Efficient construction of a linkage map and haplotypes for Mentha suaveolens using sequence capture.

The sustainability of many crops is hindered by the lack of genomic resources and a poor understanding of natural genetic diversity. Particularly, application of modern breeding requires high-density linkage maps that are integrated into a highly contiguous reference genome. Here, we present a rapid method for deriving haplotypes and developing linkage maps, and its application to Mentha suaveolens, one of the diploid progenitors of cultivated mints. Using sequence-capture via DNA hybridization to target single nucleotide polymorphisms (SNPs), we successfully genotyped ∼5000 SNPs within the genome of >400 individuals derived from a self cross. After stringent quality control, and identification of nonredundant SNPs, 1919 informative SNPs were retained for linkage map construction. The resulting linkage map defined a total genetic space of 942.17 cM divided among 12 linkage groups, ranging from 56.32 to 122.61 cM in length. The linkage map is in good agreement with pseudomolecules from our preliminary genome assembly, proving this resource effective for the correction and validation of the reference genome. We discuss the advantages of this method for the rapid creation of linkage maps.

A horizon scan of priorities for coastal marine microbiome research.

Research into the microbiomes of natural environments is changing the way ecologists and evolutionary biologists view the importance of microorganisms in ecosystem function. This is particularly relevant in ocean environments, where microorganisms constitute the majority of biomass and control most of the major biogeochemical cycles, including those that regulate Earth's climate. Coastal marine environments provide goods and services that are imperative to human survival and well-being (for example, fisheries and water purification), and emerging evidence indicates that these ecosystem services often depend on complex relationships between communities of microorganisms (the 'microbiome') and the environment or their hosts - termed the 'holobiont'. Understanding of coastal ecosystem function must therefore be framed under the holobiont concept, whereby macroorganisms and their associated microbiomes are considered as a synergistic ecological unit. Here, we evaluate the current state of knowledge on coastal marine microbiome research and identify key questions within this growing research area. Although the list of questions is broad and ambitious, progress in the field is increasing exponentially, and the emergence of large, international collaborative networks and well-executed manipulative experiments are rapidly advancing the field of coastal marine microbiome research.

Two Y-chromosome-encoded genes determine sex in kiwifruit.

Dioecy, the presence of male and female individuals, has evolved independently in multiple flowering plant lineages1-3. Although theoretical models for the evolution of dioecy, such as the 'two-mutations' model, are well established4,5, little is known about the specific genes determining sex and their evolutionary history3. Kiwifruit, a major tree crop consumed worldwide, is a dioecious species. In kiwifruit we previously identified a Y-encoded sex-determinant candidate gene acting as the suppressor of feminization (SuF), named Shy Girl (SyGI)6. Here, we identify a second Y-encoded sex-determinant that we named Friendly Boy (FrBy), which exhibits strong expression in tapetal cells. Gene-editing and complementation analyses in Arabidopsis thaliana and Nicotiana tabacum indicated that FrBy acts for the maintenance of male (M) functions, independently of SyGI, and that these functions are conserved across angiosperm species. We further characterized the genomic architecture of the small (<1 megabase pairs (Mb)) male-specific region of the Y chromosome (MSY), which harbours only two genes expressed extensively in developing gynoecia and androecia, respectively: SyGI and FrBy. Re-sequencing of the genome of a natural hermaphrodite kiwifruit revealed that this individual is genetically male but carries deletion(s) of parts of the Y chromosome, including SyGI. Additionally, expression of FrBy in female kiwifruit resulted in hermaphrodite plants. These results clearly indicate that Y-encoded SyGI and FrBy act independently as the SuF and M factors in kiwifruit, respectively, and provide insight into not only the evolutionary path leading to a two-factor sex-determination system, but also a new breeding approach for dioecious species.

Characterization of shifts of koala (Phascolarctos cinereus) intestinal microbial communities associated with antibiotic treatment.

