Odorant-binding Protein 10 From Bradysia odoriphaga (Diptera: Sciaridae) Binds Volatile Host Plant Compounds.

Bradysia odoriphaga (Diptera: Sciaridae) is a major insect pest of seven plant families including 30 commercial crops in Asia. The long-term use of chemical pesticides leads to problems such as insect resistance, environmental issues, and food contamination. Against this background, a novel pest control method should be developed. In insects, odorant-binding proteins (OBPs) transport odor molecules, including pheromones and plant volatiles, to olfactory receptors. Here, we expressed and characterized the recombinant B. odoriphaga OBP BodoOBP10, observing that it could bind the sulfur-containing compounds diallyl disulfide and methyl allyl disulfide with Ki values of 8.01 µM and 7.00 µM, respectively. Homology modeling showed that the BodoOBP10 3D structure was similar to that of a typical OBP. Both diallyl disulfide and methyl allyl disulfide bound to the same site on BodoOBP10, mediated by interactions with six hydrophobic residues Met70, Ile75, Thr89, Met90, Leu93, and Leu94, and one aromatic residue, Phe143. Furthermore, silencing BodoOBP10 expression via RNAi significantly reduced the electroantennogram (EAG) response to diallyl disulfide and methyl allyl disulfide. These findings suggest that BodoOBP10 should be involved in the recognition and localization of host plants.

Consumption of Supplementary Inulin Modulates Milk Microbiota and Metabolites in Dairy Cows with Subclinical Mastitis.

The milk microbiota and mediated metabolites directly affect the health of the udder in dairy cows. Inulin, a dietary prebiotic, can modulate the profile of gastrointestinal microbiota. However, whether the inulin intake affects the milk microbial population and metabolites remains unknown. In this study, 40 subclinical mastitis (SCM) cows were randomly divided into 5 groups. Five inulin addition doses, 0, 100, 200, 300, and 400 g/day per cow, based on the same basal diet, were supplemented. The experiments lasted for 8 weeks. The results showed lower relative abundance of mastitis-causing and proinflammation microbes in milk (i.e., Escherichia-Shigella, Pseudomonas, Rhodococcus, Burkholderia-Caballeronia-Paraburkholderia, etc.) and higher abundances of probiotics and commensal bacteria, such as Lactobacillus, Bifidobacterium, etc., in the cows fed 300 g/day inulin compared to that in the control group. Meanwhile, the levels of arachidonic acid proinflammatory mediators (leukotriene E3, 20-carboxy-leukotriene B4, and 12-Oxo-c-LTB3) and phospholipid metabolites were reduced, and the levels of compounds with antibacterial and anti-inflammatory potential (prostaglandin A1, 8-iso-15-keto-prostaglandin E2 [PGE2], etc.) and participating energy metabolism (citric acid, l-carnitine, etc.) were elevated. These data suggested that inulin intake might modulate the microflora and metabolite level in extraintestinal tissue, such as mammary gland, which provided an alternative for the regulation and mitigation of SCM. IMPORTANCE The profile of the microbial community and metabolic activity in milk are the main determinants of udder health status and milk quality. Recent studies have demonstrated that diet could directly modulate the mammary gland microbiome. Inulin is a probiotic dietary fiber which can improve the microbiota population in the gastrointestinal tract. However, whether inulin intake can further regulate the profile of the microbiota and metabolic activities in milk remains unclear. In subclinical mastitic cows, we found that inulin supplementation could reduce the abundance of Escherichia-Shigella, Pseudomonas, Rhodococcus, and Burkholderia-Caballeronia-Paraburkholderia and the levels of (±)12, 13-DiHOME, leukotriene E3 and 20-carboxy-leukotriene B4 etc., while it elevated the abundance of Lactobacillus, Bifidobacterium, and Muribaculaceae, as well as the levels of prostaglandin A1 (PGA1), 8-iso-15-keto-PGE2, benzoic acid, etc. in milk. These data suggest that inulin intake affects the profile of microorganisms and metabolites in milk, which provides an alternative for the regulation of mastitis.

Discrepancies among healthy, subclinical mastitic, and clinical mastitic cows in fecal microbiome and metabolome and serum metabolome.

