Spatially resolved transcriptomic profiling of placental development in dairy cow

The placenta plays a crucial role in successful mammalian reproduction.Ruminant animals possess a semi-invasive placenta characterized by a highly vascularized structure formed by maternal endometrial caruncles and fetal placental cotyledons,essential for full-term fetal development.The cow placenta harbors at least two trophoblast cell populations:uninucleate(UNC)and binucleate(BNC)cells.However,the limited capacity to elucidate the transcriptomic dynamics of the placental natural environment has resulted in a poor understanding of both the molecular and cellular interactions between trophoblast cells and niches,and the molecular mechanisms governing trophoblast differentiation and functionalization.To fill this knowledge gap,we employed Stereo-seq to map spatial gene expression patterns at near single-cell resolution in the cow placenta at 90 and 130 days of gestation,attaining high-resolution,spatially resolved gene expression profiles.Based on clustering and cell marker gene expression analyses,key transcription factors,including YBX1 and NPAS2,were shown to regulate the heterogeneity of trophoblast cell subpopulations.Cell communication and trajectory analysis provided a framework for understanding cell-cell interactions and the differentiation of trophoblasts into BNCs in the placental microenvironment.Differential analysis of cell trajectories identified a set of genes involved in regulation of trophoblast differentiation.Additionally,spatial modules and co-variant genes that help shape specific tissue structures were identified.Together,these findings provide foundational insights into important biological pathways critical to the placental development and function in cows.

Repeated inoculation with rumen fluid accelerates the rumen bacterial transition with no benefit on production performance in postpartum Holstein dairy cows

Background The dairy cow’s postpartum period is characterized by dramatic physiological changes,therefore imposing severe challenges on the animal for maintaining health and milk output.The dynamics of the ruminal microbiota are also tremendous and may play a crucial role in lactation launch.We aim to investigate the potential benefits of early microbial intervention by fresh rumen microbiota transplantation(RMT)and sterile RMT in postpartum dairy cows.Twelve fistulated peak-lactation dairy cows were selected to be the donors for rumen fluid collection.Thirty postpartum cows were divided into 3 groups as the transplantation receptors respectively receiving 10 L fresh rumen fluid(FR),10 L sterile rumen fluid(SR),or 10 L saline(CON)during 3 d after calving.Results Production performance,plasma indices,plasma lipidome,ruminal microbiome,and liver transcriptome were recorded.After fresh and sterile RMT,we found that the molar proportion of propionic acid was increased on d 7 in the FR and SR groups and the bacterial composition was also significantly changed when compared with the CON group.A similarity analysis showed that the similarities between the CON group and FR or SR group on d 7 were 48.40%or 47.85%,whereas the similarities between microbiota on d 7 and 21 in the FR and SR groups were 68.34%or 66.85%.Dry matter intake and feed efficiency were not affected by treatments.Plasmaβ-hydroxybutyrate concentration in the FR group was decreased and significantly different lipids mainly included phosphatidylcholine and lysophosphatidylcholine containing polyunsaturated fatty acids.Hepatic transcriptomics analysis indicated acutephase response pathways were upregulated in the SR group.Conclusions Our study suggests that RMT can shorten the transition process of the ruminal microbiota of postpartum dairy cows with no benefit on dry matter intake or feed efficiency.Inoculation with rumen fluid may not be a useful approach to promote the recovery of postpartum dairy cows.

Maternal supplementation with n-3 fatty acids affects placental lipid metabolism, inflammation, oxidative stress, the endocannabinoid system, and the neonate cytokine concentrations in dairy cows

