Population genetics of the endangered limpet Patella ferruginea (Gastropoda: Patellidae): taxonomic, conservation and evolutionary considerations

The limpet Patella ferruginea is the most endangered marine invertebrate in the western Mediterranean rocky shores, whereas Patella caerulea is the most common Mediterranean limpet. From Pleistocene to historic age, P. ferruginea was distributed around the whole Mediterranean basin, since the shells of this species are a characteristic feature of Palaeolithic and Neolithic deposits in this area. However, its Mediterranean range has progressively contracted to a few restricted areas. The ancient origin of the species (18 Ma) and the present geographical isolation among relic populations could have led to a great genetic difference among populations, taking into account the supposedly low dispersal ability of the species. However, we have observed a few genetic differences among populations and a 'star phylogeny' of COI haplotypes from the 41 individuals of P. ferruginea analysed; a similar pattern has also been observed for the seven individuals of P. caerulea studied. Genetic evidences show a possible bottleneck event on late Pleistocene that affected the two species. The results have an important implication on the future management of this endangered species. Additionally, two different morphological types of P. ferruginea have been described by Payraudeau in 1826 : lamarcki and rouxi forms. Clear morphological differences occur between these two types and some authors pointed out the hypothesis about the existence of two different species. The results of the present study conclude that the two different forms of P. ferruginea are ecotypes, rather than different species or subspecies, and intermediate steps are an ecological range instead of hybridization events among different species.     

濒危帽贝铁锈笠螺的活动节律和采食行为的初步观察

铁锈笠螺是地中海最濒危的海洋无脊椎动物,对其生物学知之甚少,缺少对其活动节律和采食行为的了解。使用环氧树脂Eporai1127(原位标记了20个不同外壳长度的个体,并在每个外壳上标有不同的数字,作者在白昼或黑夜的高潮和低潮期收集了有关数据。可能由于云斑厚纹蟹(Pachygrapsus marmoratus)的捕食影响,铁锈笠螺白天的活动和运动多于夜间,但铁锈笠螺采食行为似乎仅限于高潮期。此外,汹涌的海面条件诱导了铁锈笠螺的活动和运动。     

Effects of competition on an endangered limpet Patella ferruginea (Gastropoda: Patellidae): Implications for conservation

Spatial and trophic competitive interactions were analysed for the seldom-studied limpet Patella ferruginea (which is considered as an “extinction risk”). Spatial patterns of distribution of P. ferruginea were studied using quadrats of 1 × 1 m² at several transects along the coastline, whereas trophic interactions were studied through manipulations of densities of P. ferruginea and Patella caerulea in experimental plots located at the mid-shore. Physical parameters were measured inside each plot to determine the microenvironmental differences between fenced and unfenced plots. Fenced plots of 50 × 50 cm² were used with six different treatments. Mortality was recorded every 15 days and length was measured monthly during the 3.5 months of the experiment. At the beginning and end of the experiment, lengths and weights of limpets were recorded and chlorophyll concentration was measured by spectrophotometric analysis of rock surface. Both species showed increased mortality and reduced size and weight in treatments with greater densities. High intraspecific competition was detected in P. caerulea compared with P. ferruginea whereas asymmetrical interspecific competition was observed. The results suggest that P. caerulea is the superior competitor, possibly due to differences in radular morphology and mobility between the two species. Microalgal food assessment showed that it is probably a limiting resource, especially for P. caerulea, although this species has a lower biomass than P. ferruginea. The high intraspecific competition observed in P. caerulea could be responsible for the coexistence of both species. The present study may be useful in helping to conserve P. ferruginea, which is presently under serious threat of extinction, because the results show that experimental fences are a viable means of concentrating breeding individuals.     

Patterns of spatial genetic structuring in the endangered limpet Patella ferruginea: implications for the conservation of a Mediterranean endemic

Patella ferruginea Gmelin, 1791 is an endangered marine gastropod endemic to the Western Mediterranean. Its range is restricted to the Sardinian-Corsican region (SCR), North Africa, a few scattered sites in Southern Spain, and Sicily. Inter-simple sequence repeat (ISSR) markers and three different mitochondrial DNA (mtDNA) regions, Cytochrome c Oxidase subunit I, 12S (small-subunit ribosomal RNA gene) and 16S (large-subunit ribosomal RNA gene), were used to investigate the presence of genetic population structuring. The mtDNA sequences showed very low levels of genetic differentiation. Conversely, ISSRs showed the presence of two main genetic groups, corresponding to Spain, North Africa and Sicily and the SCR. The SCR was further split into two subgroups. The ISSR results suggest that, on a regional scale, the genetic structure of P. ferruginea is mainly determined by the restriction of gene flow by dispersal barriers. On a more local scale human harvesting may play a crucial role in population structuring by increasing the effect of genetic drift.     

