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Endemic infections with the common avian pathogen Salmonella enterica subspecies enterica serovar Typhimurium (Salmonella Typhimurium) may incur a significant cost on the host population. In this study, we determined the potential of endemic Salmonella infections to reduce the reproductive success of blue (Cyanistes caeruleus) and great (Parus major) tits by correlating eggshell infection with reproductive parameters. The fifth egg of each clutch was collected from nest boxes in 19 deciduous forest fragments. Out of the 101 sampled eggs, 7 Salmonella Typhimurium isolates were recovered. The low bacterial prevalence was reflected by a similarly low serological prevalence in the fledglings. In this study with a relatively small sample size, presence of Salmonella did not affect reproductive parameters (egg volume, clutch size, number of nestlings and number of fledglings), nor the health status of the fledglings. However, in order to clarify the impact on health and reproduction a larger number of samples have to be analyzed. Phage typing showed that the isolates belonged to the definitive phage types (DT) 193 and 99, and multi-locus variable number tandem repeat analysis (MLVA) demonstrated a high similarity among the tit isolates, but distinction to human isolates. These findings suggest the presence of passerine-adapted Salmonella strains in free-ranging tit populations with host pathogen co-existence. [ABSTRACT FROM AUTHOR]

Listeriosis is a rare but severe disease, mainly caused by Listeria monocytogenes. This study shows the results of the laboratory-based surveillance of Listeriosis in Belgium over the period 1985–2014. Besides the incidence and some demographic data we present also more detailed microbiological and molecular characteristics of human strains isolated since 2000. The strains from the latter period were compared to food and animal strains from the same period. Our study shows that different food matrices were commonly contaminated with L. monocytogenes presenting the same PFGE profile as in patient’s isolates. Since 1985, we observed a significant decrease in incidence of the Materno-Neonatal cases (from 0.15 to 0.04 cases /100,000 inhabitants-year), which is probably to be attributed to active prevention campaigns targeting pregnant women. Despite the strengthening of different control measures by the food industry, the incidence of non-Materno-Neonatal listeriosis increased in Belgium (from 0.3 to 0.7 cases /100,000 inhabitants-year), probably due to the rise of highly susceptible patients in an aging population. This significant increase found in non-Materno-Neonatal cases (slope coefficient 7.42%/year, P<0.0001) can be attributed to significant increase in incidence of isolates belonging to serovars 1/2a (n = 393, slope coefficient 6.62%/year, P<0.0001). Although resistance to antimicrobials is rare among L. monocytogenes isolates, a trend to increasing MIC values is evident with chloramphenicol, amoxicillin, tetracycline and ciprofloxacin. We show that fluoroquinolone resistance is not linked to chromosomal mutations, but caused by a variety of efflux pumps. Our study also shows that huge majority of known underlying pathologies (426 out of 785 cases) were cancers (185/426, 43.1%) and haematological malignancies (75/185, 40.5%). Moreover the risk population is susceptible to low levels of contamination in food stressing the need of prevention campaigns specifically targeting these persons. [ABSTRACT FROM AUTHOR]

In recent years, genome sequencing of filamentous fungi has revealed a high proportion of specialised metabolites with growing pharmaceutical interest. However, detecting such metabolites through in silico genome analysis does not necessarily guarantee their expression under laboratory conditions. However, one plausible strategy for enabling their production lies in modifying the growth conditions. Devising a comprehensive experimental design testing in different culture environments is time-consuming and expensive. Therefore, using in silico modelling as a preliminary step, such as Genome-Scale Metabolic Network (GSMN), represents a promising approach to predicting and understanding the observed specialised metabolite production in a given organism. To address these questions, we reconstructed a new high-quality GSMN for the Penicillium rubens Wisconsin 54-1255 strain, a commonly used model organism. Our reconstruction, iPrub22, adheres to current convention standards and quality criteria, incorporating updated functional annotations, orthology searches with different GSMN templates, data from previous reconstructions, and manual curation steps targeting primary and specialised metabolites. With a MEMOTE score of 74% and a metabolic coverage of 45%, iPrub22 includes 5,192 unique metabolites interconnected by 5,919 reactions, of which 5,033 are supported by at least one genomic sequence. Of the metabolites present in iPrub22, 13% are categorised as belonging to specialised metabolism. While our high-quality GSMN provides a valuable resource for investigating known phenotypes expressed in P. rubens, our analysis identifies bottlenecks related, in particular, to the definition of what is a specialised metabolite, which requires consensus within the scientific community. It also points out the necessity of accessible, standardised and exhaustive databases of specialised metabolites. These questions must be addressed to fully unlock the potential of natural product production in P. rubens and other filamentous fungi. Our work represents a foundational step towards the objective of rationalising the production of natural products through GSMN modelling., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Nègre et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