Background The placenta plays a crucial role in supporting and influencing fetal development.We compared the effects of prepartum supplementation with omega-3(n-3)fatty acid(FA)sources,flaxseed oil(FLX)and fish oil(FO),on the expression of genes and proteins related to lipid metabolism,inflammation,oxidative stress,and the endocannabinoid system(ECS)in the expelled placenta,as well as on FA profile and inflammatory response of neonates.Late-pregnant Holstein dairy cows were supplemented with saturated fat(CTL),FLX,or FO.Placental cotyledons(n=5)were collected immediately after expulsion,and extracted RNA and proteins were analyzed by RTPCR and proteomic analysis.Neonatal blood was assessed for FA composition and concentrations of inflammatory markers.Results FO increased the gene expression of fatty acid binding protein 4(FABP4),interleukin 10(IL-10),catalase(CAT),cannabinoid receptor 1(CNR1),and cannabinoid receptor 2(CNR2)compared with CTL placenta.Gene expression of ECS-enzyme FA-amide hydrolase(FAAH)was lower in FLX and FO than in CTL.Proteomic analysis identified 3,974 proteins;of these,51–59 were differentially abundant between treatments(P≤0.05,|fold change|≥1.5).Top canonical pathways enriched in FLX vs.CTL and in FO vs.CTL were triglyceride metabolism and inflammatory processes.Both n-3 FA increased the placental abundance of FA binding proteins(FABPs)3 and 7.The abundance of CNR1 cannabinoid-receptor-interacting-protein-1(CNRIP1)was reduced in FO vs.FLX.In silico modeling affirmed that bovine FABPs bind to endocannabinoids.The FLX increased the abundance of inflammatory CD44-antigen and secreted-phosphoprotein-1,whereas prostaglandin-endoperoxide synthase 2 was decreased in FO vs.CTL placenta.Maternal FO enriched neonatal plasma with n-3 FAs,and both FLX and FO reduced interleukin-6 concentrations compared with CTL.Conclusion Maternal n-3 FA from FLX and FO differentially affected the bovine placenta;both enhanced lipid metabolism and modulated oxidative stress,however,FO increased some transcriptional ECS components,possibly related to the increased FABPs.Maternal FO induced a unique balance of pro-and anti-inflammatory components in the placenta.Taken together,different sources of n-3 FA during late pregnancy enhanced placental immune and metabolic processes,which may affect the neonatal immune system.

Effect of Extending Voluntary Waiting Period on Health of Holstein Friesian Dairy Cows

The aim of this of this experimental study to investigate the effect of 3 voluntary waiting periods (VWP) on health of dairy cows. A total of 100 lactations of 100 Holstein Friesian cows with high milk production (average 9.442 ± 620 kg) were randomly (based on the ear tags digits of cows) distributed to 1 to 3 experimental groups with VWP of 50 (VWPG50;n = 32), 100 (VWPG100;n = 34) or 150 days (VWPG150;n = 34). Observations relating to diseases of the cows were made by the farm veterinarian and research personnel. Were defined eight disease: puerperal fever, retained fetal membranes, displaced abomasum, metritis, clinical ketosis, clinical mastitis, ovarian cysts, and claw diseases. The experimental days (ED) were from 4 days in milk (DIM) to 100 days after the next calving. The experiment was managed at Dairy Research Farm ?imnic-Craiova January 2018 to December 2022. A clear set of clinical signs were used to define a case at the diseases without the need for laboratory confirmation. Values of Lactational incidence risk (LIR) median postpartum days at diagnosis and pairwise comparison of proportion between VWPs were calculated. LIR for puerperal fiver, retained fetal membranes, displaced abomasum, clinical ketosis, clinical mastitis, ovarian cysts and claw disease were 4;5;2;8;8;and 4% respectively. Numerically, cows with VWP of 100 and150 days had more disease cases compared with VWP of 50 days. The differences between experimental groups of cows regarding proportions of cows with disease cases were not statistically significant. Any of the eight disorders reported in this study was analyzed independently to other health problems. For all enrolled cows (n = 100) VWP was extended until 100 or 150 days postpartum with no effect on the lactational incidence risk for the eight disorders analyzed.

