mucus production in the intertidal gastropod,Patella vulgata L. This species uses mucus primarily for adhesionand locomotion. Pedal mucus production rates were measured inaerial environments of 70% relative humidity (RH) and 100% RHand in seawater. Animals in 70% RH did not move, while thosein 100% RH and seawater were mobile. Animals in 100% RH showedthe highest mucus production rate. Those in seawater secretedabout 75% of the mucus produced in 100% RH and those in 70%RH only about 40% of that produced in 100% RH. Significant logarithmicrelationships between mucus production rate and whole dry weightof animal, and mucus production rate and shell length were foundin all cases. The mucus term in the energy budget was calculated as 23% ofingestion both for individuals and for a specific population.This is much higher than the estimate of 4% of Wright &Hartnoll (1981). We have recalculated an energy budget for Patellavulgata to include the substantial mucus term. The recalculatedbudget is structurally different from the original budget. Thissuggests conclusions drawn from budgets not including a measuredmucus term should be regarded with caution. (Received 12 September 1989; accepted 13 December 1989)     

A FIELD TECHNIQUE FOR RECORDING THE ACTIVITY OF LIMPETS

A field technique for recording both temporal and spatial parametersof activity in limpets was tested on a Tyrrhenian populationof Patella rustica Linnaeus which shows definite homing behaviour.Timing of foraging excursions was recorded using reed switchesactivated by a magnet glued to the shell apex. The ‘lsquo;athome-away’ status of each animal was continuously screenedthroughout one month by interfacing the sensors with a computer.Moreover, individual excursions were minutely reconstructedby using a moto-graphic technique which combined LED-tracking(nocturnal activity) and instantaneous photography (daytimemovements). An original wave-meter was designed to monitor thesea-status during the study period. The quality of the temporaland spatial data obtained with this technique is evaluated,and further improvements of the technique are discussed. (Received 24 October 1989; accepted 25 January 1990)     

COMPARISON OF THE STRUCTURE OF THE PALLIAL TENTACLES OF SEVEN SPECIES OF SOUTH AFRICAN PATELLID LIMPET

The ultrastructure of the pallial tentacles of seven speciesof patellid limpet is described. The tip of most of the tentaclesexamined bears a crown of long cilia, whereas the shaft of thetentacles has small tufts (5–10 µm diameter) ofshorter cilia. Sections through the ciliated tufts show themto be composed of several cells, each bearing cilia. The ciliacontain 5–7 central microtubules and therefore do nothave the conventional 9 + 2 arrangement of microtubules. Nerveprocesses run from the base of each ciliated cell to a nervebundle in the centre of the tentacle, suggesting a sensory function.Estimates of densities of ciliated tufts suggest that the territoriallimpets (Patella cochlear and P. longicosta) have the greaternumber of tufts. Electron dense plate-like structures are foundin the centre of the pallial tentacles of Patella cochlear,P. longicosta, P. granularis, P. barbara and Helcion pruinosus.Each plate is about 0.2 µm wide and is surrounded by adouble membrane. It is suggested that these may play a rolein scattering or reflecting light and thus form part of thedermal light sensing ability of these animals. (Received 26 January 1987;     

Recent changes in the distribution of a marine gastropod,Patella rustica Linnaeus, 1758, and their relationship to unusual climatic events

Aim Recent colonization of northern Portuguese shores by Patella rustica Linnaeus, 1758, led to the bridging of a historical gap in the distribution known since the 1900s. Long‐term oceanographic data collected over the last half‐century were examined in order to detect possible mechanisms for the observed change in its distribution. Location This study was carried out along the entire Portuguese coastline, from 41°50′ to 37°06′ N. Time‐series of hydrographical variables (sea surface temperature and salinity) were derived for the Atlantic coast of the Iberian Peninsula. Methods Abundance and size‐frequency distributions of the newly observed limpet populations were compared with those from well‐established populations in southern Portugal. Anomalies were computed for sea surface temperature (1950–2000) and sea surface salinity (1958–2001) data, covering the whole Atlantic coast of the Iberian Peninsula. An upwelling index (1967–2005) was derived for a single location within the distributional gap of P. rustica. Split moving window analysis was performed to detect significant discontinuities in hydrographical data sets. Results Patella rustica has gradually been expanding in northern Iberia, and in the late 1990s the historical gap in distribution in northern Portugal was bridged. Size‐frequency distribution differed between historical and recent populations, the latter lacking small‐sized individuals. At the same time, several anomalous oceanographic events occurred off the Portuguese coast and were probably related to this expansion. Main conclusions Although sea surface temperature might be a major determinant of the reproductive success of P. rustica and hence its dispersal potential, it is more likely that a coincidence of several factors occurring in the late 1990s provided exceptional conditions that allowed the geographical expansion of this species.     

