Abalone Gut Research Articles

Genomic and metabolic insights into the first host-associated isolate of Psychrilyobacter.

Although gut bacteria are vital to their hosts, few studies have focused on marine animals. Psychrilyobacter is frequently related to various marine animals, but its interaction with host remains unknown due to the lack of host-associated isolate or genomic information. Here, we combined cultivation-independent and cultivation-dependent methods to uncover the potential roles of Psychrilyobacter in the host abalone. The high-throughput sequencing and literature compiling results indicated that Psychrilyobacter is widely distributed in marine and terrestrial ecosystems with both host-associated and free-living lifestyles, but with a strong niche preference in the guts of marine invertebrates, especially abalone. By in vitro enrichment that mimicked the gut inner environment, the first host-related pure culture of Psychrilyobacter was isolated from the abalone intestine. Phylogenetic, physiological, and biochemical characterizations suggested that it represents a novel species named Psychrilyobacter haliotis B1. Carbohydrate utilization experiments and genomic evidence indicated that B1 can utilize diverse host-food-related monosaccharides and disaccharides but not polysaccharides, implying its potential role in the downstream fermentation instead of the upstream food degradation in the gut. Particularly, this strain showed potential to colonize the gut and benefit the host via different strategies, such as the short-chain fatty acids generation by fermenting peptides and/or amino acids, and the putative production of diverse vitamins and antibiotics to support the host growth and antipathogenicity. To our knowledge, strain B1 represents the first host-related pure culture of Psychrilyobacter; genomic and metabolic evidence showed some beneficial characteristics of the dominant gut anaerobe to the host. IMPORTANCE Psychrilyobacter is a globally distributed bacterial genus and with an inhabiting preference for guts of marine invertebrates. Due to the difficulty of cultivation and the limited genomic information, its role in host remains largely unknown. We isolated the first host-associated Psychrilyobacter species from abalone gut and uncovered its functional potential to the host through different mechanisms. Our findings provide some insights into the understanding of host-microbe interactions on a core taxon with the marine invertebrates, and the isolate may have an application potential in the protection of marine animals.

Hierarchical Microbial Functions Prediction by Graph Aggregated Embedding.

Matching 16S rRNA gene sequencing data to a metabolic reference database is a meaningful way to predict the metabolic function of bacteria and archaea, bringing greater insight to the working of the microbial community. However, some operational taxonomy units (OTUs) cannot be functionally profiled, especially for microbial communities from non-human samples cultured in defective media. Therefore, we herein report the development of Hierarchical micrObial functions Prediction by graph aggregated Embedding (HOPE), which utilizes co-occurring patterns and nucleotide sequences to predict microbial functions. HOPE integrates topological structures of microbial co-occurrence networks with k-mer compositions of OTU sequences and embeds them into a lower-dimensional continuous latent space, while maximally preserving topological relationships among OTUs. The high imbalance among KEGG Orthology (KO) functions of microbes is recognized in our framework that usually yields poor performance. A hierarchical multitask learning module is used in HOPE to alleviate the challenge brought by the long-tailed distribution among classes. To test the performance of HOPE, we compare it with HOPE-one, HOPE-seq, and GraphSAGE, respectively, in three microbial metagenomic 16s rRNA sequencing datasets, including abalone gut, human gut, and gut of Penaeus monodon. Experiments demonstrate that HOPE outperforms baselines on almost all indexes in all experiments. Furthermore, HOPE reveals significant generalization ability. HOPE's basic idea is suitable for other related scenarios, such as the prediction of gene function based on gene co-expression networks. The source code of HOPE is freely available at https://github.com/adrift00/HOPE.

The intestinal microbiome of Australian abalone, Haliotis laevigata and Haliotis laevigata × Haliotis rubra, over a 1-year period in aquaculture.