Koalas (Phascolarctos cinereus) are arboreal marsupials native to Australia that eat a specialized diet of almost exclusively eucalyptus leaves. Microbes in koala intestines are known to break down otherwise toxic compounds, such as tannins, in eucalyptus leaves. Infections by Chlamydia, obligate intracellular bacterial pathogens, are highly prevalent in koala populations. If animals with Chlamydia infections are received by wildlife hospitals, a range of antibiotics can be used to treat them. However, previous studies suggested that koalas can suffer adverse side effects during antibiotic treatment. This study aimed to use 16S rRNA gene sequences derived from koala feces to characterize the intestinal microbiome of koalas throughout antibiotic treatment and identify specific taxa associated with koala health after treatment. Although differences in the alpha diversity were observed in the intestinal flora between treated and untreated koalas and between koalas treated with different antibiotics, these differences were not statistically significant. The alpha diversity of microbial communities from koalas that lived through antibiotic treatment versus those who did not was significantly greater, however. Beta diversity analysis largely confirmed the latter observation, revealing that the overall communities were different between koalas on antibiotics that died versus those that survived or never received antibiotics. Using both machine learning and OTU (operational taxonomic unit) co-occurrence network analyses, we found that OTUs that are very closely related to Lonepinella koalarum, a known tannin degrader found by culture-based methods to be present in koala intestines, was correlated with a koala's health status. This is the first study to characterize the time course of effects of antibiotics on koala intestinal microbiomes. Our results suggest it may be useful to pursue alternative treatments for Chlamydia infections without the use of antibiotics or the development of Chlamydia-specific antimicrobial compounds that do not broadly affect microbial communities.

Community-Level Differences in the Microbiome of Healthy Wild Mallards and Those Infected by Influenza A Viruses.

Waterfowl, especially ducks and geese, are primary reservoirs for influenza A viruses (IAVs) that evolve and emerge as important pathogens in domestic animals and humans. In contrast to humans, where IAVs infect the respiratory tract and cause significant morbidity and mortality, IAVs infect the gastrointestinal tract of waterfowl and cause little or no pathology and are spread by fecal-oral transmission. For this reason, we examined whether IAV infection is associated with differences in the cloacal microbiome of mallards (Anas platyrhyncos), an important host of IAVs in North America and Eurasia. We characterized bacterial community composition by sequencing the V4 region of 16S rRNA genes. IAV-positive mallards had lower species diversity, richness, and evenness than IAV-negative mallards. Operational taxonomic unit (OTU) cooccurrence patterns were also distinct depending on infection status. Network analysis showed that IAV-positive mallards had fewer significant cooccurring OTUs and exhibited fewer coassociation patterns among those OTUs than IAV-negative mallards. These results suggest that healthy mallards have a more robust and complex cloacal microbiome. By combining analytical approaches, we identified 41 bacterial OTUs, primarily representatives of Streptococcus spp., Veillonella dispar, and Rothia mucilaginosa, contributing to the observed differences. This study found that IAV-infected wild mallards exhibited strong differences in microbiome composition relative to noninfected mallards and identified a concise set of putative biomarker OTUs. Using Random Forest, a supervised machine learning method, we verified that these 41 bacterial OTUs are highly predictive of infection status. IMPORTANCE Seasonal influenza causes 3 to 5 million severe illnesses and 250,000 to 500,000 human deaths each year. While pandemic influenza viruses emerge only periodically, they can be devastating-for example, the 1918 H1N1 pandemic virus killed more than 20 million people. IAVs infect the respiratory tract and cause significant morbidity and mortality in humans. In contrast, IAVs infect the gastrointestinal tract of waterfowl, producing little pathology. Recent studies indicated that viruses can alter the microbiome at the respiratory and gastrointestinal mucosa, but there are no reports of how the microbiota of the natural host of influenza is affected by infection. Here we find that the mallard microbiome is altered during IAV infection. Our results suggest that detailed examination of humans and animals infected with IAVs may reveal individualized microbiome profiles that correspond to health and disease. Moreover, future studies should explore whether the altered microbiome facilitates maintenance and transmission of IAVs in waterfowl populations.

Elucidating the role of AII amacrine cells in glutamatergic retinal waves.

