Transient co-expression with three <Emphasis Type="Italic">O</Emphasis>-glycosylation enzymes allows production of GalNAc-<Emphasis Type="Italic">O</Emphasis>-glycosylated Granulocyte-Colony Stimulating Factor in <Emphasis Type="Italic">N. benthamiana</Emphasis>

Background

Expression of economically relevant proteins in alternative expression platforms, especially plant expression platforms, has gained significant interest in recent years. A special interest in working with plants as bioreactors for the production of pharmaceutical proteins is related to low production costs, product safety and quality. Among the different properties that plants can also offer for the production of recombinant proteins, protein glycosylation is crucial since it may have an impact on pharmaceutical functionality and/or stability.

Results

The pharmaceutical glycoprotein human Granulocyte-Colony Stimulating Factor was transiently expressed in Nicotiana benthamiana plants and subjected to mammalian-specific mucin-type O-glycosylation by co-expressing the pharmaceutical protein together with the glycosylation machinery responsible for such post-translational modification.

Conclusions

The pharmaceutical glycoprotein human Granulocyte-Colony Stimulating Factor can be expressed in N. benthamiana plants via agroinfiltration with its native mammalian-specific mucin-type O-glycosylation.

     

The Genus <Emphasis Type="Italic">Fragaria </Emphasis>in Croatia

Only three native species of genus Fragaria (F. moschata, F. vesca and F. viridis) are recorded in three regions of Croatia. These species as well as many of their hybrids, are, or once were, cultivated for their edible fruits. The majority of cultivated strawberries in Europe belong to garden strawberries F. x ananassa (hybrids of F. chiloensis and F. virginiana). The most expanded wild strawberry species in Croatia is a woodland strawberry (F. vesca) whose berries are gathered seasonally as wild edible fruits. They often contain higher amount of nutrients and bioactive compounds in comparison to cultivated varieties. The research on the genus Fragaria species distribution in Croatia has not been carried out, and so is the case with many others wild growing fruit species in Croatia. By summing up a number of individual citations and observations, it is possible to get a perspective regarding the current state of their distribution.     

Chlorophyll <Emphasis Type="Italic">a</Emphasis> fluorescence,under half of the adaptive growth-irradiance,for high-throughput sensing of leaf-water deficit in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> accessions

Background

Non-invasive and high-throughput monitoring of drought in plants from its initiation to visible symptoms is essential to quest drought tolerant varieties. Among the existing methods, chlorophyll a fluorescence (ChlF) imaging has the potential to probe systematic changes in photosynthetic reactions; however, prerequisite of dark-adaptation limits its use for high-throughput screening.

Results

To improve the throughput monitoring of plants, we have exploited their light-adaptive strategy, and investigated possibilities of measuring ChlF transients under low ambient irradiance. We found that the ChlF transients and associated parameters of two contrasting Arabidopsis thaliana accessions, Rsch and Co, give almost similar information, when measured either after ~20 min dark-adaptation or in the presence of half of the adaptive growth-irradiance. The fluorescence parameters, effective quantum yield of PSII photochemistry (Φ_PSII) and fluorescence decrease ratio (R _FD) resulting from this approach enabled us to differentiate accessions that is often not possible by well-established dark-adapted fluorescence parameter maximum quantum efficiency of PSII photochemistry (F _V/F _M). Further, we screened ChlF transients in rosettes of well-watered and drought-stressed six A. thaliana accessions, under half of the adaptive growth-irradiance, without any prior dark-adaptation. Relative water content (RWC) in leaves was also assayed and compared to the ChlF parameters. As expected, the RWC was significantly different in drought-stressed from that in well-watered plants in all the six investigated accessions on day-10 of induced drought; the maximum reduction in the RWC was obtained for Rsch (16%), whereas the minimum reduction was for Co (~7%). Drought induced changes were reflected in several features of ChlF transients; combinatorial images obtained from pattern recognition algorithms, trained on pixels of image sequence, improved the contrast among drought-stressed accessions, and the derived images were well-correlated with their RWC.

