First Report of Citrus variegation virus in Palestine Sweet Lime, as Coffee Shade in Costa Rica

Symptoms resembling those of Citrus variegation virus (CVV) were observed in old Palestine sweet lime trees (Citrus limettioides Tan.) used as coffee shade in the Central Valley in Costa Rica. The symptoms include leaf flecking, mosaic, malformation, and dwarfing. This disease was transmitted by grafting to “Valencia” sweet orange (C. sinensis L.), Etrog Citron (C. medica L.), and sweet lime under greenhouse conditions. Symptoms were severe in Palestine sweet lime all year long, but sweet orange and Etrog citron were asymptomatic under high temperature. Reverse transcription polymerase chain reaction (RT-PCR) was used to confirm the presence of CVV in young leaves from sweet lime trees collected from field, and other Citrus spp. grafted and maintained in the greenhouse at University of Costa Rica. Fresh tissue from trees infected with CVV obtained from the USDA (Riverside, California), was used as positive control. Furthermore, the sequence obtained (605 bp) was analyzed by Blastn algorithm which showed 97% homology with CVV RNA 3 (GenBank Accession No. AF434912). Additionally, small scale virus purification was carried out and isometric particles (26 to 32 nm) were observed under the electron microscope. To our knowledge, this is the first report of the presence of CVV in Costa Rica infecting Palestine sweet limes. Also, we remark on the potential of Palestine sweet lime as a host plant indicator in biological indexing of CVVMinisterio de Ciencia y Tecnología, Fondo Reinserción/[222-2004]/MICIT- CONICIT/Costa RicaUniversidad de Costa Rica/[801-A5-506]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de AgronomíaUCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Biologí

Genetic connectivity of an endangered shark across nursery areas from the Eastern Tropical Pacific

Defining demographically independent units and understanding gene flow between them is essential for managing and conserving exploited populations. The scalloped hammerhead shark, Sphyrna lewini , is a coastal semi-oceanic species found worldwide in tropical and subtropical waters. Pregnant females give birth in shallow coastal estuarine habitats that serve as nursery grounds for neonates and small juveniles, and adults move offshore and become highly migratory. We evaluated the population structure and connectivity of S. lewini in coastal areas across the Eastern Tropical Pacific (ETP) using both sequences of the mitochondrial DNA control region (mtCR) and nuclear-encoded microsatellite loci. The mtCR defined two genetically discrete geographic groups: the Mexican Pacific and the central-southern Eastern Tropical Pacific (Guatemala, Costa Rica, Panamá, and Colombia). Overall, the mtCR data showed low levels of haplotype diversity ranged from 0.000 to 0.608, while nucleotide diversity ranged from 0.000 to 0.0015. A more fine-grade population structure analysis was detected using microsatellite loci where Guatemala, Costa Rica, and Panamá differed significantly. Genetic diversity analysis with nuclear markers revealed an observed heterozygosity ranging from 0.68 to 0.71 and an allelic richness from 5.89 to 7.00. Relatedness analysis revealed that individuals within nursery areas were more closely related than expected by chance, suggesting that S. lewini may exhibit reproductive philopatric behaviour within the ETP. Findings of at least two different management units, and evidence of philopatric behaviour call for intensive conservation actions for this critically endangered species in the ETP.Universidad de Costa Rica/[801-B6-214]/UCR/Costa RicaNational Secretary of Science and Technology/[FID-156]/SENACYT/EcuadorThe Phoenix Zoo/[no.33297]//Estados UnidosPADI Foundation/[no.32809]//Estados UnidosRufford Foundation/[no.22366-1]//Reino UnidosWaitt Foundation/[no.33297]//Estados UnidosFundación Reserva Ojochal/[]//Costa RicaThe Whitley Fund for Nature/[]/WFN/Reino UnidoSandler Family Foundation/[]//Estados UnidosOsa Conservation/[]//Costa RicaUCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de BiologíaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Ciencias del Mar y Limnología (CIMAR

Estudio sobre el patrón de acumulación y multiplicación del virus del rayado fino en plantas de maíz infectadas (Zea mays L.)

