Estimation of Aquifer Transmissivity From Analysis of Long-Term Monitoring With the Thiem Solution.

The application of the Thiem equation to support the interpretation of comprehensive long-term monitoring datasets, made possible through modern datalogging technology, is presented as an alternative to constant-rate aquifer testing to obtain representative transmissivity estimates in settings where controlled hydraulic testing may be impractical. Water levels logged at regular intervals can be readily converted to average water levels over time periods corresponding to periods of known pumping rates. By regressing average water levels during multiple time periods of known but variable withdrawal rates, steady-state conditions can be approximated and Thiem's solution applied to estimate transmissivity, without performance of a constant-rate aquifer test. Although the application is limited to settings where changes in aquifer storage are negligible, by regressing long data sets to parse interferences the method may characterize aquifer conditions over a much wider radius than short-term, non-equilibrium tests. As with all aquifer testing, informed interpretation is critical to identifying and resolving aquifer heterogeneities and interferences.

Differential analysis of the haemolymph proteome of Carcinus maenas parasitized by Sacculina carcini (Cirripeda, Rhizocephala) reveals potential mechanisms of parasite control.

Sacculina carcini is an endoparasite of the green crab, Carcinus maenas. This parasite induces behavioural changes in its host and affects its metabolism by inhibiting moulting and reproduction. Using a proteomic approach in mass spectrometry, we studied the haemolymph proteomes of healthy and parasitized wild green crabs from Brittany, France to identify proteins that are differentially expressed as a consequence of parasitization. We also investigated specific proteins involved in reproduction, moulting, and immunity. We detected 77 proteins for females and 53 proteins for males that were differentially present between the healthy and parasitized crabs, some of which were sex-specific. Detection of these differentially expressed proteins suggests that the parasite can inhibit and promote different aspects of the immune response of the host. Sacculina appears to inhibit host melanisation for self-protection, while promoting the presence of immune factors, such as antimicrobial peptides to cope with possible bacterial superinfections. Moreover, one protein, juvenile hormone esterase-like carboxylesterase, was 17-times more abundant in parasitized crabs than in healthy crabs and may be responsible for inhibiting moulting and reproduction in parasitized crabs, thus ensuring the success of Sacculina reproduction.

PepTraq: a toolbox for in silico data mining and fast sequence filtering.

The development of de novo sequencing tools has led to the massive production of genomes and transcriptomes from many unconventional animal models. To tackle this huge flow of data, PepTraq brings together many functionalities generally scattered in multiple tools, so that sequences can be filtered on the basis of multiple criteria. It is particularly suitable for the identification of non-annotated transcripts, re-annotation, extraction of secretomes, neuropeptidomes, targeted search for peptides and proteins, preparing specific proteomics/peptidomics fasta files for mass spectrometry (MS) applications, MS data processing, etc. PepTraq is developed in Java, and is available as a desktop application that can be downloaded from https://peptraq.greyc.fr . It is also available as a web application at the same URL for processing small files (10-20 MB). The source code is open under a CeCILL-B licence.

Structural and Functional Characterization of Orcokinin B-like Neuropeptides in the Cuttlefish (Sepia officinalis).

The cuttlefish (Sepia officinalis) is a Cephalopod mollusk that lives in the English Channel and breeds in coastal spawning grounds in spring. A previous work showed that the control of egg-laying is monitored by different types of regulators, among which neuropeptides play a major role. They are involved in the integration of environmental cues, and participate in the transport of oocytes in the genital tract and in the secretion of capsular products. This study addresses a family of neuropeptides recently identified and suspected to be involved in the control of the reproduction processes. Detected by mass spectrometry and immunocytochemistry in the nerve endings of the accessory sex glands of the females and ovary, these neuropeptides are also identified in the hemolymph of egg-laying females demonstrating that they also have a hormone-like role. Released in the hemolymph by the sub-esophageal mass, a region that innervates the genital tract and the neurohemal area of the vena cava, in in vitro conditions these neuropeptides modulated oocyte transport and capsular secretion. Finally, in silico analyses indicated that these neuropeptides, initially called FLGamide, had extensive structural homology with orcokinin B, which motivated their name change.

