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Endophytes play an important role in plant development, survival, and establishment, but their temporal dynamics in young conifer plants are still largely unknown. In this study, the bacterial community was determined by metabarcoding of the 16S rRNA gene in the rhizoplane, roots, and aerial parts of 1- and 5-month-old seedlings of natural populations of Abies religiosa (Kunth) Schltdl. & Cham. In 1-month-old seedlings, Pseudomonas dominated aerial parts (relative abundance 71.6%) and roots (37.9%). However, the roots exhibited significantly higher bacterial species richness than the aerial parts, with the dissimilarity between these plant sections mostly explained by the loss of bacterial amplification sequence variants. After 5 months, Mucilaginibacter dominated in the rhizoplane (9.0%), Streptomyces in the roots (12.2%), and Pseudomonas in the aerial parts (18.1%). The bacterial richness and community structure differed significantly between the plant sections, and these variations were explained mostly by 1-for-1 substitution. The relative abundance of putative metabolic pathways significantly differed between the plant sections at both 1 and 5 months. All the dominant bacterial genera (e.g., Pseudomonas and Burkholderia-Caballeronia-Paraburkholderia) have been reported to have plant growth-promoting capacities and/or antagonism against pathogens, but what defines their role for plant development has still to be determined. This investigation improves our understanding of the early plant-bacteria interactions essential for natural regeneration of A. religiosa forest.

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The resilience of forests to drought events has become a major natural resource sustainability concern, especially in response to climate change. Yet, little is known about the legacy effects of repeated droughts, and tree species ability to respond across environmental gradients. In this study, we used a tree-ring database (121 sites) to evaluate the overall resilience of tree species to drought events in the last century. We investigated how climate and geography affected the response at the species level. We evaluated temporal trends of resilience using a predictive mixed linear modeling approach. We found that pointer years (e.g., tree growth reduction) occurred during 11.3% of the 20th century, with an average decrease in tree growth of 66% compared to the previous period. The occurrence of pointer years was associated with negative values of the Standardized Precipitation Index (SPI, 81.6%) and Palmer Drought Severity Index (PDSI, 77.3%). Tree species differed in their resilience capacity, however, species inhabiting xeric conditions were less resistant but with higher recovery rates (e.g., Abies concolor, Pinus lambertiana, and Pinus jeffreyi). On average, tree species needed 2.7 years to recover from drought events, with extreme cases requiring more than a decade to reach pre-drought tree growth rates. The main abiotic factor related to resilience was precipitation, confirming that some tree species are better adapted to resist the effects of droughts. We found a temporal variation for all tree resilience indices (scaled to 100), with a decreasing resistance (-0.56 by decade) and resilience (-0.22 by decade), but with a higher recovery (+1.72 by decade) and relative resilience rate (+0.33 by decade). Our results emphasize the importance of time series of forest resilience, particularly by distinguishing the species-level response in the context of legacy of droughts, which are likely to become more frequent and intense under a changing climate.

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Secondary contact of recently diverged species may have several outcomes, ranging from rampant hybridization to reinforced reproductive isolation. In plants, selfing tolerance and disjunct reproductive phenology may lead to reproductive isolation at contact zones. However, they may also evolve under both allopatric or parapatric frameworks and originate from adaptive and/or neutral forces. Inferring the historical demography of diverging taxa is thus a crucial step to identify factors that may have led to putative reproductive isolation. We explored various competing demographypotheses to account for the rapid divergence of a fir species complex (Abies flinckii-A. religiosa) distributed in "sky-islands" across central Mexico (i.e., along the Trans-Mexican Volcanic Belt; TMVB). Despite co-occurring in two independent sympatric regions (west and centre), these taxa rarely interbreed because of disjunct reproductive phenologies. We genotyped 1147 single nucleotide polymorphisms, generated by GBS (genotyping by sequencing), across 23 populations, and compared multiple scenarios based on the geological history of the TMVB. The best-fitting model revealed one of the most rapid and complete speciation cases for a conifer species-pair, dating back to ~1.2 million years ago. Coupled with the lack of support for stepwise colonization, our coalescent inferences point to an early cessation of interspecific gene flow under parapatric speciation; ancestral gene flow during divergence was asymmetrical (mostly from western firs into A. religiosa) and exclusive to the most ancient (i.e., central) contact zone. Factors promoting rapid reproductive isolation should be explored in other slowly evolving species complexes as they may account for the large tropical and subtropical diversity.