Tesis (Magister Scientiae)--Universidad de Costa Rica. Comisión del Programa de Estudios de Posgrado en Biología, 1992.UCR::Vicerrectoría de Investigación::Sistema de Estudios de Posgrado::Ciencias Básicas::Maestría Académica en Biologí

Genetic diversity analysis of the endangered slipper orchid Phragmipedium longifolium in Costa Rica

Phragmipedium longifolium is an endangered terrestrial orchid. In Costa Rica, these plants are found growing in small and isolated patches, some of them consisting of just four to six individuals. Information about the genetic variability within and among populations is very important for the conservation of this endangered species. A total of 160 samples were collected in six locations and analyzed with amplified fragment length polymorphism technique. The genetic diversity of P. longifolium in Costa Rica (Hw = 0.1711) is high and differentiation among sampled locations is moderate (upt = 0.2013) in comparison with results of studies in some other terrestrial orchid species using the same technique. The percentage of polymorphic loci was, on average, 51.5. The analysis of molecular variance (AMOVA) indicated the main genetic variation was within sampled locations (80%), even though the variation among locations was also significant. In situ conservation is recommended because, in addition to protecting habitat and avoiding fragmentation, mycorrhizal fungi and pollinators are also protected. Close proximity between populations is required to maintain high genetic variability through a gene flow continuum. It is suggested that conservation of patches with higher genetic variability be prioritized as many are located in unprotected areas. A germplasm bank for ex situ conservation has been established in the living collection at Lankester Botanical Garden using the plants collected in this study. Finally, a search for new locations of this species is also suggested.Universidad de Costa Rica/[814-A6-107]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Jardín Botánico Lankester (JBL)UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM

Molecular data indicate that Rhytidhysteron rufulum (ascomycetes, Patellariales) in Costa Rica consists of four distinct lineages corroborated by morphological and chemical characters

Rhytidhysteron rufulum is a poorly known, common, pantropical species, capable of utilizing different substrata and occupying diverse habitats, and is the only species of its genus in Costa Rica. We have employed molecular, morphological, and chemical data to assess the variability and differentiation of R. rufulum in Costa Rica, including sites from the Pacific and Atlantic coast. Phylogenetic analyses of nuclear ITS rDNA sequences revealed the presence of four distinct lineages in the R. rufulum complex. Re-examination of the morphology and anatomy showed differences between these lineages in ascomatal, ascal, and ascospore size that have previously been regarded as intraspecific variations. In addition, there was a correlation between molecular phylogenies and chemical components as determined by hplc and nuclear magnetic resonance (NMR). Two lineages (clades I and II) produced the palmarumycins MK-3018, CJ-12372, and CR1, whereas clade III produced dehydrocurvularin, and clade IV unidentified compounds. Our results based on a polyphasic approach contradict previous taxonomic interpretations of one morphologically variable species.Ministerio de Ambiente y Energía/[R-CM-INBIO-05-2006-OT]/MINAE/Costa RicaUCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de BiologíaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Químic

Analysis of genetic diversity of Cucurbita moschata (D.) germplasm accessions from Mesoamerica revealed by PCR SSCP and chloroplast sequence data

The aim of this research was the genetic characterization of 218 accessions of Cucurbita moschata Duchesne, a squash, and its relationship with morphological characteristics of agronomic interest, which are part of the international collection conserved at Tropical Agricultural Research and Higher Education Center (CATIE), Costa Rica. The majority of the accessions came from Mexico and Central America; single genotypes from Curaζao, Colombia, Peru and the Russian Federation were also included. The polymerase chain reaction (PCR) and single strand conformation polymorphism analysis (SSCP) were used for the analysis of the regions amplified with ITS1–ITS2 nuclear primers and tRNL–F chloroplast primers. Haplotypes were constructed according to band patterns in SSCP gels. Twenty-five haplotypes were found using the ITS1–ITS2 markers, and 24 haplotypes were found with the tRNL–F markers. Unique haplotypes were found with both markers. Two individuals of each tRNL–F haplotype were sequenced. The results indicated a high level of genetic diversity in CATIE squash collection. Using ITS1–ITS2 primers, it was found that the number of haplotypes was independent of the geographical source of the accession, and haplotypes were distributed randomly throughout the study area. Mexico had the highest values of total heterozygosity (HE), genetic diversity (H) and Shannon index (I) while Panama showed the lowest values. Sequences obtained from tRNL–F intergenic marker showed the highest diversity index values were present in the group of additional sequences and Mexico, and lower values were observed for Nicaragua, Guatemala and Panama. PCoA based on morphological data showed three groups and by ANOSIM (R) all group differences were significant. Results obtained in this study suggest that high diversity is a characteristic of C. moschata from Mesoamerica.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Biologí