Feeding Calcium Salts of Linseed Oil on Metabolic Status and Reproductive Traits in Grazing Dairy Cows

This experiment aimed to evaluate the effect of calcium salts of linseed oil (rich in alpha-linolenic acid (ALA, C18:3n-3) on metabolic and reproductive traits in high-producing dairy cows under grazing. Thirty-six Holstein dairy cows were randomly assigned, in a complete block design, to receive ALA supplementation (0.85 kg∙day−1 of calcium salts of linseed oil) or to remain as untreated control (CON). The concentrate was formulated to offer the same amount of energy across treatments (CON cows received an extra kg of corn to compensate for the higher energy density of ALA treatment). A PMR + Alfalfa pasture was offered to all cows at the same time. A fixed time artificial insemination (FTAI) at 80 DIM, preceded by a Presynch plus Ovsynch protocol was implemented for the first service and later, on return to estrus, heat detection and artificial insemination (AI) were performed. Pregnancy diagnosis was checked at 30, 42, 60, and 90 d after AI. Blood and milk samples were taken biweekly. Treatment affected plasma cholesterol concentration (160.36 vs. 186.70 mg∙dl−1, p = 0.03, for ALA and CON, respectively) and on size of corpus luteum (CL, 17.6 vs. 13.7 mm, p = 0.02, for ALA and CON, respectively). Supplementation tended (p = 0.136) to increase conception rate by 200 DIM (81.69% vs. 55.43% in ALA and CON cows, respectively). However, treatment had no effect (p > 0.05) on body weight (BW), body condition score (BCS), and circulating levels of beta-hydroxybutyrate (BHBA), glucose, insulin, growth hormone (GH) and insulin-like growth factor (IGF-I). Our results suggest that supplementation with calcium salts of linseed oil could enhance ovarian function without affecting energy metabolism in early lactation dairy cows.

Effects of Oral Administration of Propylene Glycol and Ca-P-Mg Mixture on Milk Quality and SCC of Dairy Cows after Birth

In order to further study the postpartum care technology of dairy cows,the effects of oral administration of propylene glycol or Ca-P-Mg mixture and compound oral rehydration salts on milk quality and SCC of dairy cows after birth were investigated. The results showed that the milk fat and dry matter contents in the dairy cows administered with Ca-P-Mg mixture and compound oral rehydration salts were higher than those in the dairy cows administered with propylene glycol（P 〈0.05）. However, there were no significant differences in milk sugar and protein contents and SCC between the two administration groups.

Association of residual feed intake with abundance of ruminal bacteria and biopolymer hydrolyzing enzyme activities during the peripartal period and early lactation in Holstein dairy cows

Background: Residual feed intake(RFI) in dairy cattle typically calculated at peak lactation is a measure of feed efficiency independent of milk production level. The objective of this study was to evaluate differences in ruminal bacteria, biopolymer hydrolyzing enzyme activities, and overall performance between the most-and the leastefficient dairy cows during the peripartal period. Twenty multiparous Holstein dairy cows with daily ad libitum access to a total mixed ration from d-10 to d 60 relative to the calving date were used. Cows were classified into most-efficient(i.e. with low RFI, n = 10) and least-efficient(i.e. with high RFI, n = 10) based on a linear regression model involving dry matter intake(DMI), fat-corrected milk(FCM), changes in body weight(BW), and metabolic BW.Results: The most-efficient cows had ~ 2.6 kg/d lower DMI at wk 4, 6, 7, and 8 compared with the least-efficient cows. In addition, the most-efficient cows had greater relative abundance of total ruminal bacterial community during the peripartal period. Compared with the least-efficient cows, the most-efficient cows had 4-fold greater relative abundance of Succinivibrio dextrinosolvens at d-10 and d 10 around parturition and tended to have greater abundance of Fibrobacter succinogenes and Megaspheara elsdenii. In contrast, the relative abundance of Butyrivibrio proteoclasticus and Streptococcus bovis was lower and Succinimonas amylolytica and Prevotella bryantii tended to be lower in the most-efficient cows around calving. During the peripartal period, the most-efficient cows had lower enzymatic activities of cellulase, amylase, and protease compared with the least-efficient cows.Conclusions: The results suggest that shifts in ruminal bacteria and digestive enzyme activities during the peripartal period could, at least in part, be part of the mechanism associated with better feed efficiency in dairy cows.

