Persistence, Seasonal Dynamics and Pathogenic Potential of Vibrio Communities from Pacific Oyster Hemolymph.
PLoS One. 2014;9(4):e94256
Authors: Wendling CC, Batista FM, Wegner KM
Bacteria of the genus Vibrio occur at a continuum from free-living to symbiotic life forms, including opportunists and pathogens, that can contribute to severe diseases, for instance summer mortality events of Pacific oysters Crassostrea gigas. While most studies focused on Vibrio isolated from moribund oysters during mortality outbreaks, investigations of the Vibrio community in healthy oysters are rare. Therefore, we characterized the persistence, diversity, seasonal dynamics, and pathogenicity of the Vibrio community isolated from healthy Pacific oysters. In a reciprocal transplant experiment we repeatedly sampled hemolymph from adult Pacific oysters to differentiate population from site-specific effects during six months of in situ incubation in the field. We characterized virulence phenotypes and genomic diversity based on multilocus sequence typing in a total of 70 Vibrio strains. Based on controlled infection experiments we could show that strains with the ability to colonize healthy adult oysters can also have the potential to induce high mortality rates on larvae. Diversity and abundance of Vibrio varied significantly over time with highest values during and after spawning season. Vibrio communities from transplanted and stationary oysters converged over time, indicating that communities were not population specific, but rather assemble from the surrounding environment forming communities, some of which can persist over longer periods.
PMID: 24728233 [PubMed - as supplied by publisher]
Scholar Alert: New citations to my articles
Scholar Alert: New citations to my articles
Toll signal transduction pathway in bivalves: complete cds of intermediate elements and related gene transcription levels in hemocytes of immune stimulated Mytilus galloprovincialis.
Dev Comp Immunol. 2014 Apr 4;
Authors: Toubiana M, Rosani U, Giambelluca S, Cammarata M, Gerdol M, Pallavicini A, Venier P, Roch P
Based on protein domain structure and organization deduced from mRNA contigs, 15 transcripts of the Toll signaling pathway have been identified in the bivalve, Mytilus galloprovincialis. Identical searches performed on publicly available M. edulis ESTs revealed 11 transcripts, whereas searches performed in genomic and new transcriptome sequences of the Pacific oyster, Crassostrea gigas, identified 21 Toll-related transcripts. The remarkable molecular diversity of TRAF and IKK coding sequences of C. gigas, suggests that the sequence data inferred from Mytilus cDNAs may not be exhaustive. Most of the Toll pathway genes were constitutively and ubiquitously expressed in M. galloprovincialis, although at different levels, and clearly induced after in vivo injection with bacteria. Such over-transcription was more rapid and intense with Gram-negative than with Gram-positive bacteria. Injection of a fungus modulated the transcription of few Toll pathway genes, with the induction levels of TLR/MyD88 complex being always less intense. Purified LPS and β-glucans had marginal effect whereas peptidoglycans were ineffective. At the moment, we found no evidence of an IMD transcript in bivalves. In conclusion, mussels possess a complete Toll pathway which can be triggered either by Gram-positive or Gram-negative bacteria.
PMID: 24709052 [PubMed - as supplied by publisher]
Evolution of the tyrosinase gene family in bivalve molluscs: independent expansion of the mantle gene repertoire.
Acta Biomater. 2014 Apr 2;
Authors: Aguilera F, McDougall C, Degnan BM
Tyrosinase is a copper-containing enzyme that mediates the hydroxylation of monophenols and oxidation of o-diphenols to o-quinones. This enzyme is involved in a variety of biological processes, including pigment production, innate immunity, wound healing, and exoskeleton fabrication and hardening (e.g. arthropod skeleton and mollusc shell). Here we show that the tyrosinase gene family has undergone large expansions in pearl oysters (Pinctada spp.) and the Pacific oyster (Crassostrea gigas). Phylogenetic analysis reveals that pearl oysters possess at least four tyrosinase genes that are not present in the Pacific oyster. Likewise, C. gigas has multiple tyrosinase genes that are not orthologous to the Pinctada genes, indicating that this gene family has expanded independently in these bivalve lineages. Many of the tyrosinase genes in these bivalves are expressed at relatively high levels in the mantle, the organ responsible for shell fabrication. Detailed comparisons of tyrosinase gene expression in different regions of the mantle in two closely-related pearl oysters, P. maxima and P. margaritifera, reveals that recently-evolved orthologous tyrosinase genes can have markedly different expression profiles. The expansion of tyrosinase genes in these oysters and their co-option into the mantle’s gene regulatory network is consistent with mollusc shell formation being underpinned by a rapidly evolving transcriptome.