We constructed a panel of S. pombe strains expressing DNA polymerase ε variants associated with cancer, specifically POLES297F, POLEV411L, POLEL424V, POLES459F, and used these to compare mutation rates determined by canavanine resistance with other selective methods. Canavanine-resistance mutation rates are broadly similar to those seen with reversion of the ade-485 mutation to adenine prototrophy, but lower than 5-fluoroorotic acid (FOA)-resistance rates (inactivation of ura4+ or ura5+ genes). Inactivation of several genes has been associated with canavanine resistance in S. pombe but surprisingly whole genome sequencing showed that 8/8 spontaneous canavanine-resistant mutants have an R175C mutation in the any1/arn1 gene. This gene encodes an α-arrestin-like protein involved in mediating Pub1 ubiquitylation of target proteins, and the phenotypic resistance to canavanine by this single mutation is similar to that shown by the original "can1-1" strain, which also has the any1R175C mutation. Some of the spontaneous mutants have additional mutations in arginine transporters, suggesting that this may marginally increase resistance to canavanine. The any1R175C strain showed internalisation of the Cat1 arginine transporter as previously reported, explaining the canavanine-resistance phenotype., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Pai et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Photoconvertible proteins are powerful tools widely used in cellular biology to study cell dynamics and organelles. Over the past decade, photoconvertible proteins have also been used for developmental biology applications to analyze cell lineage and cell fate during embryonic development. One of these photoconvertible proteins called Kaede, from the stony coral Trachyphyllia geoffroyi, undergoes irreversible photoconversion from green to red fluorescence when illuminated with UV light. Undertaking a cell tracing approach using photoconvertible proteins can be challenging when using unconventional animal models. In this protocol, we describe the use of Kaede to track specific cells during embryogenesis of the cephalochordate Branchiostoma lanceolatum. This protocol can be adapted to other unconventional models, especially marine animals., Competing Interests: The authors have declared that no competing interests exist.

Aim: To assess the progress on the implementation of Non-Communicable Diseases (NCD) related policies and legislations in the Pacific Island Countries and Territories (PICTs)., Materials and Methods: The Pacific Monitoring Alliance for NCD Action (MANA) Dashboard was used to assess the progress on the implementation. The MANA Dashboard includes 31 indicators across four different domains such as leadership and governance; preventative policies and legislations; health system response programs; and monitoring This progress assessment was conducted between 2019 and 2020 for all 21 PICTs. The data were analyzed and compared with the baseline status (2018) report and presented across four different domains of the MANA dashboard., Results: This progress assessment found that PICTs overall have made advancements in a number of areas, particularly the establishment of a national multi-sectoral NCD taskforce; implementation of referenced approaches to restrict trans-fat in the food supply in national documents; and fiscal measures to affect access and availability to less healthy foods and drinks. However, the strengths of actions varied across PICTs, and most are categorised as low strengths. Measures which had the most limited progress in implementation include policy and legislation that restrict alcohol advertising; tobacco industry interference; marketing of foods and non-alcoholic beverages to children; and marketing for breast milk substitutes., Conclusions: This progress assessment further highlights that while PICTs continue to make progress, NCD policy and legislation gaps still exist, both in terms of weaknesses of existing measures and areas that have had little attention to-date. These require urgent actions to scale up NCD related policies and legislation at regional and national level., Competing Interests: The authors have declared that no competing interests exist.

At-sea behaviour of seabirds have received significant attention in ecology over the last decades as it is a key process in the ecology and fate of these populations. It is also, through the position of top predator that these species often occupy, a relevant and integrative indicator of the dynamics of the marine ecosystems they rely on. Seabird trajectories are recorded through the deployment of GPS, and a variety of statistical approaches have been tested to infer probable behaviours from these location data. Recently, deep learning tools have shown promising results for the segmentation and classification of animal behaviour from trajectory data. Yet, these approaches have not been widely used and investigation is still needed to identify optimal network architecture and to demonstrate their generalization properties. From a database of about 300 foraging trajectories derived from GPS data deployed simultaneously with pressure sensors for the identification of dives, this work has benchmarked deep neural network architectures trained in a supervised manner for the prediction of dives from trajectory data. It first confirms that deep learning allows better dive prediction than usual methods such as Hidden Markov Models. It also demonstrates the generalization properties of the trained networks for inferring dives distribution for seabirds from other colonies and ecosystems. In particular, convolutional networks trained on Peruvian boobies from a specific colony show great ability to predict dives of boobies from other colonies and from distinct ecosystems. We further investigate accross-species generalization using a transfer learning strategy known as 'fine-tuning'. Starting from a convolutional network pre-trained on Guanay cormorant data reduced by two the size of the dataset needed to accurately predict dives in a tropical booby from Brazil. We believe that the networks trained in this study will provide relevant starting point for future fine-tuning works for seabird trajectory segmentation., Competing Interests: The authors have declared that no competing interests exist.

Somatic and germline mutations in the proofreading domain of the replicative DNA polymerase ε (POLE-exonuclease domain mutations, POLE-EDMs) are frequently found in colorectal and endometrial cancers and, occasionally, in other tumours. POLE-associated cancers typically display hypermutation, and a unique mutational signature, with a predominance of C > A transversions in the context TCT and C > T transitions in the context TCG. To understand better the contribution of hypermutagenesis to tumour development, we have modelled the most recurrent POLE-EDM (POLE-P286R) in Schizosaccharomyces pombe. Whole-genome sequencing analysis revealed that the corresponding pol2-P287R allele also has a strong mutator effect in vivo, with a high frequency of base substitutions and relatively few indel mutations. The mutations are equally distributed across different genomic regions, but in the immediate vicinity there is an asymmetry in AT frequency. The most abundant base-pair changes are TCT > TAT transversions and, in contrast to human mutations, TCG > TTG transitions are not elevated, likely due to the absence of cytosine methylation in fission yeast. The pol2-P287R variant has an increased sensitivity to elevated dNTP levels and DNA damaging agents, and shows reduced viability on depletion of the Pfh1 helicase. In addition, S phase is aberrant and RPA foci are elevated, suggestive of ssDNA or DNA damage, and the pol2-P287R mutation is synthetically lethal with rad3 inactivation, indicative of checkpoint activation. Significantly, deletion of genes encoding some translesion synthesis polymerases, most notably Pol κ, partially suppresses pol2-P287R hypermutation, indicating that polymerase switching contributes to this phenotype., Competing Interests: The authors have declared that no competing interests exist.