Adipose tissue lipolysis and remodeling during the transition period of dairy cows

Elevated concentrations of plasma fatty acids in transition dairy cows are significantly associated with increased disease susceptibility and poor lactation performance. The main source of plasma fatty acids throughout the transition period is lipolysis from adipose tissue depots. During this time, plasma fatty acids serve as a source of calories mitigating the negative energy balance prompted by copious milk synthesis and limited dry matter intake.Past research has demonstrated that lipolysis in the adipose organ is a complex process that includes not only the activation of lipolytic pathways in response to neural, hormonal, or paracrine stimuli, but also important changes in the structure and cellular distribution of the tissue in a process known as adipose tissue remodeling. This process involves an inflammatory response with immune cell migration, proliferation of the cellular components of the stromal vascular fraction, and changes in the extracellular matrix. This review summarizes current knowledge on lipolysis in dairy cattle, expands on the new field of adipose tissue remodeling, and discusses how these biological processes affect transition cow health and productivity.

Influence on Cellular Signal Transduction Pathway in Dairy Cow Mammary Gland Epithelial Cells by Galactopoietic Compound Isolated from Vaccariae segetalis

The galactopoietic mechanism of Vaccaria segetalis is still unknown. Understanding dibutyl phthalate （DBP） separated from Vaccaria segetalis on the expression of lactation signal transduction genes of mammary gland epithelial cells, including prlr, erα, akt1, socs2, pparγ and elf5, will be helpful to reveal the molecular mechanism. Western blot and qRT- PCR were used to study the change of prlr, erα, akt, socs2, pparγ, and elf5 expression at mRNA and protein level. Co- localization expression of prolactin receptor （PRLR） and estrogen receptor α （ERα） was observed by immunofluorescence; the expression changes of miRNAs （21, 125b, 143, and 195） and the secretion of β-casein and lactose were detected by qRT-PCR and RP-HPLC. The results showed that Vaccaria segetalis active compound had similar fuctions as estrogen and/or prolactin （PRL） in dairy cow mammary gland epithelial cells （DCMECs）, increased the expressions of prlr, erα, akt1, and elf5 genes, while repressed pparγ expressions. DBP promoted socs2 mRNA expression, but its protein expressions were repressed. Furthermore, both DBP and PRL could repress the expressions of miRNA-125b, miRNA-143 and miRNA- 195 in DCMECs. DBP could repress the expression of miRNA-21, while the influence of PRL on miRNA-21 was not certain. DBP could promote the lactation ability of DCMECs by regulating the ER and PRLR cellular signal transduction pathway.

Prepartum body conditions affect insulin signaling pathways in postpartum adipose tissues in transition dairy cows

Background: Overconditioned dairy cows are susceptible to excessive lipolysis and increased insulin resistance during the transition period.The associations among body fat reserve,insulin resistance,and lipolysis in adipose tissues(AT) remain to be elucidated.Therefore,this study aimed to investigate whether excessive fat reserves influence the insulin signaling pathway in AT postpartum.Results: Twenty multiparous dairy cows were selected and assigned to one of two groups,according to prepartum body condition score(BCS): Control group(BCS = 3.0–3.5;n = 10) and Overconditioned group(BCS ≥ 4.0;n = 10).Blood samples were collected on days-14,-7,-4,-2,-1,0,1,2,4,7,and 14 relative to parturition.Subcutaneous AT were collected on day 2 following parturition for quantitative real-time polymerase chain reaction and western blot analyses.No differences were observed between the two groups in serum glucose,non-esterified fatty acids,β-hydroxybutyric acid,tumor necrosis factor(TNF)α,insulin,or leptin concentrations during the experimental period.Compared with the control cows,the overconditioned cows had lower serum triglyceride levels and higher adiponectin concentrations.In the AT postpartum,insulin receptor mRNA and protein levels were lower in the overconditioned cows than in the control cows,and no differences were found in glucose transporter 4 mRNA.Compared with the control cows,the overconditioned cows had lower mRNA levels of TNFα and higher mRNA levels of peroxisome proliferator-activated receptor gamma(PPARγ) in AT postpartum.The phosphorylated protein kinase B(AKT) content and phosphorylation rate of AKT were increased in the overconditioned cows compared with the control cows,which suggested that the downstream insulin signaling in AT was affected.Conclusions: In the present study,transition dairy cows with higher BCS did not show more fat mobilization.The changes of insulin signaling pathway in AT postpartum of overconditioned cows may be partly related to the expression of PPARγ and TNFα,and the secretion of adiponectin.

