title: Emerging SARS-CoV-2 variants reduce neutralization sensitivity to convalescent sera and monoclonal antibodies

abstract: SARS-CoV-2 Spike-specific antibodies contribute the majority of the neutralizing activity in most convalescent human sera. Two SARS-CoV-2 variants, N501Y.V1 (also known as B.1.1.7 lineage or VOC-202012/01) and N501Y.V2 (B.1.351 lineage), reported from the United Kingdom and South Africa, contain several mutations in the receptor binding domain of Spike and are of particular concern. To address the infectivity and neutralization escape phenotypes potentially caused by these mutations, we used SARS-CoV-2 pseudovirus system to compare the viral infectivity, as well as the neutralization activities of convalescent sera and monoclonal antibodies (mAbs) against SARS-CoV-2 variants. Our results showed that N501Y Variant 1 and Variant 2 increase viral infectivity compared to the reference strain (wild-type, WT) in vitro. At 8 months after symptom onset, 17 serum samples of 20 participants (85%) retaining titers of ID50 >40 against WT pseudovirus, whereas the NAb titers of 8 samples (40%) and 18 samples (90%) decreased below the threshold against N501Y.V1 and N501Y.V2, respectively. In addition, both N501Y Variant 1 and Variant 2 reduced neutralization sensitivity to most (6/8) mAbs tested, while N501Y.V2 even abrogated neutralizing activity of two mAbs. Taken together the results suggest that N501Y.V1 and N501Y.V2 reduce neutralization sensitivity to some convalescent sera and mAbs.


title: The developmental stage of the medulloblastoma cell-of-origin restricts Hedgehog pathway usage and drug sensitivity

abstract:
Sonic Hedgehog (SHH) medulloblastoma originates from the cerebellar granule neuron progenitor (CGNP) lineage that depends on Hedgehog signaling for its perinatal expansion. While SHH tumors exhibit overall deregulation of this pathway, they also show patient age-specific aberrations.

To investigate if the developmental stage of the CGNP can account for these age-specific lesions, we analyzed developing murine CGNP transcriptomes and observed highly dynamic gene expression as function of age. Cross-species comparison with human SHH medulloblastoma showed partial maintenance of these expression patterns, and highlighted low primary cilium expression as hallmark of infant medulloblastoma and early embryonic CGNPs. This coincided with reduced responsiveness to upstream Shh pathway component Smoothened, while sensitivity to downstream components Sufu and Gli was retained.

Together, these findings can explain the preference for SUFU mutations in infant medulloblastoma and suggest that drugs targeting the downstream SHH pathway will be most appropriate for infant patients.

category: cancer biology


title: Computational ligands to VKORC1s and CYPs. Could they predict new anticoagulant rodenticides?

abstract: Anticoagulant-resistance in rodents and anticoagulant off-target effects are some of the world-wide problems of increasing concern. To search for new anticoagulant rodenticide candidates we have computationally explored some of the rat genes previously implicated in resistance to actual anticoagulants. In particular, we searched among hundreds of anticoagulant-similar chemotypes those binding rat wild-type VKORC1 (the best-known anticoagulant target, a Vitamin K-recycling enzyme), VKORC1L1 (a VKORC1-related enzyme), Cytochrome P450 CYP enzymes (some of the most important enzymes implicated in detoxification) and anticoagulant-resistant VKORC1-mutants (to minimize propensity to resistance). Results predicted new VKORC1 leads with binding-scores in the low nM range (high binding-affinities) predicting hydroxycoumarin- and naphtoquinone-like chemotypes. We then selected top-leads with additional high binding-scores to more than three anticoagulant-related CYPs, suggesting minimal detoxification rates and therefore maximal anticoagulation expectatives. A downsized list of top top-leads maintaining VKORC1 low-binding scores to anticoagulant resistant mutants, was finally proposed for experimental validation. The combination of different rat targets for computational studies, could be used to search for unrelated chemotypes, for reduction of off-target environmental anticoagulant impacts, and/or as new tools to explore anticoagulant molecular mechanisms.

category: pharmacology and toxicology


title: A novel micro-computerized tomography method reveals Drosophila cardiac hypertrophy caused by upregulated store-operated calcium entry

abstract:
Heart failure is often preceded by pathological cardiac hypertrophy, a thickening of the heart musculature driven by complex gene regulatory and signaling processes. The Drosophila heart has great potential as a genetic model for deciphering the underlying mechanisms of cardiac hypertrophy. However, current methods for evaluating hypertrophy of the Drosophila heart are laborious and difficult to carry out reproducibly. Here we demonstrate that micro-computerized tomography (microCT) is an accessible, highly reproducible method for non-destructive, quantitative analysis of Drosophila heart morphology and size. To validate our microCT approach for analyzing Drosophila cardiac hypertrophy, we show that expression of constitutively active Ras (Ras85DV12), previously shown to cause hypertrophy of the fly heart, results in significant thickening of both adult and larval heart walls when measured from microCT images. We then show using microCT analysis that genetic upregulation of store-operated Ca2+ entry (SOCE) driven by expression of constitutively active Stim (StimCA) or Orai (OraiCA) proteins also results in significant hypertrophy of the Drosophila heart, through a process that specifically depends on Orai Ca2+ influx channels. Intravital imaging of heart contractility revealed significantly reduced end diastolic dimensions in StimCA and OraiCA expressing hearts, consistent with the hypertrophic phenotype. These results demonstrate that increased SOCE activity is an important driver of hypertrophic cardiomyocyte growth, and demonstrate how microCT analysis combined with tractable genetic tools in Drosophila can be used to delineate molecular signaling processes that underlie cardiac hypertrophy and heart failure.

NEW AND NOTEWORTHYGenetic analysis of cardiac hypertrophy in Drosophila holds immense potential for the discovery of new therapeutic targets to prevent and treat heart failure. However, this potential has been hindered by a lack of rapid and effective methods for analysis of heart size in flies. Here we demonstrate that analysis of the Drosophila heart with micro-computerized tomography yields accurate and highly reproducible heart size measurements that can be used to efficiently analyze heart growth and cardiac hypertrophy in Drosophila.

category: physiology


title: Analysis of Drosophila cardiac hypertrophy by micro-computerized tomography for genetic dissection of heart growth mechanisms


title: A duplicated copy of the meiotic gene ZIP4 preserves up to 50% pollen viability and grain number in polyploid wheat

abstract:
O_LIAlthough most flowering plants are polyploid, little is known of how the meiotic process evolved to stabilise and preserve polyploid fertility. On wheat polyploidisation, the major meiotic gene ZIP4 on chromosome 3B duplicated onto 5B and subsequently diverged. This 5B meiotic gene copy (TaZIP4-B2) was recently shown to promote homologous pairing, synapsis and crossover, and suppress homoeologous crossover. We therefore suspected that these stabilising effects on meiosis could be important for the preservation of wheat polyploid fertility.
C_LIO_LIA CRISPR Tazip4-B2 mutant was exploited to assess the contribution of the 5B duplicated ZIP4 copy in maintaining pollen viability and grain setting.
C_LIO_LIAnalysis demonstrated abnormalities in 56% of meiocytes in the Tazip4-B2 mutant, with micronuclei in 50% of tetrads, reduced size in 48% of pollen grains and a near 50% reduction in grain number. Further studies showed that most of the reduced grain number resulted from pollination with less viable pollen, suggesting that the stabilising effect of TaZIP4-B2 on meiosis has a greater consequence in subsequent male, rather than female gametogenesis.
C_LIO_LIThese studies reveal the extraordinary value of the wheat chromosome 5B TaZIP4-B2 duplication to agriculture and human nutrition. Future studies should assess whether different TaZIP4-B2 alleles exhibit variable effects on meiotic stabilisation and/or resistance to temperature change.
C_LI

category: plant biology


title: Pyrimidine nucleotide availability is essential for efficient photosynthesis, ROS scavenging, and organelle development

abstract:
De novo synthesis of pyrimidines is an essential and highly conserved pathway in all organisms. A peculiarity in plants is the localization of the first committed step, catalyzed by aspartate transcarbamoylase (ATC), in chloroplasts. By contrast, the third step in the pathway is catalyzed by dihydroorotate dehydrogenase (DHODH) localized in mitochondria in eukaryotes, including plants. To unravel pathway- and organelle specific functions, we analyzed knock-down mutants in ATC and DHODH in detail. ATC knock-downs were most severely affected, exhibiting low levels of pyrimidine metabolites, a low energy state, reduced photosynthetic capacity and accumulated reactive oxygen species (ROS). Furthermore, we observed altered leaf morphology and chloroplast ultrastructure in the mutants. Although less affected, DHODH knock-down mutants showed impaired seed germination and altered mitochondrial ultrastructure. Our results point to an integration of de novo pyrimidine synthesis and cellular energy states via photosynthesis and mitochondrial respiration. These findings highlight the likelihood of further regulatory roles for ATC and DHODH in pathways located in the corresponding organelles.