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Viscum album extract (VA) is a complementary treatment in cancer, with in vitro and in vivo cytotoxic effects on several tumor types when applied in phytochemical doses. However, highly diluted ethanolic homeopathic preparations' effects and mechanisms need further study. Aims:To assess the in vitro effects of highly diluted VA from the subspecies V. album abietis and V. album album at different potency levels in different dilution ratios on murine melanoma cells. Methodology:The VA mother tinctures (MT)from Abies alba (MTA) and Quercus robur (MTQ) were prepared with summer and winter samples, harvested in Switzerland. They were submitted to homeopathic ethanolic maceration and a subsequent dynamization process. MTA, MTQ and the following respective potencies were tested in B16F10 murine cells: 3x, 12x, 30x, 6cH, 12cH, 200cH, 2LM, 3LM, and 5LM. Dynamized water, dynamized and non-dynamized ethanol, and carboplatin were used as control groups. The mitochondrial activity and cell viability analysis were performed at 1, 24, 48, and 72 hours by in vitro incubation. MTA and MTQ harvested in summer, as well as 12x, 200cH and 5LM potencies were also tested to cell apoptosis and necrosis markers, reactive oxygens species (ROS) production, inflammatory cytokines profile, cell morphology, and migratory capacity. Results and discussion: MTA and MTQ induced a decrease in cell metabolism and higher cytotoxicity within 1 hour, with significant morphological changes and increased production of ROS and inflammatory cytokines. Both homeopathic dilutions 12x and 5LM showed an influence on cell metabolism, cell replication, and oxidative stress modulation with inflammatory cytokines, mitosis, and migration pattern changes. On the other hand, Quercus robur and Abies alba 200cH showed increased on cytotoxicity and ROS levels, respectively. Conclusion:The in vitro effects of Viscum album homeopathic solutions in melanoma cells highlight the promising antitumoral potential and reinforce the need for further research to better understanding their mechanisms of action.

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Constructing phylogenetic relationships among closely related species is a recurrent challenge in evolutionary biology, particularly for long-lived taxa with large effective population sizes and uncomplete reproductive isolation, like conifers. Conifers further have slow evolutionary rates, which raises the question of whether adaptive or non/adaptive processes were predominantly involved when they rapidly diversified after migrating from temperate regions into the tropical mountains. Indeed, fine-scale phylogenetic relationships within several conifer genus remain under debate. Here, we studied the phylogenetic relationships of endemic firs (Abies, Pinaceae) discontinuously distributed in the montane forests from the Southwestern United States to Guatemala, and addressed several hypotheses related to adaptive and non-adaptive radiations. We derived over 80 K SNPs from genotyping by sequencing (GBS) for 45 individuals of nine Mesoamerican species to perform phylogenetic analyses. Both Maximum Likelihood and quartets-inference phylogenies resulted in a well-resolved topology, showing a single fir lineage divided in four subgroups that coincided with the main mountain ranges of Mesoamerica; thus having important taxonomic implications. Such subdivision fitted a North-South isolation by distance framework, in which non-adaptive allopatric processes seemed the rule. Interestingly, several reticulations were observed within subgroups, especially in the central-south region, which may explain past difficulties for generating infrageneric phylogenies. Further evidence for non-adaptive processes was obtained from analyses of 21 candidate-gene regions, which exhibited diminishing values of πa/πs and Ka/Ks with latitude, thus indicating reduced efficiency of purifying selection towards the Equator. Our study indicates that non-adaptive allopatric processes may be key generators of species diversity and endemism in the tropics.

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Background: Pretreatment is the critically important step for the production of ethanol from lignocelluloses. In this study, hardwood birch (Betula pendula) and softwood spruce (Norway spruce) woods were pretreated with a newly synthesized morpholinium ionic liquid, 1-H-3-methylmorpholinium chloride ([HMMorph][Cl]), followed by enzymatic hydrolysis and fermentation to ethanol. Results: [HMMorph][Cl] was synthesized using inexpensive raw materials, i.e., hydrochloric acid and N-methyl morpholine, following a simple process. The influence of pretreatment time (2, 3, 5, and 8 h) and temperature (120 and 140°C) in terms of hydrolysis efficiency was investigated. Glucose yields from enzymatic hydrolysis were improved from 13.7% to 45.7% and 12.9% to 51.8% after pretreatment of birch and spruce woods, respectively, under optimum pretreatment conditions (i.e., at 140°C for 3 h) as compared to those from pristine woods. Moreover, the yields of ethanol production from birch and spruce were increased to 34.8% and 44.2%, respectively, while the yields were negligible for untreated woods. Conclusions: This study demonstrated the ability of [HMMorph][Cl] as an inexpensive agent to pretreat both softwood and hardwood.