The limpet Patella vulgata L. at night in air: effective feeding on Ascophyllum nodosum monocultures and stranded seaweeds

Opportunistically, the limpet Patella vulgata will switch frommicrophagous grazing to feeding on attached macroalgae or, anovel finding, on accumulations of stranded seaweeds. Macroalgalfood is ingested primarily at night during emersion, when thethalli come to lie on the rock surface and are sufficientlydamp. Monocultures of Ascophyllum nodosum are not immune tothe destructive effects of limpets as previously thought. Patellavulgata can graze them down to complete disappearance, thuspushing Ascophyllum back to refuges where P. vulgata is absentor rare. Such refuges were found along a wide range from verysheltered to strongly wave-exposed rocky shores in Brittany,France. By a combination of its macrophagous and microphagousfeeding habits, P. vulgata can control mid-tidal macroalgalassemblages in a similar fashion to the action of sea urchinsupon kelp communities inhabiting the lower intertidal and shallowsublittoral zones of rocky shores. (Received 30 October 2006; accepted 25 May 2007)     

Usefulness of marine protected areas as tools for preserving the highly endangered limpet,Patella ferruginea,in the Mediterranean Sea

Patella ferruginea is the most endangered endemic marine invertebrate of western Mediterranean rocky shores. From 2012 to 2015, we performed a census and an ecological study for this gastropod mollusc in the Zembra Archipelago National Park in Tunisia, where a large population still remains. In 1986, a total of 20,000 individuals were estimated, with 0.7?ind/m², 4.4?cm adult mean size and low recruitment rate. In 2009, the estimated population reached 38,559 individuals with 2.65?ind/m², 5.42?cm adult mean size and high recruitment rate. During the present work, 43,790 individuals were estimated in 2012, 27,359 in 2013, 33,170 in 2014 and 40,404 in 2015, with an average density ranging from 0.1 to 8.5 individuals per square metre, but reaching as many as 25?ind/m² on some shores. This makes the archipelago one of the most important Mediterranean ‘hot spots’ for P. ferruginea, as it has a large and well-conserved population of this species. Variability of the population size is influenced by various biotic and abiotic factors, but is principally impacted by poaching of the species. The poachers target individuals larger than 6?cm in length, and the sex ratio study has shown that over 70% of individuals become females at that size. In addition, high recruitment rates were recorded in 2012 and 2013, but these decreased in 2014 and 2015, although population numbers show the opposite pattern. Moreover, growth rate is low and P. ferruginea may need many years to reach a large adult size. The loss of reproductive potential due to poaching inevitably causes a significant imbalance in the limpet population. Our results indicate the importance of the National Park for P. ferruginea and underline the need for this species to be protected and monitored over a long period to provide the most appropriate management measures.     

Isolation and characterisation of polymorphic microsatellite markers for the endangered ferreous limpet <Emphasis Type="Italic">Patella ferruginea</Emphasis> (Gastropoda,Patellidae)

The limpet Patella ferruginea is one of the most endangered marine invertebrates on western Mediterranean rocky shores. We have isolated and characterised 11 polymorphic microsatellites markers to provide new tools to investigate genetic structure and gather information necessary for the proper management of this severely threatened species. The number of alleles per locus ranged from 2 to 16 (mean; Na = 8.37) with an average observed heterozygosity of 0.64 (He = 0.66). The levels of polymorphism uncovered at these loci suggest that they should be useful for population genetic studies, parentage analysis and assessment of connectivity among protected areas. None of the pairwaise comparisons among loci showed significant linkage disequilibrium after sequential Bonferroni correction. All but one locus (Pf-31IF1) conformed to HW equilibrium. Further investigation revealed that departures at Pf-31IF1 were not caused by null alleles. Results from cross-species amplifications suggest that some of these loci may also be useful for Patella tenuis (two loci) P. ulyssiponensis (one locus) and P. piperata (three loci).     