The digestive tract microbiome aids its host in food digestion, immune system function, development, and health. In marine organisms and habitats, microbiome research is limited. In Australia, abalone aquaculture is an economically important industry; however, little is known about the relationship between abalone and their microbiome. This study investigated the intestinal microbiomes of hybrid, or Tiger, (Haliotis laevigata x H. rubra) and greenlip (Haliotis laevigata) abalone over a one-year production period, grown at two commercial farms. Abalone samples (n = 30 for each species) were collected every three months, for a total of five sampling periods, mimicking seasonal change (i.e. Autumn, Winter, Spring, Summer, Autumn) for each farm. The intestinal samples were sequenced (16S rRNA gene, Illumina MiSeq) and the intestinal microbiomes were analysed. The intestinal microbiomes of hybrid and greenlip abalone were dominated by four genera: Psychrilyobacter, Vibrio, Mycoplasma, and to a lesser extent Formosa. The dominance of the digestive tract by four genera (≥88%) indicates that the abalone gut is a niche environment with unique selective pressures. The intestinal microbiomes were significantly impacted by the sampling date, which is associated with seasonal changes and abalone age.

Draft Genome Sequence of Agarivorans sp. Strain Toyoura001, Isolated from an Abalone Gut.

Agarivorans sp. strain Toyoura001 is a bacterium isolated from the gut of a wild abalone, Haliotis discus hannai Here, we report the draft genome sequence of strain Toyoura001, which consists of 60 contigs comprising 4.67 Mb and 4,257 protein-coding genes.

Effects of dietary supplementation of probiotics on growth, immune responses, and gut microbiome of the abalone Haliotis diversicolor

There is growing interest in the use of probiotics as an alternative strategy for the promotion of growth rates and prevention of diseases in aquaculture. In this study, abalone Haliotis diversicolor were fed with different probiotic-supplemented diets for 180 days, containing Bacillus stratosphericus A3440, Phaeobacter daeponensis AP1220, and a mixture of the two strains. Probiotic supplementation significantly increased shell length and wet weight of juvenile abalone (p < 0.05). Compared with those non-treatment group, trypsin and lipase activities were increased significantly in the gastrointestinal tract of animals fed with A3440 or bacterial mixture (p < 0.05). A subsequent ten-day of challenging trial against Vibrio harveyi, the animals with AP1220 had ~90% survival rate as against ~60% survival rate for the non-treated animals. Likewise, in situ glutathione peroxidase, catalase, alkaline phosphatase, acid phosphatase, and superoxide dismutase assays showed that the probiotics significantly improved immune response of H. diversicolor (p < 0.05). 13,622 OTUs were obtained from abalone intestine microbiome by 16S rRNA high-throughput sequencing. Microbial diversity indices and richness estimates in the non-treated group declined before and after Vibrio infection trial. Conversely, these indexes enhanced to different levels in probiotics treatments, and the A3440 group restored and reached the highest value. Cluster analysis showed that probiotic could re-establish and subsequently balance the gut microbial community after the challenge trial. Furthermore, these potential probiotics had a positive effect on the number of beneficial endogenous bacteria in the abalone gut, such as bacilli and actinobacterial species. Thus, dietary administration of B. stratosphericus, P. daeponensis and their mixture could improve the nutrient and health status of abalone H. diversicolor.

The effect of low-level kelp supplementation on digestive enzyme activity levels in cultured abalone Haliotis midae fed formulated feeds

Previous studies have found that macroalgal inclusion in formulated diets for cultured abalone improves feed utilisation, and it was suggested that seaweed-associated bacteria supply enzymes that aid in the animal's digestion. In the present study, we hypothesised that kelp supplementation in formulated feed affects the profile of digestive enzymes in farmed Haliotis midae. Commercially reared sub-adult abalone fed a kelp-supplemented (Ecklonia maxima; 0.88% dry weight) or kelp-free control feed were collected for analysis of the amylase, alginate lyase, laminarinase, fucoidanase and protease activities in the gut. Levels of polysaccharidase and protease activity did not significantly differ between the diet treatments. However, enzyme-activity levels were more variable in abalone fed the kelp-free diet as compared with those fed the kelp-supplemented diet (coefficients of variation: 73%, 48.3–60.2% and 31.9% [control diet] versus 42.7%, 13.6–33.8% and 14.6% [KS diet] for amylase, macroalgal polysaccharidases and acid protease activity, respectively). We suggest that the presence of dietary kelp modulates the abalone's gut bacteria and their nutrient exchange. Proteomic identification of proteins in abalone gut sections showed that exogenous enzymes associated with the gut microbiome aid in bacterial utilisation of digested molecules, whereas abalone endogenous enzymes degrade the macronutrients in formulated feeds.

