Trophic Position of the White Worm (Enchytraeus albidus) in the Context of Digestive Enzyme Genes Revealed by Transcriptomics Analysis.

To assess the impact of Enchytraeidae (potworms) on the functioning of the decomposer system, knowledge of the feeding preferences of enchytraeid species is required. Different food preferences can be explained by variations in enzymatic activities among different enchytraeid species, as there are no significant differences in the morphology or anatomy of their alimentary tracts. However, it is crucial to distinguish between the contribution of microbial enzymes and the animal's digestive capacity. Here, we computationally analyzed the endogenous digestive enzyme genes in Enchytraeus albidus. The analysis was based on RNA-Seq of COI-monohaplotype culture (PL-A strain) specimens, utilizing transcriptome profiling to determine the trophic position of the species. We also corroborated the results obtained using transcriptomics data from genetically heterogeneous freeze-tolerant strains. Our results revealed that E. albidus expresses a wide range of glycosidases, including GH9 cellulases and a specific digestive SH3b-domain-containing i-type lysozyme, previously described in the earthworm Eisenia andrei. Therefore, E. albidus combines traits of both primary decomposers (primary saprophytophages) and secondary decomposers (sapro-microphytophages/microbivores) and can be defined as an intermediate decomposer. Based on assemblies of publicly available RNA-Seq reads, we found close homologs for these cellulases and i-type lysozymes in various clitellate taxa, including Crassiclitellata and Enchytraeidae.

Genome-wide association study identified five quantitative trait loci and two candidate genes for digestive traits in Suhuai pigs.

This work aimed to identify markers and candidate genes underlying porcine digestive traits. In total, 331 pigs were genotyped by 80 K Chip data or 50 K Chip data. For apparent neutral detergent fiber digestibility, a total of 19 and 21 candidate single nucleotide polymorphisms (SNP) were respectively identified using a genome-wide efficient mixed-model association algorithm and linkage-disequilibrium adjusted kinship. Among them, three quantitative trait locus (QTL) regions were identified. For apparent acid detergent fiber digestibility, a total of 16 and 17 SNPs were identified by these two methods, respectively. Of these, three QTL regions were also identified. Moreover, two candidate genes (MST1 and LATS1), which are functionally related to intestinal homeostasis and health, were detected near these significant SNPs. Taken together, our results could provide a basis for deeper research on digestive traits in pigs.

Mechanism of the toxic effects of tetracycline on blood meal digestion in Haemaphysalis longicornis.

The extensive utilization of antibiotics in the field of animal husbandry gives rise to various concerns pertaining to the environment and human health. Here, we demonstrate that the administration of tetracycline impedes blood meal digestion in the tick Haemaphysalis longicornis. Tissue sectioning, 16S rRNA high-throughput sequencing, and transcriptome sequencing of the midgut were employed to elucidate the mechanism underlying tetracycline toxicity. The treatment group consisted of engorged female ticks that were subjected to tetracycline microinjections (75 µg per tick), whereas the control group received sterile water injections. On days 2 and 4 following the injections, the tick body weight changes were assessed and the midguts were dissected and processed. Change in tick body weight in tetracycline-treated group was less than in the control group. In tetracycline-treated ticks, midgut epithelial cells were loosely connected and blood meal digestion was impaired compared to the control group. There was no significant change in midgut bacterial diversity after tetracycline treatment. On day 2 following treatment, the relative abundance of Escherichia-Shigella was significantly decreased, whereas the relative abundance of Allorhizobium was significantly increased compared to the control group. On day 4 following treatment, the relative abundance of Escherichia-Shigella, Allorhizobium, Ochrobactrum, and Acidibacter decreased significantly, whereas the relative abundance of Paraburkholderia and Pelomonas increased significantly. Tetracycline treatment also affected midgut gene expression, producing a cumulative effect wherein the differentially expressed genes (DEGs) were mostly down-regulated. KEGG enrichment pathway analysis revealed that on day 2 the up-regulated DEGs were significantly enriched in 21 pathways, including apoptosis and phagosome. Comparatively, the down-regulated DEGs were significantly enriched in 26 pathways, including N-glycan biosynthesis, lysosome, and autophagy. In contrast, on day 4 the up-regulated DEGs were significantly enriched in 10 pathways including aminoacyl-tRNA biosynthesis, ribosome biogenesis, RNA transport, and DNA replication, whereas the down-regulated differential genes were significantly enriched in 11 pathways including lysosome, peroxisome, N-glycan biosynthesis, and fatty acid synthesis. This indicates that tetracycline injection inhibited blood meal digestion by affecting midgut digestive cells, gut flora diversity, and gene expression. These findings could contribute to tick control by inhibiting blood meal digestion.