Milk production and composition and metabolic alterations in the mammary gland of heat-stressed lactating dairy cows

This experiment was conducted to investigate the effects of heat stress(HS) on the feed intake, milk production and composition and metabolic alterations in the mammary gland of dairy cows. Twenty Holstein cows were randomly assigned to one of two treatments according to a completely randomized design. Half of the cows were allocated to the HS group in August(summer season), and the other half were assigned to the HS-free group in November(autumn season). HS reduced(P<0.01) dry matter intake(DMI), milk yield, milk protein and milk urea nitrogen(MUN) of cows compared with HSfree control, but increased(P<0.01) milk somatic cell counts(SCC). We determined the HS-induced metabolic alterations and the relevant mechanisms in dairy cows using liquid chromatography mass spectrometry combined with multivariate analyses. Thirty-four metabolites were identified as potential biomarkers for the diagnosis of HS in dairy cows. Ten of these metabolites, glucose, lactate, pyruvate, lactose, β-hydroxybutyrate, citric acid, α-ketoglutarate, urea, creatine, and orotic acid, had high sensitivity and specificity for HS diagnoses, and seven metabolites were also identified as potential biomarkers of HS in plasma, milk, and liver. These substances are involved in glycolysis, lactose, ketone, tricarboxylic acid(TCA), amino acid and nucleotide metabolism, indicating that HS mainly affects lactose, energy and nucleotide metabolism in the mammary gland of lactating dairy cows. This study suggested that HS might affect milk production and composition by affecting the feed intake and substance metabolisms in the mammary gland tissue of lactating dairy cows.

Metabolic Regulation of Mammary Gland Epithelial Cells of Dairy Cow by Galactopoietic Compound Isolated from Vaccariae segetalis

In previous experiment, we isolated a compound dibutyl phthalate （DBP） from Vaccaria segetalis which had galactopoietic function on mammary gland epithelial cells of dairy cow （DCMECs）. In this experiment, we ascertained the metabolic regulation function of DBP on DCMECs. Many genes related to lactation including Stat5, AMPK, b-casein, Glut1, SREBP-1, PEPCK, and ACC were detected by real-time PCR. Furthermore, Stat5 and AMPK were detected by Western blot and immunofluorescence co-localization, respectively. The results showed that DBP stimulates the expression of Stat5 and p-Stat5, thus activates Stat5 cell signal transduction pathway and stimulates b-casein synthesis. DBP also raises the activities of Glut1 and AMPK to stimulate glucose uptake and glycometabolism and activates the expression of AMPK downstream target genes PEPCK and ACC and expression of SREBP-1 to stimulate milk fat synthesis. In addition, the activities of HK, G-6-PDH, ICDH, ATPase, and energy charges were stimulated by DBP to increase the energy metabolism level of DCMECs. The results showed DBP stimulates energy metabolism related to galactopoietic function in DCMECs.

Changes in feed intake, nutrient digestion, plasma metabolites, and oxidative stress parameters in dairy cows with subacute ruminal acidosis and its regulation with pelleted beet pulp