Mastitis is generally considered a local inflammatory disease caused by the invasion of exogenous pathogens and resulting in the dysbiosis of microbiota and metabolites in milk. However, the entero-mammary pathway theory may establish a possible link between some endogenous gut bacteria and the occurrence and development of mastitis. In the current study, we attempted to investigate differences in the gut microbiota profile and metabolite composition in gut and serum from healthy cows and those with subclinical mastitis and clinical mastitis. Compared with those of healthy cows, the microbial community diversities in the feces of cows with subclinical mastitis (SM) and clinical mastitis (CM) were lower. Lower abundance of Bifidobacterium, Romboutsia, Lachnospiraceae_NK3A20_group, Coprococcus, Prevotellaceae_UCG-003, Ruminococcus, and Alistipes, and higher abundance of the phylum Proteobacteria and the genera Escherichia-Shigella and Streptococcus were observed in CM cows. Klebsiella and Paeniclostridium were significantly enriched in the feces of SM cows. Several similarities were observed in feces and serum metabolites in mastitic cows. Higher levels of proinflammatory lipid products (20-trihydroxy-leukotriene-B4, 13,14-dihydro-15-keto-PGE2, and 9,10-dihydroxylinoleic acids) and lower levels of metabolites involved in secondary bile acids (deoxycholic acid, 12-ketolithocholic acid), energy (citric acid and 3-hydroxyisovalerylcarnitine), and purine metabolism (uric acid and inosine) were identified in both SM and CM cows. In addition, elevated concentrations of IL-1ß, IL-6, tumor necrosis factor-α and decreased concentrations of glutathione peroxidase and superoxide dismutase were detected in the serum of SM and CM cows. Higher serum concentrations of triglyceride and total cholesterol and lower concentrations of high-density lipoproteins in mastitic cows might be related to changes in the gut microbiota and metabolites. These findings suggested a significant difference in the profile of feces microbiota and metabolites in cows with different udder health status, which might increase our understanding of bovine mastitis.

Molecular characterization and functional analysis of the Halloween genes and CYP18A1 in Bemisia tabaci MED.

The ecdysteroid hormone 20-hydroxyecdysone (20E), a critical hormone in arthropods, plays an essential role in insect growth, molting and reproduction. A previous study showed that 20E is actually regulated by six P450 genes (five P450 genes belonging to the Halloween family and a CYP18A1 gene) in model insects. However, the role of the six P450 genes in Bemisia tabaci Q (also call Mediterranean, MED), an important pest of field crops, remains unclear. Here, six P450 genes were cloned by RT-PCR, and the phylogenetic tree indicated a close orthologous relationship of these P450 genes between MED and other insects. Spatiotemporal expression profiling revealed that five P450 genes (CYP18A1, CYP306A1, CYP307A2, CYP314A1 and CYP315A1) were expressed at significantly higher levels in the head than in the abdomen and thorax. Four P450 genes (CYP302A1, CYP307A2, CYP314A1 and CYP315A1) were expressed at the highest levels in males, and CYP18A1 was expressed at the highest levels in the 4th nymph stage. The molting process was delayed by approximately 1-3 days after knockdown of these genes at the 4th nymph stage, and the mean proportion of shriveled or dead insects reached 8.3% (CYP18A1), 20.8% (CYP302A1), 7.0% (CYP307A2), 31.8% (CYP306A1), 28.6% (CYP314A1) and 24.1% (CYP315A1). In addition, 20E rescued the negative effect of ds-CYP306A1, ds-CYP314A1 and ds-CYP315A1 on the eclosion rate. We concluded that these Halloween genes and CYP18A1 likely participate in the development of MED, and in particular, CYP306A1 could be used as a putative insecticide target for controlling this piercing-sucking insect.

Cloning, characterization and widespread expression analysis of testicular piRNA-like chicken RNAs.

Piwi-interacting RNAs (piRNAs) are small RNAs abundant in the germline that have been implicated in germline development and maintenance of genomic integrity across several animal species including human, mouse, rat, zebrafish and drosophila. Tens of thousands of piRNAs have been discovered, yet abundant piRNAs have still not been detected in various eukaryotic organisms. This is a report on the characterization, cloning and expression profiling of piRNA-like chicken RNAs. Here, we identified 19 piRNAs, each 23-39 nucleotides long, from chicken testis using a small RNA cDNA library and T-A cloning methods. Three different pilRNAs were selected according to size, homology and secondary structure for temporal and spatial expression by Q-PCR technology in different tissues at five growth and four development stages of Chinese indigenous Rugao chickens (RG) and introduced recessive white feather chickens (RW). We found that, consistent to other organisms, pilRNA-encoding sequences within the chicken genome were asymmetrically distributed on the chromosomes while displaying a preference for intergenic regions across the genome. Interestingly, unlike miRNAs with unique stem-loop structures (mature miRNAs form stem section and the rest form loop section), distinct secondary structures of pilRNAs were predicted. In addition, chicken pilRNAs were not only abundant in the germline but also existed in somatic tissues, where, expression levels were influenced mainly by different pilRNAs, breed and gender. Taken together, our results suggest that two distinct secondary structures exist between pilRNAs and miRNAs, which may clarify the splicing and processing mechanisms of the two small RNAs are possible different. Moreover, our results suggest that pilRNAs may not only be confined to development and maintenance of the germline but may also play important roles in somatic tissues. Additionally, different pilRNAs may be involved in the unique regulatory machinery of complex biological processes.