ONE-SENTENCE SUMMARYImpaired pyrimidine nucleotide synthesis results in a low energy state, affecting photosynthesis and organellar ultrastructure, thus leading to reduced growth, reproduction, and seed yield


title: Diversity of receptor expression in central and peripheral mouse neurons estimated from single cell RNA sequencing

abstract: Because somatosensory PNS neurons, in particular nociceptors, are specially tuned to be able to detect a wide variety of both exogenous and endogenous signals, it is widely assumed that these neurons express a greater variety of receptor genes. Because cells detect such signals via cell surface receptors, we sought to formally test the hypothesis that PNS neurons might express a broader array of cell surface receptors than CNS neurons using existing single cell RNA sequencing resources from mouse. We focused our analysis on ion channels, G-protein coupled receptors (GPCRS), receptor tyrosine kinase and cytokine family receptors. In partial support of our hypothesis, we found that mouse PNS somatosensory, sympathetic and enteric neurons and CNS neurons have similar receptor expression diversity in families of receptors examined, with the exception of GPCRs and cytokine receptors which showed greater diversity in the PNS. Surprisingly, these differences were mostly driven by enteric and sympathetic neurons, not by somatosensory neurons or nociceptors. Secondary analysis revealed many receptors that are very specifically expressed in subsets of PNS neurons, including some that are unique among neurons for nociceptors. Finally, we sought to examine specific ligand-receptor interactions between T cells and PNS and CNS neurons. Again, we noted that most interactions between these cells are shared by CNS and PNS neurons despite the fact that T cells only enter the CNS under rare circumstances. Our findings demonstrate that both PNS and CNS neurons express an astonishing array of cell surface receptors and suggest that most neurons are tuned to receive signals from other cells types, in particular immune cells.

category: neuroscience


title: Systemic administration of anti-CD20 indirectly reduces B cells in the inflamed meninges in a chronic model of central nervous system autoimmunity

abstract: Anti-CD20 B cell depleting therapies have demonstrated that B cells are important drivers of disease progress in Multiple Sclerosis, although the pathogenic mechanisms are not well understood. A population of B cells accumulates in the inflamed meninges in MS and also some chronic animal models of disease, typically adjacent to demyelinating lesions. The role of these meningeal B cells in disease is not known, nor is their susceptibility to anti-CD20 therapy. Here, we administered anti-CD20 to 2D2 IgHMOG spontaneous experimental autoimmune encephalomyelitis mice in the chronic phase of disease, after the establishment of meningeal B cell clusters. Compared to the circulation, lymph nodes, and spleen, B cell depletion from the CNS was delayed and not evident until 7d post administration of anti-CD20. Further, we did not find evidence that anti-CD20 accessed meningeal B cells directly, but rather that depletion was indirect and the result of ongoing turnover of the meningeal population and elimination of the peripheral pool from which it is sustained. The reduction of B cell numbers in the CNS coincided with less demyelination of the spinal cord white matter and also, surprisingly, an increase in the number of T cells recruited to the meninges but not parenchyma.

category: immunology


title: CCR1 regulatory variants linked to pulmonary macrophage recruitment in severe COVID-19

abstract: Genome-wide association studies have identified 3p21.31 as the main risk locus for severe disease in COVID-19 patients, although underlying biological mechanisms remain elusive. We performed a comprehensive epigenomic dissection of the 3p21.31 locus, identifying a CTCF-dependent tissue-specific 3D regulatory chromatin hub that controls the activity of several tissue-homing chemokine receptor (CCR) genes in monocytes and macrophages. Risk SNPs colocalized with regulatory elements and were linked to increased expression of CCR1, CCR2 and CCR5 in monocytes and macrophages. As excessive organ infiltration of inflammatory monocytes and macrophages is a hallmark of severe COVID-19, our findings provide a rationale for the genetic association of 3p21.31 variants with elevated risk of hospitalization upon SARS-CoV-2 infection.


title: Severe COVID-19 associated variants linked to chemokine receptor gene control in monocytes and macrophages


title: Deciphering the differential physiological and molecular requirements for conidial anastomosis tube fusion and germ tube formation in Colletotrichum gloeosporioides.

abstract: The conidia of a hemibiotrophic fungus Colletotrichum gloeosporioides can conventionally form germ tube (GT) and develop in to a fungal colony, while under certain conditions, they tend to get connected with each other through conidial anastomosis tube (CAT) so as to share the nutrients. CAT fusion is believed to be responsible for generation of genetic variations in few asexual fungi, which appears problematic for effective fungal disease management. The physiological and molecular mechanism underlying the GT versus CAT formation remained unexplored. In the present study, we have deciphered the decision switch responsible for GT formation versus CAT fusion in C. gloeosporioides. GT formation occurred at high frequency in the presence of nutrients, while CAT fusion was found to be higher in absence of nutrients. Younger conidia were found to form GT efficiently, whilst older conidia preferentially formed CAT. Whole transcriptome analysis of GT and CAT fusion revealed differential molecular requirements for these two processes. We identified 11050 and 9786 differentially expressed genes (DEGs) in GT and CAT, respectively. A total 1567 effector candidates were identified, of them 103 and 101 were uniquely secreted during GT and CAT formation respectively. Genes coding for cell wall degrading enzymes, germination, hyphal growth, host-fungus interaction and virulence were up-regulated during GT formation. Whilst, genes involved in stress response, cell wall remodelling, membrane transport, cytoskeleton, cell cycle, and cell rescue were highly up-regulated during CAT fusion. To conclude, the GT and CAT fusion were found to be mutually exclusive processes, requiring differential physiological conditions and sets of DEGs in C. gloeosporioides. This will help to understand the basic CAT biology in the genus Colletotrichum.

category: microbiology


title: Comparative genomics of emerging lineages and mobile resistomes of contemporary broiler strains of Salmonella Infantis and E. coli

abstract:
IntroductionCommensal and pathogenic strains of MDR Escherichia coli and non-typhoid strains of Salmonella represent a growing foodborne threat from foods of poultry origin. MDR strains of S. Infantis and E. coli are frequently isolated from broiler chicks and the simultaneous presence of these two enteric bacterial species would potentially allow the exchange of mobile resistance determinants.

ObjectivesIn order to understand possible genomic relations and to obtain a first insight into the potential interplay of resistance genes between enteric bacteria, we compared genomic diversity and mobile resistomes of S. Infantis and E. coli from broiler sources.

ResultsThe cgMLST analysis of 56 S. Infantis and 90 E. coli contemporary strains revealed a high genomic heterogeneity of broiler E. coli. It also allowed the first insight into the genomic diversity of the MDR clone B2 of S. Infantis, which is endemic in Hungary. We also identified new MDR lineages for S. Infantis (ST7081 and ST7082) and for E. coli (ST8702 and ST10088). Comparative analysis of antibiotic resistance genes and plasmid types revealed a relatively narrow interface between the mobile resistomes of E. coli and S. Infantis. The mobile resistance genes tet(A), aadA1 and sul1 were identified at an overall high prevalence in both species. This gene association is characteristic to the plasmid pSI54/04 of the epidemic clone B2 of S. Infantis. Simultaneous presence of these genes and of IncI and IncX plasmids in cohabitant caecal strains of E. coli and S. Infantis suggests an important role of these plasmids in a possible interplay of resistance genes between S. Infantis and E. coli in broilers.

ConclusionsThis is the first comparative genomic analysis of contemporary broiler strains of S. Infantis and E. coli. The diversity of mobile resistomes suggests that commensal E. coli could be potential reservoirs of resistance for S. Infantis, but so far only a few plasmid types and mobile resistance genes could be considered as potentially exchangeable between these two species. Among these, IncI plasmids could make the greatest contribution to the microevolution and genetic interaction between E. coli and S. Infantis.


title: Adult porcine (Sus scrofa) derived inner ear cells possessing multipotent stem/progenitor cell characteristics in in vitro cultures

abstract: The human inner ear compared with that of other mammalian species is very complex. Although the mouses cochlea is frequently studied the mouses inner ear continues to develop postnatally whilst the human inner ear is fully developed by the third month of gestation which leads one to question the applicability of findings based on research on mice to human regenerative therapies. Here, we report a novel in vitro culture of adult porcine (Sus scrofa) inner ear cells developed from post-mortem labyrinth specimens. Anatomical findings based on maximal transverse and vertical axial diameters and the length of the cochlear duct suggest that the pigs cochlea is similar to the human cochlea. In vitro cultures of porcine cochlear and vestibular cells showed the persistence of both inner ear hair cell (HC), supporting cell (SC) and stem/progenitor cell characteristics across passages up to 6 based on scanning electron microscopy, fluorescence immunocytochemistry and quantitative reverse transcription polymerase chain reaction (RT-qPCR). Our findings showed that porcine cochlear and vestibular epithelia maintained multipotent stem/progenitor cell populations into adulthood although their regenerative capacities differed across the passages. The development of a viable and reproducible method to culture porcine inner ear cells provides an important investigative tool that can be utilized to study and evaluate the pathophysiological causes and cellular consequences of human inner ear disorders.

category: cell biology


title: Modeling post-logging height growth of black spruce forests by combining airborne LiDAR and historical forestry maps in eastern Canadian boreal forest

abstract: Increase in forest disturbance due to land use as well as climate change has led to an expansion of young forests worldwide, which affects global carbon dynamics and forest management. In this study, we present a novel method that combines a single airborne LiDAR acquisition and historical harvesting maps to model height growth of post-logged black spruce-dominated forests in a 1700 km2 eastern Canadian boreal landscape. We developed a random forest model where forest height is a function of stand age, combined with environmental variables. Our results highlight the strong predictive power of this model: least-square regression between predicted and observed height of our validation dataset was very close to the 1:1 relation and strongly supported by validation metrics (R2 = 0.75; relative RMSE = 19%). Moreover, our findings indicated an ecological gradient responsible for differences in height growth at the landscape scale, with better growth rates on mesic slopes compared to badly drained soils on flat lands. With the increased availability of LiDAR data, this method is promising since it can be applied to forests across the globe that are affected by stand-replacing disturbances.

category: ecology


title: An exceptionally preserved euarthropod with unique feather-like appendages from the Chengjiang biota

abstract: Micro-CT scanning of the Cambrian euarthropod Chuandianella ovata reveals unprecedented three-dimensional soft-part details. It has an elongate uniramous antennule and a short uniramous second appendage, followed by ten homonomous biramous appendages, each comprising a short paddle-shaped exopod and a unique feather-like endopod with at least 27 podomeres each of which bears a long blade-like endite with a short terminal seta. Based on its carapace and previously known soft-part anatomy C. ovata was compared with the Burgess Shale mandibulate euarthropod Waptia. However, Waptia has recently been shown to bear specialized head appendages that are interpreted as a mandible and maxillula, posterior to which are four appendages each with five-segmented endopods. In contrast, we interpret Chuandianella as an  upper stem-group euarthropod that possessed neither a differentiated mandible nor a maxillula. Chuandianella further demonstrates that early Cambrian  upper stem-group euarthropods were experimenting with a range of different limb arrangements and morphologies.