In many developing countries, small-scale fisheries provide employment and important food security for local populations. To support resource management, the description of the spatiotemporal extent of fisheries is necessary, but often poorly understood due to the diffuse nature of effort, operated from numerous small wooden vessels. Here, in Gabon, Central Africa, we applied Hidden Markov Models to detect fishing patterns in seven different fisheries (with different gears) from GPS data. Models were compared to information collected by on-board observers (7 trips) and, at a larger scale, to a visual interpretation method (99 trips). Models utilizing different sampling resolutions of GPS acquisition were also tested. Model prediction accuracy was high with GPS data sampling rates up to three minutes apart. The minor loss of accuracy linked to model classification is largely compensated by the savings in time required for analysis, especially in a context of nations or organizations with limited resources. This method could be applied to larger datasets at a national or international scale to identify and more adequately manage fishing effort., Competing Interests: The authors have declared that no competing interests exist.

The cephalochordates amphioxus or lancelets are benthic marine animals representing the earliest divergent evolutionary lineage within chordates. Although amphioxus are present in most of the world's tropical and temperate oceans, only about thirty different species grouped into three different genera, Branchiostoma, Epigonichthys and Asymmetron have been described. In the genus Asymmetron, only two species have been characterized, although for one of them, A. lucayanum, several cryptic lineages exist. In this work we have sequenced and analyzed the mitogenome of an A. lucayanum population previously described in the Red Sea. The phylogenetic study using this complete mitogenome as well as the analysis of COI gene sequences of several individuals of this Red Sea population show that the Red Sea population is a new cryptic species. We propose to call this new species Asymmetron rubrum., Competing Interests: The authors have declared that no competing interests exist

RNA interference (RNAi) requires RNA-dependent RNA polymerases (RdRPs) in many eukaryotes, and RNAi amplification constitutes the only known function for eukaryotic RdRPs. Yet in animals, classical model organisms can elicit RNAi without possessing RdRPs, and only nematode RNAi was shown to require RdRPs. Here we show that RdRP genes are much more common in animals than previously thought, even in insects, where they had been assumed not to exist. RdRP genes were present in the ancestors of numerous clades, and they were subsequently lost at a high frequency. In order to probe the function of RdRPs in a deuterostome (the cephalochordate Branchiostoma lanceolatum), we performed high-throughput analyses of small RNAs from various Branchiostoma developmental stages. Our results show that Branchiostoma RdRPs do not appear to participate in RNAi: we did not detect any candidate small RNA population exhibiting classical siRNA length or sequence features. Our results show that RdRPs have been independently lost in dozens of animal clades, and even in a clade where they have been conserved (cephalochordates) their function in RNAi amplification is not preserved. Such a dramatic functional variability reveals an unexpected plasticity in RNA silencing pathways., Competing Interests: The authors have declared that no competing interests exist.

[This corrects the article DOI: 10.1371/journal.pone.0117950.].

Whole genome sequencing represents a promising new technology for subtyping of bacterial pathogens. Besides the technological advances which have pushed the approach forward, the last years have been marked by considerable evolution of the whole genome sequencing data analysis methods. Prior to application of the technology as a routine epidemiological typing tool, however, reliable and efficient data analysis strategies need to be identified among the wide variety of the emerged methodologies. In this work, we have compared three existing SNP-based subtyping workflows using a benchmark dataset of 32 Salmonella enterica subsp. enterica serovar Typhimurium and serovar 1,4,[5],12:i:- isolates including five isolates from a confirmed outbreak and three isolates obtained from the same patient at different time points. The analysis was carried out using the original (high-coverage) and a down-sampled (low-coverage) datasets and two different reference genomes. All three tested workflows, namely CSI Phylogeny-based workflow, CFSAN-based workflow and PHEnix-based workflow, were able to correctly group the confirmed outbreak isolates and isolates from the same patient with all combinations of reference genomes and datasets. However, the workflows differed strongly with respect to the SNP distances between isolates and sensitivity towards sequencing coverage, which could be linked to the specific data analysis strategies used therein. To demonstrate the effect of particular data analysis steps, several modifications of the existing workflows were also tested. This allowed us to propose data analysis schemes most suitable for routine SNP-based subtyping applied to S. Typhimurium and S. 1,4,[5],12:i:-. Results presented in this study illustrate the importance of using correct data analysis strategies and to define benchmark and fine-tune parameters applied within routine data analysis pipelines to obtain optimal results.

S 47445 is a novel positive allosteric modulator of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptors (AMPA-PAM). S 47445 enhanced glutamate's action at AMPA receptors on human and rat receptors and was inactive at NMDA and kainate receptors. Potentiation did not differ among the different AMPA receptors subtypes (GluA1/2/4 flip and flop variants) (EC50 between 2.5-5.4 μM), except a higher EC50 value for GluA4 flop (0.7 μM) and a greater amount of potentiation on GluA1 flop. A low concentration of S 47445 (0.1 μM) decreased receptor response decay time of GluA1flop/GluA2flip AMPA receptors and increased the sensitivity to glutamate. Furthermore, S 47445 (0.1 and 0.3 μM) in presence of repetitive glutamate pulses induced a progressive potentiation of the glutamate-evoked currents from the second pulse of glutamate confirming a rapid-enhancing effect of S 47445 at low concentrations. The potentiating effect of S 47445 (1 μM) was concentration-dependently reversed by the selective AMPA receptor antagonist GYKI52466 demonstrating the selective modulatory effect of S 47445 on AMPA receptors. Using an AMPA-kainate chimera approach, it was confirmed that S 47445 binds to the common binding pocket of AMPA-PAMs. S 47445 did not demonstrate neurotoxic effect against glutamate-mediated excitotoxicity in vitro, in contrast significantly protected rat cortical neurons at 10 μM. S 47445 was shown to improve both episodic and spatial working memory in adult rodents at 0.3 mg/kg, as measured in the natural forgetting condition of object recognition and T-maze tasks. Finally, no deleterious effect on spontaneous locomotion and general behavior was observed up to 1000 mg/kg of S 47445 given acutely in rodents, neither occurrence of convulsion or tremors. Collectively, these results indicate that S 47445 is a potent and selective AMPA-PAM presenting procognitive and potential neuroprotective properties. This drug is currently evaluated in clinical phase 2 studies in Alzheimer's disease and in Major Depressive Disorder.