Genetic Diversity of Xylella fastidiosa Strains from Costa Rica, São Paulo, Brazil, and United States

Copyright 2007 American Phytopathology Society Journals.The diversity of 42 Xylella fastidiosa strains from Costa Rica, São Paulo, Brazil, and the United States were analyzed using the sequence of the 16S rRNA gene by variable number of tandem repeat (VNTR) fragment analysis and by restriction fragment length polymorphisms (RFLP) of a specific polymerase chain reaction (PCR)-amplification product using enzyme CfoI. Limited variability in the sequence of the 16S rRNA gene was observed and, although the separation was not absolute, most strains from Costa Rica clustered with strains from the United States and not with strains from São Paulo. The PCR-RFLP produced different patterns of DNA bands. The same pattern was shared by strains from Costa Rica, the United States, and two coffee strains from São Paulo, but a different pattern was observed in six coffee and orange strains from Brazil. In all, 32 amplification products were scored in the VNTR fragment analysis. The total variation observed among the X. fastidiosa strains had significant (P < 0.001) contributions from both geography and host origin as inferred by Nei’s values of genetic diversity and WINAMOVA statistics. The strains from Costa Rica were isolated from diseased grapevines, coffee, and sweet orange and these strains grouped together and could be distinguished from strains from grapevine from the United States or from either coffee or sweet orange from São Paulo. The strains tested from Costa Rica are most likely of local origin, although the possibility that they have been introduced along with horticultural crops cannot be excluded. In either case, they are examples of independent selection of strains of X. fastidiosa affecting coffee and sweet orange. Greater genetic similarity was observed between strains from Costa Rica and the United States than with those from São Paulo.Fundación CRUSAUniversidad de Costa Rica/[801-A2-528]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM

Phylogenetic reassessment of Specklinia and its allied genera in the pleurothallidinae (Orchidaceae)

The phylogenetic relationships within Specklinia (Pleurothallidinae; Orchidaceae) and related genera are re-evaluated using Bayesian analyses of nrITS and chloroplast matK sequence data of a wide sampling of species. Specklinia is found paraphyletic in the DNA based trees, with species alternatively assigned to Muscarella proven distinct, monophyletic and easily recognizable. Specklinia as such includes about 100 morphologically highly diverse species. Their phenotypic differences have prompted the creation of up to eleven generic names within this relatively small group. Here we show not only that these morphologically divergent species are closely related, but also that they can still be recognized by certain conserved morphological traits. The genera Acostaea, Areldia, Empusella, Cucumeria, Gerardoa, Pseudoctomeria, Sarcinula, Sylphia, Tribulago and Tridelta are found embedded within Specklinia, and therefore reduced under the synonymy of the latter. Specklinia is confirmed as sister to a clade that includes Platystele, Scaphosepalum and Teagueia. Five well-supported subgenera are proposed for Specklinia and are characterized both geographically and morphologically. The species belonging to each subgenus are listed. Incaea is synonymized with Dryadella, while Rubellia is reduced under Platystele. New combinations for several species are proposed. The criteria for the generic delimitation of Specklinia and other genera in the Pleurothallidinae are discussed.The Alberta Mennega foundation///Países BajosUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Jardín Botánico Lankester (JBL

Catharanthus roseus (Apocynaceae) naturalmente infectada con diversos fitoplasmas en Costa Rica