Cloning and expression characterization of the chicken Piwil1 gene.

Piwi gene involves in the germline stem cells self-renewal, transposon silencing and post-transcriptional gene regulation in the majority of organisms; however, the biological function of Piwi gene in poultry remains unclear. Here we cloned the Piwi-like 1 (Piwil1) gene and characterized its expression in the Langshan chickens during the development. The results showed that the PIWIL1 protein was the homolog of mice MIWI and human HIWI proteins (100 % identity), and encoded a cytoplasmic protein including the two conserved domains PAZ and PIWI. In males, the expression of Piwil1 gene showed a bimodal distribution in the gonads during embryogenesis with peaks at embryonic 14.5 and 17.5-18.5 days respectively. After puberty, the expression of Piwil1 gene increased sharply and reached a high level at the sexual maturity. The mRNA expression of Piwil1 gene at 27 weeks of age is 35-40 times that of 0 week of age, indicating that the high expression of Piwil1 gene was essential to maintain the spermatogenesis. In females, the expression of Piwil1 gene showed a unimodal distribution in the embryonic gonads. A strong peak appeared at E16.5-17.5d when the primary oocytes have entered the prophase I of meiosis. Subsequently, the expression of Piwil1 gene decreased gradually and kept at the low level during the embryogenesis. So Piwil1 gene was likely to play an important role during the meiosis I. This report filled in partly the gap of the Piwi gene researches in poultry and defined our research directions in future.

Fitness cost of spinosad resistance related to vitellogenin in Frankliniella occidentalis (Pergande).

BACKGROUND: The western flower thrips Frankliniella occidentalis, a worldwide agricultural pest, has developed resistance to an array of insecticides. Spinosad resistance confers an apparent fitness cost in F. occidentalis. In the present study, we compared the reproductive capacities, ovary development, and the expression of the vitellogenin (Vg) gene in spinosad-susceptible (Ivf03) and -resistant (NIL-R) near isogenetic lines of F. occidentalis in order to clarify the reason for the fitness cost in spinosad resistance. RESULTS: The NIL-R strain exhibited a 17.9% decrease in fecundity (eggs laid per female) as compared to the Ivf03 strain, and the ovariole was significantly shortened by 2.8% in the NIL-R strain relative to the Ivf03 strain. Compared to the Ivf03 strain, the expression levels of Vg mRNA and protein were downregulated by 33.7% and 32.9% in the NIL-R strain, respectively. Moreover, interference with the Vg gene significantly reduced the expression levels of Vg mRNA and protein, and decreased ovariole length, survival rates and the fecundity of both strains. CONCLUSION: The results indicate that the downregulated expression of Vg may contribute to the reduction of ovariole length and consequently to a fitness cost in spinosad-resistant F. occidentalis. The results not only increase our understanding of the evolution of insecticide resistance, but also could contribute to the formulation of control strategy of F. occidentalis. © 2022 Society of Chemical Industry.

Chemosensory protein 4 is required for Bradysia odoriphaga to be olfactory attracted to sulfur compounds released from Chinese chives.

Bradysia odoriphaga (Diptera: Sciaridae) is a serious pest of Chinese chives cultivated in China. Chemosensory proteins (CSPs) are important components of insect olfactory systems that capture and bind environmental semiochemicals which are then transported to olfactory receptors. Despite their importance, the mechanism of olfaction and related behavioral processes in B. odoriphaga have not been characterized. Here, we found that BodoCSP4 has an important olfactory function. RT-qPCR indicated that BodoCSP4 expression was highest in the heads (antennae removed) of adult males, followed by the antennae of adult males. Competitive binding assays with 33 ligands indicated that BodoCSP4 binds well with methyl allyl disulfide, diallyl disulfide, and n-heptadecane; the corresponding dissolution constants (Ki) were as high as 5.71, 5.71, and 6.85 µM, respectively. 3D-structural and molecular docking indicated that BodoCSP4 has five α-helices and surrounds the ligand with certain hydrophobic residues including Leu60, Leu63, Leu64, Ala67, Val28, Ile30, Ile33, Leu34, and Val86, suggesting these residues help BodoCSP4 bind to ligands. Silencing of BodoCSP4 significantly decreased the attraction of B. odoriphaga males to diallyl disulfide and n-heptadecane but not to methyl allyl disulfide in Y-tube olfaction assays. These results increase our understanding of how BodoCSP4 contributes to host and female localization by B. odoriphaga males.