Detection and Localisation of the Abalone Probiotic Vibrio midae SY9 and Its Extracellular Protease, VmproA, within the Digestive Tract of the South African Abalone, Haliotis midae

Probiotics have been widely reported to increase the growth rate of commercially important fish and shellfish by enhancing the digestion of ingested feed through the production of extracellular enzymes such as proteases and alginases. In order to investigate this further, the objective of this study was to localise the bacterial probiont Vibrio midae SY9 and one of the extracellular proteases it produces in the digestive tract of the South African abalone Haliotis midae. This was accomplished by inserting a promotorless gfp gene into the chromosome of the bacterium which was incorporated in an artificial, fishmeal-based abalone feed. In situ histological comparison of abalone fed either a basal diet or the basal diet supplemented with V. midae SY9::Tn10.52 using a cocktail of DNA probes to the gfp gene localised the probiont to the crop/stomach and intestinal regions of the H. midae digestive tract. Generally, the ingested probiotic bacterium occurred in association with feed and particulate matter within the crop/stomach and intestinal regions, as well as adhered to the wall of the crop/stomach. Histological immunohistochemical examination using polyclonal anti-VmproA antibodies localised an extracellular protease produced by V. midae SY9 to the H. midae crop/stomach and intestine where it appeared to be associated with feed and/or other particulate matter in the abalone gut. Thus the data suggests that V. midae SY9 colonises and/or adheres to the mucous lining of the abalone gut. Furthermore, the close association observed between the bacterium, its extracellular protease and ingested feed particles supports the theory that V. midae SY9 elevates in situ digestive enzyme levels and thus enhances feed digestion in farmed abalone.

Modulation of gut microbiota associated with abalone Haliotis gigantea by dietary administration of host-derived Pediococcus sp. Ab1

In the present study, we evaluated how dietary administration of host-derived Pediococcus sp. Ab1 has an effect on the abalone gut microbiota using a culture-dependent method and 16S rRNA gene library analysis. The culturable lactic acid bacteria number in the probiotic sample was 105 higher than that in the non-probiotic sample, and we speculate that this significant increase was due to colonization of Ab1 into abalone gut. The result of a culture-dependent method showed that the proportion of Vibrio halioticoli clade, which is known to be a beneficial resident bacterium to abalone, was much higher in the probiotic sample than in the non-probiotic sample. 16S rRNA gene clone sequences revealed that gut microbiota in the probiotic sample was obviously diverse compared to the non-probiotic sample, probably due to improvement of the gut environment by Ab1 colonization. In addition, some beneficial bacteria–like sequences such as V. halioticoli were only found in the probiotic sample. These results suggest that the dietary administration of Ab1 to abalone gut has a great effect on modulation of not only culturable but also unculturable gut microbiota. Our results are useful for future investigations into understanding the effect of probiotics on gut microbiota.

Changes in gut-associated flora and bacterial digestive enzymes during the development stages of abalone (Haliotis diversicolor)

