Strategies for ex situ conservation of Centaurea cineraria subsp. circae (Asteraceae), an endemic plant from Lazio (Italy)

Centaurea cineraria subsp. circae is an endemic plant with a distribution area limited to Circeo mountain (Lazio, Italy), whose population was estimated in a very low number of individuals. The aim of this work was to investigate ex situ conservation strategies such as achene collection and in vitro plant propagation, which will permit to carry out restoration programmes. The test carried out on the achenes demonstrated that only 5.5% of them were morphologically healthy. Seed germination tests showed that seeds do not display dormancy and that germination does not require pre-treatments. The higher germination rate (67.5%) was observed under a photoperiod of 12/12 h (light/dark) and temperature regime +20/+10°C. The in vitro studies demonstrated that micropropagation, acclimatization and the transfer outdoors of C. cineraria subsp. circae are not particularly difficult: 74% of shoot explants in a Murashige and Skoog (MS) medium added with 0.5 mg/l benzylaminopurine and 2 mg/l kinetin formed multiple shoots; 100% of shoots rooted in the MS medium added with 0.5 mg/l indole-3-butyric acid and over 90% survived the acclimatization phase. After been transferred outdoors, the totality of in vitro-propagated plants bloomed and appeared morphologically indistinguishable from wild plants. Preliminary chemical analyses showed a similar profile for in vitro-propagated and wild plants.     

Conservation genetics of the rare Iberian endemic Cheirolophus uliginosus (Asteraceae)

Cheirolophus uliginosus is a rare species, endemic to the south‐western Iberian Peninsula, and listed as a characteristic taxon from the temperate Atlantic wet heaths, a priority habitat for conservation by the European Union. The conservation status of this species in most of its distribution area is poorly known, but, in recent times, some populations have disappeared and there has been a reduction in the number of individuals in others. In this context, we analysed the effects of population size on genetic diversity, revealing that genetic erosion and inbreeding depression could be having a significant impact on smaller populations. Furthermore, we studied the patterns of genetic structure and variability at the species level, finding a strikingly low within‐population diversity and high among‐population genetic differentiation. Finally, the genetic structure analyses suggested a long and complex phylogeographical history of C. uliginosus in the region, in agreement with the climate relict status proposed for this species. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 179 , 157–171.     

Overcoming seed dormancy in ex situ plant germplasm conservation programmes; an example in the endemic Argyranthemum (Asteraceae: Anthemideae) species from the Canary Islands

Germplasm of 21 diverse Argyranthemum taxa was collected from contrasting ecological zones in the Canary Islands. Seed dormancy was considerable in the majority of taxa. Extensive investigations, based on a germination test procedure algorithm for Asteraceae, with achenes from ray and disc florets of five contrasting taxa identified a procedure to promote full (85%) germination of the seeds from both ray and disc florets of all five taxa; viz, excision of the seeds from the achenes, followed by testing at 15°C with 2.6×10^-3 m GA₃ co-applied. Subsequent tests showed that this regime was effective in promoting full germination in seeds from both ray and disc florets of the remaining 16 taxa. The results are discussed in the context of ex situ plant germplasm conservation.     

Karyological studies in Tripleurospermum (Asteraceae, Anthemideae) from north-east Anatolia

Chromosome number and morphology in 14 taxa belonging to 19 populations of Tripleurospermum Sch. Bip. were studied using karyological and numerical taxonomical techniques. Data on chromosome measurements were analysed using cluster analysis. Chromosome numbers of these taxa are 2 n  = 2 x  = 18, 4 x  = 36 and 5 x  = 42–48. Seven records are new, two are not consistent with previous counts, and the remainder confirm the very limited previous data (one to three records). A new ploidy level (pentaploidy) is reported for the first time for the genus. Some correlations between ploidy levels and morphological characters are noted and several systematic and evolutionary aspects of the genus are discussed in the light of karyological data.  © 2004 The Linnean Society of London, Botanical Journal of the Linnean Society , 146 , 427–438.     