category: evolutionary biology


title: The limits of evolutionary convergence in sympatry: reproductive interference and developmental constraints leading to local diversity in aposematic signals

abstract: Mutualistic interactions between defended species represent a striking case of evolutionary convergence in sympatry, driven by the increased protection against predators brought by mimicry in warning traits. However, such convergence is often limited: sympatric defended species frequently display different or imperfectly similar warning traits. The phylogenetic distance between sympatric species may indeed prevent evolution towards the exact same signal. Moreover, warning traits are also involved in mate recognition, so that trait convergence might result in heterospecific courtship and mating. Here, we develop a mathematical model to investigate the strength and direction of evolution of warning trait in defended species with different ancestral traits. Specifically, we determine the effect of phenotypic distances between ancestral trait states of sympatric defended species and of costs of heterospecific sexual interactions on imperfect mimicry and trait divergence. Our analytical results confirm that reproductive interference and historical constraints limit the convergence of warning traits, leading to either complete divergence, or imperfect mimicry. Our model reveals that imperfect mimicry evolves only when ancestral trait values differ between species, due to historical constraints and highlights the importance of female and predator discriminations in the evolution of such imperfect mimicry. Our study thus provides new predictions on how reproductive interference interacts with historical constraints and may promote the emergence of novel warning traits, enhancing mimetic diversity.


title: The limits of evolutionary convergence in sympatry: reproductive interference and historical constraints leading to local diversity in warning traits


title: Exploring the natural origins of SARS-CoV-2

abstract:
The lack of an identifiable intermediate host species for the proximal animal ancestor of SARS-CoV-2, and the large geographical distance between Wuhan and where the closest evolutionary related coronaviruses circulating in horseshoe bats (Sarbecoviruses) have been identified, is fuelling speculation on the natural origins of SARS-CoV-2. We have comprehensively analysed phylogenetic relations between SARS-CoV-2, and the related bat and pangolin Sarbecoviruses sampled so far. Determining the likely recombination events reveals a highly reticulate evolutionary history within this group of coronaviruses. Clustering of the inferred recombination events is non-random with evidence that Spike, the main target for humoral immunity, is beside a recombination hotspot likely driving antigenic shift in the ancestry of bat Sarbecoviruses. Coupled with the geographic ranges of their hosts and the sampling locations, across southern China, and into Southeast Asia, we confirm horseshoe bats, Rhinolophus, are the likely SARS-CoV-2 progenitor reservoir species. By tracing the recombinant sequence patterns, we conclude that there has been relatively recent geographic movement and co-circulation of these viruses ancestors, extending across their bat host ranges in China and Southeast Asia over the last 100 years or so. We confirm that a direct proximal ancestor to SARS-CoV-2 is yet to be sampled, since the closest relative shared a common ancestor with SARS-CoV-2 approximately 40 years ago. Our analysis highlights the need for more wildlife sampling to (i) pinpoint the exact origins of SARS-CoV-2s animal progenitor, and (ii) survey the extent of the diversity in the related Sarbecoviruses phylogeny that present high risk for future spillover.

HighlightsO_LIThe origin of SARS-CoV-2 can be traced to horseshoe bats, genus Rhinolophus, with ranges in both China and Southeast Asia.
C_LIO_LIThe closest known relatives of SARS-CoV-2 exhibit frequent transmission among their Rhinolophus host species.
C_LIO_LISarbecoviruses have undergone extensive recombination throughout their evolutionary history.
C_LIO_LIAccounting for the mosaic patterns of these recombinants is important when inferring relatedness to SARS-CoV-2.
C_LIO_LIBreakpoint patterns are consistent with recombination hotspots in the coronavirus genome, particularly upstream of the pike open reading frame with a coldspot in S1.
C_LI


title: Exploring the natural origins of SARS-CoV-2 in the light of recombination


title: Fast but not furious: a streamlined selection method for genome edited cells

abstract:
In the last decade, Transcription Activator-Like Effector Nucleases (TALEN) and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) based genome engineering have revolutionized our approach to biology. Due to their high efficiency and ease of use, the development of custom knock-out and knock-in animal or cell models is now within reach for almost every laboratory. Nonetheless, the generation of genetically modified cells often requires a selection step, usually achieved by antibiotics or fluorescent markers. The choice of the selection marker is based on the available laboratory resources, such as cell types, and parameters like time and cost should also be taken into consideration. Here, we present a new and fast strategy called MAGECS (magnetic-activated genome edited cell sorting), to select genetically modified cells based on the ability to magnetically sort surface antigens (i.e. tCD19) present in Cas9 positive cells. By using MAGECS, we successfully generated and isolated genetically modified human induced pluripotent stem cells (hiPSCs), primary human fibroblasts, SH-SY5Y neuroblast-like cells, HaCaT and HEK293T cells.

Our strategy expands the genome editing toolbox by offering a fast, cheap, and an easy to use alternative to the available selection methods.

category: genomics


title: Performance of a scalable extraction-free RNA-seq method

abstract: RNA sequencing enables high-contents/high-complexity measurements in small molecule screens performed on biological samples. Whereas the costs of DNA sequencing and the complexity of RNA-seq library preparation and analysis have decreased consistently, RNA extraction remains a significant bottleneck for RNA-seq of hundreds of samples in parallel. Direct use of cell lysate for RNA-seq library prep is common in single cell RNA-seq but not in bulk RNA-seq protocols. Recently published protocols suggest that direct lysis is compatible with simplified RNA-seq library prep. Here, we evaluate the performance of a bulk RNA-seq library prep protocol optimized for analysis of many samples of adherent cultured cells in parallel. We combine a low-cost direct lysis buffer compatible with cDNA synthesis ("in-lysate cDNA synthesis") with Smart-3SEQ and examine the effects of calmidazolium and fludrocortisone-induced perturbation of primary human dermal fibroblasts. We compared this method to normalized purified RNA inputs from matching samples followed by Smart-3SEQ or Illumina TruSeq library prep. Our results show that whereas variable RNA inputs for each sample in the in-lysate cDNA synthesis protocol result in variable sequencing depth, this had minimal effect on data quality, measurement of gene expression patterns, or generation of differentially expressed gene lists. We found that in-lysate cDNA synthesis combined with Smart-3SEQ RNA-seq library prep allows generation of high-quality data when compared to library prep with extracted RNA, or when compared to Illumina TruSeq. Our data show that small molecule screens using RNA-seq are feasible at low reagent and time costs.


title: Abundant secreted hydrolytic enzymes and secondary metabolite gene clusters in genomes of the Botryosphaeriaceae reflect their role as important plant pathogens

abstract: The Botryosphaeriaceae are important plant pathogens, but unique in their ability to establish asymptomatic infections that persist for extended periods in a latent state. In this study, we used comparative analyses to consider elements that might shed light on the genetic basis of the interactions of these fungi with their plant hosts. For this purpose, we characterised secreted hydrolytic enzymes, secondary metabolite biosynthetic gene clusters and considered general trends in genomic architecture using all available Botryosphaeriaceae genomes, and selected Dothideomycetes genomes. The Botryosphaeriaceae genomes were rich in carbohydrate-active enzymes (CAZymes), proteases, lipases and secondary metabolic biosynthetic gene clusters (BGCs) compared to other Dothideomycete genomes. The genomes of Botryosphaeria, Macrophomina, Lasiodiplodia and Neofusicoccum, in particular, had gene expansions of the major constituents of the secretome, notably CAZymes involved in plant cell wall degradation. The Botryosphaeriaceae genomes were shown to have moderate to high GC contents and most had low levels of repetitive DNA. The genomes were not compartmentalized based on gene and repeat densities, but genes of secreted enzymes were slightly more abundant in gene-sparse regions. The abundance of secreted hydrolytic enzymes and secondary metabolite BGCs in the genomes of Botryosphaeria, Macrophomina, Lasiodiplodia, and Neofusicoccum were similar to those in necrotrophic plant pathogens, but also endophytes of woody plants. The results provide a foundation for future comparative genomic analyses and hypothesis to explore the mechanisms underlying Botryosphaeriaceae host-plant interactions.


title: Identification of 13 independent genetic loci associated with cognitive resilience in healthy aging in 330,097 individuals in the UK Biobank.

abstract: Cognitive resilience is the ability to withstand the negative effects of stress on cognitive functioning and is important for maintaining quality of life while aging. UK Biobank (UKB) does not have direct measurements of the same cognitive phenotype at distal timepoints. Therefore, we used number of education years (EY) as a proxy phenotype for past cognitive performance. Current cognitive performance was determined based on processing speed. This approach captured an average time span of 40 years between past and current cognitive performance in 330,097 individuals. A confounding factor was that EY is highly polygenic and masked the genetics of resilience. To overcome this, we employed Genomics Structural Equation Modelling (GenomicSEM) to perform a GWAS-by-subtraction using two GWAS, one GWAS of EY and resilience and a second GWAS of EY but not resilience. Subtracting one from the other generated a GWAS of Resilience. Replication of this approach was shown using independent discovery and replication samples within UKB and the full UKB GWAS results show significant genetic correlation with a GWAS of cognitive change in the independent Health and Retirement Study (N=9,526; P=1.5x10-3). We found 13 independent genetic loci for Resilience. Functional analyses showed enrichment in several brain regions and involvement of specific cell types, including GABAergic neurons (P=6.59x10-8) and glutamatergic neurons (P=6.98x10-6) in the cortex. Gene-set analyses implicated the biological process "neuron differentiation" (P=9.7x10-7) and the cellular component "synaptic part" (P=2.14x10-6). Mendelian randomization analysis showed a causative effect of white matter volume on cognitive resilience. These results enhance neurobiological understanding of resilience.


title: Thirteen independent genetic loci associated with preserved processing speed in a study of cognitive resilience in 330,097 individuals in the UK Biobank


title: Quantifying the impact of electric fields on single-cell motility

abstract:
Cell motility in response to environmental cues forms the basis of many developmental processes in multicellular organisms. One such environmental cue is an electric field (EF), which induces a form of motility known as electrotaxis. Electrotaxis has evolved in a number of cell types to guide wound healing, and has been associated with different cellular processes, suggesting that observed electrotactic behaviour is likely a combination of multiple distinct effects arising from the presence of an EF. In order to determine the different mechanisms by which observed electrotactic behaviour emerges, and thus to design EFs that can be applied to direct and control electrotaxis, researchers require accurate quantitative predictions of cellular responses to externally-applied fields. Here, we use mathematical modelling to formulate and parametrise a variety of hypothetical descriptions of how cell motility may change in response to an EF. We calibrate our model to observed data using synthetic likelihoods and Bayesian sequential learning techniques, and demonstrate that EFs bias cellular motility through only one of a selection of hypothetical mechanisms. We also demonstrate how the model allows us to make predictions about cellular motility under different EFs. The resulting model and calibration methodology will thus form the basis for future data-driven and model-based feedback control strategies based on electric actuation.