Spontaneous retinal activity mediated by glutamatergic neurotransmission-so-called "Stage 3" retinal waves-drives anti-correlated spiking in ON and OFF RGCs during the second week of postnatal development of the mouse. In the mature retina, the activity of a retinal interneuron called the AII amacrine cell is responsible for anti-correlated spiking in ON and OFF α-RGCs. In mature AIIs, membrane hyperpolarization elicits bursting behavior. Here, we postulated that bursting in AIIs underlies the initiation of glutamatergic retinal waves. We tested this hypothesis by using two-photon calcium imaging of spontaneous activity in populations of retinal neurons and by making whole-cell recordings from individual AIIs and α-RGCs in in vitro preparations of mouse retina. We found that AIIs participated in retinal waves, and that their activity was correlated with that of ON α-RGCs and anti-correlated with that of OFF α-RGCs. Though immature AIIs lacked the complement of membrane conductances necessary to generate bursting, pharmacological activation of the M-current, a conductance that modulates bursting in mature AIIs, blocked retinal wave generation. Interestingly, blockade of the pacemaker conductance Ih, a conductance absent in AIIs but present in both ON and OFF cone bipolar cells, caused a dramatic loss of spatial coherence of spontaneous activity. We conclude that during glutamatergic waves, AIIs act to coordinate and propagate activity generated by BCs rather than to initiate spontaneous activity.

Copper is an endogenous modulator of neural circuit spontaneous activity.

For reasons that remain insufficiently understood, the brain requires among the highest levels of metals in the body for normal function. The traditional paradigm for this organ and others is that fluxes of alkali and alkaline earth metals are required for signaling, but transition metals are maintained in static, tightly bound reservoirs for metabolism and protection against oxidative stress. Here we show that copper is an endogenous modulator of spontaneous activity, a property of functional neural circuitry. Using Copper Fluor-3 (CF3), a new fluorescent Cu(+) sensor for one- and two-photon imaging, we show that neurons and neural tissue maintain basal stores of loosely bound copper that can be attenuated by chelation, which define a labile copper pool. Targeted disruption of these labile copper stores by acute chelation or genetic knockdown of the CTR1 (copper transporter 1) copper channel alters the spatiotemporal properties of spontaneous activity in developing hippocampal and retinal circuits. The data identify an essential role for copper neuronal function and suggest broader contributions of this transition metal to cell signaling.

Modafinil augments oscillatory power in middle frequencies during rule selection.

Control-related cognitive processes are associated with cortical oscillations and modulated by catecholamine neurotransmitters. It remains unclear how catecholamine systems modulate control-related oscillations. We tested modafinil effects on rule-related 4-30 Hz oscillations, with double-blind, placebo-controlled (within-subjects) testing of 22 healthy adults, using EEG during cognitive control task performance. EEG data underwent time-frequency decomposition with Morlet wavelets to determine power of 4-30 Hz oscillations. Modafinil enhanced oscillatory power associated with high-control rule selection in theta, alpha, and beta ranges, with a frontotemporal topography and minimal effects during rule maintenance. Augmentation of catecholamine signaling enhances middle-frequency cortical oscillatory power associated with rule selection, which may subserve diverse subcomponent processes in proactive cognitive control.

A role for correlated spontaneous activity in the assembly of neural circuits.

Before the onset of sensory transduction, developing neural circuits spontaneously generate correlated activity in distinct spatial and temporal patterns. During this period of patterned activity, sensory maps develop and initial coarse connections are refined, which are critical steps in the establishment of adult neural circuits. Over the last decade, there has been substantial evidence that altering the pattern of spontaneous activity disrupts refinement, but the mechanistic understanding of this process remains incomplete. In this review, we discuss recent experimental and theoretical progress toward the process of activity-dependent refinement, focusing on circuits in the visual, auditory, and motor systems. Although many outstanding questions remain, the combination of several novel approaches has brought us closer to a comprehensive understanding of how complex neural circuits are established by patterned spontaneous activity during development.

Extrasynaptic glutamate and inhibitory neurotransmission modulate ganglion cell participation during glutamatergic retinal waves.

During the first 2 wk of mouse postnatal development, transient retinal circuits give rise to the spontaneous initiation and lateral propagation of depolarizations across the ganglion cell layer (GCL). Glutamatergic retinal waves occur during the second postnatal week, when GCL depolarizations are mediated by ionotropic glutamate receptors. Bipolar cells are the primary source of glutamate in the inner retina, indicating that the propagation of waves depends on their activation. Using the fluorescence resonance energy transfer-based optical sensor of glutamate FLII81E-1µ, we found that retinal waves are accompanied by a large transient increase in extrasynaptic glutamate throughout the inner plexiform layer. Using two-photon Ca(2+) imaging to record spontaneous Ca(2+) transients in large populations of cells, we found that despite this spatially diffuse source of depolarization, only a subset of neurons in the GCL and inner nuclear layer (INL) are robustly depolarized during retinal waves. Application of the glutamate transporter blocker dl-threo-ß-benzyloxyaspartate (25 µM) led to a significant increase in cell participation in both layers, indicating that the concentration of extrasynaptic glutamate affects cell participation in both the INL and GCL. In contrast, blocking inhibitory transmission with the GABAA receptor antagonist gabazine and the glycine receptor antagonist strychnine increased cell participation in the GCL without significantly affecting the INL. These data indicate that during development, glutamate spillover provides a spatially diffuse source of depolarization, but that inhibitory circuits dictate which neurons within the GCL participate in retinal waves.