Comparative transcriptome analysis revealed omnivorous adaptation of the small intestine of Melinae.

As the main digestive organ, the small intestine plays a vital role in the digestion of animals. At present, most of the research on animal feeding habits focuses on carnivores and herbivores. However, the mechanism of feeding and digestion in omnivores remains unclear. This study aims to reveal the molecular basis of the omnivorous adaptive evolution of Melinae by comparing the transcriptome of the small intestines of Asian Badgers (Meles leucurus) and Northern Hog Badgers (Arctonyx albogularis). We obtained high-quality small intestinal transcriptome data from these two species. Key genes and signalling pathways were analysed through Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and other databases. Research has mainly found that orthologous genes related to six enzymes have undergone adaptive evolution. In addition, the study also found three digestion-related pathways (cGMP-PKG, cAMP, and Hippo). They are related to the digestion and absorption of nutrients, the secretion of intestinal fluids, and the transport of food through the small intestine, which may help omnivorous animals adapt to an omnivorous diet. Our study provides insight into the adaptation of Melinae to omnivores and affords a valuable transcriptome resource for future research.

Transcriptional Response in the Digestive Gland of the King Scallop (Pecten maximus) After the Injection of Domoic Acid.

Some diatom species of the genus Pseudo-nitzschia produce the toxin domoic acid. The depuration rate of domoic acid in Pecten maximus is very low; for this reason, king scallops generally contain high levels of domoic acid in their tissues. A transcriptomic approach was used to identify the genes differentially expressed in the P. maximus digestive gland after the injection of domoic acid. The differential expression analysis found 535 differentially expressed genes (226 up-regulated and 309 down-regulated). Protein-protein interaction networks obtained with the up-regulated genes were enriched in gene ontology terms, such as vesicle-mediated transport, response to stress, signal transduction, immune system process, RNA metabolic process, and autophagy, while networks obtained with the down-regulated genes were enriched in gene ontology terms, such as response to stress, immune system process, ribosome biogenesis, signal transduction, and mRNA processing. Genes that code for cytochrome P450 enzymes, glutathione S-transferase theta-1, glutamine synthase, pyrroline-5-carboxylate reductase 2, and sodium- and chloride-dependent glycine transporter 1 were among the up-regulated genes. Therefore, a stress response at the level of gene expression, that could be caused by the domoic acid injection, was evidenced by the alteration of several biological, cellular, and molecular processes.

Expression Profiles of Digestive Genes in the Gut and Salivary Glands of Tarnished Plant Bug (Hemiptera: Miridae).

Host plant preference of agricultural pests may shift throughout the growing season, allowing the pests to persist on wild hosts when crops are not available. Lygus Hahn (Hemiptera: Miridae) bugs are severe pests of cotton during flowering and fruiting stages, but can persist on alternative crops, or on weed species. Diversity of digestive enzymes produced by salivary glands and gut tissues play a pivotal role in an organism's ability to utilize various food sources. Polyphagous insects produce an array of enzymes that can process carbohydrates, lipids, and proteins. In this study, the digestive enzyme repertoire of the tarnished plant bug, Lygus lineolaris (Palisot de Beauvois), was identified by high-throughput sequencing followed by cDNA cloning and sequencing. This study identified 87 digestive genes, including 30 polygalacturonases (PG), one ß-galactosidase, three α-glucosidases, six ß-glucosidases, 28 trypsin-like proteases, three serine proteases, one apyrase-like protease, one cysteine protease, 12 lipases, and two transcripts with low similarity to a xylanase A-like genes. RNA-Seq expression profiles of these digestive genes in adult tarnished plant bugs revealed that 57 and 12 genes were differentially expressed in the salivary gland and gut (≥5-fold, P ≤ 0.01), respectively. All polygalacturonase genes, most proteases, and two xylanase-like genes were differentially expressed in salivary glands, while most of the carbohydrate and lipid processing enzymes were differentially expressed in the gut. Seven of the proteases (KF208689, KF208697, KF208698, KF208699, KF208700, KF208701, and KF208702) were not detected in either the gut or salivary glands.

