Post-pollination paternal reproductive success in Populus nigra: a male affair

Populus nigra is a riparian forest species that is considered vulnerable in several parts of its natural range due to habitat loss and fragmentation. Resulting small population sizes are a source of demographic stochasticity, loss of genetic diversity, and inbreeding which can be strongly enhanced if adult trees exhibit contrasted and stable differential reproductive success (DRS). This study was designed to test for the existence of post-pollination paternal DRS in P. nigra and its stability across female genotypes. An equal-weight pollen mix was used to hand-pollinate four female genotypes, and the paternity of the resulting seeds was elucidated using SSR markers. Pollen germination rates and number of grains per weight unit were taken into account as potential predictors of the DRS observed. Using a four-pollen mix allowed testing male-by-female interactions statistically. Although most parental genotypes originated from a single population along the Loire River in France, one pollen contributor was the Lombardy poplar (P. nigra var. “Italica”), an ornamental male cultivar supposed to originate from the Middle East and present worldwide. Moreover, two of the females involved were identified as spontaneous F ₁s from this clone. The material studied was thus genetically diverse enough to detect male-by-female interactions should they be common in the species. Significant post-pollination paternal DRS was found which could not be explained by the pollen characteristics measured. The most significant result was the absence of male-by-female interactions on the paternal DRS observed. These results are discussed in terms of in situ conservation strategies. They may also have implications in terms of evolution, a stable paternal DRS across females being a prerequisite for sexual selection to occur.     

Cytosolic hydroxymethyldihydropterin pyrophosphokinase/dihydropteroate synthase from Arabidopsis thaliana: a specific role in early development and stress response

In plants, 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase/7,8-dihydropteroate synthase (mitHPPK/DHPS) is a bifunctional mitochondrial enzyme, which catalyzes the first two consecutive steps of tetrahydrofolate biosynthesis. Mining the Arabidopsis genome data base has revealed a second gene encoding a protein that lacks a potential transit peptide, suggesting a cytosolic localization of the isoenzyme (cytHPPK/DHPS). When the N-terminal part of the cytHPPK/DHPS was fused to green fluorescent protein, the fusion protein appeared only in the cytosol, confirming the above prediction. Functionality of cytHPPK/DHPS was addressed by two parallel approaches: first, the cytHPPK/DHPS was able to rescue yeast mutants lacking corresponding activities; second, recombinant cytHPPK/DHPS expressed and purified from Escherichia coli displayed both HPPK and DHPS activities in vitro. In contrast to mitHPPK/DHPS, which was ubiquitously expressed, the cytHPPK/DHPS gene was exclusively expressed in reproductive tissue, more precisely in developing seeds as revealed by histochemical analysis of a transgenic cytHPPK/DHPS promoter-GUS line. In addition, it was observed that expression of cytHPPK/DHPS mRNA was induced by salt stress, suggesting a potential role of the enzyme in stress response. This was supported by the phenotype of a T-DNA insertion mutant in the cytHPPK/DHPS gene, resulting in lower germination rates as compared with the wild-type upon application of oxidative and osmotic stress.     

Measuring cortisol in fish scales to study stress in wild tropical tuna

Environmental Biology of Fishes - Cortisol is recognized as a physiological indicator of stress in fish. However, this hormone is typically measured in plasma samples. In this study, cortisol...     

Interactions between multiple helminths and the gut microbiota in wild rodents

The gut microbiota is vital to host health and, as such, it is important to elucidate the mechanisms altering its composition and diversity. Intestinal helminths are host immunomodulators and have evolved both temporally and spatially in close association with the gut microbiota, resulting in potential mechanistic interplay. Host–helminth and host–microbiota interactions are comparatively well-examined, unlike microbiota–helminth relationships, which typically focus on experimental infection with a single helminth species in laboratory animals. Here, in addition to a review of the literature on helminth–microbiota interactions, we examined empirically the association between microbiota diversity and composition and natural infection of multiple helminth species in wild mice (Apodemus flavicollis), using 16S rRNA gene catalogues (metataxonomics). In general, helminth presence is linked with high microbiota diversity, which may confer health benefits to the host. Within our wild rodent system variation in the composition and abundance of gut microbial taxa associated with helminths was specific to each helminth species and occurred both up- and downstream of a given helminth''s niche (gut position). The most pronounced helminth–microbiota association was between the presence of tapeworms in the small intestine and increased S24–7 (Bacteroidetes) family in the stomach. Helminths clearly have the potential to alter gut homeostasis. Free-living rodents with a diverse helminth community offer a useful model system that enables both correlative (this study) and manipulative inference to elucidate helminth–microbiota interactions.     

AN EVALUATION OF THE HYBRID SPECIATION HYPOTHESIS FOR XIPHOPHORUS CLEMENCIAE BASED ON WHOLE GENOME SEQUENCES

Once thought rare in animal taxa, hybridization has been increasingly recognized as an important and common force in animal evolution. In the past decade, a number of studies have suggested that hybridization has driven speciation in some animal groups. We investigate the signature of hybridization in the genome of a putative hybrid species, Xiphophorus clemenciae, through whole genome sequencing of this species and its hypothesized progenitors. Based on analysis of this data, we find that X. clemenciae is unlikely to have been derived from admixture between its proposed parental species. However, we find significant evidence for recent gene flow between Xiphophorus species. Although we detect genetic exchange in two pairs of species analyzed, the proportion of genomic regions that can be attributed to hybrid origin is small, suggesting that strong behavioral premating isolation prevents frequent hybridization in Xiphophorus. The direction of gene flow between species is potentially consistent with a role for sexual selection in mediating hybridization.     

