Neural stem cells characterization in the vagal complex of adult ovine brain: A combined neurosphere assay/RTqPCR approach.

Neural stem cells are the effectors of adult neurogenesis, which occurs in discrete restricted areas of adult mammalian brain. In ovine species, like in rodents, in vivo incorporation of labeled DNA precursor led to characterize neurogenic proliferation in the subventricular zone and progeny migration and differentiation into the olfactory bulb. The present study addresses directly the existence of neural stem cells in the neurogenic niche of the vagal centre (area postrema) by in vitro neurosphere assay and RT-qPCR of specific markers on ex-vivo adult tissue explants, comparatively with the canonical neurogenic niche: the subventricular zone (SVZ) of the forebrain. Explants defined from the neuroanatomical patterns of in vivo BrdU incorporation yielded expandable and self-renewing spheres from both SVZ and AP. Within SVZ though, the density of sphere-forming cells was higher in ventral SVZ (SVZ-V) than in its latero-dorsal (SVZ-D) and lateral (SVZ-L) regions, which differs from the distributions of neural stem cells in mouse and swine brains. Consistently, RT-qPCR of the biomarker of neural stem cells, Sox2, yields highest expression in SVZ-V ahead of SVZ-D, SVZ-L and AP. These results are discussed with regard to previously published dynamics of adult ovine neurogenesis in vivo, and in light of corresponding features in other mammalian species. This confirms existence of neurogenetic plasticity in the vagal complex of adult mammals.

Detection of the protistan parasite, Haplosporidium costale in Crassostrea gigas oysters from the French coast: A retrospective study.

The parasite Haplosporidium costale is known to infect and cause mortality in the oyster Crassostrea virginica in the USA. Decades after its first description in the 1960s, this parasite was detected in Crassostrea gigas in the USA and China. However, it presented a low prevalence and no mortality was associated with it. More recently, in 2019, H. costale was detected in France in a batch of moribund oysters. In order to observe how long this parasite has been present on French coasts, from Normandy to Thau lagoon, a retrospective investigation was conducted on 871 adult and spat oyster batches from 2004 to 2020. To allow rapid detection on a large panel of samples, a real-time PCR for the H. costale actin gene was developed. This method allowed the detection of H. costale DNA in adults from 2005 and in spat from 2008. The H. costale prevalence in spat appeared higher than in adults over the years studied, 14.59 % compared to 6.50 %, respectively. All samples presenting positive results were then sequenced on two targets, H. costale rRNA and actin genes. The actin gene sequencing highlighted the presence of two H. costale strains. Adult C. gigas as well as spat batches coming from hatcheries and DNA controls from C. virginica all presented with the Profile 1 H. costale strain. The Profile 2 H. costale strain was detected only in C. gigas spat coming from natural sources. These observations suggest a correlation between the origin of oysters and H. costale strains which may have been caused by commercial imports between Japan, USA and France back to the 1970s. Over the positive samples studied, only few batches (n = 3) suffered mortalities which could be hypothesized to be caused by H. costale, all presenting the Profile 1 H. costale strain.

Protein expression reveals a molecular sexual identity of avian primordial germ cells at pre-gonadal stages.

In poultry, in vitro propagated primordial germ cells (PGCs) represent an important tool for the cryopreservation of avian genetic resources. However, several studies have highlighted sexual differences exhibited by PGCs during in vitro propagation, which may compromise their reproductive capacities. To understand this phenomenon, we compared the proteome of pregonadal migratory male (ZZ) and female (ZW) chicken PGCs propagated in vitro by quantitative proteomic analysis using a GeLC-MS/MS strategy. Many proteins were found to be differentially abundant in chicken male and female PGCs indicating their early sexual identity. Many of the proteins more highly expressed in male PGCs were encoded by genes localised to the Z sex chromosome. This suggests that the known lack of dosage compensation of the transcription of Z-linked genes between sexes persists at the protein level in PGCs, and that this may be a key factor of their autonomous sex differentiation. We also found that globally, protein differences do not closely correlate with transcript differences indicating a selective translational mechanism in PGCs. Male and female PGC expressed protein sets were associated with differential biological processes and contained proteins known to be biologically relevant for male and female germ cell development, respectively. We also discovered that female PGCs have a higher capacity to uptake proteins from the cell culture medium than male PGCs. This study presents the first evidence of an early predetermined sex specific cell fate of chicken PGCs and their sexual molecular specificities which will enable the development of more precise sex-specific in vitro culture conditions for the preservation of avian genetic resources.