珍稀濒危植物迁地保护策略中植物营养问题的探讨

营养缺乏、过量或不平衡都会直接导致植物生长繁殖障碍甚至死亡,通过改良土壤或补充矿质元素来解决这些问题是植物营养学研究的主要内容。这一学科的研究范围已经从传统人工生态系统发展到自然生态系统中。但总的来说,这一学科还没有真正渗透到珍稀濒危物种的迁地保护领域中来。中国各植物园目前正在开展大规模的珍稀濒危植物迁地保护工作,但人们都只重视了物种的引入,却忽视了它们在迁地保护过程中业已存在和可预见的生长繁殖障碍,对迁地保护植物的研究很少涉及其生长繁殖所需的营养条件。针对这一现状,本文提出了目前应开展的几项工作:(1)明确优先开展植物营养学研究的物种范围;(2)物种原产地土壤与植物化学元素背景值的分析;(3)迁地保护物种营养状况动态监测与营养诊断。该研究思路的提出,无论是从植物营养学与植物保育学的学科交叉角度,还是从珍稀濒危物种迁地保护保育策略的理论探索与解决实际问题的角度,都具有十分重要的意义。     

Reproductive status of endemic felid species in Latin American zoos and implications for ex situ conservation

Reproductive evaluations were conducted on 185 male cats representing eight endemic Latin American species that were maintained in 44 zoos and private facilities in 12 Latin American countries. Reproductive assessments (testicular measures, ejaculate quality, and blood testosterone/cortisol concentration) were used to establish normative values for large‐ and small‐sized cats in Latin American collections. Data also were analyzed using multiple regression to study the impact of proven breeder status, diet, and various animal housing combinations. Most felids (>95%) in the survey were of wild‐born origin, and <20% had produced offspring in captivity. Larger felids had bigger testes and produced more semen, but tended to produce low‐sperm‐density ejaculates. The ejaculates of small felids were more sperm‐concentrated, but contained fewer total spermatozoa. Sperm motility was unrelated to species size, and certain species (puma, margay, tigrina, and jaguarundi) consistently produced few (<40%) normal sperm forms. Across species, >50% of males had low sperm counts (<1 million total sperm per ejaculate). Among large cats (jaguars and pumas), proven breeders had larger (P<0.05) testes, greater semen volume, and more normal sperm than nonbreeders. Males on adequate diets had higher (P<0.05) circulating cortisol. Among small‐sized felids, proven breeders had higher (P<0.05) testosterone, and males housed alone or paired with a conspecific female had more (P<0.05) total sperm per ejaculate and greater (P<0.05) seminal and testicular volumes. Fifty‐nine ejaculates (potentially representing ～100 artificial insemination (AI) or 26,000 in vitro fertilization (IVF) procedures) were cryopreserved for a felid genome resource bank. In conclusion, breeding success and reproductive traits for many endemic felids in Latin American zoos appear to be suboptimal, and likely would benefit from improvements in diet and exhibitry. Technology transfer and continued training of zoo staff and scientists in Latin American countries are essential if these zoos are to achieve their tremendous conservation potential for felids and other threatened endemic species. Zoo Biol 22:421–441, 2003. © 2003 Wiley‐Liss, Inc.     

Pollen morphology of some endemic Turkish Centaurea L. (Asteraceae, section Phaloletis) and their taxonomic value

In this study, the detailed pollen morphological structures of some endemic Turkish species of Centaurea amaena Boiss., C. antalyense H. Duman & A. Duran, C. aphrodisea Boiss., C. hierapolitana Boiss., C. luschaniana Heimerl, C. lycia Boiss., C. tossiensis Freyn. Et Sint., and C. wagenitzii Hub.-Mor. (Asteraceae, section Phaloletis) were studied under light microscopy (LM) and scanning electron microscopy (SEM) for the first time. LM and SEM investigations showed that the pollen grains of eight taxa are more or less spheroidal-subprolate, the amb triangular and tricolporate. The exine sculpture is tectate, microechinate-scabrate in the pollen of Centaurea taxa. Spinules are less dense in Centaurea amaena, C. antalyense, C. hierapolitana, and C. lycia, but they are more dense in C. aphrodisea, C. luschaniana, C. tossiensis, and C. wagenitzii. Spinule dimensions are different from each other. The exine has one layer of columellae beneath the spines. We determined all taxa that have the Helianthoid type. Exine sctructure and sculpture as well as spine density and dimensions in Asteraceae are the most reliable characteristics for discriminating taxa.     