Phytoplasmas (class Mollicutes) are causal agents of plant diseases with an economic impact on crops or threatening local biodiversity. A survey was conducted from 2012 to 2016 on infected Catharanthus roseus plants that exhibited symptoms reminiscent of phytoplasma infection throughout Costa Rica. A total of 73 plants were collected exhibiting symptoms such as virescence, phyllody, axillary proliferation, little leaf, leaf malformation, chlorosis, or yellowing. All samples were tested by nested PCR using phytoplasma universal and specific primer pairs. Phytoplasma infection was detected in 52 (71.2 %) of the plants collected. Phytoplasmas of six subgroups belonging to 16Sr groups I, III, IX, XIII and XV were identified based on sequencing and in silico RFLP analyses. ´Candidatus Phytoplasma asteris´ (16SrI) was the predominant group among the positive samples (n = 30) showing variety of symptoms and wide distribution from sea level to ca. 1 400 masl in six of the seven Costa Rican provinces. Group 16SrIII was the second most abundant (14 samples); and the remaining three groups were seldom found in C. roseus (8 samples). Moreover, group 16SrXIII phytoplasma was detected for the first time in the country. To the best of our knowledge, this is the first report of natural infection of C. roseus with phytoplasma subgroups 16SrI-B, 16SrI-P, 16SrIII-F, 16SrIX-F, 16SrXIII-A, and 16SrXV-B in Costa Rica and Central America.Catharanthus roseus (Apocynaceae) naturalmente infectada con diversos fitoplasmas en Costa Rica. Los fitoplasmas (clase Mollicutes) son agentes causales de enfermedades de plantas que provocan pérdidas económicas o amenazan la biodiversidad local. Una recolecta de plantas de Catharanthus roseus que mostraban síntomas de posible infección con fitoplasmas se realizó en diferentes lugares de Costa Rica desde 2012 a 2016. Un total de 73 plantas fueron recolectadas con síntomas tales como viriscencia, filodia, brotación axilar múltiple, reducción foliar, deformación foliar, clorosis, y amarillamiento. Todas las muestras fueron evaluadas mediante PCR anidado usando los pares de imprimadores universales y específicos para fitoplasmas. Infección por fitoplasmas se detectó en 52 (71.2 %) de las muestras. Fitoplasmas de seis subgrupos dentro de los grupos 16Sr I, III, IX, XIII y XV fueron identificados basados en secuenciación del ADN y análisis de polimorfismos de restricción (RFLP) in silico. El grupo predominante encontrado en las muestras positivas (n = 30) fue el 16SrI (‘Candidatus Phytoplasma asteris’), éste mostró variedad de síntomas y amplia distribución desde el nivel del mar hasta casi los 1 400 m.s.n.m. en seis de las siete provincias de Costa Rica. El grupo 16SrIII fue el segundo más abundante (14 muestras); y los restantes tres grupos se encontraron en pocas muestras de C. roseus (8 muestras). Además, fitoplasmas del grupo 16SrXIII se detectaron por primera vez en el país. De acuerdo a nuestro conocimiento, este es el primer informe de infección natural de C. roseus con fitoplasmas de los subgrupos 16SrI-B, 16SrI-P, 16SrIII-F, 16SrIX-F, 16SrXIII-A y 16SrXV-B en Costa Rica y Centroamérica.Universidad de Costa Rica/[801-A1-801]/UCR/Costa RicaUniversidad de Costa Rica/[801-B3-091]/UCR/Costa RicaUniversidad de Costa Rica/[801-B7-138]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM

Phenolic variation among Chamaecrista nictitans subspecies and varieties revealed through UPLC-ESI(-)-MS/MS chemical fingerprinting

Introduction Comparative analysis of metabolic features of plants has a high potential for determination of quality control of active ingredients, ecological or chemotaxonomic purposes. Specifically, the development of efficient and rapid analytical tools that allow the differentiation among species, subspecies and varieties of plants is a relevant issue. Here we describe a multivariate model based on LC–MS/MS fingerprinting capable of discriminating between subspecies and varieties of the medicinal plant Chamaecrista nictitans, a rare distributed species in Costa Rica. Methods Determination of the chemical fingerprint was carried out on a LC–MS (ESI-QTOF) in negative ionization mode, main detected and putatively identified compounds included proanthocyanidin oligomers, several flavonoid C- and O-glycosides, and flavonoid acetates. Principal component analysis (PCA), partial least square-discriminant analysis (PLS-DA) and cluster analysis of chemical profiles were performed. Results Our method showed a clear discrimination between the subspecies and varieties of Chamaecrista nictitans, separating the samples into four fair differentiated groups: M1 = C. nictitans ssp. patellaria; M2 = C. nictitans ssp. disadena; M3 = C. nictitans ssp. nictitans var. jaliscensis and M4 = C. nictitans ssp. disadena var. pilosa. LC–MS/MS fingerprint data was validated using both morphological characters and DNA barcoding with ITS2 region. The comparison of the morphological characters against the chemical profiles and DNA barcoding shows a 63% coincidence, evidencing the morphological similarity in C. nictitans. On the other hand, genetic data and chemical profiles grouped all samples in a similar pattern, validating the functionality of our metabolomic approach. Conclusion The metabolomic method described in this study allows a reliably differentiation between subspecies and varieties of C. nictitans using a straightforward protocol that lacks extensive purification steps.Universidad de Costa Rica/[809-B3-082]/UCR/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigaciones en Productos Naturales (CIPRONA)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de QuímicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Biologí