Conclusions

We demonstrate here that ChlF transients and associated parameters measured even in the presence of low ambient irradiance preserved its features comparable to that of measured after dark-adaptation and discriminated the accessions having differential geographical origin; further, in combination with combinatorial image analysis tools, these data may be readily employed for early sensing and mapping effects of drought on plant’s physiology via easy and fully non-invasive means.

     

<Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of safflower and the efficient recovery of transgenic plants via grafting

Background

Safflower (Carthamus tinctorius L.) is a difficult crop to genetically transform being susceptible to hyperhydration and poor in vitro root formation. In addition to traditional uses safflower has recently emerged as a broadacre platform for the production of transgenic products including modified oils and pharmaceutically active proteins. Despite commercial activities based on the genetic modification of safflower, there is no method available in the public domain describing the transformation of safflower that generates transformed T₁ progeny.

Results

An efficient and reproducible protocol has been developed with a transformation efficiency of 4.8% and 3.1% for S-317 (high oleic acid content) and WT (high linoleic acid content) genotypes respectively. An improved safflower transformation T-DNA vector was developed, including a secreted GFP to allow non-destructive assessment of transgenic shoots. Hyperhydration and necrosis of Agrobacterium-infected cotyledons was effectively controlled by using iota-carrageenan, L-cysteine and ascorbic acid. To overcome poor in vitro root formation for the first time a grafting method was developed for safflower in which ~50% of transgenic shoots develop into mature plants bearing viable transgenic T₁ seed. The integration and expression of secreted GFP and hygromycin genes were confirmed by PCR, Southern and Western blot analysis. Southern blot analysis in nine independent lines indicated that 1-7 transgenes were inserted per line and T₁ progeny displayed Mendelian inheritance.

Conclusions

This protocol demonstrates significant improvements in both the efficiency and ease of use over existing safflower transformation protocols. This is the first complete method of genetic transformation of safflower that generates stably-transformed plants and progeny, allowing this crop to benefit from modern molecular applications.

     

High-level diterpene production by transient expression in <Emphasis Type="Italic">Nicotiana benthamiana</Emphasis>

Background

Characterization of plant terpene synthases is typically done by production of recombinant enzymes in Escherichia coli. This is often difficult due to solubility and codon usage issues. Furthermore, plant terpene synthases which are targeted to the plastids, such as diterpene synthases, have to be shortened in a more or less empirical approach to improve expression. We report here an optimized Agrobacterium-mediated transient expression assay in Nicotiana benthamiana for plant diterpene synthase expression and product analysis.

Results

Agrobacterium-mediated transient expression of plant diterpene synthases in N. benthamiana led to the accumulation of diterpenes within 3 days of infiltration and with a maximum at 5 days. Over 50% of the products were exported onto the leaf surface, thus considerably facilitating the analysis by reducing the complexity of the extracts. The robustness of the method was tested by expressing three different plant enzymes, cembratrien-ol synthase from Nicotiana sylvestris, casbene synthase from Ricinus communis and levopimaradiene synthase from Gingko biloba. Furthermore, co-expression of a 1-deoxy-D-xylulose-5-phosphate synthase from tomato and a geranylgeranyl diphosphate synthase from tobacco led to a 3.5-fold increase in the amount of cembratrien-ol produced, with maximum yields reaching 2500 ng/cm².

Conclusion

With this optimized method for diterpene synthase expression and product analysis, a single infiltrated leaf of N. benthamiana would be sufficient to produce quantities required for the structure elucidation of unknown diterpenes. The method will also be of general use for gene function discovery, pathway reconstitution and metabolic engineering of diterpenoid biosynthesis in plants.

     

Evaluation of parameters affecting switchgrass tissue culture: toward a consolidated procedure for <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of switchgrass (<Emphasis Type="Italic">Panicum virgatum</Emphasis>)

Background

Switchgrass (Panicum virgatum), a robust perennial C4-type grass, has been evaluated and designated as a model bioenergy crop by the U.S. DOE and USDA. Conventional breeding of switchgrass biomass is difficult because it displays self-incompatible hindrance. Therefore, direct genetic modifications of switchgrass have been considered the more effective approach to tailor switchgrass with traits of interest. Successful transformations have demonstrated increased biomass yields, reduction in the recalcitrance of cell walls and enhanced saccharification efficiency. Several tissue culture protocols have been previously described to produce transgenic switchgrass lines using different nutrient-based media, co-cultivation approaches, and antibiotic strengths for selection.