Population genetics of the endangered Canarian endemic Atractylis arbuscula (Asteraceae): implications for taxonomy and conservation

We use the information provided by 36 RAPD loci and 15 morphological traits to describe and construe the population differentiation in A. arbuscula, a Canarian endemic Asteraceae threatened with extinction that exhibits a disjunct distribution in the islands of Gran Canaria (var. schizogynophylla) and Lanzarote (var. arbuscula). Our evaluation of morphological characters and the extent of RAPD differentiation found sets the stage for a taxonomic restructuring to hoist both var. arbuscula and var. schizogynophylla to subspecies category. Our genetic results suggest that fragmentation and generally low population sizes are jeopardising the survival of this species through a predominance of stochastic processes in microevolutionary dynamics, especially in Gran Canaria, where subpopulations exhibit the lowest levels of genetic variation and gene flow. If, as most Asteraceae, these plants have a sporophytic self-incompatibility system, the scarce reproductive turnover observed in the much smaller subpopulations from Gran Canaria (ssp. schizogynophylla) is possibly more influenced by a deficiency of S-alleles (that would be provoking the unavailability of compatible mating types, and a cascade of deleterious effects associated with the Allee effect) than by a reduced pollinator visitation frequency. Based on the depauperated values of genetic parameters for this subspecies, urgent mixed subpopulation reinforcements and reintroductions (with specimens belonging to the same subspecies) seem advisable on the grounds that they might allow the isolated habitat remnants to retrieve from inbreeding through the introduction of a new stock of S-alleles and the subsequent production of genotypes that may have a higher selective value.     

Effective seed harvesting strategies for the ex situ genetic diversity conservation of rare tropical tree populations

Tropical rain forests harbor a high diversity of tree species, a large portion of which are considered rare and threatened by anthropogenic factors such as land-use change. Addressing the global need for ex situ rescue populations of such species, we investigated whether certain methods of wild-source seed collection harvest greater amounts of genetic variation from the source population while minimizing the costs expended. We used an individual-based computer program (NEWGARDEN) to conduct simulation trials examining which aspects of seed procurement provided the most cost-effective methods for maximal rare allele capture for single isolated populations of rare tropical tree species. Aspects examined included the geometric pattern of harvesting, location of seed collections within the spatial distribution of the population (central vs. peripheral), geometric partitioning of sampling regions (contiguous vs. subdivided), gene dispersal distance, linear travelling costs, and ratio of seeds collected to plants sampled. Results demonstrated that ratio of seeds collected to plants sampled had the highest effect on genetic diversity harvested relative to sampling location or geometric pattern of collection, explaining 82 and 79% of the variance in alleles harvested, respectively. Collecting from subdivided peripheral sampling regions and populations having long distance gene dispersal led to increases of up to 39 and 51% in the amount of genetic variation conserved respectively. These results suggest that ex situ conservation efforts for rare tropical trees can benefit by collecting fewer seeds from each of a larger number of trees, sampling from peripheral subdivided regions of the population rather than in a contiguous centrally located one under the transect or nearest neighbor strategies, and adjusting harvesting based on the dispersal biology of the source population. This study underscores the need for more information regarding the dispersal characteristics of the target species and demonstrates the value of predictive modeling for planning seed collection projects.     

Germination responses to GA3 and short‐time chilling of three endemic species: Tripleurospermum pichleri,Cirsium leucopsis and Senecio olympicus (Asteraceae)

In the present study, the germination characteristics of three endemic species from Turkey, Tripleurospermum pichleri (Boiss.) Bornm., Cirsium leucopsis D.C and Senecio olympicus Boiss. (Asteraceae), were investigated. Germination was studied for fresh seeds, for seeds subjected to short‐time chilling (15 days, moist +4°C), to GA₃ (100, 150 and 250 ppm) and a combination of chilling and GA₃; in all cases seeds were incubated either at 20/10°C day/night with light daytime or at 20°C in darkness with daily short‐time dim light (DSDL). In C. leucopsis seeds, all GA₃ treatments enhanced the final germination percentages. The mean germination time (MGT) of C. leucopsis was lower under DSDL than with photoperiod. The chilling treatment with GA₃ in DSDL significantly increased germination in S. olympicus seeds (from 45 to 87%). Germination increased to 55% in T. pichleri by chilling under photoperiod compared with 32% by chilling followed by DSDL. In conclusion, these three co‐existing endemic Asteraceae species have different germination behaviours; something that should be taken into account for ex situ propagation. However, an efficient way to germinate all species is to use 250 ppm GA₃ and 20/10°C with photoperiod.     