Starch and Cellulose Degradation in the Rumen and Applications of Metagenomics on Ruminal Microorganisms.

Carbohydrates (e.g., starch and cellulose) are the main energy source in the diets of dairy cows. The ruminal digestion of starch and cellulose is achieved by microorganisms and digestive enzymes. In order to improve their digestibility, the microbes and enzymes involved in starch and cellulose degradation should be identified and their role(s) and activity known. As existing and new analytical techniques are continuously being developed, our knowledge of the amylolytic and cellulolytic microbial community in the rumen of dairy cows has been evolving rapidly. Using traditional culture-based methods, the main amylolytic and cellulolytic bacteria, fungi and protozoa in the rumen of dairy cows have been isolated. These culturable microbes have been found to only account for a small fraction of the total population of microorganisms present in the rumen. A more recent application of the culture-independent approach of metagenomics has acquired a more complete genetic structure and functional composition of the rumen microbial community. Metagenomics can be divided into functional metagenomics and sequencing-based computational metagenomics. Both approaches have been applied in determining the microbial composition and function in the rumen. With these approaches, novel microbial species as well as enzymes, especially glycosyl hydrolases, have been discovered. This review summarizes the current state of knowledge regarding the major amylolytic and cellulolytic microorganisms present in the rumen of dairy cows. The ruminal amylases and cellulases are briefly discussed. The application of metagenomics technology in investigating glycosyl hydrolases is provided and the novel enzymes are compared in terms of glycosyl hydrolase families related to amylolytic and cellulolytic activities.

Changes in the Profile of Fecal Microbiota and Metabolites as Well as Serum Metabolites and Proteome After Dietary Inulin Supplementation in Dairy Cows With Subclinical Mastitis.

The occurrence and development of mastitis is linked to dysbiostic gastrointestinal microbiota. Inulin is a dietary prebiotic that improves the profile of intestinal flora. Our previous study showed that inulin supplementation could improve the ruminal microbes of subclinical mastitis (SCM) cows. The current study attempted to further investigate the response of hindgut (fecal) microbiome and metabolites, serum metabolism, and protein expression to inulin in the in SCM cows. Different levels of inulin (0, 100, 200, 300, and 400 g/day per cow) were supplemented in SCM cows. Compared with control group, Bacteroides and Bifidobacteria were increased, and Paeniclostridium, Ruminococcaceae, Coprococcus, and Clostridia were decreased in the feces of inulin groups, and accompanied with elevated propionate and butyrate concentrations, while secondary bile acid (SBA) metabolites were increased and proinflammatory lipid oxidation products were dropped in both feces and serum. In serum, inulin intake suppressed the levels of triglyceride (TG) and low-density lipoprotein (LDL). Serum proteome analysis found that CD44 antigen, phosphatidylinositol-glycan-specific phospholipase D, apolipoprotein A-II, and superoxide dismutase [Cu-Zn] were upregulated, while cathelicidin-1, haptoglobin, serpin A3, inter-alpha-trypsin inhibitor heavy chain H4 were downregulated in inulin groups. These findings suggested further evidence for inulin supplementation in amelioration of inflammatory symptoms in SCM cows, which might provide alternative treatment for mastitis.

Glucogenic and lipogenic diets affect in vitro ruminal microbiota and metabolites differently.

This study was conducted to evaluate the effects of two glucogenic diets (C: ground corn and corn silage; S: steam-flaked corn and corn silage) and a lipogenic diet (L: sugar beet pulp and alfalfa silage) on the ruminal bacterial and archaeal structures, the metabolomic products, and gas production after 48 h in vitro fermentation with rumen fluid of dairy cows. Compared to the C and S diets, the L dietary treatment leaded to a lower dry matter digestibility (DMD), lower propionate production and ammonia-nitrogen concentration. The two glucogenic diets performed worse in controlling methane and lactic acid production compared to the L diet. The S diet produced the greatest cumulative gas volume at any time points during incubation compared to the C and L diet. The metabolomics analysis revealed that the lipid digestion especially the fatty acid metabolism was improved, but the amino acid digestion was weakened in the L treatment than in other treatments. Differences in rumen fermentation characteristics were associated with (or resulting from) changes in the relative abundance of bacterial and archaeal genera. The rumen fluid fermented with L diet had a significantly higher number of cellulolytic bacteria, including the genera of Ruminococcus, Butyrivibrio, Eubacterium, Lachnospira, unclassified Lachnospiraceae, and unclassified Ruminococcaceae. The relative abundances of amylolytic bacteria genera including Selenomonas_1, Ruminobacter, and Succinivibrionaceae_UCG-002 were higher in samples for diets C and S. The results indicated that the two glucogenic diets leaded to a higher relative abundance of bacteria which functions in succinate pathway resulting in a higher propionate production. The steam-flaked corn diet had a higher gas production and lower level of metabolites in fatty acids and amino acids. Most highly abundant bacteria were observed to be not sensitive to dietary alterations of starch and fiber, except for several amylolytic bacteria and cellulolytic bacteria. These finding offered new insights on the digesting preference of ruminal bacteria, which can assist to improve the rumen functioning.

