Stable isotope analysis of vegetation history and land use change at Laguna Santa Elena in southern Pacific Costa Rica

Vegetation History and Archaeobotany - Laguna Santa Elena (8.9290° N, 82.9257° W, 1055 m a.s.l.) is a small lake in the Diquís archaeological sub-region of southern Pacific...     

Fatty acid activation and utilization by Alistipes finegoldii,a representative Bacteroidetes resident of the human gut microbiome

Members of the Bacteroidetes phylum, represented by Alistipes finegoldii, are prominent anerobic, Gram-negative inhabitants of the gut microbiome. The lipid biosynthetic pathways were analyzed using bioinformatic analyses, lipidomics, metabolic labeling and biochemistry to characterize exogenous fatty acid metabolism. A. finegoldii only produced the saturated fatty acids. The most abundant lipids were phosphatidylethanolamine (PE) and sulfonolipid (SL). Neither phosphatidylglycerol nor cardiolipin are present. PE synthesis is initiated by the PlsX/PlsY/PlsC pathway, whereas the SL pathway is related to sphingolipid biosynthesis. A. finegoldii incorporated medium-chain fatty acids (≤14 carbons) into PE and SL after their elongation, whereas long-chain fatty acids (≥16 carbons) were not elongated. Fatty acids >16 carbons were primarily incorporated into the 2-position of phosphatidylethanolamine at the PlsC step, the only biosynthetic enzyme that utilizes long-chain acyl-ACP. The ability to assimilate a broad-spectrum of fatty acid chain lengths present in the gut environment is due to the expression of two acyl-acyl carrier protein (ACP) synthetases. Acyl-ACP synthetase 1 had a substrate preference for medium-chain fatty acids and synthetase 2 had a substrate preference for long-chain fatty acids. This unique combination of synthetases allows A. finegoldii to utilize both the medium- and long-chain fatty acid nutrients available in the gut environment to assemble its membrane lipids.     

Phosphatidic acid produced by phospholipase D is required for hyphal cell-cell fusion and fungal-plant symbiosis

Although lipid signaling has been shown to serve crucial roles in mammals and plants, little is known about this process in filamentous fungi. Here we analyze the contribution of phospholipase D (PLD) and its product phosphatidic acid (PA) in hyphal morphogenesis and growth of Epichloë festucae and Neurospora crassa, and in the establishment of a symbiotic interaction between E. festucae and Lolium perenne. Growth of E. festucae and N. crassa PLD deletion strains in axenic culture, and for E. festucae in association with L. perenne, were analyzed by light-, confocal- and electron microscopy. Changes in PA distribution were analyzed in E. festucae using a PA biosensor and the impact of these changes on the endocytic recycling and superoxide production investigated. We found that E. festucae PldB, and the N. crassa ortholog, PLA-7, are required for polarized growth and cell fusion and contribute to ascospore development, whereas PldA/PLA-8 are dispensable for these functions. Exogenous addition of PA rescues the cell-fusion phenotype in E. festucae. PldB is also crucial for E. festucae to establish a symbiotic association with L. perenne. This study identifies a new component of the cell-cell communication and cell fusion signaling network for hyphal morphogenesis and growth of filamentous fungi.     

PEATmoss (Physcomitrella Expression Atlas Tool): a unified gene expression atlas for the model plant Physcomitrella patens

Physcomitrella patens is a bryophyte model plant that is often used to study plant evolution and development. Its resources are of great importance for comparative genomics and evo‐devo approaches. However, expression data from Physcomitrella patens were so far generated using different gene annotation versions and three different platforms: CombiMatrix and NimbleGen expression microarrays and RNA sequencing. The currently available P. patens expression data are distributed across three tools with different visualization methods to access the data. Here, we introduce an interactive expression atlas, Physcomitrella Expression Atlas Tool (PEATmoss), that unifies publicly available expression data for P. patens and provides multiple visualization methods to query the data in a single web‐based tool. Moreover, PEATmoss includes 35 expression experiments not previously available in any other expression atlas. To facilitate gene expression queries across different gene annotation versions, and to access P. patens annotations and related resources, a lookup database and web tool linked to PEATmoss was implemented. PEATmoss can be accessed at https://peatmoss.online.uni-marburg.de     