SIGNIFICANCEElectrotaxis is attracting much interest and development as a technique to control cell migration due to the precision of electric fields as actuation signals. However, precise control of electrotactic migration relies on an accurate model of how cell motility changes in response to applied electric fields. We present and calibrate a parametrised stochastic model that accurately replicates experimental single-cell data and enables the prediction of input-output behaviour while quantifying uncertainty and stochasticity. The model allows us to elucidate and quantify how electric fields perturb the motile behaviour of the cell. This model and the associated simulation-based calibration methodology will be central to future developments in the control of electrotaxis.

category: biophysics


title: MYCN-induced nucleolar stress drives an early senescence-like transcriptional program in hTERT-immortalized RPE cells

abstract:
MYCN is an oncogenic driver in neural crest-derived neuroblastoma and medulloblastoma. To better understand the early effects of MYCN activation in a neural-crest lineage context, we profiled the transcriptome of immortalized human retina pigment epithelial cells with inducible MYCN activation. Gene signatures associated with elevated MYC/MYCN activity were induced after 24 h of MYCN activation, which attenuated but sustained at later time points. Unexpectedly, MYCN activation was accompanied by reduced cell growth. Gene set enrichment analysis revealed a senescence-like signature with strong induction of p53 and p21 but in the absence of canonical hallmarks of senescence such as beta-galactosidase positivity, suggesting incomplete cell fate commitment. When scrutinizing the putative drivers of this growth attenuation, differential gene expression analysis identified several regulators of nucleolar stress. This process was also reflected by phenotypic correlates such as cytoplasmic granule accrual and nucleolar coalescence. Hence, we propose that the induction of MYCN congests the translational machinery, causing nucleolar stress and driving cells into a transient pre-senescent state. Our findings shed new light on the early events induced by MYCN activation and may help unravelling which factors are required for cells to tolerate unscheduled MYCN overexpression during early malignant transformation.

HighlightsO_LIActivation of MYCN attenuates proliferation in RPE1 cells
C_LIO_LIGrowth arrest is associated with an early senescence-like transcriptional signature
C_LIO_LITranscriptional and phenotypic evidence of nucleolar stress
C_LIO_LICDKN1A upregulation in G2 phase primes cells for faltering in subsequent G1
C_LI


title: Direct imaging of anthrax intoxication in animals reveals shared and individual functions of CMG-2 and TEM-8 in cellular toxin entry

abstract:
The virulence of Bacillus anthracis is linked to the secretion of anthrax lethal toxin and anthrax edema toxin. These binary toxins consist of a common cell-binding moiety, protective antigen (PA), and the enzymatic moieties, lethal factor (LF) and edema factor (EF). PA binds either of two specific cell surface receptors, capillary morphogenesis protein-2 (CMG-2) or tumor endothelial marker-8 (TEM-8), which triggers the binding, endocytosis, and cytoplasmic translocation of LF and EF. The cellular distribution of functional TEM-8 and CMG-2 receptors during anthrax toxin intoxication in animals is not fully elucidated. Herein, we describe a novel assay to image anthrax toxin intoxication in live animals, and we use the assay to visualize TEM-8- and CMG-2-dependent intoxication. Specifically, we generated a chimeric protein consisting of the N-terminal domain of LF fused to a nuclear localization signal-tagged Cre recombinase (LFn-NLS-Cre). When PA and LFn-NLS-Cre were co-administered to transgenic mice that ubiquitously express a red fluorescent protein in the absence of Cre activity and a green fluorescent protein in the presence of Cre activity, anthrax toxin intoxication could be visualized at single-cell resolution by confocal microscopy. By using this assay, we show that CMG-2 is critical for intoxication in the liver and heart, whereas TEM-8 is required for full intoxication in the kidney and spleen. Other tissues examined were largely unaffected by single deficiences in either receptor, suggesting extensive overlap in TEM-8 and CMG-2 expression. The novel assay will be useful for basic and clinical/translational studies of Bacillus anthracis infection and for identifying on- and off-targets for reengineered toxin variants in the clinical development of cancer treatments.

BackgroundAssays for imaging of anthrax toxin intoxication in animals are not available.

ResultsAnthrax toxin-Cre fusions combined with fluorescent Cre reporter mice enabled imaging of anthrax toxin intoxication in animals.

ConclusionShared and distinct functions of toxin receptors in cellular entry were uncovered. Significance. A simple and versatile assay for anthrax toxin intoxication is described.

category: pathology


title: Identification of a filamentous form of kunitz protease inhibitor in Asteraceae

abstract: Filamentous structures were observed in purified extracts from chrysanthemum, gerbera, sunflower and zinnia. When purified filament proteins were subjected to SDS-PAGE, the major protein associated with filaments from all three species has an apparent molecular mass of {approx}25 kDa. Protein bands from chrysanthemum, gerbera, and zinnia were subjected to N-terminal protein sequencing while proteins from sunflower were sequenced by CID MS/MS. All of the sequences shared highest similarity to the kunitz trypsin inhibitor family. The sequencing results indicated that the proteins lacked the signal sequences. We tested the gerbera filament protein for glycosylation and found that it was a glycoprotein. Together these results indicate that the filaments are composed of mature KTI protein. This is the first report of a KTI assembling into filaments and the first report of a filament forming Asteraceae enzyme.


title: Prioritizing and characterizing functionally relevant genes across human tissues

abstract: Knowledge of genes that are critical to a tissues function remains difficult to ascertain and presents a major bottleneck toward a mechanistic understanding of genotype-phenotype links. Here, we present the first machine learning model - FUGUE - combining transcriptional and network features, to predict tissue-relevant genes across 30 human tissues. FUGUE achieves an average cross-validation auROC of 0.86 and auPRC of 0.50 (expected 0.09). In three independent datasets, FUGUE accurately distinguishes tissue or cell type-specific genes, significantly outperforming the conventional metric based on tissue-specific expression alone. Comparison of tissue-relevant transcription factors across tissue recapitulate their developmental relationships. Interestingly, the tissue-relevant genes cluster on the genome within topologically associated domains and furthermore, are highly enriched for differentially expressed genes in the corresponding cancer type. We provide the prioritized gene lists in 30 human tissues and an open-source software to prioritize genes in a novel context given multi-sample transcriptomic data.

category: systems biology


title: Bacterial galactosemia is caused by cytoplasmic interference of an essential cell wall biosynthesis glycosyltransferase

abstract:
Bacterial galactosemia or "galactose death," triggered by incomplete galactose metabolism, was first discovered in Escherichia coli and Salmonella six decades ago, and later in many other microorganisms, yet the mechanism for the toxicity and subsequent cell death remains unclear. In Bacillus subtilis, galactosemia is manifested by a buildup of uridine-diphosphate-galactose (UDP-Gal) and a strong toxicity phenotype characterized by cell shape abnormality and rapid cell lysis. Here we present evidence that in B. subtilis, the toxicity is due to inhibition of cell wall biosynthesis through interference of the essential glycosyltransferase MurG that carries out lipid II synthesis from lipid I and uridine-diphosphate-N-acetyl-glucosamine (UDP-GlcNAc). Single-molecule imaging reveals real-time inhibition of cell wall biosynthesis and MurG activities in cells exhibiting toxicity. We further show that in vitro, MurG is able to utilize UDP-Gal as a substrate generating a "toxic" lipid II, causing a potential poisoning effect on peptidoglycan crosslinking. Evidence also suggests a similar mechanism in Vibrio cholerae and Staphylococcus aureus. Lastly, a strong synergistic lethality was seen in S. aureus wild-type cells treated with both galactose and sub-lethal doses of cell-wall antibiotics. Our study provides mechanistic explanation of the toxicity associated with bacterial galactosemia and its potential application in antibacterial solutions.

SignificanceGalactosemia is a potentially fatal genetic disorder due to incomplete galactose metabolism, found in both eukaryotic and prokaryotic organisms. The molecular mechanisms of galactosemia-associated toxicity remain unclear in all cases. Here we present evidence that in the bacterium Bacillus subtilis, the toxicity is due to interference of an essential glycosyltransferase, MurG, which concerts lipid I to lipid II during peptidoglycan biosynthesis, by a nucleotide sugar derived from galactose metabolism. This interference leads to a halt of cell wall biosynthesis and structural defects causing rapid cell lysis. Our evidence also suggests a similar mechanism in other bacteria such as Staphylococcus aureus and Vibrio cholerae. Our study may help solve the long-time puzzle of bacterial galactosemia first uncovered six decades ago.


title: Quantifying cell transitions in C. elegans with data-fitted landscape models

abstract: Increasing interest has emerged in new mathematical approaches that simplify the study of complex differentiation processes by formalizing Waddingtons landscape metaphor. However, a rational method to build these landscape models remains an open problem. Building on pioneering work by Corson and Siggia (2012, 2017) we study vulval development in C. elegans by developing a framework based on Catastrophe Theory (CT) and approximate Bayesian computation (ABC) to build data-fitted landscape models. We first identify the candidate qualitative landscapes, and then use CT to build the simplest model consistent with the data, which we quantitatively fit using ABC. The resulting model suggests that the underlying mechanism is a quantifiable two-step decision controlled by EGF and Notch-Delta signals, where a non-vulval/vulval decision is followed by a bistable transition to the two vulval states. This new model fits a broad set of data and makes several novel predictions.

category: developmental biology


title: Quantifying and cataloguing unknown sequences within human microbiomes

abstract: Advances in genome sequencing technologies and lower costs have enabled the exploration of a multitude of known and novel environments and microbiomes. This has led to an exponential growth in the raw sequence data that is deposited in online repositories. Metagenomic and metatranscriptomic data sets are typically analysed with regards to a specific biological question. However, it is widely acknowledged that these data sets are comprised of a proportion of sequences that bear no similarity to any currently known biological sequence, and this so-called  dark matter is often excluded from downstream analyses. In this study, a systematic framework was developed to assemble, identify, and measure the proportion of unknown sequences present in distinct human microbiomes. This framework was applied to forty distinct studies, comprising 963 samples, and covering ten different human microbiomes including fecal, oral, lung, skin and circulatory system microbiomes. The framework was used to determine the proportion of taxonomically unknown sequences present within samples, and to compare such sequences both within and across assembled metagenomes. We found that whilst the human microbiome is one of the most extensively studied, on average 2% of assembled sequences have not yet been taxonomically defined. However, this proportion varied extensively among different microbiomes and was as high as 25% for skin and oral microbiomes that have more interactions with the environment. The publicly available data sets used have not previously been systematically mined to quantify and compare such dark matter. Typically, these unknown sequences are found in several microbiomes and potentially belong to unidentified novel microbes that we interact with on a daily basis. A cross-study comparison led to the identification of similar unknown sequences in different samples and/or microbiomes. A rate of taxonomic characterisation of 1.64% of unknown sequences being characterised per month was calculated from these taxonomically unknown sequences discovered in this study. Additionally, the approach led to the discovery of several potentially novel viral genomes that bear no similarity to sequences in the public databases. Both our computational framework and the novel unknown sequences produced are publicly available for future cross-referencing.