Background: Bloodstream infections (BSI) caused by Salmonella Typhi and invasive non-Typhoidal Salmonella (iNTS) frequently affect children living in rural sub-Saharan Africa but data about incidence and serotype distribution are rare., Objective: The present study assessed the population-based incidence of Salmonella BSI and severe malaria in a Health and Demographic Surveillance System in a rural area with seasonal malaria transmission in Nanoro, Burkina Faso., Methods: Children between 2 months-15 years old with severe febrile illness were enrolled during a one-year surveillance period (May 2013-May 2014). Thick blood films and blood cultures were sampled and processed upon admission. Population-based incidences were corrected for non-referral, health seeking behavior, non-inclusion and blood culture sensitivity. Adjusted incidence rates were expressed per 100,000 person-years of observations (PYO)., Results: Among children < 5 years old, incidence rates for iNTS, Salmonella Typhi and severe malaria per 100,000 PYO were 4,138 (95% Confidence Interval (CI): 3,740-4,572), 224 (95% CI: 138-340) and 2,866 (95% CI: 2,538-3,233) respectively. Among those aged 5-15 years, corresponding incidence rates were 25 (95% CI: 8-60), 273 (95% CI: 203-355) and 135 (95% CI: 87-195) respectively. Most iNTS occurred during the peak of the rainy season and in parallel with the increase of Plasmodium falciparum malaria; for Salmonella Typhi no clear seasonal pattern was observed. Salmonella Typhi and iNTS accounted for 13.3% and 55.8% of all 118 BSI episodes; 71.6% of iNTS (48/67) isolates were Salmonella enterica serovar Typhimurium and 25.4% (17/67) Salmonella enterica serovar Enteritidis; there was no apparent geographical clustering., Conclusion: The present findings from rural West-Africa confirm high incidences of Salmonella Typhi and iNTS, the latter with a seasonal and Plasmodium falciparum-related pattern. It urges prioritization of the development and implementation of Salmonella Typhi as well as iNTS vaccines in this setting.

Salmonella is frequently found in poultry and represent an important source for human gastrointestinal infections worldwide. The aim of this study was to investigate the prevalence, genotypes and antimicrobial resistance of Salmonella serotypes in broilers from Ecuador. Caeca content from 388 at random selected broiler batches were collected in 6 slaughterhouses during 1 year and analyzed by the ISO 6579/Amd1 protocol for the isolation for Salmonella. Isolates were serotyped and genotypic variation was acceded by pulsed field gel electrophoresis. MIC values for sulfamethoxazole, gentamicin, ciprofloxacin, ampicillin, cefotaxime, ceftazidime, tetracycline, streptomycin, trimethropim, chloramphenicol, colistin, florfenicol, kanamycin and nalidixic acid were obtained. Presence of blaCTX-M, blaTEM, blaSHV and blaCMY; and mcr-1 plasmid genes was investigated in resistant strains to cefotaxime and colistin respectively. Prevalence at batch level was 16.0%. The most common serotype was S. Infantis (83.9%) followed by S. Enteritidis (14.5%) and S. Corvallis (1.6%). The pulsed field gel electrophoresis analysis showed that S. Corvallis, S. Enteritidis and S. Infantis isolates belonged to 1, 2 and 12 genotypes respectively. S. Infantis isolates showed high resistance rates to 12 antibiotics ranging from 57.7% (kanamycin) up to 98.1% (nalidixic acid and sulfamethoxazole). All S. Enteritidis isolates showed resistance to colistin. High multiresistant patterns were found for all the serotypes. The blaCTX-M gene was present in 33 S. Infantis isolates while mcr-1 was negative in 10 colistin resistant isolates. This study provides the first set of scientific data on prevalence and multidrug-resistant Salmonella coming from commercial poultry in Ecuador.

We present the results of a longitudinal surveillance study (1995-2014) on fluoroquinolone resistance (FQ-R) among Belgian non-invasive Streptococcus pneumoniae isolates (n = 5,602). For many years, the switch to respiratory fluoroquinolones for the treatment of (a)typical pneumonia had no impact on FQ-R levels. However, since 2011 we observed a significant decrease in susceptibility towards ciprofloxacin, ofloxacin and levofloxacin with peaks of 9.0%, 6.6% and 3.1% resistant isolates, respectively. Resistance to moxifloxacin arised sporadically, and remained <1% throughout the entire study period. We observed classical topoisomerase mutations in gyrA (n = 25), parC (n = 46) and parE (n = 3) in varying combinations, arguing against clonal expansion of FQ-R. The impact of recombination with co-habiting commensal streptococci on FQ-R remains marginal (10.4%). Notably, we observed that a rare combination of DNA Gyrase mutations (GyrA&#95;S81L/GyrB&#95;P454S) suffices for high-level moxifloxacin resistance, contrasting current model. Interestingly, 85/422 pneumococcal strains display MICCIP values which were lowered by at least four dilutions by reserpine, pointing at involvement of efflux pumps in FQ-R. In contrast to susceptible strains, isolates resistant to ciprofloxacin significantly overexpressed the ABC pump PatAB in comparison to reference strain S. pneumoniae ATCC 49619, but this could only be linked to disruptive terminator mutations in a fraction of these. Conversely, no difference in expression of the Major Facilitator PmrA, unaffected by reserpine, was noted between susceptible and resistant S. pneumoniae strains. Finally, we observed that four isolates displayed intermediate to high-level ciprofloxacin resistance without any known molecular resistance mechanism. Focusing future molecular studies on these isolates, which are also commonly found in other studies, might greatly assist in the battle against rising pneumococcal drug resistance.