Regulation of gene expression encoding the digestive α-amylase in the larvae of Colorado potato beetle, Leptinotarsa decemlineata (Say) in response to plant protein extracts.

Considering the relevance of insect α-amylases and natural α-amylase inhibitors present in plants to protect against insect damage, we investigated the effect of white bean and rapeseed protein extracts on digestive α-amylase gene expression of the Colorado potato beetle, Leptinotarsa decemlineata (Say). For this purpose, in vitro and in vivo trials were performed to determine the inhibitory activity of seed proteins on the third and fourth instar larvae. In both trials, the significant inhibitory effect of each extracts on the third and fourth instar larval α-amylase activity and considerable mortality in treatments were observed compared to control trials. In the RT-qPCR, expression ratio demonstrated that the α-amylase gene of two different larval stages grown on both proteins treated leaves had significantly differentiated expression and was up-regulated in third instar larvae and down-regulated in fourth instar larvae compared to control. Results suggest that the hyper-production of α-amylase in third instar larvae is elicited to compensate for the enzyme activity inhibition at an earlier stage and also down-regulation suggests the existence of a negative feedback of plant proteins on the last instar larvae via impaired food intake and digestive α-amylase activity in Colorado potato beetle. Therefore, disruption of the insect's digestive physiology by plant defensive proteins can be considered in the development of innovative controlling methods of this crucial potato pest.

Gene networks for three feed efficiency criteria reveal shared and specific biological processes.

BACKGROUND: French beef producers suffer from the decrease in profitability of their farms mainly because of the continuous increase in feed costs. Selection for feed efficiency in beef cattle represents a relevant solution to face this problem. However, feed efficiency is a complex trait that can be assessed by three major criteria: residual feed intake (RFI), residual gain (RG) and feed efficiency ratio (FE), which involve different genetic determinisms. An analysis that combines phenotype and whole-genome sequence data provides a unique framework for genomic studies. The aim of our study was to identify the gene networks and the biological processes that are responsible for the genetic determinism that is shared between these three feed efficiency criteria. RESULTS: A population of 1477 French Charolais young bulls was phenotyped for feed intake (FI), average daily gain (ADG) and final weight (FW) to estimate RFI, RG and FE. A subset of 789 young bulls was genotyped on the BovineSNP50 single nucleotide polymorphism (SNP) array and imputed at the sequence level using RUN6 of the 1000 Bull Genomes Project. We conducted a genome-wide association study (GWAS) to estimate the individual effect of 8.5 million SNPs and applied an association weight matrix (AWM) approach to analyse the results, one for each feed efficiency criterion. The results highlighted co-association networks including 626 genes for RFI, 426 for RG and 564 for FE. Enrichment assessment revealed the biological processes that show the strongest association with RFI, RG and FE, i.e. digestive tract (salivary, gastric and mucin secretion) and metabolic processes (cellular and cardiovascular). Energetic functions were more associated with RFI and FE and cardio-vascular and cellular processes with RG. Several hormones such as apelin, glucagon, insulin, aldosterone, the gonadotrophin releasing hormone and the thyroid hormone were also identified, and these should be tested in future studies as candidate biomarkers for feed efficiency. CONCLUSIONS: The combination of network and pathway analyses at the sequence level led to the identification of both common and specific mechanisms that are involved in RFI, RG and FE, and to a better understanding of the genetic determinism underlying these three criteria. The effects of the genes involved in each of the identified processes need to be tested in genomic evaluations to confirm the potential gain in reliability of using functional variants to select animals for feed efficiency.

De novo assembly and discovery of genes related to blood digestion in the transcriptome of Dermanyssus gallinae (Acari: Dermanyssidae).