category: bioinformatics


title: Characterisation of the RNA-interference pathway as a Tool for Genetics in the Nascent Phototrophic Endosymbiosis, Paramecium bursaria

abstract: Endosymbiosis was fundamental for the evolution of eukaryotic complexity. Endosymbiotic interactions can be dissected through forward and reverse-genetic experiments, such as RNA-interference (RNAi). However, distinguishing small (s)RNA pathways in a eukaryote-eukaryote endosymbiotic interaction is challenging. Here, we investigate the repertoire of RNAi pathway protein-encoding genes in the model nascent endosymbiotic system, Paramecium bursaria-Chlorella spp. Using comparative genomics and transcriptomics supported by phylogentics, we identify essential proteome components of the small interfering (si)RNA, scan (scn)RNA, and internal eliminated sequence (ies)RNA pathways. Our analyses reveal that copies of these components have been retained throughout successive whole genome duplication (WGD) events in the Paramecium clade. We then validate feeding-induced siRNA-based RNAi in P. bursaria via knock-down of the splicing factor, u2af1, which we show to be crucial to host growth. Finally, using simultaneous knock-down paradox controls to rescue the effect u2af1 knock-down, we demonstrate that feeding-induced RNAi in P. bursaria is dependent upon a core pathway of host-encoded Dcr1, Piwi and Pds1 components. Our experiments confirm the presence of a functional, host-derived RNAi pathway in P. bursaria that generates 23-nt siRNA, validating use of the P. bursaria-Chlorella spp. system to investigate the genetic basis of a nascent endosymbiosis.

category: molecular biology


title: Identification of genes associated with productivity traits and salinity tolerance from activation tagged lines of rice

abstract:
World-wide crop productivity is hugely impacted by diverse eco-environmental conditions. In the present investigation, activation tagged (AT) lines of rice endowed with improved agronomic attributes have been analyzed for tolerance to salinity stress besides identification of genes associated with these attributes. Under salinity stress conditions, AT lines exhibited increased seed germination rates, improved plant growth and development at vegetative and reproductive stages as compared to wild-type (WT) plants. Furthermore, AT lines disclosed enhanced plant water content, photosynthetic efficiency, stomatal conductance, water use efficiency and maximum quantum yield when compared to WT plants, leading to improved yields and delayed onset of stress symptoms. Moreover, AT lines revealed effective antioxidant systems causing decreased accumulation of reactive oxygen species and delayed salinity stress symptoms compared to WT plants. Reduced accumulation of malondialdehyde with concomitant increases in proline and soluble sugars of AT lines further endorsing their improved stress tolerance levels. TAIL and qRT-PCR analyses of AT lines revealed Ds element integrations at different loci and respective overexpression of identified candidate genes involved in various aspects of plant development and stress tolerance. Accordingly, the AT lines plausibly serve as a rare genetic resource for fortifying stress tolerance and productivity traits of elite rice cultivars.

HighlightActivation tagged lines of rice endowed with improved agronomic attributes have been analyzed for tolerance to salinity stress besides identification and expression analysis of genes associated with these attributes.


title: Human genetic analyses of organelles highlight the nucleus in age-related trait heritability

abstract: Most age-related human diseases are accompanied by a decline in cellular organelle integrity, including impaired lysosomal proteostasis and defective mitochondrial oxidative phosphorylation. An open question, however, is the degree to which inherited variation in or near genes encoding each organelle contributes to age-related disease pathogenesis. Here, we evaluate if genetic loci encoding organelle proteomes confer greater-than-expected age-related disease risk. As mitochondrial dysfunction is a "hallmark" of aging, we begin by assessing nuclear and mitochondrial DNA loci near genes encoding the mitochondrial proteome and surprisingly observe a lack of enrichment across 24 age-related traits. Within nine other organelles, we find no enrichment with one exception: the nucleus, where enrichment emanates from nuclear transcription factors. In agreement, we find that genes encoding several organelles tend to be "haplosufficient," while we observe strong purifying selection against heterozygous protein-truncating variants impacting the nucleus. Our work identifies common variation near transcription factors as having outsize influence on age-related trait risk, motivating future efforts to determine if and how this inherited variation then contributes to observed age-related organelle deterioration.


title: Human genetic analyses of organelles highlight the nucleus, but not the mitochondrion, in age-related trait heritability


title: Historical disturbances to rivers appear to constrain contemporary distribution of a river-dependent frog,

abstract: Frogs dependent on lotic environments are sensitive to disturbances that alter the hydrology (e.g., water impoundments), substrate (e.g., debris torrents), and riparian vegetation (e.g., wildfires) of river ecosystems. Although rivers are often very dynamic, disturbances can push environmental baselines outside of narrowly defined ecological tolerances under which a species evolved. Short-lived lotic-dependent organisms, restricted to movements within the water or the riparian corridor, are at risk of local extirpations owing to such disturbances if they fragment and isolate affected populations from recolonizing source populations. In Oregon, USA, the foothill yellow-legged frog (Rana boylii) is at its northernmost range margin and has experienced an approximately 41% range contraction compared to their historical distribution. To inform conservation and management, we used species distribution models to identify environmentally suitable watersheds based on intrinsic baseline environmental variables, and then examined potential effects of human-caused alterations to rivers, including splash dams used to ferry timber downstream prior to 1957, large water impoundments, and adjacency to agricultural croplands. We used machine-learning in program Maxent and three different river layers that varied in extent and location of mapped rivers but contained distinct information to produce species distribution models which we then combined into a single ensemble model. Stream order, annual precipitation, and precipitation frequency were the highest ranked baseline environmental variables in most models. Watersheds with highly suitable baseline conditions in our ensemble model were negatively correlated with anthropogenic disturbances to rivers. Foothill yellow-legged frogs appeared to be sensitive to human-caused disturbances to rivers, perhaps indicative of their narrow ecological tolerance to in-river conditions. We do not anticipate variables in our model to change much through time. Rather, for conservation we identified potential legacy (spash dams) and ongoing human-caused disturbances that are more likely to change conditions for the species in the short- and long-term.


title: Effect of landscape heterogeneity on diurnal raptor community richness and diversity in Jammu Shiwaliks, Jammu and Kashmir

abstract: In this study, we examined the richness and diversity of diurnal raptors along with their foraging guilds across different land uses in a sub-tropical landscape during December 2016 to November 2018. A total of 80 vantage points, 19 line transects and 36 road transects were sampled in 33 sites in six different habitat types in the study area where we recorded 3409 individuals of 29 diurnal raptors in 2 orders and 3 families. Significant variation in bird abundance was observed among different habitat types, farmlands being more specious followed by pure forests, water bodies and forest-farmland interfaces. Among the seasons, summers recorded higher abundance followed by winter, monsoon and post-monsoon. A low diversity value (H'=2.22) however was observed for the whole study area with mean monthly highest recorded during February (H'=2.44) and least during June (H'=1.85). Most of the raptors observed for their food types and foraging were predators (n=22) and rest were carrion feeders (n=22). Fourteen among all observed diurnal raptors were winter visitors and 13 residents with 9 reported globally threatened. A moderately high richness of diurnal raptors substantiate high conservation value of these habitats especially the forest patches and farmlands and thus calls for effective management strategies for the conservation and proliferation of raptors in sub-tropical areas of Jammu region.


title: Risk and protective factors of feline tooth resorption in 8115 Finnish cats

abstract: Tooth resorption (TR) is one of the most common dental diseases in cats. It is a painful disease characterised by progressive dental destruction, which eventually results in loss of teeth. The aetiology of the disease remains unclear, but associations with old age, breed, other oral and dental diseases, and certain environmental factors have been suspected. We collected health and environmental information of 8115 Finnish cats from 41 breeds through an online survey for cat owners to study TR in Finland with an observational study. The frequency of TR that the owner reported as veterinarian-diagnosed was 3.9% in the entire data (316/8115) and 15% in cats that the owner reported as having veterinarian-diagnosed oral or dental disease (316/2070). We determined the risk factors and breed variation of feline TR by logistic regression with the entire data and a case-control study in a more defined population drawn from the entire data. In the entire data, we defined those cats that the owners had reported as being diagnosed with TR by a veterinarian as TR-cats and compared them to all other cats. In the case-control study, we included only cats with owner-reported veterinarian-diagnosed oral and dental diseases. We defined cats with TR-diagnosis made under sedation as TR-cases. Other veterinarian-diagnosed TR-cats were excluded. Cats with other oral and dental diseases were used as controls. The model with the entire population included 308 TR-cats compared with 7508 cats, and the case-control study included 198 TR-cats and 1683 controls. Although the diagnoses were reportedly made by a veterinarian, the owner answered the survey and might have understood, remembered, or interpreted the diagnoses in a way that could affect the results. The results indicated an increasing risk of TR with age. Dental calculus, gingivitis, and periodontitis were potential risk factors for TR. These findings and the interaction between dental calculus and gingivitis suggest that inflammatory changes caused by dental calculus increase the risk of TR. Keeping food available constantly was a potential protective factor. We found that Cornish Rex, European, and Ragdoll breeds were at higher risk for TR. The observed differences between breeds highlight a genetic contribution.