Background: Invasive meningococcal disease (IMD) is a major cause of bacterial meningitides and septicaemia. This study shows the results of the laboratory-based surveillance of IMD in Belgium over the period 1997-2012., Methods: The results are based on microbiological and molecular laboratory surveillance of 2997 clinical isolates of N. meningitides received by the Belgian Meningococcal Reference Centre (BMRC) over the period 1997-2012., Results: Serogroup B has always been a major cause of meningococcal disease in Belgium, with P3.4 as most frequent serotype till 2008, while an increase in non-serotypable strains has been observed in the last few years. Clonal complexes cc-41/44 and cc-269 are most frequently observed in serogroup B strains. In the late nineties, the incidence of serogroup C disease increased considerably and peaked in 2001, mainly associated with phenotypes C:2a:P1.5,2, C:2a:P1.5 and C:2a:P1.2 (ST-11/ET-37 clonal complex). The introduction of the meningococcal C conjugate vaccine has been followed by an 88% significant decrease in serogroup C disease from 2001 to 2004 nationally, yet sharper in Flanders (92%) compared to Wallonia (77%). Since 2008 a difference in incidence of serogroup C was observed in Flanders (0-0.1/100,000) versus Wallonia (0.1-0.3/100,000)., Conclusion: This study showed the change in epidemiology and strain population over a 16 years period spanning an exhaustive vaccination campaign and highlights the influence of regional vaccination policies with different cohorts sizes on short and long-term IMD incidences.

During vertebrate development, the paraxial mesoderm becomes segmented, forming somites that will give rise to dermis, axial skeleton and skeletal muscles. Although recently challenged, the "clock and wavefront" model for somitogenesis explains how interactions between several cell-cell communication pathways, including the FGF, RA, Wnt and Notch signals, control the formation of these bilateral symmetric blocks. In the cephalochordate amphioxus, which belongs to the chordate phylum together with tunicates and vertebrates, the dorsal paraxial mesendoderm also periodically forms somites, although this process is asymmetric and extends along the whole body. It has been previously shown that the formation of the most anterior somites in amphioxus is dependent upon FGF signalling. However, the signals controlling somitogenesis during posterior elongation in amphioxus are still unknown. Here we show that, contrary to vertebrates, RA and FGF signals act independently during posterior elongation and that they are not mandatory for posterior somites to form. Moreover, we show that RA is not able to buffer the left/right asymmetry machinery that is controlled through the asymmetric expression of Nodal pathway actors. Our results give new insights into the evolution of the somitogenesis process in chordates. They suggest that RA and FGF pathways have acquired specific functions in the control of somitogenesis in vertebrates. We propose that the "clock and wavefront" system was selected specifically in vertebrates in parallel to the development of more complex somite-derived structures but that it was not required for somitogenesis in the ancestor of chordates.

Introduction: In April and May 2014, two suspected egg-related outbreaks of Salmonella enterica subsp. enterica serovar Enteritidis (S. Enteritidis) were investigated by the Belgian National Reference Laboratory of Foodborne Outbreaks. Both the suspected food and human isolates being available, and this for both outbreaks, made these the ideal case study for a retrospective whole genome sequencing (WGS) analysis with the goal to investigate the feasibility of this technology for outbreak investigation by a National Reference Laboratory or National Reference Centre without thorough bioinformatics expertise., Methods: The two outbreaks were originally investigated epidemiologically with a standard questionnaire and with serotyping, phage typing, multiple-locus variable-number of tandem repeats analysis (MLVA) and antimicrobial susceptibility testing as classical microbiological methods. Retrospectively, WGS of six outbreak isolates was done on an Illumina HiSeq. Analysis of the WGS data was performed with currently available, user-friendly software and tools, namely CLC Genomics Workbench, the tools available on the server of the Center for Genomic Epidemiology and BLAST Ring Image Generator (BRIG)., Results: To all collected human and food outbreak isolates, classical microbiological investigation assigned phage type PT4 (variant phage type PT4a for one human isolate) and MLVA profile 3-10-5-4-1, both of which are common for human isolates in Belgium. The WGS analysis confirmed the link between food and human isolates for each of the outbreaks and clearly discriminated between the two outbreaks occurring in a same time period, thereby suggesting a non-common source of contamination. Also, an additional plasmid carrying an antibiotic resistance gene was discovered in the human isolate with the variant phage type PT4a., Discussion: For the two investigated outbreaks occurring at geographically separated locations, the gold standard classical microbiological subtyping methods were not sufficiently discriminative to distinguish between or assign a common origin of contamination for the two outbreaks, while WGS analysis could do so. This case study demonstrated the added value of WGS for outbreak investigations by confirming and/or discriminating food and human isolates between and within outbreaks. It also proved the feasibility of WGS as complementary or even future replacing (sub)typing method for the average routine laboratory.