Effects of monochromatic lights on the growth performance, carcass characteristics, eyeball development, oxidation resistance, and cecal bacteria of Pekin ducks.

OBJECTIVE: Light is a significant component of housing environment in commercial poultry industry. This study was conducted to investigate whether Pekin ducks perform better under monochromatic lights than under white light with respect to their growth performance, carcass quality, eyeball development, oxidation resistance, and cecal bacterial communities. METHODS: A total of 320 one-day-old male Pekin ducklings were randomly distributed into five rooms with different light treatments, white, red, yellow, green, and blue light. Each room consisted of 4 replicated pens with 16 ducklings per pen. RESULTS: Blue light significantly decreased fat deposition by decreasing abdominal fat. Long wavelength light, such as red, green, and yellow light, considerably increased the back-to-front eyeball diameter and the red light potentially enlarged the side-to-side eyeball diameter. Besides, the blue light had adverse effects on the oxidation resistance status in terms of increasing the product malonaldehyde of lipid oxidation and decreasing the plasma concentration of total superoxide dismutase. The phyla of Firmicutes had the greatest abundance in the green and blue treatments, while Bacteroidetes in blue treatment was the least. The genus of Faecalibacterium was significantly lower under the red light. CONCLUSION: The high risk of cecal health status and decreased anti-oxidation activity were observed under blue light. Red, yellow, and green light might increase the risk of oversized eyeball and cecal illness. Therefore, monochromatic lights compared to white light did not show advantages on the performance of housing ducks, it turns out that the white light is the best light condition for grow-out ducks.

Dietary supplementation with inulin improves lactation performance and serum lipids by regulating the rumen microbiome and metabolome in dairy cows.

This study investigated the effects of inulin on rumen fermentation parameters, ruminal microbiome and metabolites, as well as lactation performance and serum indexes in dairy cows. Sixteen Holstein dairy cows with similar body conditions were randomly divided into 2 groups (n = 8 per group), with inulin addition at 0 and 200 g/d per cow. The experiment lasted for 6 weeks, including a 1-week adaptation period and a 5-week treatment period. At the end of the experimental period, the milk, serum and rumen fluid were sampled and analyzed. The microbiome and metabolome in the rumen fluid were analyzed via 16S rRNA sequencing and untargeted metabolomics, respectively. The results showed that supplementation with inulin (200 g/d per cow) increased the milk yield (P = 0.001), milk protein (P = 0.032), lactose rate (P = 0.004) and proportion of saturated fatty acids (SFA) in milk (P < 0.001), but decreased the proportion of unsaturated fatty acids (USFA) (P = 0.041). Rumen pH (P = 0.040) and the concentration of NH3-N (P = 0.024) were decreased; however, acetate (P < 0.001), propionate (P = 0.003), butyrate (P < 0.001) and lactic acid (LA) (P = 0.043) were increased. The total cholesterol (TC) (P = 0.008) and triglycerides (TG) (P = 0.01) in serum were also reduced. Additionally, inulin addition elevated the relative abundance of several beneficial symbiotic and short-chain fatty acid (SCFA)-producing bacteria, such as Muribaculaceae (false discovery rate [FDR]-adjusted P < 0.01), Acetitomaculum (FDR-adjusted P = 0.043), and Butyrivibrio (FDR-adjusted P = 0.036), while elevating the levels of L-lysine (FDR-adjusted P = 4.24 × 10-3), L-proline (FDR-adjusted P = 0.0158), and L-phenylalanine (FDR-adjusted P = 0.027). In contrast, several pathogens and ruminal bacteria abundant in high-fat diets, such as Escherichia-Shigella (FDR-adjusted P = 0.022), Erysipelotrichaceae __UCG-004 (FDR-adjusted P < 0.01) and RF39 (FDR-adjusted P = 0.042) were decreased along with the reduction of lysophosphatidylcholine (LysoPC) (18:1 (9Z)) (FDR-adjusted P = 1.03 × 10-3), LysoPC (16:0) (FDR-adjusted P = 0.0108), LysoPC (18:2 (9Z, 12Z)) (FDR-adjusted P = 1.65 × 10-3) and 8-methylnonenoate. In conclusion, dietary inulin supplementation could increase the relative abundance of commensal microbiota and SCFA-producing bacteria, upregulate amino acidmetabolism and downregulate lipid metabolism in the rumen of dairy cows, which might further improve lactation performance and the level of serum lipids.