The kinesin of the flagellum attachment zone in Leishmania is required for cell morphogenesis,cell division and virulence in the mammalian host

Leishmania parasites possess a unique and complex cytoskeletal structure termed flagellum attachment zone (FAZ) connecting the base of the flagellum to one side of the flagellar pocket (FP), an invagination of the cell body membrane and the sole site for endocytosis and exocytosis. This structure is involved in FP architecture and cell morphogenesis, but its precise role and molecular composition remain enigmatic. Here, we characterized Leishmania FAZ7, the only known FAZ protein containing a kinesin motor domain, and part of a clade of trypanosomatid-specific kinesins with unknown functions. The two paralogs of FAZ7, FAZ7A and FAZ7B, display different localizations and functions. FAZ7A localizes at the basal body, while FAZ7B localizes at the distal part of the FP, where the FAZ structure is present in Leishmania. While null mutants of FAZ7A displayed normal growth rates, the deletion of FAZ7B impaired cell growth in both promastigotes and amastigotes of Leishmania. The kinesin activity is crucial for its function. Deletion of FAZ7B resulted in altered cell division, cell morphogenesis (including flagellum length), and FP structure and function. Furthermore, knocking out FAZ7B induced a mis-localization of two of the FAZ proteins, and disrupted the molecular organization of the FP collar, affecting the localization of its components. Loss of the kinesin FAZ7B has important consequences in the insect vector and mammalian host by reducing proliferation in the sand fly and pathogenicity in mice. Our findings reveal the pivotal role of the only FAZ kinesin as part of the factors important for a successful life cycle of Leishmania.     

Giant clams discriminate threats along a risk gradient and display varying habituation rates to different stimuli

It is often beneficial for animals to discriminate between different threats and to habituate to repeated exposures of benign stimuli. While much is known about risk perception in vertebrates and some invertebrates, risk perception in marine invertebrates is less extensively studied. One method to study risk perception is to habituate animals to a series of exposures to one stimulus, and then present a novel stimulus to test if it transfers habituation. Transfer of habituation is seen as a continued decrease in response while lack of transfer is seen either by having a similar or greater magnitude response. We asked whether giant clams (Tridacna maxima) discriminate between biologically relevant types of threats along a risk gradient. Giant clams retract their mantle and close their shell upon detecting a threat. While closed, they neither feed nor photosynthesize, and prior work has shown that the cost of being closed increases as the duration of their response increases. We recorded a clam's latency to emerge after simulated threats chosen to represent a risk gradient: exposure to a small shading event, a medium shading event, a large shading event (chosen to simulate fish swimming above them), tapping on their shell and touching their mantle (chosen to simulate different degrees of direct attack). Although these stimuli are initially perceived as threatening, we expected clams to habituate to them because they are ultimately non‐damaging and it would be costly for clams to remain closed for extended periods of time when there is no threat present. Clams had different initial latencies to emerge and different habituation rates to these treatments, and they did not transfer habituation to higher risk stimuli and to some lower risk stimuli. These results suggest that clams discriminated between these stimuli along a risk gradient and the lack of habituation transfer shows that the new stimulus was perceived as a potential threat. This study demonstrates that sessile bivalves can discern between levels of predatory threat. These photosynthetic clams may benefit from being able to categorize predator cues for efficient energy allocation.     

First efficient CRISPR‐Cas9‐mediated genome editing in Leishmania parasites

Protozoan pathogens that cause leishmaniasis in humans are relatively refractory to genetic manipulation. In this work, we implemented the CRISPR‐Cas9 system in Leishmania parasites and demonstrated its efficient use for genome editing. The Cas9 endonuclease was expressed under the control of the Dihydrofolate Reductase‐Thymidylate Synthase (DHFR‐TS) promoter and the single guide RNA was produced under the control of the U6snRNA promoter and terminator. As a proof of concept, we chose to knockout a tandemly repeated gene family, the paraflagellar rod‐2 locus. We were able to obtain null mutants in a single round of transfection. In addition, we confirmed the absence of off‐target editions by whole genome sequencing of two independent clones. Our work demonstrates that CRISPR‐Cas9‐mediated gene knockout represents a major improvement in comparison with existing methods. Beyond gene knockout, this genome editing tool opens avenues for a multitude of functional studies to speed up research on leishmaniasis.     

UVA-induced cyclobutane pyrimidine dimers form predominantly at thymine-thymine dipyrimidines and correlate with the mutation spectrum in rodent cells

Ligation-mediated PCR was employed to quantify cyclobutane pyrimidine dimer (CPD) formation at nucleotide resolution along exon 2 of the adenine phosphoribosyltransferase (aprt) locus in Chinese hamster ovary (CHO) cells following irradiation with either UVA (340–400 nm), UVB (295–320 nm), UVC (254 nm) or simulated sunlight (SSL; λ > 295 nm). The resulting DNA damage spectrum for each wavelength region was then aligned with the corresponding mutational spectrum generated previously in the same genetic target. The DNA sequence specificities of CPD formation induced by UVC, UVB or SSL were very similar, i.e., in each case the overall relative proportion of this photoproduct forming at TT, TC, CT and CC sites was ~28, ~26, ~16 and ~30%, respectively. Furthermore, a clear correspondence was noted between the precise locations of CPD damage hotspots, and of ‘UV signature’ mutational hotspots consisting primarily of C→T and CC→TT transitions within pyrimidine runs. However, following UVA exposure, in strong contrast to the above situation for UVC, UVB or SSL, CPDs were generated much more frequently at TT sites than at TC, CT or CC sites (57% versus 18, 11 and 14%, respectively). This CPD deposition pattern correlates well with the strikingly high proportion of mutations recovered opposite TT dipyrimidines in UVA- irradiated CHO cells. Our results directly implicate the CPD as a major promutagenic DNA photoproduct induced specifically by UVA in rodent cells.