title: Feline tooth resorption findings from a questionnaire survey on 8115 Finnish cats and a case-control study for a more defined population drawn from the survey


title: Inspiratory muscle training for enhancing repeated-sprint ability: A pilot study

abstract: This pilot study examined the effect of inspiratory muscle training (IMT) on repeated-sprint ability and vastus lateralis reoxygenation. Ten recreationally trained subjects were randomly divided into two groups to complete 4 weeks of IMT or Sham (placebo) training. Pre- and post-intervention, a repeated-sprint ability (RSA) test was performed in both normoxia and hypoxia (FiO2 {approx} 14.5%). Vastus lateralis reoxygenation (VLreoxy), defined as peak to minimum amplitude deoxyhaemoglobin for each sprint/recovery cycle, was assessed during all trials using near-infrared spectroscopy. For total work performed, power analysis revealed that for small, medium and large effects (Cohens f), sample sizes of n = 8, 16 and 90 respectively, are required to achieve a power of 80% at an  level of 0.05. Maximal inspiratory mouth pressure increased in IMT by 36.5%, 95% CI [20.9, 61.6] and by 2.7%, 95% CI [-4.46, 8.8] in Sham. No clear difference in the change of work completed during the sprints between groups were observed in normoxia (Sham -0.805 kJ, 95% CI [-3.92, 0.39]; IMT -2.06 kJ, 95% CI [-11.5, 4.96]; P = 0.802), or hypoxia (Sham -3.09 kJ, 95% CI [-7, 0.396]; IMT 0.354 kJ, 95% CI [-1.49, 2.1]; P = 0.802). VLreoxy in IMT increased by 9.34%, 95% CI [5.15, 13.7] in normoxia only. In conclusion, despite a large increase in IMT, this was only associated with a small effect on RSA in our pilot study cohort. Owing to a potentially relevant impact of training the inspiratory musculature, future studies should include a sample size of at least 16-20 to detect moderate to large effects on RSA.


title: High variation in immune responses and parasite phenotypes in naturally acquired Trypanosoma cruzi infection in a captive non-human primate breeding colony in Texas, USA

abstract:
Trypanosoma cruzi, the causative agent of human Chagas disease, is endemic to the southern region of the United States where it routinely infects many host species. The indoor/outdoor housing configuration used in many non-human primate research and breeding facilities in the southern U.S. provides the opportunity for infection by T. cruzi and thus provides source material for in-depth investigation of host and parasite dynamics in a natural host species under highly controlled and restricted conditions. For cynomolgus macaques housed at such a facility, we used a combination of serial blood quantitative PCR (qPCR) and hemoculture to confirm infection in >92% of seropositive animals, although each method alone failed to detect infection in >20% of cases. Parasite isolates obtained from 43 of the 64 seropositive macaques were of 2 broad genetic types (discrete typing units, (DTUs) I and IV); both within and between these DTU groupings, isolates displayed a wide variation in growth characteristics and virulence, elicited host immune responses, and susceptibility to drug treatment in a mouse model. Likewise, the macaques displayed a diversity in T cell and antibody response profiles that rarely correlated with parasite DTU type, length of infection, or age of the primate. This study reveals the complexity of infection dynamics, parasite phenotypes, and immune response patterns that can occur in a primate group, despite being housed in a uniform environment at a single location, and the limited time period over which the T. cruzi infections were established.

Author SummaryWe evaluated naturally occurring infections of Trypanosoma cruzi, the causative agent of human Chagas disease, in an indoor/outdoor primate colony at a breeding facility in Texas, USA. Using serial quantitative PCR and hemoculture, we confirmed infection in 92% of the 64 seropositive animals, but neither of these two methods confirmed more than 80% of the cases. Parasites by hemoculture fell into two genetic groups (discrete typing units I and IV), and displayed large variation in growth characteristics, elicited cellular and humoral immune responses as well as virulence and drug susceptibility when tested in mice. EKG abnormalities were found in 13 out of 51 qPCR-positive macaques. Our results demonstrate the complexity of these infection parameters in this colony in spite of the uniform and geographically constrained housing conditions of the macaques.


title: Forecasting the future risk of dengue epidemics facing climate change in New Caledonia, South Pacific.

abstract:
BackgroundDengue dynamics result from the complex interaction between the virus, the host and the vector, all being under the influence of the environment. Several studies have explored the link between climate and dengue outbreaks in New Caledonia. None of them have explored the evolution of the dengue outbreak risk facing climate change.

Methodology/Principal FindingsIn this study we chose the threshold time dependent reproduction number (Rt) as the modeling target to focus on time frames suitable for outbreak growths. A weekly statistical model of dengue outbreak risk (i.e. dengue outbreak probability) based on climate variables was developed using support vector machines (SVM) and then used in combination with CMIP5 projections of rainfall and temperature to estimate the future evolution of seasonal weekly risk and the inter-annual yearly risk of dengue outbreak up to the year 2100. The weekly risk of dengue outbreak is estimated using the number of days with maximal temperature exceeding 30.8{degrees}C during 80 days preceding the predicted week and the mean of precipitation during 60 days preceding the predicted week. According to the SVM model and to the worst greenhouse gas emission scenario projection (RCP8.5), the time frame suitable for epidemic growth will gain one month starting in November instead of December and the yearly risk of dengue outbreak occurrence increases regularly up to 2100 and reach a probability of 1 around 2080, making the dynamic of dengue fever endemic in New Caledonia.

Conclusions/SignificanceA complete method to assess seasonal and inter annual variability of the risk of dengue outbreaks with respect to climate change is proposed. We conclude that climate change is likely to increase the risk of dengue in New-Caledonia (the other non climatic parameters remaining constant) in terms of both frequency of outbreak and temporal spread of the outbreak.

Author summaryDengue virus is transmitted to human through the bite of an Aedes mosquito vector. Dengue fever is a worldwide public health concern, especially on tropical and subtropical countries. Over the last decade, the toll of dengue fever has increased in New Caledonia, raising questions about the future of the disease in this French island territory located in the South Pacific. Climate has a strong influence on dengue through its influence on the ecology of the vector and the viral cycle. Several studies have explored the link between climate and dengue in New Caledonia, with the aim of explaining and predicting dengue outbreaks. None of these studies have explored the possible outcome climate change will have on the risk of dengue fever in New Caledonia. This is the goal of this study, through projections of rainfall and temperature and the selection of an appropriate prediction target for our statistical model, we assess the climate-induced risk of dengue outbreaks up to the 2100 horizon. We prove that the inter-annual risk of dengue outbreaks in New Caledonia will raise, according to all the greenhouse gas emission scenarios and according to the high emission scenario, dengue fever will become an endemic disease in New Caledonia.


title: Effect of hydroxyapatite microspheres, amoxicillin-hydroxyapatite and collagen-hydroxyapatite composites on human dental pulp-derived mesenchymal stem cells

abstract: The aim of this study was to evaluate the in vitro behavior of human dental pulp mesenchymal stem cells (hDPSCs) cultured on scaffolds of three hydroxyapatite-based materials: hydroxyapatite microspheres [HAp]; amoxicillin-hydroxyapatite composite [Amx-HAp]; and collagen-hydroxyapatite composite [Col-HAp]. These hydroxyapatites (HAps) were synthesized through three methods: microwave hydrothermal, hydrothermal reactor (teflon pouches), and precipitation, respectively. We performed an in vitro experimental study using dental pulp stem cells obtained from samples of third molars and characterized by immunophenotypic analysis. Cells were cultured on scaffolds with osteogenic differentiation medium and were maintained for 21 days. Cytotoxicity analysis and migration assay of hDPSCs were evaluated. Each experiment was performed in triplicate. Data analysis was performed using Kruskal-Wallis test and Dunns post-hoc test. After 21 days of induction, no differences in genes expression were observed. hDPSCs highly expressed the collagen IA and the osteonectin at the mRNA, which indicated these genes plays an important role in odontogenesis regardless of induction stimulus. Cytotoxicity assay using hDPSCs demonstrated that Col-HAp group presented a number of non-viable cells statistically lower than the control group (p=0.03). In the migration assay after 24h, biomaterials HAp, Amx-HAp, and Col-HAp revealed the same migration behavior for hDPSCs observed to the positive control. Col-HAp also provided a statistically significant higher migration of hDPSCs than HAp (p=0.02). The migration results in 48h for HAp, Amx-HAp, and Col-HAp was intermediate from those achieved by control groups. There was no statistical difference between positive control and Col-HAp (p>0.05). In general, Col-HAp scaffold showed better features for these dynamic parameters of cell viability and cell migration capacities for hDPSCs, leading to suitable adhesion, proliferation, and differentiation of this osteogenic lineage. These data present high clinical importance because Col-HAp can be used in a wide variety of therapeutic areas, including ridge preservation, minor bone augmentation, and periodontal regeneration.


title: Coral Reefs of the Perhentian Islands: Abundance Levels and Distribution Patterns of Hosting Sea Anemones, and Implications for Citizen-Science Monitoring.

abstract: Around the Perhentian Islands, coral reefs have been undergoing significant degradation, as is reported annually through citizen-science coral reef monitoring programmes. Typical methodologies included in citizen-science surveys overlook the hosting sea anemones contribution to lowered live coral cover (LCC) on reef sites. As sea anemones compete with corals for suitable substrate, nutrients, and light availability, the current study was designed to investigate hosting sea anemone abundance and distribution patterns independently. In addition to identifying and assessing sites of localised sea anemone dominance, sea anemones were examined to identify factors that positively influence reproductive success and growth rate, including formations, hosting status, and resident Amphiprion species. In total, 379 sea anemone samples were analysed over 1600 m2 of Perhentian reef, making this study the first known independent investigation into sea anemone abundance patterns around the Perhentian Islands. Statistical analysis revealed that at site Village Reef, sea anemone abundance was higher than would be expected in healthy reef settings, and sea anemone cover was negatively impacting LCC. At second research site Teluk Keke, such analysis resulted in marginally insignificant results, potentially due to temporal onset differences or pressures exerted by other coral competitors. The most dominant sea anemone species, Heteractis magnifica (N=352), was analysed to test whether factors linked to higher reproductive success and growth rate were present in larger sea anemones. Results corroborate that larger sea anemones were significantly more likely to be actively hosting and were more often encountered in cluster formations. The current study employed a citizen-science method to explore its feasibility as a survey tool to identify reef sites undergoing dominance shifts, such as could be further developed and incorporated into currently used citizen-science monitoring programmes.


