Lise-Marie Pigneur

Physiological and proteomic evidences that domestication process differentially modulates the immune status of juvenile Eurasian perch (Perca fluviatilis) under chronic confinement stress.

The current study aimed to evaluate the influence of domestication process on the stress response and subsequent immune modulation in Eurasian perch juveniles (Perca fluviatilis) submitted to chronic confinement. Briefly, F1 and F4 generations were confined into small-size tanks and sampled 7 and 55 days after stocking. Cortisol and glucose levels as well as lysozyme activity and immunoglobulin level were evaluated in the serum. Spleen Somatic Index and spleen ROS production were also measured. A proteomic analysis was performed on serum sampled on day 7. Finally, both generations were genetically characterized using a microsatellite approach. Globally, results revealed that chronic confinement did not elicit a typical stress response but resulted in a prolonged immune stimulation. Proteomic results suggested that domestication process influenced the immune status of perch submitted to chronic confinement as the F1 confined fish displayed lower abundance of C3 complement component, transferrin and Apolipoprotein E. Microsatellite data showed a strong genetic drift as well as reduced genetic diversity, allelic number and heterozygosity along with domestication process. The present work is the first to report that fish under domestication can develop an immune response, assessed by a combined approach, following recurrent challenges imposed by captive environment despite a reduced genetic variation.

Bcl-2 and Caspase 3 mRNA levels in the testes of gudgeon, Gobio gobio, exposed to ethinylestradiol (EE2).

Apoptosis inhibition has been reported in the male reproductive tract of teleost fish exposed to 17beta-estrogen or estrogen-like compounds. In order to understand the molecular mechanisms of cell death inhibition, this study examined 2 genes involved in the apoptotic pathway, Bcl-2 and Caspase 3, an anti-apoptotic and a pro-apoptotic genes, respectively. Partial cDNA sequences of Bcl-2 and Caspase 3 were cloned from gudgeon (Gobio gobio), a common European cyprinid fish. To follow mRNA levels of Bcl-2 and Caspase 3 under xenoestrogen exposure, we first performed an in vitro experiment on fish testis exposed to the most potent xenoestrogen found in the environment, ethinylestradiol (EE2). We further studied mRNA expression of both genes in the testis of fish exposed to xenoestrogens in situ. In the in vitro experiment, fragments of gudgeon testis were exposed for 21 days to 10(-3), 10(-2), 10(-1), 1 and 10 microg/L of EE2, as well as to positive (10(-1) microg/L of E2) and ethanol control medium. Results showed a significant induction of Bcl-2 mRNA at 10(-1) microg/L (p<0.05). Surprisingly, Caspase 3, a cell death effector, displayed the same profile as observed for the anti-apoptotic gene Bcl-2. In the experiment on wild gudgeon exposed from birth to an estrogenic sewage treatment plant effluent, the mRNA expression of Bcl-2 and Caspase 3 in feminized fish (ovotestis) was not significantly different due to high variability of expression between individuals. At the current state of knowledge on spermatogenesis disruption in teleost fish, in vitro studies seem better adapted than in situ investigations to enlighten the molecular pathway of apoptosis inhibition in testis exposed to xenoestrogens.

Bacterial Community Composition in Three Freshwater Reservoirs of Different Alkalinity and Trophic Status

In order to investigate the factors controlling the bacterial community composition (BCC) in reservoirs, we sampled three freshwater reservoirs with contrasted physical and chemical characteristics and trophic status. The BCC was analysed by 16S rRNA gene amplicon 454 pyrosequencing. In parallel, a complete dataset of environmental parameters and phytoplankton community composition was also collected. BCC in the analysed reservoirs resembled that of epilimnetic waters of natural freshwater lakes with presence of Actinobacteria, Alpha- and Betaproteobacteria, Cytophaga–Flavobacteria–Bacteroidetes (CFB) and Verrucomicrobia groups. Our results evidenced that the retrieved BCC in the analysed reservoirs was strongly influenced by pH, alkalinity and organic carbon content, whereas comparatively little change was observed among layers in stratified conditions.