The gut of abalone contains a diverse population of bacteria which contribute to food digestion and nutrient absorption. To investigate the development of gut microflora in the small abalone Haliotis diversicolor and the effect of various diets on the bacterial communities, we analyzed the composition of gut microbial flora and the bacterial digestive enzymes in connection with four important development stages, i.e., larva, early-juvenile, late-juvenile and grow-out. Among these cultivated bacteria, the Firmicutes group was isolated from both the larval and early-juvenile stages; the CFB (Cytophaga/Flexibacteria/Bacterioides) group was detected in early juveniles, late juveniles and grow-out; the γ-proteobacteria existed throughout the four development stages. The isolated Bacillus sp. and Pseudoalteromonas sp. dominated in the larvae and were rarely detected in other stages. Subsequently, the genus Vibrio predominated in the latter three phases after the first feeding. Cluster analysis of the DGGE (Denaturing Gradient Gel Electrophoresis) patterns was basically congruent with the result of culture-dependent method in which the gut bacterial community indicated the successional change in the development stages. And the artificial diets contribute to the change of gut bacterial composition to some extent. The enzyme assay showed that the gut bacteria isolated from the early juveniles produced the greatest diverse digestive enzymes compared with those isolated from other development stages. Most cultivated bacteria in each development stage showed protease, lipase or alginate-degrading activities which could help the farmed abalone H. diversicolor to digest diets. All results showed that a unique but interrelated bacterial flora was developed in the abalone gut which was coincident with the change of diets.

Improved gut environment of abalone Haliotis gigantea through Pediococcus sp. Ab1 treatment

This study demonstrated that host-derived Pediococcus sp. Ab1, a potential probiotic, can colonize the gut of abalone ( Haliotis gigantea) for 12 days from cessation of feeding with Ab1-supplemented commercial diet. The effective colonization by Ab1 of H. gigantea fed a supplemented diet led to the alteration of microbial populations, increased in situ alginate lyase activity, and increased volatile short chain fatty acids (VSCFAs) production in the abalone gut. Vibrio halioticoli clade dominated the gut microflora of abalone fed an Ab1-supplemented diet, while other Vibrio species were dominant in the abalone fed an unsupplemented commercial diet alone. The same species of bacteria were found in the gut of abalone fed diet with and without supplementation; however, the representation of the V. halioticoli clade was much higher in the gut of abalone fed the Ab1-supplemented diet compared to the gut of abalone fed the unsupplemented commercial diet. The alginate lyase activity and production of formate, a VSCFA, were also higher in the gut of abalone fed the Ab1-supplemented diet compared to the gut of abalone fed unsupplemented commercial diet. These results indicate that the host-derived Pediococcus sp. Ab1 has significant potential as an effective probiotic, especially in the improvement of the gut microflora and nutrition status in abalone aquaculture.

Activity of digestive enzymes along the gut of juvenile red abalone, Haliotis rufescens, fed natural and balanced diets

Two carbohydrases (cellulase, lysozyme), three proteases (trypsin, aminopeptidase and non-specific protease), a non-specific lipase, and semiquantitative tests of 19 digestive enzymes were assayed in different gut sections of juvenile red abalone, Haliotis rufescens, in order to identify the regions where digestion takes place and investigate the extent to which diet composition can modify the digestive capacity of abalone. The abalone were fed either fresh kelp (K) or balanced diets containing 25 or 38% crude protein for 6 months. Enzyme assays were carried out on different sections of the abalone's gut at the end of this period. On a weight-specific basis, the digestive gland was the site containing most of the enzymes. On a protein-specific basis, two main digestion regions were identified: the digestive gland-stomach region that is characterized by high activities of cellulase and lysozyme, chymotrypsin and protease, and the mouth-intestine region with a typically high activity of lipase and amino peptidase. Significant dietary effects were observed on the activity of enzymes, especially in the digestive gland. Abalone fed with 25 and 38% crude protein diets exhibited higher cellulase (39.8 ± 4.6 and 14.2 ± 0.8 mU mg − 1 protein, respectively) and lysozyme activities (88.0 ± 20.4 and 56.6 ± 15.7 U, respectively) than those fed with fresh kelp (5.5 ± 0.7 mU mg − 1 protein and 17.1 ± 1.8 U). In contrast, higher protease activity was found in kelp-fed organisms (234.1 ± 20.4 μg product/mg protein) than those fed the 25 and 38% crude protein diets (109.5 ± 20.7 and 119.5 ± 20.5 μg product/mg protein, respectively). Semiquantitative API ZYM assays resulted in no clear food-specific effects on the activity of carbohydrases, proteases, ester hydrolases or phosphohydrolases, yet organ-specific differences were conspicuous in various cases, and generally agreed with quantitative results. It is suggested that the increased carbohydrase activity exhibited by organisms fed the balanced diets resulted from a combination of an increased number of resident bacteria in the abalone's gut and facilitated contact between dietary substrates and digestive cells. The present results indicate that H. rufescens can adjust their enzyme levels in order to maximize the acquisition of dietary protein and carbohydrates. This characteristic can be advantageously used to search for suitable diets in abalone aquaculture.