GIS-facilitated ex situ conservation of the rare Greek endemic Campanula incurva Aucher: Seed germination requirements and effect of growth regulators on in vitro proliferation and rooting

In order to develop conservation protocols for Campanula incurva, the geographical information systems (GIS) were used to unveil its ecological requirements; this facilitated the selection of substrates and of appropriate temperatures for cultivation and guided propagation experiments and acclimatization. Seed germination was tested under (i) dark, (ii) 16-h photoperiod, (iii) immersion in 400 ppm gibberellic acid (GA₃) followed by incubation at dark, and (iv) immersion in 400 ppm GA₃ followed by incubation at 16-h photoperiod (all at 21 ± 1°C). Dormancy was not detected. Germination exceeded 85% in 10 days. Shoot tips were established in vitro in Murashige and Skoog (MS) medium with 1 μM 6-benzyladenine (BA) and 0.1 μM indole-3-butyric acid (IBA). The effect of 1–8 μM BA and 1–8 μM kinetin on shoot proliferation was studied. Moreover, 8 μM BA was combined with 0, 1, 5, and 10 μM IBA to investigate effects of cytokinin/auxin. The highest number of microshoots/explant (4.03) was obtained with 8 μM BA. Microshoots were transferred to half strength MS and full strength MS media with 0, 0.5, 1, 5, and 10 μM IBA to evaluate their root induction ability. Half strength MS medium with 5 μM IBA resulted in 100% rooting (16.80 average number of roots/microshoot). Plantlets produced were successfully acclimatized.     

Genetic variation relevant for the conservation of the narrow endemic Senecio carbonensis (Asteraceae) from the southern Andes

We studied morphological variation, reproductive biology and genetic structure of Senecio carbonensis, a narrow endemic from high elevations of the southern Andes, and compared it to Senecio peteroanus, a closely related widespread congeneric species. Using ANOVAs and discriminant analysis we showed that populations of S. carbonensis were more similar to each other in their morphology, had comparatively limited reproductive capacity and presented lower plant density and more reduced plant cover than populations of S. peteroanus. Similar high genetic variation was found at the population and species level in both species, based on isozyme variation at 14 and 11 putative gene loci that were resolved in S. carbonensis and S. peteroanus, respectively. The two species were genetically distinct. However, the small genetic distance between populations of each species suggests that either they were recently founded and that genetic drift did not have time to promote divergence, or that its effects are masked by significant current gene flow. These results show that ecological and reproductive characteristics of S. carbonensis may limit its extension and abundance. In contrast, high genetic variation at the population level is probably favored by self‐incompatibility. Despite the restricted geographical distribution and low plant density of S. carbonensis populations, genetic characteristics do not seem to be limiting their long‐term persistence. This information suggests that populations of S. carbonensis are not at risk of extinction as a result of genetic factors, although it is important to study and monitor population dynamics to further assess the degree of recruitment through time.     

Microbial diversity: ex situ conservation of Indonesian microorganisms

This paper discusses the role of microbial resource collections in the ex situ conservation of microbial diversity and, in particular, the activities of a recently completed Darwin Initiative project to facilitate collection, identification and exploitation of Indonesian microbial diversity. This helped revitalize the collection of the Lembaga Ilmu Pengetahuea Indonesia (Indonesian Institute of Sciences), LIPI, Research and Development Centre for Biology, Bogor, Indonesia. It is suggested that a focus on microorganisms beneficial to man is essential to enable ex situ collections to be a useful tool in the conservation and sustainable utilization of microbial diversity. This task is enormous and far beyond the capabilities of one collection and its achievement will necessitate a network of collections.     

Failed protective effort of ex situ conservation of River Vistula trout (Salmo trutta) in Sweden

Environmental Biology of Fishes - Ex situ conservation comprises some of the oldest and best-known conservation methods and it has been applied for different fish stocks. This study describes...     