Electrochemical device based on nonspecific DNAzyme for the high-accuracy determination of Ca2+ with Pb2+ interference.

Calcium is one of the most abundant and indispensable elements in biology, as it is a vital component of nerves, bones, and muscles and maintains the excitability of normal neuromuscular muscles. However, it may be harmful to the human body and even damage the organs if the calcium content exceeds the standard value by several times. To evaluate the level of calcium ions (Ca2+), an electrochemical biosensor (FET/SWNTs/Cazyme) was developed using a nonspecific DNAzyme with high stability, which combined the unique advantage of field-effect transistors and single-walled carbon nanotubes, while being easy-to-use and having excellent sensitivity. The incubation time and voltage after optimization were 15 min and +0.02 V. The nonspecific DNAzyme-based biosensor was sensitive to Ca2+, but it was also interfered with by Pb2+, which affected the detection accuracy. To solve this shortcoming, an electrochemical device was proposed, in which FET/SWNTs/Cazyme combined with other specific biosensors for Pb2+, and then established some data processing models were established through support vector machine regression (SVMR) and artificial neural network fitting (ANNF). For the optimal SVMR, the electrochemical device can determine the Ca2+ concentration in the range of 7.5-1000 µM with a detection limit of 5.48 µM. Finally, the prepared electrochemical device was employed to detect the Ca2+ in different milk and water samples.

Dietary Supplementation of Inulin Ameliorates Subclinical Mastitis via Regulation of Rumen Microbial Community and Metabolites in Dairy Cows.

Subclinical mastitis (SCM) is one of the highly infectious diseases in dairy cows with the characteristics of high incidence and nonvisible clinical symptoms. The gastrointestinal microbiota is closely related to mastitis. Inulin is a prebiotic fiber with functions in improving intestinal microbial communities and enhancing the host's immunity. However, the impact of dietary inulin on the rumen inner environment remains unknown. The current study investigated whether inulin could relieve SCM by affecting the profiles of ruminal bacterial and metabolites in dairy cows. Inulin inclusion rates were 0, 100, 200, 300, and 400 g/day per cow, respectively. Inulin increased milk yield, milk protein, and lactose and reduced the somatic cell counts (SCC) in milk. In serum, the concentration of proinflammatory cytokines, such as interleukin-6 (IL-6), IL-8, tumor necrosis factor α (TNF-α), and malondialdehyde (MDA) were decreased, and IL-4 and superoxide dismutase (SOD) were increased. Meanwhile, inulin increased the concentration of propionate, butyrate, and lactic acid (LA), while it decreased NH3-N in rumen. The propionate- and butyrate-producing bacteria (e.g., Prevotella and Butyrivibrio) and several beneficial commensal bacteria (e.g., Muribaculaceae and Bifidobacterium) as well as metabolites related to energy and amino acid metabolism (e.g., melibiose and l-glutamate) were increased. However, several proinflammatory bacteria (e.g., Clostridia UCG-014, Streptococcus, and Escherichia-Shigella) were decreased, accompanied by the downregulation of lipid proinflammatory metabolites, for example, ceramide(d18:0/15:0) [Cer(d18:0/15:0)] and 17-phenyl-18,19,20-trinor-prostaglandin E2. In the current study, the above indicators showed the best response in the 300 g/day inulin group. Overall, dietary supplementation of inulin could alleviate inflammatory responses in cows with SCM through improving the rumen inner environment. IMPORTANCE The correlation between mastitis and the gastrointestinal microbiome in dairy cows has been demonstrated. Regulating the profile of rumen microorganisms may contribute to remission of subclinical mastitis (SCM). Supplementation of inulin in the diets of cows with SCM could increase the abundance of short-chain fatty acid (SCFA)-producing bacteria and beneficial commensal bacteria in rumen and meanwhile the levels of amino acids and energy metabolism. Conversely, the abundance of ruminal bacteria and metabolites with proinflammatory effects were decreased. Our study suggests that the improvement of the rumen internal environment by inulin supplementation could ameliorate inflammatory responses during SCM in dairy cows and thus improve lactation performance and milk quality. Our results provide a theoretical basis for regulation measures of SCM in dairy cows.