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Abies religiosa forests in central Mexico are the only overwinter refuge of the monarch butterfly and provide important ecosystem services. These forests have lost 55% of their original area and as a consequence, diversity and biotic interactions in these ecosystems are in risk. The aim of this study was to compare the soil fungal diversity and community structure in the Abies religiosa forests and surrounding Pinus montezumae, Pinus hartwegii, and coniferous mixed forest plant communities to provide data on ecology of mycorrhizal interactions for the assisted migration of A. religiosa. We sampled soil from five coniferous forests, extracted total soil DNA, and sequenced the ITS2 region by Illumina MiSeq. The soil fungi community was integrated by 1746 taxa with a species turnover ranging from 0.280 to 0.461 between sampling sites. In the whole community, the more abundant and frequent species were Russula sp. (aff. olivobrunnea), Mortierella sp.1, and Piloderma sp. (aff. olivacearum). The ectomycorrhizal fungi were the more frequent and abundant functional group. A total of 298 species (84 ectomycorrhizal) was shared in the five conifer forests; these widely distributed species were dominated by Russulaceae and Clavulinaceae. The fungal community composition was significantly influenced by altitude and the lowest species turnover happened between the two A. religiosa forests even though they have different soil types. As Pinus montezumae forests have a higher altitudinal distribution adjacent to A. religiosa and share the largest number of ectomycorrhizal fungi with it, we suggest these forests as a potential habitat for new A. religiosa populations.

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Plantas nodrizas son aquellas especies vegetales útiles para la sobrevivencia y convivencia de otras especies. Abies guatemalensis Rehder (pinabete), es una especie endémica en peligro de extinción y para asegurar su sobrevivencia es necesario estudiar sus plantas nodrizas. Se colectaron seis especies de plantas nodrizas provenientes de bosque Los Cuervos, Ixchiguán, San Marcos, se secaron a la sombra en un horno, hasta obtener un porcentaje de humedad < 10%. Se obtuvieron extractos etanólicos por percolación y concentración empleando rotavapor; los aceites senciales fueron extraídos por hidrodestilación con Neoclevenger. Para los análisis de actividad biológica se realizaron ensayos contra bacterias, hongos, levaduras, larvas de insectos, nauplios de Artemia salina y actividad antioxidante. Los aceites esenciales presentaron un porcentaje de rendimiento < 0.3%. Cinco de los seis extractos etanólicos demostraron baja actividad antimicrobiana y larvicida (CIM: 1 mg/mL). Respecto a la actividad antioxidante por DPPH, Acaena elongata (CI50 de 0.21 [0.019, 0.23] mg/mL) y Rubus trilobus (CI50 0.32 [0.31, 0.33] mg/mL) presentaron los mejores resultados y Buddleja megalocephala (CI50 0.75 [0.72, 0.77] mg/mL) presentó actividad moderada. Estos hallazgos estimulan a continuar la investigación de plantas nodrizas para identificar las moléculas responsables de la actividad antioxidante y definir su posible aplicación como antioxidantes para la prevención o tratamiento de patologías humanas o para la preservación de alimentos o uso cosmético, permitiendo que las comunidades conserven y aprovechen de manera sostenible dichas especies.

Nurse plants are species useful for the survival and coexistence of other plants. Abies guatemalensis Rehder (fir) is an endemic species in danger of extinction and in order to ensure its survival, it is necessary to study their nurse plants. Six species of nurse plants were collected from Los Cuervos forest, Ixchiguán, San Marcos, they were dried in the shade and in an oven until a moisture content < 10% was obtained. Ethanol extracts were obtained by percolation followed by concentration in rotavapor; essential oils were obtained by hydrodistillation with Neoclevenger. Assays for biological activity were established against bacteria, fungi, yeasts, insect larvae, nauplii of Artemia salina and antioxidant activity. The essential oils yields were < 0.3%. Extracts have little antimicrobial and larvicidal activity with a MIC 1 mg/mL of the ethanol extracts. The free radical trapping activity assay evaluated by DPPH, yielded good results: Acaena elongata (IC50 of 0.21 [0.019, 0.23] mg/mL) and Rubus trilobus (IC50 0.32 [0.31, 0.33] mg/mL) and moderate activity in Buddleja megalocephala (IC50 0.75 [0.72, 0.77] mg/mL). These findings stimulate research on nurse plants to identify the molecules responsible for the antioxidant activity and to define its possible application for the prevention or treatment of human pathologies or for the preservation of food or cosmetic use, allowing the communities to preserve and take advantage on a sustainable way.