title: Dopamine transporter is a master regulator of dopaminergic neural network connectivity

abstract: Dopaminergic neurons of the substantia nigra (SNC) and ventral tegmental area (VTA) exhibit spontaneous firing activity. The dopaminergic neurons in these regions have been shown to exhibit differential sensitivity to neuronal loss and psychostimulants targeting dopamine transporter. However, it remains unclear whether these regional differences scale beyond individual neuronal activity to regional neuronal networks. Here we utilized live-cell calcium imaging to show that network connectivity greatly differs between SNC and VTA regions with higher incidence of hub-like neurons in the VTA. Specifically, the frequency of hub-like neurons was significantly lower in SNC dopamine neurons than in the adjacent VTA, consistent with the interpretation of a lower network resilience to SNC neuronal loss. We tested this hypothesis when activity of an individual dopaminergic neuron is suppressed, through whole-cell patch clamp electrophysiology, in either SNC, or VTA networks. Neuronal loss in the SNC decreased network clustering, whereas the larger number of hub-neurons in the VTA overcompensated by increasing network clustering in the VTA. We further show that network properties are regulatable via a dopamine transporter but not a D2 receptor dependent mechanism. Our results demonstrate novel regulatory mechanisms of functional network topology in dopaminergic brain regions.


title: Category-specific item encoding in the medial temporal lobe and beyond: The role of reward.

abstract: Forming new memories is a fundamental part of human life, and the medial temporal lobe (MTL) is central to memory formation. Recent research suggests that within MTL, the perirhinal and parahippocampal cortices (PRC, PHC) process object and scene memory, respectively, whereas the hippocampus (HC) is agnostic to stimulus category. It is unclear, however, whether MTL category specificity extends to item encoding. Furthermore, MTL does not act in isolation: Reward-related memories are formed in interplay with the dopaminergic midbrain (substantia nigra/ventral tegmental area, SNVTA) and amygdala (AMY), but it is unclear whether reward modulates neural item encoding in a category-specific way. To address these questions, we had 39 healthy volunteers (27 for all memory-based analyses) undergo functional magnetic resonance imaging while they solved an incidental encoding task, which paired objects or scenes with high or low reward, followed by a next-day surprise recognition test. Behaviourally, high reward preferably enhanced object memory. Importantly, neural activity in PRC and PHC reflected item encoding of objects and scenes, respectively. Moreover, AMY encoding effects were selective for high-reward objects, with a similar pattern in PRC. SNVTA and HC showed no clear evidence of item encoding. The behavioural and neural asymmetry of reward-related encoding effects may be conveyed through an anterior-temporal memory system, including AMY and PRC, potentially in interplay with the ventromedial prefrontal cortex (vmPFC).


title: Genetically independent phenotype analysis identifies LPA and VCAM1 as drug targets for human ageing

abstract: The length and quality of life is important to us all, yet identification of promising drug targets for human ageing using genetics has had limited success. Here, we combine six large European-ancestry genome-wide association studies (GWAS) of human ageing traits--healthspan, father and mother lifespan, exceptional longevity, frailty index, and self-rated health--in a principal component framework that maximises their shared genetic architecture. The first principal component (GIP1) is more heritable than the original studies and shows strong genetic correlations with length of life as well as multiple indices of mental and physical wellbeing. We identify 27 genomic regions associated with GIP1, and provide additional, independent evidence for an effect on human ageing for loci near HTT and MAML3 using a study of Finnish and Japanese subject survival. Across the genome, GIP1 associations are enriched in genes involved in haem metabolism and pathways related to transcription, neurogenesis, homeostasis, proteolysis, intracellular signalling, immunity, and the muscle system. Finally, using proteome-wide two-sample Mendelian randomisation and colocalisation, we provide robust evidence for a detrimental effect of blood levels of apolipoprotein(a) (LPA) and vascular cell adhesion molecule 1 (VCAM1) on GIP1. Together, our results demonstrate that combining multiple ageing traits using genetic principal components enhances power to detect biological targets for human ageing.


title: Brain Connectivity meets Reservoir Computing

abstract:
The connectivity of Artificial Neural Networks (ANNs) is different from the one observed in Biological Neural Networks (BNNs). Can the wiring of actual brains help improve ANNs architectures? Can we learn from ANNs about what network features support computation in the brain when solving a task?

ANNs architectures are carefully engineered and have crucial importance in many recent performance improvements. On the other hand, BNNs exhibit complex emergent connectivity patterns. At the individual level, BNNs connectivity results from brain development and plasticity processes, while at the species level, adaptive reconfigurations during evolution also play a major role shaping connectivity.

Ubiquitous features of brain connectivity have been identified in recent years, but their role in the brains ability to perform concrete computations remains poorly understood. Computational neuroscience studies reveal the influence of specific brain connectivity features only on abstract dynamical properties, although the implications of real brain networks topologies on machine learning or cognitive tasks have been barely explored.

Here we present a cross-species study with a hybrid approach integrating real brain connectomes and Bio-Echo State Networks, which we use to solve concrete memory tasks, allowing us to probe the potential computational implications of real brain connectivity patterns on task solving.

We find results consistent across species and tasks, showing that biologically inspired networks perform as well as classical echo state networks, provided a minimum level of randomness and diversity of connections is allowed. We also present a framework, bio2art, to map and scale up real connectomes that can be integrated into recurrent ANNs. This approach also allows us to show the crucial importance of the diversity of interareal connectivity patterns, stressing the importance of stochastic processes determining neural networks connectivity in general.


title: Social prediction modulates activity of macaque superior temporal cortex

abstract: The ability to attribute thoughts to others, also called theory of mind (TOM), has been extensively studied. Computationally, the basis of TOM in humans has been interpreted within the predictive coding framework and associated with activity in the temporo-parietal junction (TPJ). However, the evolutionary origins of these human mindreading abilities have been challenged since the concept was coined. Here we identify a brain region in the Rhesus macaque that shares computational properties with the human TPJ. We revealed, using a non-linguistic task and functional magnetic resonance imaging, that activity in a region of the macaque middle superior temporal cortex was specifically modulated by the predictability of social interactions. As in human TPJ, this region could be distinguished from other temporal regions involved in face processing. Our result suggests the existence of a precursor for the theory of mind ability in the last common ancestor of human and old-world monkeys.


title: Presynaptic NMDA receptors facilitate short-term plasticity and BDNF release at hippocampal mossy fiber synapses

abstract: Neurotransmitter release is a highly controlled process by which synapses can critically regulate information transfer within neural circuits. While presynaptic receptors -typically activated by neurotransmitters and modulated by neuromodulators- provide a powerful way of fine-tuning synaptic function, their contribution to activity-dependent changes in transmitter release remains poorly understood. Here, we report that presynaptic NMDA receptors (preNMDARs) at mossy fiber boutons in the rodent hippocampus can be activated by physiologically relevant patterns of activity and selectively enhance short-term synaptic plasticity at mossy fiber inputs onto CA3 pyramidal cells and mossy cells, but not onto inhibitory interneurons. Moreover, preNMDARs facilitate brain-derived neurotrophic factor (BDNF) release and contribute to presynaptic calcium rise. Taken together, our results indicate that by increasing presynaptic calcium, preNMDARs fine tune mossy fiber neurotransmission and can control information transfer during dentate granule cell burst activity that normally occur in vivo.


title: Envelope reconstruction of speech and music highlights unique tracking of speech at low frequencies

abstract: The human brain tracks amplitude fluctuations of both speech and music, which reflects acoustic processing in addition to the processing of higher-order features and ones cognitive state. Comparing neural tracking of speech and music envelopes can elucidate stimulus-general mechanisms, but direct comparisons are confounded by differences in their envelope spectra. Here, we use a novel method of frequency-constrained reconstruction of stimulus envelopes using EEG recorded during passive listening. We expected to see music reconstruction match speech in a narrow range of frequencies, but instead we found that speech was reconstructed better than music for all frequencies we examined. Additionally, speech envelope tracking at low frequencies, below 1 Hz, was uniquely associated with increased weighting over parietal channels. Our results highlight the importance of low-frequency speech tracking and its origin from speech-specific processing in the brain.


title: Pre-diabetes linked microRNA miR-193b-3p targets PPARGC1A and increases lipid accumulation in hepatocytes: potential biomarker of fatty liver disease

abstract: Distinct plasma microRNA profiles associate with different disease features. Elevated plasma microRNA hsa-miR-193b-3p has been reported in patients with pre-diabetes where early liver dysmetabolism plays a crucial role. MicroRNA target databases revealed that hsa-miR-193b-3p could potentially target PPARGC1A, a master switch transcriptional co-activator that orchestrates the expression of genes involved in multiple metabolic pathways. In this study we evaluated the effects of hsa-miR-193b-3p in the human HepG2 hepatocyte cell line. We show that hsa-miR-193b-3p targets PPARGC1A/PGC1 mRNA 3UTR and reduces its expression. Overexpression of hsa-miR-193b-3p under hyperglycemia resulted in increased accumulation of intracellular lipid droplets; and quantitative real-time RT-PCR of a panel of 24 genes revealed significant disruption in the expression of genes coordinating glucose uptake, glycolysis, pyruvate metabolism, fatty acid synthesis, fatty acid oxidation, triglyceride synthesis, VLDL secretion, insulin signalling, and mitochondrial biogenesis, dynamics and function. These results provide in vitro evidence that microRNA hsa-miR-193b-3p downregulates PPARGC1A/PGC1, disrupts downstream metabolic pathways and leads to the accumulation of intracellular lipids in hepatocytes. We propose that microRNA hsa-miR-193b-3p may have potential as a clinically relevant plasma biomarker for metabolic associated fatty liver disease in a pre-diabetic dysglycemic context.

category: biochemistry


title: The CLASSY family controls tissue-specific DNA methylation patterns in Arabidopsis

abstract: DNA methylation shapes the epigenetic landscape of the genome, plays critical roles in regulating gene expression, and ensures transposon silencing. As evidenced by the numerous defects associated with aberrant DNA methylation landscapes, establishing proper tissue-specific methylation patterns is critical. Yet, how such differences arise remains a largely open question in both plants and animals. Here we demonstrate that CLASSY1-4 (CLSY1-4), four locus-specific regulators of DNA methylation that are differentially expressed during plant development, play major roles in controlling tissue-specific DNA methylation patterns. Depending on the tissue, the genetic requirements for specific CLSYs differ significantly and, on a global scale, certain clsy mutants are sufficient to largely shift the epigenetic landscape between tissues. Together, these findings not only reveal substantial epigenetic diversity between tissues, but assign these changes to specific CLSY proteins, revealing how locus-specific targeting combined with tissue-specific expression enables the CLSYs to generate epigenetic diversity during plant development.