Dermanyssus gallinae is an economically important blood-feeding ectoparasite affecting layer chicken farms in many countries. Similar to other blood-feeding arthropods, the blood-meal digestion plays a key role in the survival and reproduction of D. gallinae. The knowledge of the genes involved in blood digestion processes may provide new targets for drug and vaccine against the red mites. In the present study, we sequenced and de novo assembled the transcriptomes of unfed and fed adult red mites using Illumina RNA sequencing (RNA-seq) technology. Up to 40,506 unigenes were assembled, and 13,018 unigenes were identified and annotated. GO analysis of the annotated unigenes clustered into three main GO terms. The dominant GO terms of biological processes were cellular process and metabolic process, the prevailing GO terms of the cellular component were cell part and membrane part, and the dominant GO terms of molecular functions were catalytic and binding activities. Up to 6,443 annotated sequences were assigned to 246 active pathways by KEGG analysis. Differentially expressed genes (DEGs) analysis identified 2,877 unigenes with upregulated 2,094 and downregulated 783 in fed female mites compared with unfed female mites. The biological function of these DEGs was further investigated using the KEGG and GO databases. The upregulated DEGs were potentially involved in nutrient metabolism, highlighting their importance in red mite biology. Quantitative reverse transcription real-time PCR (qRT-PCR) validated that the expression levels of the selected six upregulated DEGs were consistent with those in RNA-seq, indicating that the transcriptomic data are reliable. The present study provides valuable and fundamental knowledge that improves our understanding of the physiology of D. gallinae digestion at a molecular level. Moreover, these transcriptomic data will facilitate the identification of novel function genes and candidate antigens for the development of effective vaccines or drug targets to control D. gallinae.

Genome-Wide Analyses Provide Insights into the Scavenging Lifestyle of the Striped Hyena (Hyaena hyaena).

Hyenas (family Hyaenidae) occupy a variety of different niches, of which the striped hyena (Hyaena hyaena) scavenges mainly on the carcasses of animals. We compared its genome with the genomes of nine other mammals, focusing on similarities and differences in chemoreception, detoxification, digestive, and immune systems. The results showed that the striped hyena's immune and digestive system-related gene families have significantly expanded, which was likely to be an adaptive response to its scavenging lifestyle. In addition, 88 and 26 positive selected genes (PSGs) were identified in the immune system and digestive system, respectively, which may be the molecular basis for immune defense system to effectively resist pathogen invasion. Functional enrichment analysis of PSGs revealed that most of them were involved in the immune regulation process. Among them, eight specific missense mutations were found in two PSGs (MHC class II antigen DOA and MHC class II antigen DOB), suggesting important reorganization of the immune system in the striped hyena. Moreover, we identified one cathelicidin gene and four defensin genes in the striped hyenas by genome mining, which have high-efficiency and broad-spectrum antimicrobial activity. Of particular interest, a striped hyena-specific missense mutation was found in the cathelicidin gene. PolyPhen-2 classified the missense mutation as a harmful mutation, which may have aided in immune adaptation to carrion feeding. Our genomic analyses on the striped hyena provided insights into its success in the adaptation to the scavenging lifestyle.

Identifying genes for resistant starch, slowly digestible starch, and rapidly digestible starch in rice using genome-wide association studies.

BACKGROUND: The digestibility of starch is important for the nutritive value of staple food. Although several genes are responsible for resistant starch (RS) and slowly digestible starch (SDS), gaps persist concerning the molecular basis of RS and SDS formation due to the complex genetic mechanisms of starch digestibility. OBJECTIVES: The objective of this study was to identify new genes for starch digestibility in rice and interprete the genetic mechanisms of RS and SDS by GWAS. METHODS: Genome-wide association studies were conducted by associating the RS and SDS phenotypes of 104 re-sequenced rice lines to an SNP dataset of 2,288,867 sites using a compressed mixed linear model. Candidate genes were identified according to the position of the SNPs based on data from the MSU Rice Genome Annotation Project. RESULTS: Seven quantitative trait loci (QTLs) were detected to be associated with the RS content, among which the SNP 6 m1765761 was located on Waxy. Starch branching enzymes IIa (BEIIa) close to QTL qRS-I4 was detected and further identified as a specific candidate gene for RS in INDICA. Two QTLs were associated with SDS, and the LOC_Os09g09360 encoding lipase was identified as a causal gene for SDS. CONCLUSIONS: GWAS is a valid strategy to genetically dissect the formation of starch digestion properties in rice. RS formation in grains is dependent on the rice type; lipid might also contribute to starch digestibility and should be an alternative factor to improve rice starch digestibility.

Digestion-related proteins in the tobacco hornworm, Manduca sexta.