Identification of a New Set of Polypeptidic Sex Pheromones from Cuttlefish (Sepia officinalis).

The common English Channel cuttlefish (Sepia officinalis) reproduces every year on very localized coastal spawning areas after a west-east horizontal migration of several tens of kilometers (80-200 km). The massive arrival of spawners on the coasts of west Cotentin and the Bay of Seine is suspected to be driven by the action of sex pheromones expressed and secreted by the genitals of sexually mature females. The present study aims to verify the existence of polypeptide pheromones, of a higher molecular weight than those described above. Their size could confer them a wider range of action than that of the previously identified peptide pheromones. The implementation of an experimental strategy combining transcriptomics and proteomics with functional tests and an in silico study led to the identification of a cocktail of pheromones with molecular weights ranging between 22 and 26 kDa. Proteomic analyses combined to functional tests revealed partial pheromone release in the environment, and their accumulation in the outer capsule of the egg, suggesting the eggs as pheromone diffusers, also able to induce stimulation by contact when the eggs are handled by females.

Rapid Cleanup of a Perchlorate Plume from Fireworks.

Perchlorate was detected in a municipal wellfield in Evart, Michigan in April 2015. Perchlorate concentrations were detected initially in six of the City's wells at concentrations ranging up to 20 µg/L. An investigation to identify the source determined that the perchlorate was from fireworks launched during the annual 4th of July show held at the fairgrounds located upgradient from the wellfield. The use of approximately 600 kg of fireworks during the annual display resulted in an annual loading of approximately 4 kg of perchlorate to groundwater. An aggressive groundwater extraction system began operation in June 2016 to restore water quality in the affected aquifer, and the 2016 fireworks display was relocated to a location outside the capture zone of the water supply wells. Within 18 months average perchlorate concentrations in the water supply wells had been reduced to about 0.6 µg/L. The extraction system continued to operate through the end of 2019, by which time the average perchlorate concentrations in the water supply wells were reduced to 0.2 µg/L. In 2019, approximately 0.4 kg of perchlorate were removed from the aquifer, about one-half of the amount removed in 2018, reflecting the slow leaching of perchlorate of fireworks residuals from vadose zone soils.

School Engagement Among Youth Entering Probation.

Strong school engagement is crucial for school success among adolescents and particularly important for reducing recidivism. Yet, little is known about school engagement among youth serving probation while attending community schools. This study tested the multivariate associations between risk and promotive factors with three components of school engagement (behavioral, cognitive, and emotional). The study's sample was derived from 5,378 intake assessments (23.6% female) of youth entering juvenile probation in a Pacific Northwest county who were assessed as either moderate or high risk for recidivism. The racial composition of the sample was predominantly White or European American (56.0%) and Black or African American (24.2%) and ranged in age from 10 to 19 years old (M = 15.5, SD = 1.46). The results suggest that dimensions of school engagement can be strengthened by increased relational and skill-building supports throughout youth's social ecologies. The promotive factors of prosocial attitudes and prosocial community connections were significantly associated with increased school engagement. The implications of these findings are discussed regarding opportunities and strategies that promote school engagement for youth on probation.

In-Depth In Silico Search for Cuttlefish (Sepia officinalis) Antimicrobial Peptides Following Bacterial Challenge of Haemocytes.

Cuttlefish (Sepia officinalis) haemocytes are potential sources of antimicrobial peptides (AMPs). To study the immune response to Vibrio splendidus and identify new AMPs, an original approach was developed based on a differential transcriptomic study and an in-depth in silico analysis using multiple tools. Two de novo transcriptomes were retrieved from cuttlefish haemocytes following challenge by V. splendidus or not. A first analysis of the annotated transcripts revealed the presence of Toll/NF-κB pathway members, including newly identified factors such as So-TLR-h, So-IKK-h and So-Rel/NF-κB-h. Out of the eight Toll/NF-κB pathway members, seven were found up-regulated following V. splendidus challenge. Besides, immune factors involved in the immune response were also identified and up-regulated. However, no AMP was identified based on annotation or conserved pattern searches. We therefore performed an in-depth in silico analysis of unannotated transcripts based on differential expression and sequence characteristics, using several tools available like PepTraq, a homemade software program. Finally, five AMP candidates were synthesized. Among them, NF19, AV19 and GK28 displayed antibacterial activity against Gram-negative bacteria. Each peptide had a different spectrum of activity, notably against Vibrio species. GK28-the most active peptide-was not haemolytic, whereas NF19 and AV19 were haemolytic at concentrations between 50 and 100 µM, 5 to 10 times higher than their minimum inhibitory concentration.