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El presente reporte de caso tuvo como objetivo generar el modelo de negocios para el pinabete (Abies guatemalensis Rehder) en restauración, no se pretende hacer un análisis profundo de la producción de árboles de pinabete, únicamente mostrar que la producción se puede combinar con la conservación. En la región alta del altiplano occidental de Guatemala hay muchas tierras degradadas fuera de bosques, donde la sucesión vegetal ha promovido el crecimiento de especies arbustivas, útiles como nodrizas en el establecimiento de pinabete para protegerlas de heladas y lograr establecimiento mayor a 80 %, lo cual ya se ha puesto en práctica por varios silvicultores locales. Se realizó un análisis técnico financiero sobre la producción de pinabete con doble propósito: especie comercial y para la restauración de áreas degradadas. El modelo de negocios se estableció para 0.25 ha, con un precio de venta por árbol de Q. 300, con y sin incentivos forestales, considerando establecer previamente plantas arbustivas. Los resultados indican que con una inversión de Q. 45,227 durante ocho años a una tasa de descuento del 13 %, el Valor Actual Neto (VAN) toma valores entre Q. 398,576 a 742,020 con una Tasa Interna de Rendimiento (TIR) entre 30 y 50 %. A la vez se preparó un plan de negocios utilizando la metodología Canvas, que establece un plan estratégico que permite visualizar lo que actualmente se está realizando y qué situaciones hay que mejorar. El modelo propuesto puede utilizarse de una forma mixta para la conservación al dejar al menos un 30 % de los árboles, la otra parte se puede utilizar para comercialización a partir del octavo al 12do año de forma sostenible.

The present case study aimed to generate the business model for the Guatemalan fir (Abies guatemalensis Rehder) under restoration process. In the highlands of the altiplano region in western Guatemala there are many degraded lands outside of forests, where the ecological succession has promoted the growth of shrub species that help to the establishment of the fir and to protect it from frost. This condition also helped the specie to achieve more than 80 % of survival rate. The method has been implemented by several local silvicultors. A technical and financial analysis was carried out on the production of fir with two objectives: production of commercial Christmas trees and for the restoration of degraded areas. The business model was established for an area of 0.25 ha, with an estimated market price per tree of Q. 300, with and without forestry incentives, assuming previously establish shrub plants. The results indicate that with an investment of Q. 45,227 for eight years at a discount rate of 13 %, the Net Present Value (NPV) takes values between Q. 398,576 to 742,020 with an Internal Rate of Return (IRR) ranging between 30 and 50 %. At the same time, a business plan was prepared using the Canvas methodology, which establishes a strategic plan that allows visualizing what is currently being done and what areas need improvement. The proposed model can be used in a mixed way for conservation by leaving at least 30 % of the trees; the other part can be used for trading the trees from the eighth to the twelfth years old in a sustainable way.

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Abies koreana is an endemic and rare species from Korea and is classified as endangered by the International Union for Conservation of Nature. Although the genetic diversity assessment for current population of A. koreana needs to be performed urgently, no microsatellite markers have been developed for this species. In the present study, we developed 22 novel polymorphic microsatellite loci and the characteristics of these loci were determined in A. koreana as well as in Abies nephrolepis, the most closely related species, and these loci were compared with previously reported microsatellite markers developed for the Abies genus. Genomic sequence (161 Mbp; 325,776 reads) was obtained from one individual of A. koreana using Roche 454 GS-FLX Titanium sequencing and 19,258 repeat motifs were identified from it. A total of 288 primer pairs with high copy numbers of di-repeat motifs were evaluated for amplification in A. koreana and A. nephrolepis. A total of 71 primer pairs successfully amplified fragments, of which 22 showed polymorphisms in A. koreana and A. nephrolepis. The average expected diversity was 0.767 and 0.717 in A. koreana and A. nephrolepis, respectively; these heterozygosity levels were moderate compared to the previously reported microsatellite loci from Abies species. This is the first set of microsatellite markers developed for A. koreana as well as A. nephrolepis and further population genetic studies of both species and genetic delimitation can be carried out for the species conservation and management.

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