PMID: 24704693 [PubMed - as supplied by publisher]
Scholar Alert: New citations to my articles
The immunomodulation of inducible hydrogen sulfide in Pacific oyster Crassostrea gigas.
Dev Comp Immunol. 2014 Mar 31;
Authors: Sun Z, Wang L, Zhang T, Zhou Z, Jiang Q, Yi Q, Yang C, Qiu L, Song L
Hydrogen sulfide (H2S) is an important gasotransmitter, which plays indispensable roles in cardiovascular, nervous and immune systems of vertebrates. However, the information about the immunomodulation of H2S in invertebrates is still very limited. In the present study, the temporal expression profile of cystathionine γ lyase in oyster Crassostrea gigas (CgCSE) was investigated after the oysters were stimulated by lipopolysaccharide. The expression levels of CgCSE mRNA transcripts in hemocytes increased significantly at 12 h (1.31-fold of the PBS group, P < 0.05) after LPS stimulation. The immunomodulation of inducible H2S in oyster was examined by monitoring the alterations of both cellular and humoral immune parameters in response to the stimulations of LPS, LPS+Na2S and LPS+propargylglycine (PAG). The total hemocyte counts (THC) and hemolymph PO activity increased significantly after LPS stimulation, and the increase could be further enhanced by adding PAG, while inhibited by appending Na2S. The phagocytosis activity of hemocytes was also increased firstly after LPS treatment, and the increase was enhanced by adding Na2S but inhibited after appending PAG. The anti-bacterial activity in hemolymph increased at 3 h post LPS treatment, and then decreased after adding PAG. The total SOD activity of hemolymph was also elevated at 6 h post LPS treatment, and the elevated activity was depressed by adding Na2S. These results collectively indicated that H2S might play crucial roles in the immune response of oyster via modulating the turnover and phagocytosis of hemocytes, and regulating the anti-bacterial activity and proPO activation in the hemolymph.
PMID: 24699445 [PubMed - as supplied by publisher]
And our final installment of highlights from the last year…
3) Anesthesia in mature oysters
In shellfish research, there are many situations in which tissues of individual animals need to be sampled multiple times, necessitating nonlethal sampling methods. In particular, an accurate way to effectively measure fecundity, one of the primary metrics of fitness in our project, would be highly beneficial. We needed a nonlethal method to remove broods from female Olympia oysters to enable direct measurement of fecundity, and so we explored the possibility of oyster anesthesia. Although relaxation methods have been developed for a number of oyster species, none were directly applicable or effective for the Olympia oyster, Ostrea lurida. We optimized an anesthetization method to induce gaping in the Olympia oyster. We tested concentrations of 25-100 g/L MgSO4 and exposure durations of up to three hours on adult Olympia oysters, and measured response, recovery, and survival. To stimulate opening of the valves to ensure exposure to the anesthetic, we also tested temperature increase and ambient air exposure as pre-treatments. We found the optimal concentration to be 85 g/L in 50% seawater, which resulted in 45% of the animals anaesthetized in two hours; the remaining oysters did not open their shells and were therefore not exposed. We also found pre-treatment with 30 minutes of air exposure and a temperature increase of 10°C to increase the proportion of oysters that opened their shells for exposure to the treatment. We observed no adverse effects of treatment on subsequent recovery or survival. Future work will assess the influence of microalgae presence on the proportion of open oysters, efficiency of brood removal, and the effect of brood removal on survival. We are planning to use this method to measure fecundity and success in conservation efforts. In addition, this method can accommodate nonlethal tissue sampling.