How organisms move and disperse is crucial to understand how population dynamics relates to the spatial heterogeneity of the environment. Random walk (RW) models are typical tools to describe movement patterns. Whether Lévy or alternative RW better describes forager movements is keenly debated. We get around this issue using the Generalized Pareto Distribution (GPD). GPD includes as specific cases Normal, exponential and power law distributions, which underlie Brownian, Poisson-like and Lévy walks respectively. Whereas previous studies typically confronted a limited set of candidate models, GPD lets the most likely RW model emerge from the data. We illustrate the wide applicability of the method using GPS-tracked seabird foraging movements and fishing vessel movements tracked by Vessel Monitoring System (VMS), both collected in the Peruvian pelagic ecosystem. The two parameters from the fitted GPD, a scale and a shape parameter, provide a synoptic characterization of the observed movement in terms of characteristic scale and diffusive property. They reveal and quantify the variability, among species and individuals, of the spatial strategies selected by predators foraging on a common prey field. The GPD parameters constitute relevant metrics for (1) providing a synthetic and pattern-oriented description of movement, (2) using top predators as ecosystem indicators and (3) studying the variability of spatial behaviour among species or among individuals with different personalities.

Surveillance of Salmonella enterica subsp. enterica serovar Enteritidis is generally considered to benefit from molecular techniques like multiple-locus variable-number of tandem repeats analysis (MLVA), which allow early detection and confinement of outbreaks. Here, a surveillance study, including phage typing, antimicrobial susceptibility testing and MLVA on 1,535 S. Enteritidis isolates collected between 2007 and 2012, was used to evaluate the added value of MLVA for public health surveillance in Belgium. Phage types PT4, PT8, PT21, PT1, PT6, PT14b, PT28 and PT13 dominate the Belgian S. Enteritidis population. The isolates of S. Enteritidis were most frequently susceptible to all antibiotics tested. 172 different MLVA profiles were detected, of which 9 frequent profiles included 67.2% of the S. Enteritidis population. During a serial passage experiment on selected isolates to investigate the in vitro stability of the 5 MLVA loci, no variations over time were observed indicating that the MLVA profiles were stable. The MLVA profile of isolates originating from different outbreaks in the Democratic Republic of the Congo (DRC) between 2010 and 2011 were distinct from any of the MLVA profiles found in Belgian isolates throughout the six year observational period and demonstrates that MLVA improves public health surveillance of S. Enteritidis. However, MLVA should be complemented with other subtyping methods when investigating outbreaks is caused by the most common MLVA profile.

Ochrobactrum intermedium is considered as an emerging human environmental opportunistic pathogen with mild virulence. The distribution of isolates and sequences described in literature and databases showed frequent association with human beings and polluted environments. As population structures are related to bacterial lifestyles, we investigated by multi-locus approach the genetic structure of a population of 65 isolates representative of the known natural distribution of O. intermedium. The population was further surveyed for genome dynamics using pulsed-field gel electrophoresis and genomics. The population displayed a clonal epidemic structure with events of recombination that occurred mainly in clonal complexes. Concerning biogeography, clones were shared by human and environments and were both cosmopolitan and local. The main cosmopolitan clone was genetically and genomically stable, and grouped isolates that all harbored an atypical insertion in the rrs. Ubiquitism and stability of this major clone suggested a clonal succes in a particular niche. Events of genomic reduction were detected in the population and the deleted genomic content was described for one isolate. O. intermedium displayed allopatric characters associated to a tendancy of genome reduction suggesting a specialization process. Considering its relatedness with Brucella, this specialization might be a commitment toward pathogenic life-style that could be driven by technological selective pressure related medical and industrial technologies.

Surveillance of Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) is generally considered to benefit from molecular techniques like multiple-locus variable-number of tandem repeats analysis (MLVA), which allow earlier detection and confinement of outbreaks. Here, a surveillance study, including phage typing, antimicrobial susceptibility testing and the in Europe most commonly used 5-loci MLVA on 1,420 S. Typhimurium isolates collected between 2010 and 2012 in Belgium, was used to evaluate the added value of MLVA for public health surveillance. Phage types DT193, DT195, DT120, DT104, DT12 and U302 dominate the Belgian S. Typhimurium population. A combined resistance to ampicillin, streptomycin, sulphonamides and tetracycline (ASSuT) with or without additional resistances was observed for 42.5% of the isolates. 414 different MLVA profiles were detected, of which 14 frequent profiles included 44.4% of the S. Typhimurium population. During a serial passage experiment on selected isolates to investigate the in vitro stability of the 5 MLVA loci, variations over time were observed for loci STTR6, STTR10, STTR5 and STTR9. This study demonstrates that MLVA improves public health surveillance of S. Typhimurium. However, the 5-loci MLVA should be complemented with other subtyping methods for investigation of possible outbreaks with frequent MLVA profiles. Also, variability in these MLVA loci should be taken into account when investigating extended outbreaks and studying dynamics over longer periods.

One major challenge in the emerging field of movement ecology is the inference of behavioural modes from movement patterns. This has been mainly addressed through Hidden Markov models (HMMs). We propose here to evaluate two sets of alternative and state-of-the-art modelling approaches. First, we consider hidden semi-Markov models (HSMMs). They may better represent the behavioural dynamics of foragers since they explicitly model the duration of the behavioural modes. Second, we consider discriminative models which state the inference of behavioural modes as a classification issue, and may take better advantage of multivariate and non linear combinations of movement pattern descriptors. For this work, we use a dataset of >200 trips from human foragers, Peruvian fishermen targeting anchovy. Their movements were recorded through a Vessel Monitoring System (∼1 record per hour), while their behavioural modes (fishing, searching and cruising) were reported by on-board observers. We compare the efficiency of hidden Markov, hidden semi-Markov, and three discriminative models (random forests, artificial neural networks and support vector machines) for inferring the fishermen behavioural modes, using a cross-validation procedure. HSMMs show the highest accuracy (80%), significantly outperforming HMMs and discriminative models. Simulations show that data with higher temporal resolution, HSMMs reach nearly 100% of accuracy. Our results demonstrate to what extent the sequential nature of movement is critical for accurately inferring behavioural modes from a trajectory and we strongly recommend the use of HSMMs for such purpose. In addition, this work opens perspectives on the use of hybrid HSMM-discriminative models, where a discriminative setting for the observation process of HSMMs could greatly improve inference performance.