Rumen microbiome structure and metabolites activity in dairy cows with clinical and subclinical mastitis.

BACKGROUND: Due to the high prevalence and complex etiology, bovine mastitis (BM) is one of the most important diseases to compromise dairy cow health and milk quality. The shift in milk compositions has been widely investigated during mastitis, but recent studies suggested that gastrointestinal microorganism also has a crucial effect on the inflammation of other peripheral tissues and organs, including the mammary gland. However, research focused on the variation of rumen inner-environment during mastitis is still limited. Therefore, the ruminal microbial profiles, metabolites, and milk compositions in cows with different udder health conditions were compared in the present study. Furthermore, the correlations between udder health status and ruminal conditions were investigated. Based on the somatic cell counts (SCC), California mastitis test (CMT) parameters and clinical symptoms of mastitis, 60 lactating Holstein dairy cows with similar body conditions (excepted for the udder health condition) were randomly divided into 3 groups (n = 20 per group) including the healthy (H) group, the subclinical mastitis (SM) group and the clinical mastitis (CM) group. Lactation performance and rumen fermentation parameters were recorded. And rumen microbiota and metabolites were also analyzed via 16S rRNA amplicon sequencing and untargeted metabolomics, respectively. RESULTS: As the degree of mastitis increased, rumen lactic acid (LA) (P < 0.01), acetate, propionate, butyrate, valerate (P < 0.001), and total volatile fatty acids (TVFAs) (P < 0.01) concentrations were significantly decreased. In the rumen of CM cows, the significantly increased bacteria related to intestinal and oral inflammation, such as Lachnospiraceae (FDR-adjusted P = 0.039), Moraxella (FDR-adjusted P = 0.011) and Neisseriaceae (FDR-adjusted P = 0.036), etc., were accompanied by a significant increase in 12-oxo-20-dihydroxy-leukotriene B4 (FDR-adjusted P = 5.97 × 10- 9) and 10beta-hydroxy-6beta-isobutyrylfuranoeremophilane (FDR-adjusted P = 3.88 × 10- 10). Meanwhile, in the rumen of SM cows, the Ruminiclostridium_9 (FDR-adjusted P = 0.042) and Enterorhabdus (FDR-adjusted P = 0.043) were increased along with increasing methenamine (FDR-adjusted P = 6.95 × 10- 6), 5-hydroxymethyl-2-furancarboxaldehyde (5-HMF) (FDR-adjusted P = 2.02 × 10- 6) and 6-methoxymellein (FDR-adjusted P = 2.57 × 10- 5). The short-chain fatty acids (SCFAs)-producing bacteria and probiotics in rumen, including Prevoterotoella_1 (FDR-adjusted P = 0.045) and Bifidobacterium (FDR-adjusted P = 0.035), etc., were significantly reduced, with decreasing 2-phenylbutyric acid (2-PBA) (FDR-adjusted P = 4.37 × 10- 6). CONCLUSION: The results indicated that there was a significant shift in the ruminal microflora and metabolites associated with inflammation and immune responses during CM. Moreover, in the rumen of cows affected by SM, the relative abundance of several opportunistic pathogens and the level of metabolites which could produce antibacterial compounds or had a competitive inhibitory effect were all increased.

Ruminal Methanogenic Responses to the Thiamine Supplementation in High-Concentrate Diets.

BACKGROUND: Thiamine supplementation in high-concentrate diets (HC) was confirmed to attenuate ruminal subacute acidosis through promoting carbohydrate metabolism, however, whether thiamine supplementation in HC impacts methane metabolism is still unclear. Therefore, in the present study, thiamine was supplemented in the high-concentrate diets to investigate its effects on ruminal methanogens and methanogenesis process. METHODS: an in vitro fermentation experiment which included three treatments: control diet (CON, concentrate/forage = 4:6; DM basis), high-concentrate diet (HC, concentrate/forage = 6:4; DM basis) and high-concentrate diet supplemented with thiamine (HCT, concentrate/forage = 6:4, DM basis; thiamine supplementation content = 180 mg/kg DM) was conducted. Each treatment concluded with four repeats, with three bottles in each repeat. The in vitro fermentation was sustained for 48h each time and repeated three times. At the end of fermentation, fermentable parameters, ruminal bacteria and methanogens community were measured. RESULTS: HC significantly decreased ruminal pH, thiamine and acetate content, while significantly increasing propionate content compared with CON (p < 0.05). Conversely, thiamine supplementation significantly increased ruminal pH, acetate while significantly decreasing propionate content compared with HC treatment (p < 0.05). No significant difference of ruminal methanogens abundances among three treatments was observed. Thiamine supplementation significantly decreased methane production compared with CON, while no significant change was found in HCT compared with HC. CONCLUSION: thiamine supplementation in the high-concentrate diet (HC) could efficiently reduce CH4 emissions compared with high-forage diets while without causing ruminal metabolic disorders compared with HC treatment. This study demonstrated that supplementation of proper thiamine in concentrate diets could be an effective nutritional strategy to decrease CH4 production in dairy cows.