Scholar Alert: New citations to my articles
Survival, Growth and Reproduction of Cryopreserved Larvae from a Marine Invertebrate, the Pacific Oyster (Crassostrea gigas).
PLoS One. 2014;9(4):e93486
Authors: Suquet M, Labbé C, Puyo S, Mingant C, Quittet B, Boulais M, Queau I, Ratiskol D, Diss B, Haffray P
This study is the first demonstration of successful post-thawing development to reproduction stage of diploid cryopreserved larvae in an aquatic invertebrate. Survival, growth and reproductive performances were studied in juvenile and adult Pacific oysters grown from cryopreserved embryos. Cryopreservation was performed at three early stages: trochophore (13±2 hours post fertilization: hpf), early D-larvae (24±2 hpf) and late D-larvae (43±2 hpf). From the beginning (88 days) at the end of the ongrowing phase (195 days), no mortality was recorded and mean body weights did not differ between the thawed oysters and the control. At the end of the growing-out phase (982 days), survival of the oysters cryopreserved at 13±2 hpf and at 43±2 hpf was significantly higher (P<0.001) than those of the control (non cryopreserved larvae). Only the batches cryopreserved at 24±2 hpf showed lower survival than the control. Reproductive integrity of the mature oysters, formely cryopreserved at 13±2 hpf and 24±2 hpf, was estimated by the sperm movement and the larval development of their offspring in 13 crosses gamete pools (five males and five females in each pool). In all but two crosses out of 13 tested (P<0.001), development rates of the offspring were not significantly different between frozen and unfrozen parents. In all, the growth and reproductive performances of oysters formerly cryopreserved at larval stages are close to those of controls. Furthermore, these performances did not differ between the three initial larval stages of cryopreservation. The utility of larvae cryopreservation is discussed and compared with the cryopreservation of gametes as a technique for selection programs and shellfish cryobanking.
PMID: 24695576 [PubMed - in process]
More research highlights from the past year….
2) Genetic assessment of restoration aquaculture practices
Olympia oyster seed produced from wild broodstock in our restoration hatchery from two different years (2010 and 2011) were compared. Wild broodstock were collected from five locations in in the south subbasin of Puget Sound in both 2010 and 2011. Each year, approximately 500 broodstock were randomly assigned to each of 20 breeding groups. In 2010, the larvae produced by each breeding group were collected and reared separately as a seed group (n=20). In 2011, the larvae produced on a given day from all breeding groups were reared separately as a seed group (n=20). A tissue subsample from 10 individuals from each seed group was taken in 2013 and stored in 70% EtOH. From 10 randomly selected seed groups from each year, DNA was extracted from each sampled individual following standard protocols, and each individual was genotyped at 7 microsatellite loci. Raw genotype data, binned into length classes using TANDEM (Matschiner & Salzburger, 2009), were used to calculate pairwise relatedness using the maximum likelihood estimator of Queller and Goodnight (1989) as implemented in GenALEx (Peakall and Smouse 2006, 2012). Mean pairwise relatedness values within each seed group ranged from 0.07-0.28 (2010) and 0.09-0.33 (2011) (Fig. 3, open circles). Mean pairwie relatedness among 100 pairs over all seed groups (100 bootstrap replicates) did not differ significantly from zero for 2010 or 2011 (P<0.05; Fig. , solid circles). This result suggests that the partial matrix breeding design as implemented incorporates sufficient genetic diversity to obviate undo relatedness, assuming equal contribution from each breeding group. Future work will broaden comparisons to include the restoration seed, commercially produced seed, and wild groups. We will conduct allele count rarefaction (HP-Rare, Kalinowski 2005), apply the more powerful groupwise maximum likelihood relatedness estimator of Jones & Wang (2009), and test for shifts in allele frequency among groups.
Fig. 3. Mean pairwise relatedness values for offspring from ten breeding groups of Olympia oysters, Ostrea lurida, in 2010 and 2011 (open circles) and from pairs randomly sampled from each year (closed circles).