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.     

New polymorphic microsatellite markers for the limpet Patella rustica and cross-priming testing in four Patella species

Eighteen microsatellite markers have been characterized from the Patella rustica genome. An average of 10 alleles per locus and an expected heterozygosity ranging from 0.054 to 0.937 were observed in a sample of 32 wild individuals from Viana do Castelo, Portugal. Distinct cross-priming amplification rates were recovered on four additional Patella species. Three monomorphic P. rustica loci were polymorphic in other Patella species. The microsatellites developed herein could be a useful intraspecific genetic tool to undertake fine population studies in the genus Patella.     

居群遗传学原理及其在珍稀濒危植物保护中的应用

居群遗传学在珍稀濒危植物保护研究中有着重要的应用价值。本文首先介绍了居群遗传学中的几个重要概念——有效居群大小、近交繁殖、遗传漂变和基因流,然后详细叙述了居群遗传学原理在珍稀濒危植物保护中的应用途径和前景。     

广西九万山珍稀濒危植物及其保育对策

通过对广西九万山植物资源的考查和研究 ,发现该地区分布有珍稀濒危植物 1 9个科 ,2 8个属 ,共 2 9种。其中国家一级保护植物有 5种 ,国家二级保护植物有 2 4种。对它们的种类组成、区系成分、地理分布进行了分析 ,结果显示出广西九万山珍稀濒危植物不仅种类繁多 ,而且具有古老、原始、孑遗特性 ,热带性质明显 ,中国特有现象极为突出等特点。据此提出对广西九万山珍稀濒危植物的保育对策 ,以便更好地保护其生物多样性和这些珍稀濒危植物资源的可持续利用。     

濒危药用植物桃儿七的生物学特性及其保育措施

桃儿七为重要抗癌药物前体物来源物种,现正处在濒危状态,已被列为国家三级保护植物。本文全 面地综述了桃儿七的生物学特性,包括外部形态、地理分布、系统位置、生态学和群落学特性、发育生物学、繁 殖生物学、植物化学、细胞组织培养和内生真菌研究以及遗传学等研究。最后分析了引起桃儿七濒危的原因, 并提出了具体保育措施和研究展望。     

南京市珍稀濒危植物的分布与保护

对南京市珍稀濒危植物的种类、分布、生境及濒危原因进行了调查和分析,并对主要种类进行了就地保存、迁地保存或就地-迁地保存实验。调查发现,南京市共有珍稀濒危植物17种1变种,隶属9科17属,主要分布在丘陵山地;人为因素是其濒危的主要原因。对明党参(Changium smyrnioides H.Wolff.)、短穗竹〔Brachystachyumdensiflorum(Rendle)Keng〕及秤锤树(Sinojackia xylocarpa Hu)等种类的保护研究结果表明,采取一定的人为干预措施,可使南京市珍稀濒危植物的种群数量得到有效的保存和恢复。     

MtDNA diversity of the critically endangered Mekong giant catfish (Pangasianodon gigas Chevey, 1913) and closely related species: implications for conservation

Catfishes of the family Pangasiidae are an important group that contributes significantly to the fisheries of the Mekong River basin. In recent times the populations of several catfish species have declined, thought to be due to overfishing and habitat changes brought about by anthropogenic influences. The Mekong giant catfish Pangasianodon gigas Chevey, 1913 is listed as Critically Endangered on the IUCN Red List. In the present study, we assessed the level of genetic diversity of nine catfish species using sequences of the large subunit of mitochondrial DNA (16S rRNA). Approximately 570 base pairs (bp) were sequenced from 672 individuals of nine species. In all species studied, haplotype diversity and nucleotide diversity ranged from 0.118±0.101 to 0.667±0.141 and from 0.0002±0.0003 to 0.0016±0.0013, respectively. Four haplotypes were detected among 16 samples from natural populations of the critically endangered Mekong giant catfish. The results, in spite of the limited sample size for some species investigated, indicated that the level of genetic variation observed in wild populations of the Mekong giant catfish (haplotype diversity=0.350±0.148, nucleotide diversity=0.0009±0.0008) is commensurate with that of some other related species. This finding indicates that (1) wild populations of the Mekong giant catfish might be more robust than currently thought or (2) present wild populations of this species carry a genetic signature of the historically larger population(s). Findings from this study also have important implications for conservation of the Mekong giant catfish, especially in designing and implementing artificial breeding programme for restocking purposes.     

DNA分子标记技术在濒危物种保护中的应用

近20年来,随着分子生物学技术的迅猛发展,涌现出一批高效、可靠的DNA分子标记技术.本文论述了限制性片段长度多态性、微卫星DNA、随机扩增多态性DNA、扩增片段长度多态性等DNA分子标记技术的基本原理及技术特点;同时,介绍了DNA分子标记在濒危物种种群遗传学研究、致危因素分析及保护策略的制定等保护生物学方面的应用.     