title: Whole Animal Multiplexed Single-Cell RNA-Seq Reveals Plasticity of Clytia Medusa Cell Types

abstract: We present an organism-wide, transcriptomic cell atlas of the hydrozoan medusa Clytia hemisphaerica, and determine how its component cell types respond to starvation. Utilizing multiplexed scRNA-seq, in which individual animals were indexed and pooled from control and perturbation conditions into a single sequencing run, we avoid artifacts from batch effects and are able to discern shifts in cell state in response to organismal perturbations. This work serves as a foundation for future studies of development, function, and plasticity in a genetically tractable jellyfish species. Moreover, we introduce a powerful workflow for high-resolution, whole animal, multiplexed single-cell genomics (WHAM-seq) that is readily adaptable to other traditional or non-traditional model organisms.


title: Dynamics and turnover of memory CD8 T cell responses following yellow fever vaccination

abstract:
Understanding how immunological memory lasts a lifetime requires quantifying changes in the number of memory cells as well as how their division and death rates change over time. We address these questions by using a statistically powerful mixed-effects differential equations framework to analyze data from two human studies that follow CD8 T cell responses to the yellow fever vaccine (YFV-17D). Models were first fit to the frequency and division rates of YFV-specific memory CD8 T cells 42 days to 1 year post-vaccination. A different dataset, on the loss of YFV-specific CD8 T cells over three decades, was used to assess out of sample predictions of our models. The commonly used exponential and bi-exponential decline models performed relatively poorly. Models with the cell loss following a power law (exactly or approximately) were most predictive. Notably, using only the first year of data, these models accurately predicted T cell frequencies up to 30 years post-vaccination. Our analyses suggest that division rates of these cells drop and plateau at a low level (0.001 per day, ~double estimates for naive T cells) within one year following vaccination, whereas death rates continue to decline for much longer. Our results show that power laws can be predictive for T cell memory, a finding that may be useful for vaccine evaluation and epidemiological modeling. Moreover, since power laws asymptotically decline more slowly than any exponential decline, our results help explain the longevity of immune memory phenomenologically.

Author summaryImmunological memory, generated in response to infection or vaccination, may provide complete or partial protection from antigenically similar infections for the lifetime. Memory CD8 T cells are important players in protection from secondary viral infections but quantitative understanding of their dynamics in humans is limited. We analyze data from two studies where immunization with the yellow fever virus vaccine (YFV-17D) generates a mild acute infection and long-term memory. We find that: (i) the division rate of YFV-17D-specific CD8 T cells drops and stabilizes at ~ 0.1% per day during the first year following vaccination whereas the death rate declines more gradually, and (ii) the number of these cells declines approximately in accordance with a power law ({propto}time-0.82) for at least several decades following vaccination.


title: Oral epithelial cells distinguish between Candida species with high or low pathogenic potential through miRNA regulation

abstract:
Oral epithelial cells monitor microbiome composition and initiate immune response upon dysbiosis, as in case of Candida imbalances. Comparison of healthy oral epithelial cell responses revealed that the inability of C. parapsilosis to induce a robust antifungal response was due to activation of various inflammation-independent pathways, while C. albicans robustly activated inflammation cascades. Regarding posttranscriptional regulation, several miRNAs were altered by both species. For C. parapsilosis, the applied dose directly correlated with changes in transcriptomic responses. Carbohydrate metabolism, hypoxia- and cardiovascular development-related responses dominate after C. parapsilosis stimulus, whereas C. albicans altered inflammatory responses. Subsequent analyses of HIF1- and HSC-activation pathways predicted target genes through which miRNA-dependent regulation of yeast-specific functions may occur, supporting the observed responses. Thus, C. parapsilosis is recognized as a commensal at low doses by the oral epithelium; however, increased fungal burden activates different pathways, some of which overlap with inflammatory processes induced by C. albicans.

Impact statementAltered miRNA regulation discriminates between C. albicans and C. parapsilosis in human oral epithelial cells


title: Linking plasmid-based beta-lactamases to their bacterial hosts using single-cell fusion PCR

abstract: The horizonal transfer of plasmid-encoded genes allows bacteria to adapt to constantly shifting environmental pressures, bestowing functional advantages to their bacterial hosts such as antibiotic resistance, metal resistance, virulence factors, and polysaccharide utilization. However, common molecular methods such as short- and long-read sequencing of microbiomes cannot associate extrachromosomal plasmids with the genome of the host bacterium. Alternative methods to link plasmids to host bacteria are either laborious, expensive or prone to contamination. Here we present the One-step Isolation and Lysis PCR (OIL-PCR) method, which molecularly links target ARGs with the bacterial 16S rRNA gene via fusion PCR performed within an emulsion. After validating this method, we apply it to identify the bacterial hosts of three clinically relevant beta-lactamases in a neutropenic patient population who are particularly vulnerable multidrug-resistant infections. We detect novel associations of two low-abundance genera, Romboutsia and Agathobacter, with a multi-drug resistant plasmid harbored by Klebsiella pneumoniae. We put forth a robust, accessible, and high-throughput platform for sensitively surveying the bacterial hosts of mobile genes in complex microbial communities.


title: Anaerobic Benzene Biodegradation Linked to Growth of Highly Specific Bacterial Clades

abstract:
Reliance on bioremediation to remove benzene from anoxic environments has proven risky for decades but for unknown reasons. Years of research have revealed a strong link between anaerobic benzene biodegradation and the enrichment of highly specific microbes, namely Thermincola in the family Peptococcaceae and the deltaproteobacterial Candidate Sva0485 clade. Using aquifer material from Canadian Forces Base Borden, we compared five bioremediation approaches in batch microcosms. Under conditions simulating natural attenuation or sulfate biostimulation, benzene was not degraded after 1-2 years of incubation and no enrichment of known benzene-degrading microbes occurred. In contrast, nitrate-amended microcosms reported benzene biodegradation coincident with significant growth of Thermincola spp., along with a functional gene presumed to catalyze anaerobic benzene carboxylation (abcA). Inoculation with 2.5% of a methanogenic benzene-degrading consortium containing Sva0485 (Deltaproteobacteria ORM2) resulted in benzene biodegradation in the presence of sulfate or under methanogenic conditions. The presence of other hydrocarbon co-contaminants decreased rates of benzene degradation by a factor of 2-4. Tracking the abundance of the abcA gene and 16S rRNA genes specific for benzene-degrading Thermincola and Sva0485 is recommended to monitor benzene bioremediation in anoxic groundwater systems to further uncover growth rate limiting conditions for these two intriguing phylotypes.

SYNOPSISAnaerobic benzene biodegradation was accelerated by biostimulation with nitrate or by bioaugmentation under methanogenic or sulfate-reducing conditions.


title: Zinc Import Mediated by AdcABC is Critical for Colonization of the Dental Biofilm by Streptococcus mutans in an Animal Model

abstract: Trace metals are essential to all domains of life but toxic when found at high concentrations. While the importance of iron in host-pathogen interactions is firmly established, contemporary studies indicate that other trace metals, including manganese and zinc, are also critical to the infectious process. In this study, we sought to identify and characterize the zinc uptake system(s) of S. mutans, a keystone pathogen in dental caries and a causative agent of bacterial endocarditis. Different than other pathogenic bacteria, including several streptococci, that encode multiple zinc import systems, bioinformatic analysis indicated that the S. mutans core genome encodes a single, highly conserved, zinc importer commonly known as AdcABC. Inactivation of the genes coding for the metal-binding AdcA ({Delta}adcA) or both AdcC ATPase and AdcB permease ({Delta}adcCB) severely impaired the ability of S. mutans to grow under zinc-depleted conditions. Intracellular metal quantifications revealed that both mutants accumulated less zinc when grown in the presence of a sub-inhibitory concentration of a zinc-specific chelator. Notably, the {Delta}adcCB strain displayed a severe colonization defect in a rat oral infection model. Both {Delta}adc strains were hypersensitive to high concentrations of manganese, showed reduced peroxide tolerance, and formed less biofilm in sucrose-containing media when cultivated in the presence of the lowest amount of zinc that support their growth, but not when zinc was supplied in excess. Collectively, this study identifies AdcABC as the lone high affinity zinc importer of S. mutans and provides preliminary evidence that zinc is a growth-limiting factor within the dental biofilm.


title: Diversity and prevalence of colibactin- and yersiniabactin encoding mobile genetic elements in enterobacterial populations: insights into evolution and co-existence of two bacterial secondary metabolite determinants

abstract: 1 AbstractThe bacterial genotoxin colibactin interferes with the eukaryotic cell cycle by causing double-stranded DNA breaks. It has been linked to bacterially induced colorectal cancer in humans. Colibactin is encoded by a 54-kb genomic region in Enterobacteriaceae. The colibactin genes commonly co-occur with the yersiniabactin biosynthetic determinant. Investigating the prevalence and sequence diversity of the colibactin determinant and its linkage to the yersiniabactin operon in prokaryotic genomes, we discovered mainly species-specific lineages of the colibactin determinant and classified three main structural settings of the colibactin-yersiniabactin genomic region in Enterobacteriaceae. The colibactin gene cluster has a similar but not identical evolutionary track to that of the yersiniabactin operon. Both determinants could have been acquired on several occasions and/or exchanged independently between enterobacteria by horizontal gene transfer. Integrative and conjugative elements play(ed) a central role in the evolution and structural diversity of the colibactin-yersiniabactin genomic region. Addition of an activating and regulating module (clbAR) to the biosynthesis and transport module (clbB-S) represents the most recent step in the evolution of the colibactin determinant. In a first attempt to correlate colibactin expression with individual lineages of colibactin determinants and different bacterial genetic backgrounds, we compared colibactin expression of selected enterobacterial isolates in vitro. Colibactin production in the tested Klebsiella spp. and Citrobacter koseri strains was more homogeneous and generally higher than that in most of the E. coli isolates studied. Our results improve the understanding of the diversity of colibactin determinants and its expression level, and may contribute to risk assessment of colibactin-producing enterobacteria.