Food digestion is vital for the survival and prosperity of insects. Research on insect digestive enzymes yields knowledge of their structure and function, and potential targets of antifeedants to control agricultural pests. While such enzymes from pest species are more relevant for inhibitor screening, a systematic analysis of their counterparts in a model insect has broader impacts. In this context, we identified a set of 122 digestive enzyme genes from the genome of Manduca sexta, a lepidopteran model related to some major agricultural pests. These genes encode hydrolases of proteins (85), lipids (20), carbohydrates (16), and nucleic acids (1). Gut serine proteases (62) and their noncatalytic homologs (11) in the S1A subfamily are encoded by abundant transcripts whose levels correlate well with larval feeding stages. Aminopeptidases (10), carboxypeptidases (10), and other proteases (3) also participate in dietary protein digestion. A large group of 11 lipases as well as 9 esterases are probably responsible for digesting lipids in diets. The repertoire of carbohydrate hydrolases (16) is relatively small, including two amylases, three maltases, two sucrases, two α-glucosidases, and others. Lysozymes, peptidoglycan amidases, and ß-1,3-glucanase may hydrolyze peptidoglycans and glucans to harvest energy and defend the host from microbes on plant leaves. One alkaline nuclease is associated with larval feeding, which is likely responsible for hydrolyzing denatured DNA and RNA undergoing autolysis at a high pH of midgut. Proteomic analysis of the ectoperitrophic fluid from feeding larvae validated at least 131 or 89% of the digestive enzymes and their homologs. In summary, this study provides for the first time a holistic view of the digestion-related proteins in a lepidopteran model insect and clues for comparative research in lepidopteran pests and beyond.

Gene expression profiles during postnatal development of the liver and pancreas in giant pandas.

Giant pandas are unique Carnivora with a strict bamboo diet. To investigate the molecular mechanism of giant panda nutrient metabolism from newborn to adult, the gene expression profiles of giant panda liver and pancreas tissues collected from three important feeding stages were investigated using RNA-seq. We found a total of 3,211 hepatic and 3,343 pancreatic differentially expressed genes (DEGs) from three comparisons between suckling and no feeding, adult and no feeding, and adult and suckling groups. Few differences in gene-expression profiles were exhibited between no feeding and suckling groups in both tissues. GO and KEGG analyses were performed to further understand the biological functions of the DEGs. In both the liver and pancreas, genes related mainly to cell cycle processes were highly up-regulated in newborn samples whereas genes related to metabolism and immunity were up-regulated in adult giant pandas. The high expression of metabolism-related genes in adult samples probably helps to fulfill the metabolic function requirements of the liver and pancreas. In contrast, several vital genes involved in cholesterol metabolism and protein digestion and absorption were over-expressed in newborn samples. This may indicate the importance of cholesterol metabolism and protein digestion and absorption processes in giant panda infancy.

Intestinal Transcriptome Analysis Highlights Key Differentially Expressed Genes Involved in Nutrient Metabolism and Digestion in Yellowtail Kingfish (Seriola lalandi) Fed Terrestrial Animal and Plant Proteins.

This study investigated the effects of dietary terrestrial animal and plant proteins on the intestinal transcriptomes of yellowtail kingfish (YTK), Seriola lalandi, an ecologically and economically important marine species in Australia. Five diets containing fish meal (FM), poultry by-product meal (PBM), blood meal (BLM), faba bean meal (FBM) and corn gluten meal (CGM) were formulated and fed over a period of 4 weeks. The Illumina RNA-sequencing (RNA-Seq) results identified a suite of differentially expressed genes involved in nutrient metabolism and protein digestion pathways, reinforced by quantitative polymerase chain reaction (qPCR) results. These findings provide molecular support to the notion that PBM and FBM are useful raw materials in commercial diets for YTK. Using the same evidence, we have demonstrated that BLM and CGM may be less useful and their incorporation into commercial aquafeeds for this species should be done cautiously. The differentially expressed genes showed a subtle difference and high correlation with apparent nutrient digestibility of raw materials. Further, our results indicate that transcriptome profiling provides a useful tool to evaluate alternative protein sources for use in aquaculture feeds.

Broilers divergently selected for digestibility differ for their digestive microbial ecosystems.