Development of novel microsatellite markers to identify the different invasive lineages in the Corbicula complex and to assess androgenesis.

Reliable markers are needed to identify the lineages in the invasive clam genus Corbicula. Previous studies have demonstrated that mitochondrial (mt) DNA poorly resolves Corbicula phylogeny, owing to its androgenetic reproductive mode. Moreover, hybridization and mitochondrial/nuclear mismatches occur. We developed the first eleven polymorphic markers to detect these phenomena and to investigate the nuclear identity of Corbicula populations. These microsatellite loci revealed three main lineages in Western Europe. One locus allowed rapid discrimination of these three lineages on agarose gel, saving time and money. Moreover, the eleven markers were successfully cross‐amplified in the invasive Corbicula lineages found in North America.

Genetic structure of fragmented southern populations of African Cape buffalo ( Syncerus caffer caffer )

African wildlife experienced a reduction in population size and geographical distribution over the last millennium, particularly since the 19th century as a result of human demographic expansion, wildlife overexploitation, habitat degradation and cattle-borne diseases. In many areas, ungulate populations are now largely confined within a network of loosely connected protected areas. These metapopulations face gene flow restriction and run the risk of genetic diversity erosion. In this context, we assessed the “genetic health” of free ranging southern African Cape buffalo populations (S.c. caffer) and investigated the origins of their current genetic structure. The analyses were based on 264 samples from 6 southern African countries that were genotyped for 14 autosomal and 3 Y-chromosomal microsatellites. The analyses differentiated three significant genetic clusters, hereafter referred to as Northern (N), Central (C) and Southern (S) clusters. The results suggest that splitting of the N and C clusters occurred around 6000 to 8400 years ago. Both N and C clusters displayed high genetic diversity (mean allelic richness (A r ) of 7.217, average genetic diversity over loci of 0.594, mean private alleles (P a ) of 11), low differentiation, and an absence of an inbreeding depression signal (mean F IS = 0.037). The third (S) cluster, a tiny population enclosed within a small isolated protected area, likely originated from a more recent isolation and experienced genetic drift (F IS = 0.062, mean A r = 6.160, P a = 2). This study also highlighted the impact of translocations between clusters on the genetic structure of several African buffalo populations. Lower differentiation estimates were observed between C and N sampling localities that experienced translocation over the last century. We showed that the current genetic structure of southern African Cape buffalo populations results from both ancient and recent processes. The splitting time of N and C clusters suggests that the current pattern results from human-induced factors and/or from the aridification process that occurred during the Holocene period. The more recent S cluster genetic drift probably results of processes that occurred over the last centuries (habitat fragmentation, diseases). Management practices of African buffalo populations should consider the micro-evolutionary changes highlighted in the present study.

Identification and characterization of a novel multicopper oxidase from Acidomyces acidophilus with ferroxidase activity

A new multicopper oxidase gene AaMco1 was identified in Acidomyces acidophilus, a pigmented extremophile ascomycete originally isolated from acidic water. Sequence analysis revealed that it encodes a 682 amino acid protein with an apparent molecular mass of 85 kDa as determined by denaturing SDS-PAGE. Interestingly, AaMco1 has a predicted N-terminal transmembrane helix and no signal peptide. To obtain an active and soluble protein, AaMco1 was truncated at its N-terminal to remove the transmembrane helix, but even in this form the protein was found in the insoluble fraction. AaMco1 and its truncated form were then denatured, purified and renatured before characterization. Structural analysis and protein characterization by enzymatic assays indicate that AaMco1 has ferroxidase activity. AaMco1 is also able to oxidize the DMPPDA compound and could be part of a new phylogenetic cluster, the ascomycete MCOs family, described for the first time here.