-Omic Analysis of the Sepia officinalis White Body: New Insights into Multifunctionality and Haematopoiesis Regulation.

Cephalopods, like other protostomes, lack an adaptive immune system and only rely on an innate immune system. The main immune cells are haemocytes (Hcts), which are able to respond to pathogens and external attacks. First reports based on morphological observations revealed that the white body (WB) located in the optic sinuses of cuttlefish was the origin of Hcts. Combining transcriptomic and proteomic analyses, we identified several factors known to be involved in haematopoiesis in vertebrate species in cuttlefish WB. Among these factors, members of the JAK-STAT signaling pathway were identified, some of them for the first time in a molluscan transcriptome and proteome. Immune factors, such as members of the Toll/NF-κB signaling pathway, pattern recognition proteins and receptors, and members of the oxidative stress responses, were also identified, and support an immune role of the WB. Both transcriptome and proteome analyses revealed that the WB harbors an intense metabolism concurrent with the haematopoietic function. Finally, a comparative analysis of the WB and Hct proteomes revealed many proteins in common, confirming previous morphological studies on the origin of Hcts in cuttlefish. This molecular work demonstrates that the WB is multifunctional and provides bases for haematopoiesis regulation in cuttlefish.

Identification and structural characterization of the factors involved in vitellogenesis and its regulation in the African Osteoglossiforme of aquacultural interest Heterotis niloticus (Cuvier, 1829).

The African bonytongue (Heterotis niloticus) is an excellent candidate for fish farming because it has outstanding biological characteristics and zootechnical performances. However, the absence of sexual dimorphism does not favor its reproduction in captivity or the understanding of its reproductive behavior. Moreover, no molecular data related to its reproduction is yet available. This study therefore focuses on the structural identification of the different molecular actors of vitellogenesis expressed in the pituitary gland, the liver and the ovary of H. niloticus. A transcriptomic approach based on de novo RNA sequencing of the pituitary gland, ovary and liver of females in vitellogenesis led to the creation of three transcriptomes. In silico analysis of these transcriptomes identified the sequences of pituitary hormones such as prolactin (PRL), luteinizing hormone (LH) and follicle-stimulating hormone (FSH) and their ovarian receptors (PRLR, FSHR, LHR). In the liver and ovary, estrogen receptors (ER) beta and gamma, liver vitellogenins (VtgB and VtgC) and their ovarian receptors (VLDLR) were identified. Finally, the partial transcript of an ovarian Vtg weakly expressed compared to hepatic Vtg was identified based on structural criteria. Moreover, a proteomic approach carried out from mucus revealed the presence of one Vtg exclusively in females in vitellogenesis. In this teleost fish that does not exhibit sexual dimorphism, mucus Vtg could be used as a sexing biomarker based on a non-invasive technique compatible with the implementation of experimental protocols in vivo.

A standardized, innovative method to characterize the structure of aquatic protein hydrolysates.

The performances of protein hydrolysates highly depend on their peptide composition (amount, size and diversity), which itself closely depends on raw material origin and the hydrolysis parameters of the manufacturing process. The current analyses that characterize protein hydrolysates provide information on the level of hydrolysis (degree of hydrolysis, DH). However, they need additional describers to better characterize peptide profiles and product standardization. To reach this objective, we developed a fast and standardized method to characterize the abundance and the diversity of low-molecular-weight peptides in protein hydrolysates. This method innovatively combines classical HPSEC and nLC-ESI-MS analytical tools to characterize any kind of hydrolysate, whether solid or liquid, in terms of peptide level and diversity, and then merge peptides into 2D diagrams to visualize comparisons between protein hydrolysates. The targeted applications of this new tool for characterizing complex protein hydrolysates are (i) verifying the standardization of the produced products across batches, and (ii) analyzing and understanding the consequences of the modifications of the hydrolysis process on the molecular profiles of the generated peptides. The sample standardization described in this study is therefore an essential prerequisite for the functional characterization of hydrolysates in vitro.