The objectives of this study were to 1） determine the variation of nutrient digestion, plasma metabolites and oxidative stress parameters triggered by induced subacute ruminal acidosis （SARA）; and 2） evaluate the ability of pelleted beet pulp （BP） as a replacement for ground corn to alleviate SARA. Eight Holstein-Friesian cows were fed four diets during four successive17 day periods： 1） total mixed ration （TMR） containing 0% finely ground wheat （FGW） （WO）; 2） TMR containing 10% FGW （W10）; 3） TMR containing 20% FGW （W20）; and 4） TMR containing 10% BP as a replacement for 10% ground corn （BP10）. The SARA induction protocol reduced the mean ruminal pH from 6.37 to 5.94, and the minimum ruminal pH decreased from 5.99 to 5.41 from baseline to challenge period. Mean ruminal pH increased from 5.94 to 6.05, and minimum daily ruminal pH increased from 5.41 to 5.63, when BP was substituted for corn. The apparent digestibility of nutrients was not affected by the dietary treatments, except that the digestibility of neutral detergent fibre （NDF） and acid detergent fibre （ADF） was reduced in cows fed the W20 diet compared with cows fed the W0 and W10 diets, and cows fed the BP10 diet had higher NDF and ADF digestibility than the cows fed the W20 diet. Cows fed the W20 diet had a lower plasma concentration of 13-hydroxybutyrate （BHBA）, non-esterified fatty acids （NEFA）, cholesterol, triglyceride, and total antioxidative capacity （TAC）, and a higher plasma concentration of glucose, insulin, malonaldehyde （MDA）, super oxygen dehydrogenises （SOD）, and glutathione peroxidase （GSH-Px） than cows fed the W0 diet. Substitution of BP for corn increased concentrations of plasma BHBA and TAC, but decreased concentrations of plasma MDA. Our results indicate that reduction of fibre digestion; the concomitant increase of plasma glucose and insulin; the decrease of plasma BHBA, NEFA, cholesterol, and triglyceride; and changes of plasma oxidative stress parameters are highly related to SARA induced by W20 diets. These variables may be alternative candidates for SARA diagnosis. We also suggest that the substitution of BP for corn could reduce the risk of SARA, increase fibre digestion, and improve the antioxidant status in dairy cows.

Advances in fatty acids nutrition in dairy cows: from gut to cells and effects on performance

High producing dairy cows generally receive in the diet up to 5–6% of fat. This is a relatively low amount of fat in the diet compared to diets in monogastrics;however, dietary fat is important for dairy cows as demonstrated by the benefits of supplementing cows with various fatty acids(FA). Several FA are highly bioactive, especially by affecting the transcriptome;thus, they have nutrigenomic effects. In the present review, we provide an up-to-date understanding of the utilization of FA by dairy cows including the main processes affecting FA in the rumen,molecular aspects of the absorption of FA by the gut, synthesis, secretion, and utilization of chylomicrons;uptake and metabolism of FA by peripheral tissues, with a main emphasis on the liver, and main transcription factors regulated by FA. Most of the advances in FA utilization by rumen microorganisms and intestinal absorption of FA in dairy cows were made before the end of the last century with little information generated afterwards. However,large advances on the molecular aspects of intestinal absorption and cellular uptake of FA were made on monogastric species in the last 20 years. We provide a model of FA utilization in dairy cows by using information generated in monogastrics and enriching it with data produced in dairy cows. We also reviewed the latest studies on the effects of dietary FA on milk yield, milk fatty acid composition, reproduction, and health in dairy cows. The reviewed data revealed a complex picture with the FA being active in each step of the way, starting from influencing rumen microbiota, regulating intestinal absorption, and affecting cellular uptake and utilization by peripheral tissues, making prediction on in vivo nutrigenomic effects of FA challenging.

Month-Wise Prevalence of Subclinical Mastitis in Dairy Cows in Guangdong Province,China

To evaluate the month-wise prevalence of subclinical mastitis （SM） and its relations with climatic temperature and humidity, and to provide references for control and prevention of mastitis in dairy cows in Guangdong, China, California mastitis test method was used to determine the monthly prevalence of SM （MPSM, %） of lactating Holstein cows from 11 dairy farms in Guangdong Province, China, including about 516 cows per monthly examination. The average MPSM on a cow and a quarter basis were 30 and 13% respectively, and there was a positive linear correlation between monthly mean air temperature （MT, °C） and MPSM on a cow basis （r=0.763, P=0.004） or a quarter basis （r=0.577, P=0.049）, but there were no correlations between MT and MPSM on a cow or a quarter basis （P〉0.05）. There was a shift in MPSM trend that the highest MPSM （38.4%） was not for the hottest July with MT at 30.8°C, but for the later September at 27.9°C. The farms need to develop new housing conditions to control cowshed temperatures in the hot season to reduce the mastitis prevalences in the post-hot months.

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(FDRadjusted 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.

