Characterization of a C-Type Lectin Domain-Containing Protein with Antibacterial Activity from Pacific Abalone (Haliotis discus hannai).

Genes that influence the growth of Pacific abalone (Haliotis discus hannai) may improve the productivity of the aquaculture industry. Previous research demonstrated that the differential expression of a gene encoding a C-type lectin domain-containing protein (CTLD) was associated with a faster growth in Pacific abalone. We analyzed this gene and identified an open reading frame that consisted of 145 amino acids. The sequence showed a significant homology to other genes that encode CTLDs in the genus Haliotis. Expression profiling analysis at different developmental stages and from various tissues showed that the gene was first expressed at approximately 50 days after fertilization (shell length of 2.47 ± 0.13 mm). In adult Pacific abalone, the gene was strongly expressed in the epipodium, gill, and mantle. Recombinant Pacific abalone CTLD purified from Escherichia coli exhibited antimicrobial activity against several Gram-positive bacteria (Bacillus subtilis, Streptococcus iniae, and Lactococcus garvieae) and Gram-negative bacteria (Vibrio alginolyticus and Vibrio harveyi). We also performed bacterial agglutination assays in the presence of Ca2+, as well as bacterial binding assays in the presence of the detergent dodecyl maltoside. Incubation with E. coli and B. subtilis cells suggested that the CTLD stimulated Ca2+-dependent bacterial agglutination. Our results suggest that this novel Pacific abalone CTLD is important for the pathogen recognition in the gastropod host defense mechanism.

Use of a gene encoding zona pellucida 4 as a female-specific marker for early stage sexual differentiation and size dimorphism in the pacific abalone Haliotis discus hannai.

Growth rates of Pacific abalone Haliotis discus hannai are an important trait affecting the economic value in the abalone aquaculture industry. A reverse-transcription polymerase chain reaction (RT-PCR) analyses of tissues from H. discus hannai was conducted for sexually mature gonads to determine male- and female-specific target gene expression, including genes encoding zona pellucida domain 4 (zp4), sperm protein (sp) and lysin (lys), respectively. Sex-specific expression patterns of these gene expression, even in sexually immature abalone, indicate these genes can be used as sensitive and robust sex-specific molecular markers. The RT-PCR procedure was also performed to analyze tissues collected at various developmental stages (50-day intervals) beginning at fertilization to determine when sex differentiation and expression of sex-specific genes was initiated. Detection of zp4 transcript in tissues collected at 200 days post-fertilization (dpf) indicated egg-specific development starts at 150-200 dpf. To evaluate possible sex-specific differences in growth rate, there was conducting of a molecular marker-based sex identification of abalone from a population selected for rapid growth rate. In a group of large H. discus hannai, females were more prevalent than males. To assess the correlation between growth and sex, there was comparison of weights of 3-year-old Pacific abalone in specimens where there had been sex determinations by visual examination and molecular methods. The results indicated females weighed more (55.92 ± 9.38 g, n = 15) than males (43.64 ± 15.55 g, n = 6, P = 0.037), indicating females had a more rapid growth rate than males.