Cumulative Adversity Profiles Among Youth Experiencing Housing and Parental Care Instability.

This study applies cumulative adversity and stress proliferation theories to examine risk and protective resource profiles of youth with three different levels of housing and parental care instability. Data derive from a state representative sample (n=27,087) of school-based adolescent students. ANCOVA analyses identified significant differences in sociodemographic and psychosocial functioning variables for youth with 0, 1, or 2 forms of housing and parental care instability, with more deleterious functioning being observed among youth with greater levels of instability. Those experiencing either or both housing and parental care instability are more represented by males, sexual minorities, and youth of color; psychosocial risk and protective factors demonstrated consistent differences between instability groups. Dimensions of cumulative adversity operate with social marginalities (e.g., race, sexual minority status) relative to instability, with higher frequency of victimization, lower grades, diminished self-regulation capabilities and school engagement, weakened psychological health, and strained family and peer relationships. The paper discusses theorized mechanisms through which cumulative adversity conveys effects as well as implications for social work prevention and resilience-fostering strategies in schools and other youth-serving settings.

A FHIR-Enabled Streaming Sepsis Prediction System for ICUs.

Sepsis is a common disease with very costly, potentially deadly implications. Early prediction of Sepsis and initiation of antibiotic is widely considered as an important determinant of patient survival. Cross-institutional validation and implementation of algorithms for early prediction of Sepsis at a minimum require common data formats, streaming analytic platforms for timely risk assessment, and interoperable and standardized interfaces. In this work we present an open-source cloud-based analytic pipeline, which receives de-identified patient data from an interoperable server and produces real-time Sepsis risk scores for Intensive Care Unit (ICU) patients.

Design of antimicrobial peptides from a cuttlefish database.

No antimicrobial peptide has been identified in cephalopods to date. Annotation of transcriptomes or genomes using basic local alignment Search Tool failed to yield any from sequence identities. Therefore, we searched for antimicrobial sequences in the cuttlefish (Sepia officinalis) database by in silico analysis of a transcriptomic database. Using an original approach based on the analysis of cysteine-free antimicrobial peptides selected from our Antimicrobial Peptide Database (APD3), the online prediction tool of the Collection of Anti-Microbial Peptides (CAMPR3), and a homemade software program, we identified potential antibacterial sequences. Nine peptides less than 25 amino acids long were synthesized. The hydrophobic content of all nine of them ranged from 30 to 70%, and they could form alpha-helices. Three peptides possessed similarities with piscidins, one with BMAP-27, and five were totally new. Their antibacterial activity was evaluated on eight bacteria including the aquatic pathogens Vibrio alginolyticus, Aeromonas salmonicida, or human pathogens such as Salmonella typhimurium, Listeria monocytogenes, or Staphylococcus aureus. Despite the prediction of an antimicrobial potential for eight of the peptides, only two-GR21 and KT19-inhibited more than one bacterial strain with minimal inhibitory concentrations below 25 µM. Some sequences like VA20 and FK19 were hemolytic, while GR21 induced less than 10% of hemolysis on human blood cells at a concentration of 200 µM. GR21 was the only peptide derived from a precursor with a signal peptide, suggesting a real role in cuttlefish immune defense.

Dietary aquaculture by-product hydrolysates: impact on the transcriptomic response of the intestinal mucosa of European seabass (Dicentrarchus labrax) fed low fish meal diets.