Results

After evaluating the published protocols, we consolidated these approaches and optimized the process to develop a more efficient protocol for producing transgenic switchgrass. First, seed sterilization was optimized, which led to a 20% increase in yield of induced calluses. Second, we have selected a N₆ macronutrient/B₅ micronutrient (NB)-based medium for callus induction from mature seeds of the Alamo cultivar, and chose a Murashige and Skoog-based medium to regenerate both Type I and Type II calluses. Third, Agrobacterium-mediated transformation was adopted that resulted in 50–100% positive regenerated transformants after three rounds (2 weeks/round) of selection with antibiotic. Genomic DNA PCR, RT-PCR, Southern blot, visualization of the red fluorescent protein and histochemical β-glucuronidase (GUS) staining were conducted to confirm the positive switchgrass transformants. The optimized methods developed here provide an improved strategy to promote the production and selection of callus and generation of transgenic switchgrass lines.

Conclusion

The process for switchgrass transformation has been evaluated and consolidated to devise an improved approach for transgenic switchgrass production. With the optimization of seed sterilization, callus induction, and regeneration steps, a reliable and effective protocol is established to facilitate switchgrass engineering.

     

<Emphasis Type="Italic">Dactylonectria torresensis</Emphasis> as the Main Component of the Black Root Rot Complex of Strawberries and Raspberries in Northern Germany

In a long-term survey of black root rot of strawberries and raspberries in Northern Germany in 2007–2014, fungi with and without Cylindrocarpon-like anamorphs were isolated as potential pathogens. Dactylonectria torresensis was the most common species, being isolated from 18% of strawberry roots obtained from nursery plants and 37% of roots from production fields, as well as 21% and 29% (respectively) of raspberry roots. Less frequently isolated fungi with Cylindrocarpon-like anamorphs included Ilyonectria crassa, Ilyonectria sp. 2, I. pseudodestructans, I. robusta, C. obtusisporium and Ilyonectria sp. 1. Severe disease symptoms were reproduced by artificial inoculation of strawberries with D. torresensis, I. crassa and Ilyonectria sp. 2, milder symptoms with C. obtusisporium. A wide range of other root-pathogenic fungi such as Fusarium oxysporum, Verticillium dahliae, Ceratobasidium fragariae, Gnomoniopsis fructicola, Hainesia lythri, and species of Cadophora, Leptodontidium, Pythium, Phytophthora, Plectosporella, Pestalotiopsis and Truncatella were either isolated only sporadically or were not associated with black root rot symptoms, suggesting that they did not play any major role in this disease in Northern Germany. Visible disease symptoms and high frequencies of D. torresensis isolations in many batches of nursery plants indicated that these may comprise a major source of contamination of production fields. The previously unrecognised prominence of D. torresensis resolves a long-standing puzzle concerning the cause of the ongoing black root rot epidemic in Northern German strawberry and raspberry production.     

Analysis of Genetic Relationships Between Wild Roses (<Emphasis Type="Italic">Rosa</Emphasis> L. Spp.) Growing in Turkey

Rosa L. taxa are dominantly used for horticulture, food, medicinal, ornamental and aesthetic purposes. Rosa is represented with approximately 200 wild species in the world. The number of natural Rosa species in Turkey is reported as about 31. Rosa species are widely distributed in all regions in Turkey. In this study, RAPD and ISSR markers were applied to assess genetic diversity and genetic relationships among 27 species of Rosa. The UPGMA cluster was constructed using a combination of data from RAPD and ISSR markers. The dendrogram revealed two main clusters. Each cluster was divided into subgroups. This investigation showed that genetic distance was relatively significant among the species. As the results of the study close genetic relationships were found between evolutionary relations, morphplogical properties, and phytogeographical distributions of the Rosa species, The results also propose that RAPD and ISSR markers are useful tools for indicating genetic relationships among Rosa genotypes.     