Improving the digestive efficiency of broiler chickens (Gallus gallus) could reduce organic waste, increase the use of alternative feed not used for human consumption and reduce the impact of feed in production costs. By selecting chicken lines divergently for their digestive efficiency, we showed previously that digestive efficiency is under genetic control and that the two resulting divergent lines, D+ (high digestive efficiency or "digestibility +") and D- (low digestive efficiency or "digestibility -"), also differ for the abundance of specific bacteria in their caeca. Here we perform a more extensive census of the bacteria present in the digestive microbiota of 60 chickens selected for their low apparent metabolizable energy corrected for nitrogen balance (AMEn-) or high (AMEn+) digestive efficiency in a [D+ x D-] F8 progeny of 200 individuals. We sequenced the 16S rRNA genes of the ileal, jejunal and caecal microbiotas, and compared the compositions and predicted functions of microbiotas from the different intestinal segments for 20 AMEn+ and 19 AMEn- birds. The intestinal segment of origin was the main factor structuring the samples. The caecal microbiota was the most impacted by the differences in digestive efficiency, with 41 bacterial species with abundances differing between highly and poorly efficient birds. Furthermore, we predicted that the caecal microbiota of efficient birds might be enriched in genes contributing to the degradation of short chain fatty acids (SCFA) from non-starch polysaccharides. These results confirm the impact of the genetic selection led on digestibility on the caecal microbiota taxonomic composition. They open the way toward the identification of specific, causal genes of the host controlling variations in the abundances of bacterial taxons.

In Vitro Evaluation of the Effects of Commercial Prebiotic GOS and FOS Products on Human Colonic Caco-2 Cells.

Prebiotic oligosaccharides are widely used as human and animal feed additives for their beneficial effects on the gut microbiota. However, there are limited data to assess the direct effect of such functional foods on the transcriptome of intestinal epithelial cells. The purpose of this study is to describe the differential transcriptomes and cellular pathways of colonic cells directly exposed to galacto-oligosaccharides (GOS) and fructo-oligosaccharides (FOS). We have examined the differential gene expression of polarized Caco-2 cells treated with GOS or FOS products and their respective mock-treated cells using mRNA sequencing (RNA-seq). A total of 89 significant differentially expressed genes were identified between GOS and mock-treated groups. For FOS treatment, a reduced number of 12 significant genes were observed to be differentially expressed relative to the control group. KEGG and gene ontology functional analysis revealed that genes up-regulated in the presence of GOS were involved in digestion and absorption processes, fatty acids and steroids metabolism, potential antimicrobial proteins, energy-dependent and -independent transmembrane trafficking of solutes and amino acids. Using our data, we have established complementary non-prebiotic modes of action for these frequently used dietary fibers.

Dissecting the Interaction Deficiency of a Cartilaginous Fish Digestive Lipase with Pancreatic Colipase: Biochemical and Structural Insights.

A full-length cDNA encoding digestive lipase (SmDL) was cloned from the pancreas of the smooth-hound (Mustelus mustelus). The obtained cDNA was 1350 bp long encoding 451 amino acids. The deduced amino acid sequence has high similarity with known pancreatic lipases. Catalytic triad and disulphide bond positions are also conserved. According to the established phylogeny, the SmDL was grouped with those of tuna and Sparidae lipases into one fish digestive lipase cluster. The recently purified enzyme shows no dependence for bile salts and colipase. For this, the residue-level interactions between lipase-colipase are yet to be clearly understood. The structural model of the SmDL was built, and several dissimilarities were noticed when analyzing the SmDL amino acids corresponding to those involved in HPL binding to colipase. Interestingly, the C-terminal domain of SmDL which holds the colipase shows a significant role for colipase interaction. This is apt to prevent the interaction between fish lipase and the pancreatic colipase which and can provide more explanation on the fact that the classical colipase is unable to activate the SmDL.

Localized expression and inhibition effect of miR-184 on blood digestion and oviposition in Haemaphysalis longicornis (Acari: Ixodidae).