BACKGROUND: Aquaculture production is expected to double by 2030, and demands for aquafeeds and raw materials are expected to increase accordingly. Sustainable growth of aquaculture will require the development of highly nutritive and functional raw materials to efficiently replace fish meal. Enzymatic hydrolysis of marine and aquaculture raw materials could bring new functionalities to finished products. The aim of this study was to determine the zootechnical and transcriptomic performances of protein hydrolysates of different origins (tilapia, shrimp, and a combination of the two) in European seabass (Dicentrarchux labrax) fed a low fish meal diet (5%), for 65 days. RESULTS: Results were compared to a positive control fed with 20% of fish meal. Growth performances, anterior intestine histological organization and transcriptomic responses were monitored and analyzed. Dietary inclusion of protein hydrolysates in the low fish meal diet restored similar growth performances to those of the positive control. Inclusion of dietary shrimp hydrolysate resulted in larger villi and more goblet cells, even better than the positive control. Transcriptomic analysis of the anterior intestine showed that dietary hydrolysate inclusion restored a pattern of intestinal gene expression very close to the pattern of the positive control. However, as compared to the low fish meal diet and depending on their origin, the different hydrolysates did not modulate metabolic pathways in the same way. Dietary shrimp hydrolysate inclusion modulated more metabolic pathways related to immunity, while nutritional metabolism was more impacted by dietary tilapia hydrolysate. Interestingly, the combination of the two hydrolysates enhanced the benefits of hydrolysate inclusion in diets: more genes and metabolic pathways were regulated by the combined hydrolysates than by each hydrolysate tested independently. CONCLUSIONS: Protein hydrolysates manufactured from aquaculture by-products are promising candidates to help replace fish meal in aquaculture feeds without disrupting animal metabolism and performances.

Crustacean cardioactive peptides: Expression, localization, structure, and a possible involvement in regulation of egg-laying in the cuttlefish Sepia officinalis.

The cuttlefish (Sepia officinalis) is a cephalopod mollusk distributed on the western European coast, in the West African Ocean and in the Mediterranean Sea. On the Normandy coast (France), cuttlefish is a target species of professional fishermen, so its reproduction strategy is of particular interest in the context of stock management. Egg-laying, which is coastal, is controlled by several types of regulators among which neuropeptides. The cuttlefish neuropeptidome was recently identified by Zatylny-Gaudin et al. (2016). Among the 38 neuropeptide families identified, some were significantly overexpressed in egg-laying females as compared to mature males. This study is focused on crustacean cardioactive peptides (CCAPs), a highly expressed neuropeptide family strongly suspected of being involved in the control of egg-laying. We investigated the functional and structural characterization and tissue mapping of CCAPs, as well as the expression patterns of their receptors. CCAPs appeared to be involved in oocyte transport through the oviduct and in mechanical secretion of capsular products. Immunocytochemistry revealed that the neuropeptides were localized throughout the central nervous system (CNS) and in the nerve endings of the glands involved in egg-capsule synthesis and secretion, i.e. the oviduct gland and the main nidamental glands. The CCAP receptor was expressed in these glands and in the subesophageal mass of the CNS. Multiple sequence alignments revealed a high level of conservation of CCAP protein precursors in Sepia officinalis and Loligo pealei, two cephalopod decapods. Primary sequences of CCAPs from the two species were fully conserved, and cryptic peptides detected in the nerve endings were also partially conserved, suggesting biological activity that remains unknown for the time being.

Neuropeptidome of the Cephalopod Sepia officinalis: Identification, Tissue Mapping, and Expression Pattern of Neuropeptides and Neurohormones during Egg Laying.

Cephalopods exhibit a wide variety of behaviors such as prey capture, communication, camouflage, and reproduction thanks to a complex central nervous system (CNS) divided into several functional lobes that express a wide range of neuropeptides involved in the modulation of behaviors and physiological mechanisms associated with the main stages of their life cycle. This work focuses on the neuropeptidome expressed during egg-laying through de novo construction of the CNS transcriptome using an RNAseq approach (Illumina sequencing). Then, we completed the in silico analysis of the transcriptome by characterizing and tissue-mapping neuropeptides by mass spectrometry. To identify neuropeptides involved in the egg-laying process, we determined (1) the neuropeptide contents of the neurohemal area, hemolymph (blood), and nerve endings in mature females and (2) the expression levels of these peptides. Among the 38 neuropeptide families identified from 55 transcripts, 30 were described for the first time in Sepia officinalis, 5 were described for the first time in the animal kingdom, and 14 were strongly overexpressed in egg-laying females as compared with mature males. Mass spectrometry screening of hemolymph and nerve ending contents allowed us to clarify the status of many neuropeptides, that is, to determine whether they were neuromodulators or neurohormones.