Temporal variability in potential connectivity of <Emphasis Type="Italic">Vallisneria americana</Emphasis> in the Chesapeake Bay

Context

Submersed aquatic vegetation (SAV) performs water quality enhancing functions that are critical to the overall health of estuaries such as the Chesapeake Bay. However, eutrophication and sedimentation have decimated the Bay’s SAV population to a fraction of its historical coverage. Understanding the spatial distribution of and connectedness among patches is important for assessing the dynamics and health of the remaining SAV population.

Objectives

We seek to explore the distribution of SAV patches and patterns of potential connectivity in the Chesapeake Bay through time.

Methods

We assess critical distances, from complete patch isolation to connection of all patches, in a merged composite coverage map that represents the sum of all probable Vallisneria americana containing patches between 1984 and 2010 and in coverage maps for individual years within that timeframe for which complete survey data are available.

Results

We have three key findings: First, the amount of SAV coverage in any given year is much smaller than the total recently occupied acreage. Second, the vast majority of patches of SAV that are within the tolerances of V. americana are ephemeral, being observed in only 1 or 2 years out of 26 years. Third, this high patch turnover results in highly variable connectivity from year to year, dependent on dispersal distance and patch arrangement.

Conclusions

Most of the connectivity thresholds are beyond reasonable dispersal distances for V. americana. If the high turnover in habitat occupancy is due to marginal water quality, relatively small improvements could greatly increase V. americana growth and persistence.

     

How landscape,pollen intake and pollen quality affect colony growth in <Emphasis Type="Italic">Bombus terrestris</Emphasis>

Context

Abundance and diversity of bumblebees have been declining over the past decades. To successfully conserve bumblebee populations, we need to understand how landscape characteristics affect the quantity and quality of floral resources collected by colonies and subsequently colony performance.

Objectives

We therefore investigated how amount and composition of pollen collected by buff-tailed bumblebee Bombus terrestris colonies was affected by the surrounding landscape (i.e. the proportion of forest, urban, semi-natural habitats) and how they were related to colony growth.

Methods

Thirty B. terrestris colonies were placed at grassland sites differing in surrounding landscape. Colonies were established in spring when availability of flowering plants was highest, and their weight gain was monitored for 1 month. We additionally recorded the quantity and compared plant taxonomic composition and nutritional quality (i.e. amino acid composition) of pollen stored.

Results

Bumblebee colonies varied little in the pollen spectra collected despite differences in surrounding landscape composition. They collected on average 80 % of pollen from woody plants, with 34 % belonging to the genus Acer. Early colony growth positively correlated with total amount of woody pollen and protein collected and decreased with increasing proportions of semi-natural habitats and total amino acid concentrations.

Conclusions

Our results suggest that woody plant species represent highly important pollen sources for the generalist forager B. terrestris early in the season. We further show that colony growth of B. terrestris is predominantly affected by the quantity, not quality, of forage, indicating that several abundant plant species flowering throughout the bumblebees’ foraging season may cover the colonies’ nutritional needs.

     

Editing of the urease gene by CRISPR-Cas in the diatom <Emphasis Type="Italic">Thalassiosira pseudonana</Emphasis>

Background

CRISPR-Cas is a recent and powerful addition to the molecular toolbox which allows programmable genome editing. It has been used to modify genes in a wide variety of organisms, but only two alga to date. Here we present a methodology to edit the genome of Thalassiosira pseudonana, a model centric diatom with both ecological significance and high biotechnological potential, using CRISPR-Cas.

Results

A single construct was assembled using Golden Gate cloning. Two sgRNAs were used to introduce a precise 37 nt deletion early in the coding region of the urease gene. A high percentage of bi-allelic mutations (≤61.5%) were observed in clones with the CRISPR-Cas construct. Growth of bi-allelic mutants in urea led to a significant reduction in growth rate and cell size compared to growth in nitrate.