Role of a noncanonical disulfide bond in the stability,affinity, and flexibility of a VHH specific for the Listeria virulence factor InlB

A distinguishing feature of camel (Camelus dromedarius) VHH domains are noncanonical disulfide bonds between CDR1 and CDR3. The disulfide bond may provide an evolutionary advantage, as one of the cysteines in the bond is germline encoded. It has been hypothesized that this additional disulfide bond may play a role in binding affinity by reducing the entropic penalty associated with immobilization of a long CDR3 loop upon antigen binding. To examine the role of a noncanonical disulfide bond on antigen binding and the biophysical properties of a VHH domain, we have used the VHH R303, which binds the Listeria virulence factor InlB as a model. Using site directed mutagenesis, we produced a double mutant of R303 (C33A/C102A) to remove the extra disulfide bond of the VHH R303. Antigen binding was not affected by loss of the disulfide bond, however the mutant VHH displayed reduced thermal stability (T_m = 12°C lower than wild‐type), and a loss of the ability to fold reversibly due to heat induced aggregation. X‐ray structures of the mutant alone and in complex with InlB showed no major changes in the structure. B‐factor analysis of the structures suggested that the loss of the disulfide bond elicited no major change on the flexibility of the CDR loops, and revealed no evidence of loop immobilization upon antigen binding. These results suggest that the noncanonical disulfide bond found in camel VHH may have evolved to stabilize the biophysical properties of the domain, rather than playing a significant role in antigen binding.     

Flexibility of nutritional strategies within a mutualism: food availability affects algal symbiont productivity in two congeneric sea anemone species

Mutualistic symbioses are common, especially in nutrient-poor environments where an association between hosts and symbionts can allow the symbiotic partners to persist and collectively out-compete non-symbiotic species. Usually these mutualisms are built on an intimate transfer of energy and nutrients (e.g. carbon and nitrogen) between host and symbiont. However, resource availability is not consistent, and the benefit of the symbiotic association can depend on the availability of resources to mutualists. We manipulated the diets of two temperate sea anemone species in the genus Anthopleura in the field and recorded the responses of sea anemones and algal symbionts in the family Symbiodiniaceae to our treatments. Algal symbiont density, symbiont volume and photosynthetic efficiency of symbionts responded to changes in sea anemone diet, but the responses depended on the species of sea anemone. We suggest that temperate sea anemones and their symbionts can respond to changes in anemone diet, modifying the balance between heterotrophy and autotrophy in the symbiosis. Our data support the hypothesis that symbionts are upregulated or downregulated based on food availability, allowing for a flexible nutritional strategy based on external resources.     

The molecular basis of venom resistance in a rattlesnake‐squirrel predator‐prey system

Understanding how interspecific interactions mould the molecular basis of adaptations in coevolving species is a long‐sought goal of evolutionary biology. Venom in predators and venom resistance proteins in prey are coevolving molecular phenotypes, and while venoms are highly complex mixtures it is unclear if prey respond with equally complex resistance traits. Here, we use a novel molecular methodology based on protein affinity columns to capture and identify candidate blood serum resistance proteins (“venom interactive proteins” [VIPs]) in California Ground Squirrels (Otospermophilus beecheyi) that interact with venom proteins from their main predator, Northern Pacific Rattlesnakes (Crotalus o. oreganus). This assay showed that serum‐based resistance is both population‐ and species‐specific, with serum proteins from ground squirrels showing higher binding affinities for venom proteins of local snakes compared to allopatric individuals. Venom protein specificity assays identified numerous and diverse candidate prey resistance VIPs but also potential targets of venom in prey tissues. Many specific VIPs bind to multiple snake venom proteins and, conversely, single venom proteins bind multiple VIPs, demonstrating that a portion of the squirrel blood serum “resistome” involves broad‐based inhibition of nonself proteins and suggests that resistance involves a toxin scavenging mechanism. Analyses of rates of evolution of VIP protein homologues in related mammals show that most of these proteins evolve under purifying selection possibly due to molecular constraints that limit the evolutionary responses of prey to rapidly evolving snake venom proteins. Our method represents a general approach to identify specific proteins involved in co‐evolutionary interactions between species at the molecular level.     