Nutritional value, bioactivity, and application potential of Jerusalem artichoke (Helianthus tuberosus L.) as a neotype feed resource.

The large-scale development of herbivorous animal husbandry in China has increased the demand for forage products. However, due to scarce land resources and poor soil quality, forage is in short supply. In particular, high-quality forage in China heavily relies on imports. The contradiction between supply and demand for forage grass products is increasingly notable. Therefore, the development of indigenous new forage resources with a strong ecological adaptability and a high nutritional value is a key to solving this problem. Jerusalem artichoke (JA, Helianthus tuberosus L.), a perennial herb of the genus Helianthus, has advantageous growth traits such as resistance to salinity, barrenness, drought, cold, and disease. The contents of crude protein, crude fiber, and calcium in the optimal harvest period of forage-type JA straw are comparable to those of alfalfa hay at the full bloom stage and the straw of ryegrass and corn at the mature stage. Inulin in JA tubers is a functional ingredient that has prebiotic effects in the gastrointestinal tract of monogastric animals and young ruminants. In addition, some bioactive substances (e.g. flavonoids, phenolic acids, sesquiterpenes, polysaccharides, and amino acids) in JA leaves and flowers have antibacterial, anti-inflammatory, and antioxidant functions as well as toxicities to cancer cells. These functional ingredients may provide effective alternatives to antibiotics used in livestock production. In this review, we summarized the potentials of JA as a feed ingredient from the aspects of nutritional value and fermenting characteristics of the straw, the functions of physiological regulation and disease prevention of inulin in the tubers, and bioactive substances in the leaves and flowers.

Assessing Pesticide Residue and Spray Deposition in Greenhouse Eggplant Canopies to Improve Residue Analysis.

Pesticide residue analysis is critically important for ensuring food safety and regulating international trade. In a greenhouse setting, we assessed pesticide residue and exposure patterns using liquid chromatography-mass spectrometry and Allura Red tracer analysis. Our results show significant effects of eggplant canopy position and spray concentration on deposition parameters. Specifically, spray coverage, spray density, deposition, and residue were generally greater for leaves in the higher canopy. Deposition and residue, unlike spray coverage and spray density, linearly correlated with spray concentration. Pesticide residues on leaves linearly correlated with depositions on artificial targets, and a correction factor was constructed to estimate residues from tracer depositions. This comparative analysis suggests that tracer analysis using Allura Red in combination with artificial targets is a time- and cost-saving alternative to conventional residue analysis for assessing spray parameters under high-volume spraying scenarios. A better understanding of pesticide residue and spray patterns will not only enhance the safety of our food supply chain but also improve pest control efficacy within the sustainable integrated pest management framework.

Metagenomic Insights into Effects of Thiamine Supplementation on Carbohydrate-Active Enzymes' Profile in Dairy Cows Fed High-Concentrate Diets.

As the co-enzyme of pyruvate formate-lyase under ruminal anaerobic condition, thiamine plays a critical role in carbohydrate metabolism in dairy cows. The objective of this study was to investigate the impacts of thiamine supplementation on ruminal carbohydrate-active enzymes. Twelve Holstein dairy cows were randomly assigned into three dietary treatments: control diet (CON; 20% starch, dry matter (DM) basis), high-concentrate diet (HC; 33.2% starch, DM basis) and a high-concentrate diet supplemented with 180 mg thiamine/kg DM (HCT; 33.2% starch, DM basis). Dry matter intake and milk production were recorded for 21 days. Rumen fluid samples were collected, and ruminal pH and volatile fatty acids (VFAs) were measured. The metagenome sequencing technique was used to detect the genes in ruminal microorganisms and identify putative carbohydrate-active enzymes. The total abundances of carbohydrate-active enzymes and fiber-degrading enzymes were both reduced by HC with no effect on starch-degrading enzymes compared with CON. However, the fiber-degrading enzymes and starch-degrading enzymes were both increased after thiamine supplementation. These results indicated that 180 mg thiamine /kg DM might effectively improve rumen carbohydrate metabolism through increasing the abundance of ruminal carbohydrate-active enzymes and consequently balanced the rumen volatile fatty acids and rumen pH, providing a practical strategy in preventing subacute ruminal acidosis in cows offered HC.