Identification of a novel RIG-I isoform and its truncating variant in Japanese eel, Anguilla japonica.

Retinoic acid-inducible gene-I (RIG-I) is a cytoplasmic viral RNA sensor that triggers the production of type I interferons (IFNs) and proinflammatory cytokines during viral infection. RIG-I gene has been identified previously in Japanese eel, Anguilla japonica. In the present study, we have characterized a novel isoform of RIG-I (designated as AjRIG-Ib) and its truncated variant (AjRIG-Ibv). The AjRIG-Ib encodes 940 amino acids (aa) consisting of two N-terminal caspase activation and recruitment domains (CARDs), a DEX(D/H) box RNA helicase domain, and a C-terminal regulatory domain (CTD). The AjRIG-Ibv encodes a protein of 843 aa, that shares similar structural organization with AjRIG-Ib, but lacking CTD. The gene expression analyses showed that AjRIG-Ib and AjRIG-Ibv were detectable in all tissues/organs examined, and AjRIG-Ib was the predominant form. The mRNA level of AjRIG-Ibv was upregulated rapidly at 8 h after the Poly I:C injection, and the significant increase of AjRIG-Ib was observed at 16 and 24 h post-injection (hpi). Laser confocal microscopy showed that AjRIG-Ib and AjRIG-Ibv were both located in cytoplasm. In addition, the overexpression of AjRIG-Ib or AjRIG-Ibv led to the increased activity of IFN promoter in transient transfection assay. Taken together, our results indicated that AjRIG-Ib and AjRIG-Ibv may play cooperative or somewhat complementary roles in coordinating the antiviral response in fish.

Identification of four type I IFNs from Japanese eel with differential expression properties and Mx promoter inducibility.

Type I IFNs are a family of cytokines with antiviral, anti-proliferative and immune-modulatory functions. In this study, four type I IFNs (termed AjIFN1-4) have been cloned from the Japanese eel, Anguilla japonica. The open reading frames of AjIFN1-4 are 552, 534, 546 and 561 bp in length, encoding 183, 177, 181, and 186 amino acids (aa), respectively. Sequence comparison and phylogenetic analysis results revealed that AjIFN1 and AjIFN2 belong to group one (2C-containing) IFNs, while AjIFN3 and AjIFN4 belong to group two (4C-containing) IFNs. Syntenic comparison showed that chromosome block duplication and rearrangement events might have occurred at IFN loci in different teleost lineages. Expression analysis revealed the rapid induction of AjIFNl and AjIFN2 in response to poly I:C stimulation, while AjIFN3 and AjIFN4 were predominantly expressed at later time points. Two Mx promoter reporter assays were conducted to assess the Mx-inducing capability of AjIFN1-4. It is shown that the overexpression of AjIFN1-4 all promoted the luciferase activity of MxB reporter, but the activity of MxC reporter increased only in cells transfected with AjIFN1. Collectively, it is suggested that teleost IFNs were evolved independently in different lineages of fish and may function differently in teleost antiviral immunity.

The composition and structure of the intestinal microflora of Anguilla marmorata at different growth rates: a deep sequencing study.

AIMS: The aim of this study was to determine the intestinal microflora of Anguilla marmorata at different growth rates and to identify potential probiotic/pernicious bacteria. METHODS AND RESULTS: Bacterial communities from eight different eels' intestinal sites (including the intestinal contents and the intestinal mucosa) from three fish groups (three fast-, two medium-, and three stunted-growth samples), two water samples, and one diet sample were characterized by Illumina next-generation sequencing. The data revealed that the predominant genera (relative abundance of bacteria genera >1%) in the intestine of fast- and medium-growth groups were Cetobacterium, Edwardsiella, Clostridium, Lactococcus, Bacteroides, Plesiomonas and Akkermansia. The dominant genus in the stunted-growth group was Spiroplasma. Moreover, culture-associated (water and feed) environmental microbes were distinct from those present in fish intestines, and included Flavobacterium (the dominant bacteria in water) and Corynebacterium (the dominant bacteria in feed). CONCLUSIONS: Only minor differences in gut microbial communities were observed between the fast-growth group and the medium-growth group; however, significant differences were observed between the normal-growth group (including the fast-growth group and medium-growth group, which showed uninhibited growth during the rearing stage) and the stunted-growth group. Together, these data suggested that intestinal microbes were significantly associated with marbled eels' growth rate. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, we demonstrated for the first time, the intestinal bacterial communities of A. marmorata at different growth rates. Moreover, we found that the genus Spiroplasma was abundant in the guts of stunted-growth eels, which had never been noticed. Such a finding indicates that the genus Spiroplasma plays a key role associated with retardation in growth and should be controlled to recover the growth of stunted eels, which is meaningful to farmers.

Effect of level of metabolizable protein on milk production and nitrogen utilization in lactating dairy cows.

The objective of this study was to investigate the effects of the level of metabolizable protein (MP) on milk production and nitrogen utilization in Chinese Holstein dairy cows. Forty multiparous dairy cows (body weight = 590 kg; days in milk = 135; average milk yield = 30.2 kg/d) were assigned to treatments randomly within groups based on days in milk and milk production. Animals were offered diets with different levels of MP: 8.3% (diet A), 8.9% (diet B), 9.7% (diet C), and 10.4% (diet D) of dry matter. The MP level in diet A was designed to meet the current Chinese National Station of Animal Production and Health guidelines, whereas that in diet D was based on the National Research Council (2001) model. The experiment lasted for 7 wk. Milk yield and milk composition (fat, protein, and lactose) were recorded, and urea nitrogen concentrations in serum, urine, and milk were measured during the experiment. Milk yield and milk protein percentage increased as the MP increased up to 9.7% of dry matter, and then leveled off. Concentrations of nitrogen in urine, serum, and milk increased linearly as the amount of MP was increased, indicating decreased efficiency of nitrogen utilization. Milk lactose percentage and total solids percentage showed no significant differences among the 4 diets. We concluded that the optimal dietary MP level was at 9.6% of dry matter for Chinese Holstein dairy cows producing 30 kg of milk per day.