山东省大型真菌物种濒危程度与优先保育评价

近年来我国开展了大型真菌多样性及保育的研究报道。山东省为经济真菌资源大省,但是对于大型真菌濒危物种评价与保育研究的研究未见报道。通过野外调查、市场调查、民间调查及查阅相关文献,山东省共发现大型真菌64科166属435种(包括种下等级),其中野生食药用真菌182种,隶属于39科,80属,其中食用菌123种,占总种数的28.28%,药用菌90种,占总种数的20.69%,其中野生贸易真菌有60种,占总种数的13.79%,占食用菌总数的48.78%,如:白林地蘑菇Agaricus silvicola、林地蘑菇Agaricus silvaticus、灵芝Ganoderma lingzhi、灰树花Grifola frondosa、松乳菇Lactarius deliciosus、亚绒白乳菇Lactarius subvellereus、多脂鳞伞Pholiota adiposa、白蜡多年卧孔菌Perenniporia fraxinea、裂褶菌Schizophyllum commune、黏盖乳牛肝菌Suillus bovinus、点柄乳牛肝菌Suillus granulatus、红绒盖牛肝菌Xerocomellus chrysenteron、长根干蘑Xerula radicata等。在查阅文献的基础上,同时结合大型真菌的生物学、生态学特性,建立了山东省大型真菌物种濒危评价和优先保育评估体系,采用层次分析法(AHP)和专家咨询相结合的方法确定个评价层次的目标权重。对分布在该地区的175种物种的濒危程度和优先保育物种进行了评价,最终获得了山东省大型真菌珍稀物种的濒危等级(V1≥0.650,濒危种;0.650≥V1≥0.550,为脆危种;0.550≥V1≥0.440,敏感种;0.440≥V1,安全种)和优先保护级别评价标准(V1≥1.900,一级保护;1.900≥V1≥1.700,二级保护;1.700≥V1≥1.600,三级保护;1.600≥V1暂缓保护),结果发现该地有濒危种4种,占该区大型真菌总种数的0.92%,灵芝Ganoderma lingzhi、松乳菇Lactarius deliciosus、橙黄红菇Russula aurea、羊肚菌Morchella esculenta;脆危种34种,占该区大型真菌总种数的7.82%;敏感种74种,种该区大型真菌总数的17.01%;安全种63种,占该区大型真菌总数的14.48%。一级保护物种有7种,占该区总种数的1.61%,其中灵芝Ganoderma lingzhi、松乳菇Lactarius deliciosus、羊肚菌Morchella esculenta、橙黄红菇Russula aurea等4个种是人们十分喜爱的食药用菌,其中松乳菇Lactarius deliciosus、橙黄红菇Russula aurea、是外生菌根菌,尚不能够人工栽培;淡玫瑰红鹅膏Amanita pallidorosea、淡紫鸡油菌Cantharellus amethysteu、中国十字孢Crucispora sinensis、山东粉褶菌Entoloma shandongense是中国特有种和该地区特有种;二级保护物种有25种,占该区大型真菌总种数的5.75%;三级保护物种有33种,占该区大型真菌物种总种数的7.59%;暂缓保护物种有110种,占该区大型真菌总种数的25.29%。通过此项研究了解了该地区大型真菌物种的生存状态,对大型真菌资源的有效保护提供了科学依据。     

Development of the musculature in the limpet Patella (Mollusca, Patellogastropoda)

 Whole-mount technique using fluorescent-labelled phalloidin for actin staining and confocal laser scanning microscopy as well as semi-thin serial sectioning, scanning and transmission electron microscopy were applied to investigate the ontogeny of the various muscular systems during larval development in the limpets Patella vulgata L. and P. caerulea L. In contrast to earlier studies, which described a single or two larval shell muscles, the pretorsional trochophore-like larva shows no less than four different muscle systems, namely the asymmetrical main head/foot larval retractor muscle, an accessory larval retractor with distinct insertion area, a circular prototroch/velar system, and a plexus-like pedal muscle system. In both Patella species only posttorsional larvae are able to retract into the shell and to close the aperture by means of the operculum. Shortly after torsion the two adult shell muscles originate independently in lateral positions, starting with two fine muscle fibres which insert at the operculum and laterally at the shell. During late larval development the main larval retractor and the accessory larval retractor become reduced and the velar muscle system is shed. In contrast, the paired adult shell muscles and the pedal muscle plexus increase in volume, and a new mantle musculature, the tentacular muscle system, and the buccal musculature arise. Because the adult shell muscles are entirely independent from the various larval muscular systems, several current hypotheses on the ontogeny and phylogeny of the early gastropod muscle system have to be reconsidered. Received: 23 June 1998 / Accepted: 25 November 1998     