两近缘种烟草上棉铃虫种群适合度的比较研究

分别在室内和自然条件下,比较研究了普通烟草和黄花烟草两种不同寄主植物上棉铃虫生长发育、食物转化和利用及种群增长情况。结果表明： (1) 室内26±1℃、75%±5% RH和16L∶8D光周期条件下,与普通烟草相比,黄花烟草上幼虫的发育历期显著延长,1龄和2龄幼虫的存活率及雌蛾产卵量显著下降,种群净增殖率（R₀=30.5374）和内禀增长率（r_m=0.0951）减小;(2) 6龄幼虫对两种烟草的近似消化率及其相对生长率无显著差异,但对黄花烟草的利用率和转化率显著下降,相对取食量显著提高;(3) 第2代棉铃虫在普通烟田的种群增长（I=1.9922>1）比黄花烟田（I=1.1581>1）快,“烟草和其他”对种群起主要控制作用,特别是对低龄幼虫;第3代棉铃虫在普通烟田的种群增长有所减慢但仍呈上升趋势（I=1.5994>1）,而在黄花烟田则呈下降趋势（I=0.6434<1）,低龄幼虫受烟草抗性和雨水等的影响较大,高龄幼虫被病菌感染的数量增多。这些结果表明,与普通烟草相比,黄花烟草不适合棉铃虫种群的增长和繁殖。     

Molecular and Morphological Evidence Reveals a New Species in the Phyllomedusa hypochondrialis Group (Hylidae,Phyllomedusinae) from the Atlantic Forest of the Highlands of Southern Brazil

The taxonomic status of a disjunctive population of Phyllomedusa from southern Brazil was diagnosed using molecular, chromosomal, and morphological approaches, which resulted in the recognition of a new species of the P. hypochondrialis group. Here, we describe P. rustica sp. n. from the Atlantic Forest biome, found in natural highland grassland formations on a plateau in the south of Brazil. Phylogenetic inferences placed P. rustica sp. n. in a subclade that includes P. rhodei + all the highland species of the clade. Chromosomal morphology is conservative, supporting the inference of homologies among the karyotypes of the species of this genus. Phyllomedusa rustica is apparently restricted to its type-locality, and we discuss the potential impact on the strategies applied to the conservation of the natural grassland formations found within the Brazilian Atlantic Forest biome in southern Brazil. We suggest that conservation strategies should be modified to guarantee the preservation of this species.     

Molecular cytogenetic study of three common Mediterranean limpets, Patella caerulea, P. rustica and P. ulyssiponensis (Archaeogastropoda, Mollusca)

The present paper shows the results of chromosome banding and rDNA-FISH study performed on several specimens of different populations of Patella caerulea, Patella rustica and Patella ulyssiponensis. The taxonomic attribution of specimens was ascertained by the molecular phylogenetic analysis of the mitochondrial 16S rRNA gene. P. caerulea and P. rustica had 2n = 18 chromosomes with first seven of biarmed pairs and the remaining two uniarmed pairs. P. ulyssiponensis had 2n = 16 with all biarmed chromosomes. Ag-NOR loci were on the short arms of the first metacentric pair in the three studied limpets, whereas they showed a different pattern of heterochromatin distribution and composition. A chromosome mosaicism was observed in several P. caerulea specimens, which exhibited an unpaired metacentric element and loss of a telocentric pair. The obtained results suggest that in the genus Patella specific diversification was accompanied by variations in heterochromatin distribution and composition and reduction of chromosome number by Robertsonian centric fusion.     

Measuring the precision of estimates of total egg production based on plankton surveys

Methods are given to measure the effects of spatial and temporaldifferences in egg production on the precision of estimatesof total seasonal egg production derived from ichythyoplanktonsurveys. The techniques are applied to the results of largescale plankton surveys conducted in 1977 and 1979 off the northeasternUSA. For the three species analyzed (Atlantic mackerel, Scomberscombrus; silver hake, Merluccius bilinearis and yellowtailflounder, Limanda ferruginea). the surveys produced estimatesof total egg production having an average coefficient of variationequal to 31%. Estimates of spawning stock size based on theegg production estimates compared favorably with other independentassessments of stock size. ^*MARMAP Contribution MED/NEFC 83–39.     

Embryogenesis and Plantlet Formation through Direct Culture of Isolated Pollen of Nicotiana tabacum cv. Samsum and Nicotiana rustica cv. Rustica

Pollen embryos and plantlets of Nicotiana tabacum cv. Samsunand Nicotiana rustica cv. Rustica were obtained through directpollen culture without prior treatment or prior culture of anthersor buds. Isolated pollen was cultured first in a medium withoutsucrose, then transferred into Nitsch's H medium containing2% sucrose and 5 mM glutamine. The optimum medium for the initialculture was water and the optimum period of culture was ca.6 days when binucleate pollen was used. ¹ Present address: Friedrich Miescher Inst., P.O.B. 273, CH-4002Basel, Switzerland. (Received January 18, 1982; Accepted March 19, 1982)