Conclusions

CRISPR-Cas can precisely and efficiently edit the genome of T. pseudonana. The use of Golden Gate cloning to assemble CRISPR-Cas constructs gives additional flexibility to the CRISPR-Cas method and facilitates modifications to target alternative genes or species.

     

Landscape genetics of the foundational salt marsh plant species black needlerush (<Emphasis Type="Italic">Juncus roemerianus</Emphasis> Scheele) across the northeastern Gulf of Mexico

Context

Common species important for ecosystem restoration stand to lose as much genetic diversity from anthropogenic habitat fragmentation and climate change as rare species, but are rarely studied. Salt marshes, valuable ecosystems in widespread decline due to human development, are dominated by the foundational plant species black needlerush (Juncus roemerianus Scheele) in the northeastern Gulf of Mexico.

Objectives

We assessed genetic patterns in J. roemerianus by measuring genetic and genotypic diversity, and characterizing population structure. We examined population connectivity by delineating possible dispersal corridors, and identified landscape factors influencing population connectivity.

Methods

A panel of 19 microsatellite markers was used to genotype 576 samples from ten sites across the northeastern Gulf of Mexico from the Grand Bay National Estuarine Research Reserve (NERR) to the Apalachicola NERR. Genetic distances (F_ST and D_ch) were used in a least cost transect analysis (LCTA) within a hierarchical model selection framework.

Results

Genetic and genotypic diversity results were higher than expected based on life history literature, and samples structured into two large, admixed genetic clusters across the study area, indicating sexual reproduction may not be as rare as predicted in this clonal macrophyte. Digitized coastal transects buffered by 500 m may represent possible dispersal corridors, and developed land may significantly impede population connectivity in J. roemerianus.

Conclusions

Results have important implications for coastal restoration and management that seek to preserve adaptive potential by sustaining natural levels of genetic diversity and conserving population connectivity. Our methodology could be applied to other common, widespread and understudied species.

     

A simple and efficient CRISPR/Cas9 platform for induction of single and multiple,heritable mutations in barley (<Emphasis Type="Italic">Hordeum vulgare</Emphasis> L.)

Background

Genome editing of monocot plants can be accomplished by using the components of the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeat/CRISPR associated Cas9) technology specifically optimized for these types of plants. Here, we present the development of RNA-guided Cas9 system for simplex and multiplex genome editing in barley.

Results

We developed a set of customizable RNA-guided Cas9 binary vectors and sgRNA modules for simplex and multiplex editing in barley. To facilitate the design of RNA-guided Cas9 constructs, the pBract derived binary vectors were adapted to Gateway cloning and only one restriction enzyme was required for construction of the sgRNA. We designed a synthetic, codon optimized Cas9 gene containing the N terminal SV40 nuclear localization signal and the UBQ10 Arabidopsis 1st intron. Two different sgRNAs were constructed for simplex editing and one polycistronic tRNA-gRNA construct (PTG) for multiplex editing using an endogenous tRNA processing system. The RNA-guided Cas9 constructs were validated in transgenic barley plants produced by Agrobacterium-mediated transformation. The highest mutation rate was observed in simplex editing of the cytokinin oxidase/dehydrogenase HvCKX1 gene, where mutations at the hvckx1 locus were detected in 88% of the screened T₀ plants. We also proved the efficacy of the PTG construct in the multiplex editing of two CKX genes by obtaining 9 plants (21% of all edited plants) with mutations induced in both HvCKX1 and HvCKX3. Analysis of the T₁ lines revealed that mutations in the HvCKX1 gene were transmitted to the next generation of plants. Among 220 screened T₁ plants we identified 85 heterozygous and 28 homozygous mutants, most of them bearing frameshift mutations in the HvCKX1 gene. We also observed independent segregation of mutations and the Cas9-sgRNA T-DNA insert in several T₁ plants. Moreover, the knockout mutations of the Nud gene generated phenotype mutants with naked grains, and the phenotypic changes were identifiable in T₀ plants.