Karyosystematic studies on threeCrepis species (Asteraceae) endemic to Greece

This work examines the cytogeographical distribution, the morphological characters, and the karyotypes of threeCrepis species endemic to Greece (C. sibthorpiana, C. incana, andC. heldreichiana). C. sibthorpiana is diploid (2n = 2x = 8),C. incana is diploid (2n = 2x = 8) and tetraploid (2n = 4x = 16, 17), andC. heldreichiana is always dekaploid (2n = 10x = 40). The Giemsa positive bands, usually pairs of dots, are mainly centromeric inC. incana, while they are terminal inC. sibthorpiana (on the short arm of all chromosomes) and inC. heldreichiana (on both arms of all chromosomes). Intercalary C-bands are scarce and usually variable within karyotypes, individuals, and species. The most variable karyotype both in Feulgen and Giemsa preparations is that ofC. incana, in which also supernumerary chromosomes were observed, which are polysomic to standard set members. On the basis of morphological and karyological data the evolutionary relationships among the threeCrepis taxa are discussed.     

Quasi in situ: a bridge between ex situ and in situ conservation of plants

Plant conservation urgently needs a concept that would unify different aspects of population viability as parts of conservation methodology. Such unification is especially lacking for ex situ conservation. We introduce a novel conservation approach in which ex situ collections maintained in natural or semi-natural environment and preserving both neutral and adaptive genetic diversity are a part of a complementary ex situ–in situ conservation strategy. Our approach is the first that explicitly takes into account ecologically significant (i.e. adaptive) variation of plants in both ex situ and in situ conservation actions. Using this approach we provide detailed guidelines for (1) representative sampling of the populations; (2) collection maintenance; and (3) utilization for in situ actions.     

Chromosome numbers of five Turkish Centaurea L. (Asteraceae) species

Abstract

We analysed the effects of patch configuration and site history on vascular plants in ancient forests of the Eastern Po Plain, documented back to 1740. Despite their reduced size, all the forests are part of Natura 2000 Network and significantly contribute to the maintenance of a threatened habitat and support biological diversity of the Continental biogeographic region. The presence of some functional ecological plant species groups was correlated with patch configuration and age. Habitat quality, in terms of suitability for forest species, was found to be important in explaining the presence of species of high conservation value, but patch age (as an indicator for habitat quality) played a major role too. For core forest species, patch area is a redundant variable in explaining species richness relative to habitat quality and patch age and the extinction of specialists seems to occur mainly in a deterministic way. Even small forest fragments can be very important for maintaining plant species diversity, at least if they are of high habitat quality and if the forest management is appropriate. However, to achieve a long term conservation, management plans should also aim at an improvement of the anthropogenic matrix surrounding forest remnants.     

Restoration genetics of the vernal pool endemic Lasthenia conjugens (Asteraceae)

Restoration of habitat for endangered species often involves translocation of seeds or individuals from source populations to an area targeted for revegetation. Long-term persistence of a species is dependent on the maintenance of sufficient genetic variation within and among populations. Thus, knowledge and maintenance of genetic variability within rare or endangered species is essential for developing effective conservation and restoration strategies. Genetic monitoring of both natural and restored populations can provide an assessment of restoration protocol success in establishing populations that maintain levels of genetic diversity similar to those in natural populations. California’s vernal pools are home to many endangered plants, thus conservation and restoration are large components of their management. Lasthenia conjugens (Asteraceae) is a federally endangered self-incompatible vernal pool annual with gravity- dispersed seeds. Using the molecular technique of intersimple sequence repeats (ISSRs), this study assessed levels and patterns of genetic variability present within natural and restored populations of L. conjugens. At Travis Air Force Base near Fairfield, California, a vernal pool restoration project is underway. Genetic success of the ecologically based seeding protocol was examined through genetic monitoring of natural and restored populations over a three-year period. Genetic diversity remained constant across the three sampled generations. Diversity was also widely distributed across all populations. We conclude that the protocol used to establish restored populations was successful in capturing similar levels and patterns of genetic diversity to those seen within natural pools. This study also demonstrates how genetic markers can be used to inform conservation and restoration decisions.