On the verge of extinction: genetics of the critically endangered Iberian plant species, Borderea chouardii (Dioscoreaceae) and implications for conservation management

Borderea chouardii is a relictual and dioecious, strictly sexually reproducing, long-living geophyte of the Dioscoreaceae family. Previous biological and demographic studies have indicated the existence of a uniformly distributed panmictic population of this taxon at the southernmost Spanish pre-Pyrenean mountain ranges where it occurs in rather inaccessible crevices of a single limestone cliff. However, individuals of B. chouardii are spatially subdivided into two subpopulations located, respectively, on the upper and lower parts of the cliff, and vertically separated 150 m. Because of its extreme rarity, B. chouardii was the first Iberian taxon to have a specific conservation plan and has been included in several red lists under the category of critically endangered (CR). However, no previous attempts have been conducted to analyse the fine scale evolutionary mechanisms involved in its present microspatial distribution. Genetic diversity and population structure have been investigated through the analysis of neutral hypervariable markers such as simple sequence repeats (SSRs) and randomly amplified polymorphic DNAs (RAPDs) to unravel the impact of life history traits in the differentiation of the two subpopulations. Both types of molecular markers were unequivocal in distinguishing two genetically distinct groups of individuals corresponding to their spatial separation. However, SSRs detected a higher level of subpopulation differentiation (F(ST) = 0.35, R(ST) = 0.32) than RAPDs (F(ST) = 0.21). SSR data indicated significant deviation from random dispersal of genes and genotypes between the two groups, suggesting that mating occurs mainly among individuals within subpopulations, thus, favouring the divergence between the two groups. This microevolutionary differentiation scenario might have been caused by a coupled effect of past genetic drift and reproductive isolation, as a result of strong glacial age bottlenecks and inefficient dispersal system of pollen and seeds, respectively. The identification of such genetic structure in this narrow endemic prompts a modification of the management strategies of its single extant population.     

The species survival plan and the conference on reproductive strategies for endangered wildlife

The application of advanced reproduction technology to the breeding of vanishing species in captivity, as planned by the Species Survival Plan (SSP) of the American Association of Zoological Parks and Aquariums, may be crucial to practical longterm propagation of some species. However, current techniques are usually specific to domestic animals and man. Major modifications and new approaches, on a species-specific basis, will be required for broad application to specialized wild animals. Prerequisites include basic studies of unique species' physiology and basic care requirements. The current SSP program needs a major research component.     

Allozymic variation and relationships of the endangered cyprinodontid genus Valencia and its implications for conservation

Allozyme analysis was used to determine patterns of genetic variation and relationships within the genus Valencia , a group with two allopatric species, V. letourneuxi inhabiting the Balkan Peninsula, and V. hispanica which is endemic to the Iberian Peninsula. Products of 25 gene loci were analysed with AK *, FH *, LDH -1* and LDH -2* monomorphic when Fundulus heteroclitus macrolepidotus was not taken into account. Nine loci were diagnostic for both species of the genus Valencia . Levels of genetic variation of V. hispanica ( P =0·416, H ^e=0·118) and V. letourneuxi ( P =0·160, H _e=0·040) were higher than values reported previously. High population subdivision ( F _st=0·321) among V. hispanica populations indicates a clear interruption of genetic exchange among populations. High genetic variation and differentiation exhibited by populations of V. hispanica suggest that a recovery program should be carried out with natural stocks from the same localities.     

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.     

Life-history traits of the endangered river blenny Salaria fluviatilis (Asso) and their implications for conservation

The life-history characteristics of the river blenny Salaria fluviatilis in a Mediterranean stream in north-east Spain have been studied. The maximum age observed was 4+ years in both sexes, although only age groups 0+ and 1+ years were well represented. For both sexes, growth was rapid between April and June (before the summer drought) and in October. The instantaneous rate of monthly growth (April to October) was positively correlated with water depth. All fish attained sexual maturity during their first year of life. Females spawned multiple times. Spawning occurred mainly from June to August, although older females (2+ years old) began in May. The relationship between fecundity and total length recorded for older females in the spring was not significantly different from that obtained for 1 year-old females in the summer. The mean diameter of ripe oocytes, however, was greater in older females. Larger eggs that were laid during a period of good food supply (spring) might have a higher probability of survival. The results suggest that female fecundity is affected by environmental conditions. The species is characterized by fast growth, early maturity, multiple spawning and limited longevity. This may be a good life-history strategy for small fish species in unstable environments, where adult mortality is high, variable or unpredictable. Some life-history traits of the river blenny suggest that this species might be vulnerable to more severe drought conditions than those associated with the Mediterranean climate.