Data for evolutive analysis of insulin related peptides in bilaterian species.

In bilaterian species, the amino acid sequence conservation between Insulin related peptides is relatively low except for the cysteine residues involved in the disulphide bonds. In the A chain, the conserved cystein residues are included in a signature motif. Investigating the variations in this motif would give insight into the phylogenetic history of the family. The table presented in this paper contains a large set of insulin-related peptides in bilateral phylogenetic groups (deuterostomian, ecdysozoan, lophotrochozoan). NCBI databases in silico wide screening combined with bibliographic researches provided a framework for identifying and categorising the structural characteristics of these insulin related peptides. The dataset includes NCBI IDs of each sequence with hyperlinks to FASTA format. Moreover, the structural type (α, ß or γ), the A chain motif, the total number of cysteins, the C peptide cleavage mode and the potential additional domains (D or E) are specified for each sequence. The data are associated with the research article "Molecular evolution and functional characterisation of insulin-related peptides in molluscs: contributions of Crassostrea gigas genomic and transcriptomic-wide screening" [1]. The table presented here can be found at http://dx.doi.org/10.17632/w4gr8zcpk5.4#file-21c0f6a5-a3e3-4a15-86e0-e5a696458866.

Morphological and molecular criteria allow the identification of putative germ stem cells in a lophotrochozoan, the Pacific oyster Crassostrea gigas.

While our knowledge of bivalve gametogenesis recently progressed, data on early stages of gametogenesis remain to be developed, especially when dealing with germinal stem cells (GSC) and their niche in these organisms. Here, we wish to develop a strategy to identify putative GSC in Pacific oyster Crassostrea gigas based on morphological criteria combined with vasa marker expression. A histological quantitative approach, based on stereology, allowed us to identify two types of early germ cells in the germinal epithelium, one presenting round nuclei and the other irregular ones. Both early germ cell types present slightly condensed chromatin in nucleus, are vasa-positive and the Oyvlg (oyster vasa-like gene) expression in these cells is recorded throughout the whole gametogenesis process. The microenvironment of an early germ cell in oyster includes an associated somatic cell presenting an immunolabeling for BMP2/4 and a close myoid cell. In agreement with the GSC characteristics in other species, we postulate that putative germ stem cells in C. gigas correspond to the early germ cell type with irregular nucleus shape; those early germ cells with a round nucleus may consist in progenitors.

Molecular evolution and functional characterisation of insulin related peptides in molluscs: Contributions of Crassostrea gigas genomic and transcriptomic-wide screening.

Insulin Related Peptides (IRPs) belong to the insulin superfamily and possess a typical structure with two chains, B and A, linked by disulphide bonds. As the sequence conservation is usually low between members, IRPs are classified according to the number and position of their disulphide bonds. In molluscan species, the first IRPs identified, named Molluscan Insulin-related Peptides (MIPs), exhibit four disulphide bonds. The genomic and transcriptomic data screening in the Pacific oyster Crassostrea gigas (Mollusc, Bivalvia) allowed us to identify six IRP sequences belonging to three structural groups. Cg-MIP1 to 4 have the typical structure of MIPs with four disulphide bonds. Cg-ILP has three disulphide bonds like vertebrate Insulin-Like Peptides (ILPs). The last one, Cg-MILP7 has a significant homology with Drosophila ILP7 (DILP7) associated with two additional cysteines allowing the formation of a fourth disulphide bond. The phylogenetic analysis points out that ILPs may be the most ancestral form. Moreover, it appears that ILP7 orthologs are probably anterior to lophotrochozoa and ecdysozoa segregation. In order to investigate the diversity of physiological functions of the oyster IRPs, we combine in silico expression data, qPCR measurements and in situ hybridization. The Cg-ilp transcript, mainly detected in the digestive gland and in the gonadal area, is potentially involved in the control of digestion and gametogenesis. The expression of Cg-mip4 is mainly associated with the larval development. The Cg-mip transcript shared by the Cg-MIP1, 2 and 3, is mainly expressed in visceral ganglia but its expression was also observed in the gonads of mature males. This pattern suggested the key roles of IRPs in the control of sexual reproduction in molluscan species.