BACKGROUND: The hard tick Haemaphysalis longicornis (Ixodidae) is widely distributed in East Asia, China, Australia and New Zealand. It can transmit many infectious pathogens, including the causative agents of human rickettsiosis, bovine theileriosis, bovine babesiosis and canine babesiosis. Therefore, a greater understanding of H. longicornis biology might aid in the development of more effective control measures against the tick and tick-borne pathogens. METHODS: We analyzed the expression of miR-184 in different developmental stages and various tissues of H. longicornis using real-time PCR (qRT-PCR). Antagomir (Ant-184) was used to knock-down miR-184, whilst Ms-Ant and non-injected ticks were used as the negative and blank controls, respectively. We used online software tools (RNAhybrid and TargetScan) to predict the putative target genes of miR-184. RESULTS: The expression of miR-184 was highest in unfed nymphs and lowest in unfed larvae. The tissue distribution of miR-184 showed abundant expression in the midgut. To investigate the probable roles of miR-184, antagomir (Ant-184) was used to knock-down miR-184 (t(4) = 12.32, P = 0.0002). After inhibiting miR-184, other biological factors were examined in each group. The engorged body weight was significantly reduced in the treated group (Ant-184) in contrast to control groups (t(22) = 2.19, P = 0.0388). The mean duration of the egg-laying days was significantly increased (33.5 ± 1.91) and the number of eggs (t(10) = 3.147, P = 0.0137), and egg mass (t(10) = 3.4472, P = 0.0063) were significantly reduced in the treated group. During oviposition, eggs were monitored and in half of the ticks of the Ant-184 group the eggs were completely desiccated, lacked embryo development and did not hatch. We analyzed the expression of Vg proteins (Vg1, Vg2, Vg3) in semi-engorged ticks, engorged ticks, ticks at day 2 after engorgement and egg stage in Ant-184, non-injected and Ms-Ant groups, and found significant variation. CONCLUSIONS: This study provides information on the role of miR-184 in H. longicornis ticks. The data suggest that miR-184 targets Vg proteins and affects blood digestion and oviposition.

Active mode of excretion across digestive tissues predates the origin of excretory organs.

Most bilaterian animals excrete toxic metabolites through specialized organs, such as nephridia and kidneys, which share morphological and functional correspondences. In contrast, excretion in non-nephrozoans is largely unknown, and therefore the reconstruction of ancestral excretory mechanisms is problematic. Here, we investigated the excretory mode of members of the Xenacoelomorpha, the sister group to Nephrozoa, and Cnidaria, the sister group to Bilateria. By combining gene expression, inhibitor experiments, and exposure to varying environmental ammonia conditions, we show that both Xenacoelomorpha and Cnidaria are able to excrete across digestive-associated tissues. However, although the cnidarian Nematostella vectensis seems to use diffusion as its main excretory mode, the two xenacoelomorphs use both active transport and diffusion mechanisms. Based on these results, we propose that digestive-associated tissues functioned as excretory sites before the evolution of specialized organs in nephrozoans. We conclude that the emergence of a compact, multiple-layered bilaterian body plan necessitated the evolution of active transport mechanisms, which were later recruited into the specialized excretory organs.

Digestive anatomy, physiology, resting metabolism and methane production of captive maras (Dolichotis patagonum).

The digestive physiology of maras (Dolichotis patagonum) has not been investigated in detail. Maras have a particular limb anatomy facilitating a unique cursoriality among rodents. This may also have led to additional adaptations such as a reduced volume of the gastrointestinal tract. We performed macroanatomical measurements of, and determined mean particle size along, the digestive tract of 10 semi-free-ranging animals (7.04 ±â€¯1.05 kg). Additionally, we measured CH4 emission in five captive animals (7.67 ±â€¯0.98 kg) fed a diet of pelleted lucerne, and measured food intake, digestibility, and digesta mean retention time (MRT) of a solute and three particle markers (fed at <2, 10 and 20 mm particle size). The digestive tract contents represented 11.1 ±â€¯1.4% of body mass, similar to other mammals and rodents, and there was slight indication of selective small particle retention in the caecum. Secondary peaks in marker elimination patterns suggested the possibility of caecotrophy. The MRTs were 15.4 h for the solute and 13.6 h, 13.3 h and 13.3 h for the three particle markers, respectively. At a dry matter intake of 61 ±â€¯12 g kg body mass-0.75 d-1, the maras digested organic matter and neutral detergent fibre to 48 ±â€¯8% and 34 ±â€¯10%, respectively, which is in the lower range of results from horses fed on a diet with a similar fibre content. The respiratory quotient (CO2/O2) was 0.93 ±â€¯0.03, the resting metabolic rate 346 ±â€¯35 kJ kg body mass-0.75 d-1, and CH4 emissions averaged at 3.85 ±â€¯0.47 L d-1 and 14.5 ±â€¯5.2 L per kg dry matter intake; this at a CH4/CO2 ratio of 0.042 ±â€¯0.004. Thus, the methane yield was of a magnitude expected for a hypothetical ruminant of this body mass. The results are consistent with the general understanding of hystricomorph rodent digestive physiology, including caecotrophy, but do not indicate a reduction of digestive capacity to support cursoriality. These results, and those obtained from other hystricomorph rodents, suggest that CH4 production may be more prominent in rodents than previously thought.