The Toll/NF-κB pathway in cuttlefish symbiotic accessory nidamental gland.

The female genital apparatus of decapod cephalopods contains a symbiotic accessory nidamental gland (ANG) that harbors bacterial symbionts. Although the ANG bacterial consortium is now well described, the impact of symbiosis on Sepia officinalis innate immunity pathways remains unknown. In silico analysis of the de novo transcriptome of ANG highlighted for the first time the existence of the NF-κB pathway in S. officinalis. Several signaling components were identified, i.e. five Toll-like receptors, eight signaling cascade features, and the immune response target gene iNOS, previously described as being involved in the initiation of bacterial symbiosis in a cephalopod gland. This work provides a first key for studying bacterial symbiosis and its impact on innate immunity in S. officinalis ANG.

How Egg Case Proteins Can Protect Cuttlefish Offspring?

Sepia officinalis egg protection is ensured by a complex capsule produced by the female accessory genital glands and the ink bag. Our study is focused on the proteins constituting the main egg case. De novo transcriptomes from female genital glands provided essential databases for protein identification. A proteomic approach in SDS-PAGE coupled with MS unveiled a new egg case protein family: SepECPs, for Sepia officinalis Egg Case Proteins. N-glycosylation was demonstrated by PAS staining SDS-PAGE gels. These glycoproteins are mainly produced in the main nidamental glands. SepECPs share high sequence homology, especially in the signal peptide and the three cysteine-rich domains. SepECPs have a high number of cysteines, with conserved motifs involved in 3D-structure. SDS-PAGE showed that SepECPs could form dimers; this result was confirmed by TEM observations, which also revealed a protein network. This network is similar to the capsule network, and it associates these structural proteins with polysaccharides, melanin and bacteria to form a tight mesh. Its hardness and elasticity provide physical protection to the embryo. In addition, SepECPs also have bacteriostatic antimicrobial activity on GRAM- bacteria. By observing the SepECP / Vibrio aestuarianus complex in SEM, we demonstrated the ability of these proteins to agglomerate bacteria and thus inhibit their growth. These original proteins identified from the outer egg case ensure the survival of the species by providing physical and chemical protection to the embryos released in the environment without any maternal protection.

Neuropeptides encoded by the genomes of the Akoya pearl oyster Pinctata fucata and Pacific oyster Crassostrea gigas: a bioinformatic and peptidomic survey.

BACKGROUND: Oysters impart significant socio-ecological benefits from primary production of food supply, to estuarine ecosystems via reduction of water column nutrients, plankton and seston biomass. Little though is known at the molecular level of what genes are responsible for how oysters reproduce, filter nutrients, survive stressful physiological events and form reef communities. Neuropeptides represent a diverse class of chemical messengers, instrumental in orchestrating these complex physiological events in other species. RESULTS: By a combination of in silico data mining and peptide analysis of ganglia, 74 putative neuropeptide genes were identified from genome and transcriptome databases of the Akoya pearl oyster, Pinctata fucata and the Pacific oyster, Crassostrea gigas, encoding precursors for over 300 predicted bioactive peptide products, including three newly identified neuropeptide precursors PFGx8amide, RxIamide and Wx3Yamide. Our findings also include a gene for the gonadotropin-releasing hormone (GnRH) and two egg-laying hormones (ELH) which were identified from both oysters. Multiple sequence alignments and phylogenetic analysis supports similar global organization of these mature peptides. Computer-based peptide modeling of the molecular tertiary structures of ELH highlights the structural homologies within ELH family, which may facilitate ELH activity leading to the release of gametes. CONCLUSION: Our analysis demonstrates that oysters possess conserved molluscan neuropeptide domains and overall precursor organization whilst highlighting many previously unrecognized bivalve idiosyncrasies. This genomic analysis provides a solid foundation from which further studies aimed at the functional characterization of these molluscan neuropeptides can be conducted to further stimulate advances in understanding the ecology and cultivation of oysters.