Background: Co-resistance against the first-line antibiotics ampicillin, chloramphenicol and trimethoprim/sulphamethoxazole or multidrug resistance (MDR) is common in non typhoid Salmonella (NTS). Use of alternative antibiotics, such as fluoroquinolones or third generation cephalosporins is threatened by increasing resistance, but remains poorly documented in Central-Africa., Methodology/principal Findings: As part of a microbiological surveillance study in DR Congo, blood cultures were collected between 2007 and 2011. Isolated NTS were assessed for serotype and antimicrobial resistance including decreased ciprofloxacin susceptibility and extended-spectrum beta-lactamase (ESBL) production. In total, 233 NTS isolates (representing 23.6% of clinically significant organisms) were collected, mainly consisting of Salmonella Typhimurium (79%) and Salmonella Enteritidis (18%). The majority of NTS were isolated in the rainy season, and recovered from children ≤2 years old. MDR, decreased ciprofloxacin susceptibility, azithromycin and cefotaxime resistance were 80.7%, 4.3%, 3.0% and 2.1% respectively. ESBL production was noted in three (1.3%) isolates. Decreased ciprofloxacin susceptibility was associated with mutations in codon 87 of the gyrA gene, while ESBLs all belonged to the SHV-2a type., Conclusions/significance: Presence of almost full MDR among NTS isolates from blood cultures in Central Africa was confirmed. Resistance to fluoroquinolones, azithromycin and third generation cephalosporins is still low, but emerging. Increased microbiological surveillance in DR Congo is crucial for adapted antibiotic therapy and the development of treatment guidelines.

Background: Drug resistance of Salmonella enterica serovar Typhi (Salmonella Typhi) to first-line antibiotics is emerging in Central Africa. Although increased use of fluoroquinolones is associated with spread of resistance, Salmonella Typhi with decreased ciprofloxacin susceptibility (DCS) has rarely been reported in Central Africa., Methodology/principal Findings: As part of a microbiological surveillance study in the Democratic Republic of the Congo (DR Congo), Salmonella Typhi isolates from bloodstream infections were collected prospectively between 2007 and 2011. The genetic relationship of the Salmonella Typhi isolates was assessed by pulsed-field gel electrophoresis (PFGE). The antimicrobial resistance profile of the isolates was determined and mutations associated with DCS were studied. In total, 201 Salmonella Typhi isolates were collected. More than half of the Salmonella Typhi isolates originated from children and young adults aged 5-19. Thirty different PFGE profiles were identified, with 72% of the isolates showing a single profile. Multidrug resistance, DCS and azithromycin resistance were 30.3%, 15.4% and 1.0%, respectively. DCS was associated with point mutations in the gyrA gene at codons 83 and 87., Conclusions/significance: Our study describes the first report of widespread multidrug resistance and DCS among Salmonella Typhi isolates from DR Congo. Our findings highlight the need for increased microbiological diagnosis and surveillance in DR Congo, being a prerequisite for rational use of antimicrobials and the development of standard treatment guidelines.

Background: Salmonella enterica is a frequent cause of bloodstream infection (BSI) in Asia but few data are available from Cambodia. We describe Salmonella BSI isolates recovered from patients presenting at Sihanouk Hospital Centre of Hope, Phnom Penh, Cambodia (July 2007-December 2010)., Methodology: Blood was cultured as part of a microbiological prospective surveillance study. Identification of Salmonella isolates was performed by conventional methods and serotyping. Antibiotic susceptibilities were assessed using disk diffusion, MicroScan and E-test macromethod. Clonal relationships were assessed by Pulsed Field Gel Electrophoresis; PCR and sequencing for detection of mutations in Gyrase and Topoisomerase IV and presence of qnr genes., Principal Findings: Seventy-two Salmonella isolates grew from 58 patients (mean age 34.2 years, range 8-71). Twenty isolates were identified as Salmonella Typhi, 2 as Salmonella Paratyphi A, 37 as Salmonella Choleraesuis and 13 as other non-typhoid Salmonella spp. Infection with human immunodeficiency virus (HIV) was present in 21 of 24 (87.5%) patients with S. Choleraesuis BSI. Five patients (8.7%) had at least one recurrent infection, all with S. Choleraesuis; five patients died. Overall, multi drug resistance (i.e., co-resistance to ampicillin, sulphamethoxazole-trimethoprim and chloramphenicol) was high (42/59 isolates, 71.2%). S. Typhi displayed high rates of decreased ciprofloxacin susceptibility (18/20 isolates, 90.0%), while azithromycin resistance was very common in S. Choleraesuis (17/24 isolates, 70.8%). Two S. Choleraesuis isolates were extended spectrum beta-lactamase producer., Conclusions and Significance: Resistance rates in Salmonella spp. in Cambodia are alarming, in particular for azithromycin and ciprofloxacin. This warrants nationwide surveillance and revision of treatment guidelines.