Regional differences in the abiotic environment contribute to genomic divergence within a wild tomato species

The wild currant tomato Solanum pimpinellifolium inhabits a wide range of abiotic habitats across its native range of Ecuador and Peru. Although it has served as a key genetic resource for the improvement of domestic cultivars, little is known about the genetic basis of traits underlying local adaptation in this species, nor what abiotic variables are most important for driving differentiation. Here we use redundancy analysis (RDA) and other multivariate statistical methods (structural equation modelling [SEM] and generalized dissimilarity modelling [GDM]) to quantify the relationship of genomic variation (6,830 single nucleotide polymorphisms [SNPs]) with climate and geography, among 140 wild accessions. RDA, SEM and GDM each identified environment as explaining more genomic variation than geography, suggesting that local adaptation to heterogeneous abiotic habitats may be an important source of genetic diversity in this species. Environmental factors describing temporal variation in precipitation and evaporative demand explained the most SNP variation among accessions, indicating that these forces may represent key selective agents. Lastly, by studying how SNP–environment associations vary throughout the genome (44,064 SNPs), we mapped the location and investigated the functions of loci putatively contributing to climatic adaptations. Together, our findings indicate an important role for selection imposed by the abiotic environment in driving genomic differentiation between populations.     

A total crapshoot? Evaluating bioinformatic decisions in animal diet metabarcoding analyses

Metabarcoding studies provide a powerful approach to estimate the diversity and abundance of organisms in mixed communities in nature. While strategies exist for optimizing sample and sequence library preparation, best practices for bioinformatic processing of amplicon sequence data are lacking in animal diet studies. Here we evaluate how decisions made in core bioinformatic processes, including sequence filtering, database design, and classification, can influence animal metabarcoding results. We show that denoising methods have lower error rates compared to traditional clustering methods, although these differences are largely mitigated by removing low‐abundance sequence variants. We also found that available reference datasets from GenBank and BOLD for the animal marker gene cytochrome oxidase I (COI) can be complementary, and we discuss methods to improve existing databases to include versioned releases. Taxonomic classification methods can dramatically affect results. For example, the commonly used Barcode of Life Database (BOLD) Classification API assigned fewer names to samples from order through species levels using both a mock community and bat guano samples compared to all other classifiers (vsearch‐SINTAX and q2‐feature‐classifier's BLAST + LCA, VSEARCH + LCA, and Naive Bayes classifiers). The lack of consensus on bioinformatics best practices limits comparisons among studies and may introduce biases. Our work suggests that biological mock communities offer a useful standard to evaluate the myriad computational decisions impacting animal metabarcoding accuracy. Further, these comparisons highlight the need for continual evaluations as new tools are adopted to ensure that the inferences drawn reflect meaningful biology instead of digital artifacts.     

Adaptation to monogamy influences parental care but not mating behavior in the burying beetle,Nicrophorus vespilloides

The mating system is expected to have an important influence on the evolution of mating and parenting behaviors. Although many studies have used experimental evolution to examine how mating behaviors evolve under different mating systems, this approach has seldom been used to study the evolution of parental care. We used experimental evolution to test whether adaptation to different mating systems involves changes in mating and parenting behaviors in populations of the burying beetle, Nicrophorus vespilloides. We maintained populations under monogamy or promiscuity for six generations. This manipulation had an immediate impact on reproductive performance and adult survival. Compared to monogamy, promiscuity reduced brood size and adult (particularly male) survival during breeding. After six generations of experimental evolution, there was no divergence between monogamous and promiscuous populations in mating behaviors. Parents from the promiscuous populations (especially males) displayed less care than parents from the monogamous populations. Our results are consistent with the hypothesis that male care will increase with the certainty of paternity. However, it appears that this change is not associated with a concurrent change in mating behaviors.