Improved growth rate and disease resistance in farmed Haliotis midae through probiotic treatment

The use of probiotics in aquaculture is proving to be highly effective in improving disease resistance, nutrition and/or growth of cultured organisms. We have shown that Haliotis midae fed a probiotic-supplemented diet have an improved survival and growth rate compared to animals not fed probiotics. The growth rate of small (20 mm) and large (67mm) abalone was improved by 8% and 34%, respectively, in two separate eight-month farm growth trials. Furthermore, the number, and phagocytic activity, of circulating haemocytes was significantly higher ( P<0.05) in probiotic treated animals compared to non-treated animals following challenge with the pathogenic bacterium Vibrio anguillarum. Seven days after challenging with V. anguillarum, the probiotic-fed animals had 62% survival compared to 25% survival for non-treated animals. In situ protease assays showed that probiotic treatment significantly increased ( P<0.05) protease activity in the intestinal region of the digestive tract of animals fed the probiotic-supplemented feed. This correlated with a significant increase ( P<0.05) in the amount of protein digestion and absorption measured in this region of the abalone gut. Histological analysis showed that the digestive glands of animals receiving probiotics were bacteria-free, whereas the digestive glands of 70% of the animals receiving the non-supplemented feed had a high bacterial load. The microorganisms tested in this study therefore have tremendous potential as probiotics for commercially produced H. midae.

Pharmacological and histological assessment of gut muscle movement in blacklip abalone,Haliotis rubra(Leach)

We examined the basic pharmacology of abalone gut tissues for adrenergic and peptide receptors using ligand binding and by determining the pharmacological effectors of abalone gut motility in a number of gut regions. Contractile responses could not be elicited, even though we could show the existence of α-adrenergic and peptide receptors. Responses to any muscle contractile agents, including carbachol and potassium chloride, could not be found in blacklip abalone, Haliotis rubra (Leach). Histology confirmed the relative absence of muscular layers in all the tissues of the gut, indicating that previous literature reports of muscular involvement in contractile movement of gut contents did not appear to apply to this species.

The role of bacteria in the digestion of seaweed by the abalone Haliotis midae

The polysaccharolytic enzymes of the abalone Haliotis midae and its resident gut bacteria were investigated. Bacteria isolated from the abalone gut were able to degrade the polysaccharides laminarin, carboxymethylcellulose (CMC), alginate, agarose and carrageenan. Detection of alginate lyase, CMCase, laminarinase, agarase and carrageenase in the hepatopancreas, which was devoid of bacteria, indicated that H. midae produces a range of polysaccharases. The endogenous polysaccharases of abalone fed either Ecklonia maxima or Gracilaria verrucosa varied in response to diet. It is proposed that bacteria resident in the digestive system of H. midae assist in the digestion of alginate, laminarin, agarose, carrageenan and cellulose.

Strain selection and cell isolation of Ulvaria oxysperma (Kuetz.) Bliding (Chlorophyta) for net cultivation

North Carolina and Bermuda isolates of the green seaweed Ulvaria oxysperma (Kuetz.) Bliding were grown in cross-gradient light-temperature culture to determine the effects of varied photon fluence rates, temperature and photoperiod on their growth and reproduction. Results indicated significant genetic-based variation between the two populations. Spheroplast were obtained from the vegetative thallus of the North Carolina isolate following enzymatic digestion with abalone gut extract and maceration. Single cell isolates developed new cell walls in 2–3 days and attached to nylon net substrate. Cultured isolates regenerated into juvenile plants with normal development and morphology. Results are discussed in terms of the suitability of this economically important species for land-based mariculture.