Background: The basally divergent phylogenetic position of amphioxus (Cephalochordata), as well as its conserved morphology, development and genetics, make it the best proxy for the chordate ancestor. Particularly, studies using the amphioxus model help our understanding of vertebrate evolution and development. Thus, interest for the amphioxus model led to the characterization of both the transcriptome and complete genome sequence of the American species, Branchiostoma floridae. However, recent technical improvements allowing induction of spawning in the laboratory during the breeding season on a daily basis with the Mediterranean species Branchiostoma lanceolatum have encouraged European Evo-Devo researchers to adopt this species as a model even though no genomic or transcriptomic data have been available. To fill this need we used the pyrosequencing method to characterize the B. lanceolatum transcriptome and then compared our results with the published transcriptome of B. floridae., Results: Starting with total RNA from nine different developmental stages of B. lanceolatum, a normalized cDNA library was constructed and sequenced on Roche GS FLX (Titanium mode). Around 1.4 million of reads were produced and assembled into 70,530 contigs (average length of 490 bp). Overall 37% of the assembled sequences were annotated by BlastX and their Gene Ontology terms were determined. These results were then compared to genomic and transcriptomic data of B. floridae to assess similarities and specificities of each species., Conclusion: We obtained a high-quality amphioxus (B. lanceolatum) reference transcriptome using a high throughput sequencing approach. We found that 83% of the predicted genes in the B. floridae complete genome sequence are also found in the B. lanceolatum transcriptome, while only 41% were found in the B. floridae transcriptome obtained with traditional Sanger based sequencing. Therefore, given the high degree of sequence conservation between different amphioxus species, this set of ESTs may now be used as the reference transcriptome for the Branchiostoma genus.

Ward and Zahavi suggested in 1973 that colonies could serve as information centres, through a transfer of information on the location of food resources between unrelated individuals (Information Centre Hypothesis). Using GPS tracking and observations on group movements, we studied the search strategy and information transfer in two of the most colonial seabirds, Guanay cormorants (Phalacrocorax bougainvillii) and Peruvian boobies (Sula variegata). Both species breed together and feed on the same prey. They do return to the same feeding zone from one trip to the next indicating high unpredictability in the location of food resources. We found that the Guanay cormorants use social information to select their bearing when departing the colony. They form a raft at the sea surface whose position is continuously adjusted to the bearing of the largest returning columns of cormorants. As such, the raft serves as a compass signal that gives an indication on the location of the food patches. Conversely, Peruvian boobies rely mainly on personal information based on memory to take heading at departure. They search for food patches solitarily or in small groups through network foraging by detecting the white plumage of congeners visible at long distance. Our results show that information transfer does occur and we propose a new mechanism of information transfer based on the use of rafts off colonies. The use of rafts for information transfer may be common in central place foraging colonial seabirds that exploit short lasting and/or unpredictably distributed food patches. Over the past decades Guanay cormorants have declined ten times whereas Peruvian boobies have remained relatively stable. We suggest that the decline of the cormorants could be related to reduced social information opportunities and that social behaviour and search strategies have the potential to play an important role in the population dynamics of colonial animals.

Nuclear receptors (NRs) are transcription factors that are implicated in several biological processes such as embryonic development, homeostasis, and metabolic diseases. To study the role of NRs in development, it is critically important to know when and where individual genes are expressed. Although systematic expression studies using reverse transcriptase PCR and/or DNA microarrays have been performed in classical model systems such as Drosophila and mouse, no systematic atlas describing NR involvement during embryonic development on a global scale has been assembled. Adopting a systems biology approach, we conducted a systematic analysis of the dynamic spatiotemporal expression of all NR genes as well as their main transcriptional coregulators during zebrafish development (101 genes) using whole-mount in situ hybridization. This extensive dataset establishes overlapping expression patterns among NRs and coregulators, indicating hierarchical transcriptional networks. This complete developmental profiling provides an unprecedented examination of expression of NRs during embryogenesis, uncovering their potential function during central nervous system and retina formation. Moreover, our study reveals that tissue specificity of hormone action is conferred more by the receptors than by their coregulators. Finally, further evolutionary analyses of this global resource led us to propose that neofunctionalization of duplicated genes occurs at the levels of both protein sequence and RNA expression patterns. Altogether, this expression database of NRs provides novel routes for leading investigation into the biological function of each individual NR as well as for the study of their combinatorial regulatory circuitry within the superfamily., Competing Interests: Competing interests. The authors have declared that no competing interests exist.

Understanding the role of gene duplications in establishing vertebrate innovations is one of the main challenges of Evo-Devo (evolution of development) studies. Data on evolutionary changes in gene expression (i.e., evolution of transcription factor-cis-regulatory elements relationships) tell only part of the story; protein function, best studied by biochemical and functional assays, can also change. In this study, we have investigated how gene duplication has affected both the expression and the ligand-binding specificity of retinoic acid receptors (RARs), which play a major role in chordate embryonic development. Mammals have three paralogous RAR genes--RAR alpha, beta, and gamma--which resulted from genome duplications at the origin of vertebrates. By using pharmacological ligands selective for specific paralogues, we have studied the ligand-binding capacities of RARs from diverse chordates species. We have found that RAR beta-like binding selectivity is a synapomorphy of all chordate RARs, including a reconstructed synthetic RAR representing the receptor present in the ancestor of chordates. Moreover, comparison of expression patterns of the cephalochordate amphioxus and the vertebrates suggests that, of all the RARs, RAR beta expression has remained most similar to that of the ancestral RAR. On the basis of these results together, we suggest that while RAR beta kept the ancestral RAR role, RAR alpha and RAR gamma diverged both in ligand-binding capacity and in expression patterns. We thus suggest that neofunctionalization occurred at both the expression and the functional levels to shape RAR roles during development in vertebrates., Competing Interests: Competing interests. The authors have declared that no competing interests exist.