Conclusions

We demonstrated the effectiveness of an optimized RNA-guided Cas9 system that can be used for generating homozygous knockout mutants in the progeny of transgenic barely plants. This is also the first report of successful multiplex editing in barley using a tRNA processing system.

     

Shaken but not stirred: multiscale habitat suitability modeling of sympatric marten species (<Emphasis Type="Italic">Martes martes</Emphasis> and <Emphasis Type="Italic">Martes foina</Emphasis>) in the northern Iberian Peninsula

Context

Multispecies and multiscale habitat suitability models (HSM) are important to identify the environmental variables and scales influencing habitat selection and facilitate the comparison of closely related species with different ecological requirements.

Objectives

This study explores the multiscale relationships of habitat suitability for the pine (Martes martes) and stone marten (M. foina) in northern Spain to evaluate differences in habitat selection and scaling, and to determine if there is habitat niche displacement when both species coexist.

Methods

We combined bivariate scaling and maximum entropy modeling to compare the multiscale habitat selection of the two martens. To optimize the HSM, the performance of three sampling bias correction methods at four spatial scales was explored. HSMs were compared to explore niche differentiation between species through a niche identity test.

Results

The comparison among HSMs resulted in the detection of a significant niche divergence between species. The pine marten was positively associated with cooler mountainous areas, low levels of human disturbance, high proportion of natural forests and well-connected forestry plantations, and medium-extent agroforestry mosaics. The stone marten was positively related to the density of urban areas, the proportion and extensiveness of croplands, the existence of some scrub cover and semi-continuous grasslands.

Conclusions

This study outlines the influence of the spatial scale and the importance of the sampling bias corrections in HSM, and to our knowledge, it is the first comparing multiscale habitat selection and niche divergence of two related marten species. This study provides a useful methodological framework for multispecies and multiscale comparatives.

     

Protocol: a medium-throughput method for determination of cellulose content from single stem pieces of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Background

Lignocellulosic biomass is an important renewable resource for biofuels and materials. How plants synthesise cellulose is not completely understood. It is known that cellulose synthase complex (CSCs) moving in the plasma membrane synthesise the cellulose. CESA proteins are the core components of CSC. In Arabidopsis, in vitro mutagenesis of proteins followed by complementation analysis of mutants lacking the gene represents an important tool for studying any biological process, including cellulose biosynthesis. Analysis of a large number of plants is crucial for these types of studies.

Results

By using aspiration rather than centrifugation to remove liquids during various stages of protocol, we were able to increase the throughput of the method as well as minimise the sample loss. As a test case, we determined cellulose content of wild type and secondary wall cesa mutants across the length of primary shoot which was fond to be rather uniform in 7-week-old plants. Additionally, we found that the cellulose content of single mutants was comparable to the higher order mutants.

Conclusions

Here we describe a medium-throughput adaptation of Updegraff’s method that allowed us to determine cellulose content of 200 samples each week.

     

Robust genetic transformation of sorghum (<Emphasis Type="Italic">Sorghum bicolor</Emphasis> L.) using differentiating embryogenic callus induced from immature embryos

Background

Sorghum (Sorghum bicolor L.) is one of the world’s most important cereal crops grown for multiple applications and has been identified as a potential biofuel crop. Despite several decades of study, sorghum has been widely considered as a recalcitrant major crop for transformation due to accumulation of phenolic compounds, lack of model genotypes, low regeneration frequency and loss of regeneration potential through sub-cultures. Among different explants used for genetic transformation of sorghum, immature embryos are ideal over other explants. However, the continuous supply of quality immature embryos for transformation is labour intensive and expensive. In addition, transformation efficiencies are also influenced by environmental conditions (light and temperature). Despite these challenges, immature embryos remain the predominant choice because of their success rate and also due to non-availability of other dependable explants without compromising the transformation efficiency.