The role of neuronal connexins 36 and 45 in shaping spontaneous firing patterns in the developing retina.

Gap junction coupling synchronizes activity among neurons in adult neural circuits, but its role in coordinating activity during development is less known. The developing retina exhibits retinal waves--spontaneous depolarizations that propagate among retinal interneurons and drive retinal ganglion cells (RGCs) to fire correlated bursts of action potentials. During development, two connexin isoforms, connexin 36 (Cx36) and Cx45, are expressed in bipolar cells and RGCs, and therefore provide a potential substrate for coordinating network activity. To determine whether gap junctions contribute to retinal waves, we compared spontaneous activity patterns using calcium imaging, whole-cell recording, and multielectrode array recording in control, single-knock-out (ko) mice lacking Cx45 and double-knock-out (dko) mice lacking both isoforms. Wave frequency, propagation speed, and bias in propagation direction were similar in control, Cx36ko, Cx45ko, and Cx36/45dko retinas. However, the spontaneous firing rate of individual retinal ganglion cells was elevated in Cx45ko retinas, similar to Cx36ko retinas (Hansen et al., 2005; Torborg and Feller, 2005), a phenotype that was more pronounced in Cx36/45dko retinas. As a result, spatial correlations, as assayed by nearest-neighbor correlation and functional connectivity maps, were significantly altered. In addition, Cx36/45dko mice had reduced eye-specific segregation of retinogeniculate afferents. Together, these findings suggest that although Cx36 and Cx45 do not play a role in gross spatial and temporal propagation properties of retinal waves, they strongly modulate the firing pattern of individual RGCs, ensuring strongly correlated firing between nearby RGCs and normal patterning of retinogeniculate projections.

Gamma oscillatory power is impaired during cognitive control independent of medication status in first-episode schizophrenia.

Schizophrenia is characterized by impaired cognitive control associated with prefrontal cortex dysfunction, but the underlying pathophysiological mechanisms remain unknown. Higher cognitive processes are associated with cortical oscillations in the gamma range, which are also impaired in chronic schizophrenia. We tested whether cognitive control-related gamma deficits are observed in first-episode patients, and whether they are associated with antipsychotic medication exposure. Fifty-three first-episode schizophrenia patients (21 without antipsychotic medication treatment) and 29 healthy control subjects underwent electroencephalography (EEG) during performance of a preparatory cognitive control task (preparing to overcome prepotency or POP task). The first-episode schizophrenia patient group was impaired (relative to the control group) on task performance and on delay-period gamma power at each of the three subgroups of frontal electrodes. The unmedicated patient subgroup was similarly impaired compared with controls, and was not different on these measures compared with the medicated patient subgroup. In contrast, delay-period theta power was not impaired in the full patient group nor in the unmedicated patient subgroup. Impaired cognitive control-related gamma cortical oscillatory activity is present at the first psychotic episode in schizophrenia, and is independent of medication status. This suggests that altered local circuit function supporting high-frequency oscillatory activity in prefrontal cortex ensembles may serve as the pathophysiological substrate of cognitive control deficits in schizophrenia.

Google and the mind: predicting fluency with PageRank.

Human memory and Internet search engines face a shared computational problem, needing to retrieve stored pieces of information in response to a query. We explored whether they employ similar solutions, testing whether we could predict human performance on a fluency task using PageRank, a component of the Google search engine. In this task, people were shown a letter of the alphabet and asked to name the first word beginning with that letter that came to mind. We show that PageRank, computed on a semantic network constructed from word-association data, outperformed word frequency and the number of words for which a word is named as an associate as a predictor of the words that people produced in this task. We identify two simple process models that could support this apparent correspondence between human memory and Internet search, and relate our results to previous rational models of memory.