Results

We report here a robust genetic transformation method for sorghum (Tx430) using differentiating embryogenic calli (DEC) with nodular structures induced from immature embryos and maintained for more than a year without losing regeneration potential on modified MS media. The addition of lipoic acid (LA) to callus induction media along with optimized growth regulators increased callus induction frequency from 61.3 ± 3.2 to 79 ± 6.5% from immature embryos (1.5–2.0 mm in length) isolated 12–15 days after pollination. Similarly, the regeneration efficiency and the number of shoots from DEC tissue was enhanced by LA. The optimized regeneration system in combination with particle bombardment resulted in an average transformation efficiency (TE) of 27.2 or 46.6% based on the selection strategy, 25% to twofold higher TE than published reports in Tx430. Up to 100% putative transgenic shoots were positive for npt-II by PCR and 48% of events had < 3 copies of transgenes as determined by digital droplet PCR. Reproducibility of this method was demonstrated by generating ~ 800 transgenic plants using 10 different gene constructs.

Conclusions

This protocol demonstrates significant improvements in both efficiency and ease of use over existing sorghum transformation methods using PDS, also enables quick hypothesis testing in the production of various high value products in sorghum.

     

Effect of connectivity and habitat availability on the occurrence of the Chestnut-throated Huet-Huet (<Emphasis Type="Italic">Pteroptochos castaneus,</Emphasis> Rhinocryptidae) in fragmented landscapes of central Chile

Context

Although small isolated habitat patches may not be able to maintain a minimum viable population, small patches that are structurally isolated may be functionally connected if individuals can cross the gaps between them, in which case, their areas could be added to form a larger habitat patch, eventually surpassing the size threshold for holding a viable population.

Objectives

We studied whether models based on the size and isolation of habitat patches could be used to predict the distribution of the Chestnut-throated Huet-Huet (Pteroptochos castaneus) in fragmented landscapes of the coastal range of the Maule region, central Chile.

Methods

We selected seven 10,000-ha landscapes (8.4–70.7% forest cover). For each habitat patch we made 18 predictions of the presence of the species based on the combination of two thresholds: three critical patch sizes for maintaining a viable population (62.5, 125 and 250 ha) and six critical isolation distances between patches (0, 10, 50, 100, 150 and 200 m). We used playbacks in 59 sampling points to estimate the species’ presence/absence. We used logistic regressions to test whether the output of the patch-matrix models could explain part of the variation in the presence of Pteroptochos castaneus.

Results

The best predictions for the presence of P. castaneus were obtained with the most conservative scenarios (125–250 ha to 0–10 m), including a positive effect of the understory cover and a lack of effect of the forest type (native or exotic).

Conclusions

Our findings suggest that the long term persistence of P. castaneus may depend on the existence of large and/or very connected forest tracts.

     

Contiguity of landscape features pose barriers to gene flow among American marten (<Emphasis Type="Italic">Martes americana</Emphasis>) genetic clusters in the Upper Peninsula of Michigan

Context

Barriers to dispersal influence the ability of individuals to expand into new areas and can ultimately define success of reintroduction programs. American marten (Martes americana) were reintroduced to the Upper Peninsula of Michigan, USA, from multiple, genetically differentiated source populations from 1955 to 1992. Previous research found multiple genetic clusters near release sites with little admixing, suggesting barriers to dispersal exist.

Objectives

We sought to identify whether the contact zones between genetic clusters coincided with landscape features hypothesized to influence M. americana dispersal. We also investigated whether the degree of landscape contiguity within each genetic cluster differed among clusters.

Methods

We mapped cluster boundaries in M. americana genetic assignment probabilities and used correlation length as a measure of landscape contiguity between genetic clusters. We then evaluated the effects of land cover and roads on spatial genetic structure using a spatial autoregressive model.

Results

We found that gene flow was facilitated by contiguous coniferous forest and low incidence of roads. However, the strength of those relationships varied by genetic cluster. Contact zones among some genetic clusters spatially coincided with areas of high road and low conifer contiguity, compared to within-clusters.

Conclusions

In contrast to landscape genetic analyses focused on identifying barriers to gene flow, we incorporated methods that are relatively novel in landscape genetics to quantify how landscape contiguity influences spatial genetic structure. Using this method we were able to identify landscape barriers to dispersal at the genetic cluster boundaries for a reintroduced species distributed continuously across the landscape.