Effects of different sources of carbohydrates on intake, digestibility, chewing, and performance of Holstein dairy cows

To investigate the effects of different sources of carbohydrates on intake, digestibility, chewing, and performance, nine lactating Holstein dairy cows （day in milk= 100±21 d; body weight=645.7 ± 26.5 kg） were allotted to a 3 × 3 Latin square design at three 23-d periods. The three treatments included 34.91% （B）, 18.87% （BC）, and 18.86% （BB） barley that in treatment B was partially replaced with only corn or corn plus beet pulp in treatments BC and BB, respectively. The concentration of starch and neutral detergent soluble carbohydrate varied （22.2, 20.2, and 14.5; 13.6, 15.9, and 20.1% of DM in treatments B, BC, and BB, respectively）. Cows in treatment BB showed a higher DMI and improved digestibility of DM, NDF, and EE compared with treatments B or BC Ruminal pH was higher in cows fed on BB （6.83） compared with those that received B or BC treatments （6.62 and 6.73, respectively）. A lower proportion of propionate accompanied the higher pH in the BB group; however, a greater proportion of acetate and acetate： propionate ratio was observed compared with cows fed either on the B or BC diet. Moreover, cows fed on the BB diet showed the lowest ruminal passage rate and longest ruminal and total retention time. Eating time did not differ among treatments, rumination time was greater among cows fed on the BB diet compared with the others, whereas total chewing activity was greater than those fed on BC, but similar to those fed on B. The treatments showed no effect on milk yield. Partially replacing barley with corn or beet pulp resulted in an increase in milk fat and a lower protein concentration. Changing dietary NFC with that of a different degradability thus altered intake, chewing activity, ruminal environment, retention time or passage rate, and lactation performance. The results of this study showed that beet pulp with a higher NDF and a detergent-soluble carbohydrate or pectin established a more consistent ruminal mat than barley and corn, thus resulting in higher mean retention time and chewing activity, whereas no changes in 3.5% ECM and milk fat were observed.

Accuracy comparison of dry matter intake prediction models evaluated by a feeding trial of lactating dairy cows fed two total mixed rations with different forage source

Dry matter intake （DMI） prediction models of NRC （2001）, Fox et aL （2004） and Fuentes-Pila et aL （2003） were targeted in the present study, and the objective was to evaluate their prediction accuracy with feeding trial data of 32 lactating Holstein cows fed two total mixed rations with different forage source. Thirty-two cows were randomly assigned to one of two total mixed ration groups： a ration containing a mixed forage （MF） of 3.7% Chinese wildrye, 28.4% alfalfa hay and 26.5% corn silage diet and another ration containing 33.8% corn stover （CS） as unique forage source. The actual DMI was greater in MF group than in CS group （P=0.064）. The NRC model to predict DMI resulted in the lowest root mean square prediction error for both MF and CS groups （1.09 kg d-1 vs. 1.28 kg d-1） and the highest accuracy and precision based on concordance correlation coefficient for both MF and CS diet （0.89 vs. 0.87）. Except the NRC model, the other two models presented mean and linear biases in both MF and CS diets when prediction residuals were plotted against predicted DMI values （P〈0.001）. The DMI variation in MF was caused by week of lactation （55.6%）, milk yield （13.9%）, milk fat percentage （7.1%） and dietary neutral detergent fiber （13.3%）, while the variation in CS was caused by week of lactation （50.9%）, live body weight （28.2%）, milk yield （8.4%）, milk fat percentage （5.2%） and dietary neutral detergent fibre （3.8%）. In a brief, the NRC model to predict DMI is comparatively acceptable for lactating dairy cows fed two total mixed rations with different forage source.

Effects of the dietary nonfiber carbohydrate content on lactation performance, rumen fermentation, and nitrogen utilization in mid-lactation dairy cows receiving corn stover

Background: Corn stover(CS) is an abundant source of feed for livestock in China. However, it is low in nutritional value that we have been seeking technologies to improve. Previous studies show that non-fiber carbohydrate(NFC)might limit the utilization of a CS diet by lactating dairy cows. Thus, this study was conducted to investigate the lactation performance and rumen fermentation characteristics in lactating cows consuming CS with two contents of NFC compared to an alfalfa hay-containing diet. Twelve Holstein cows were used in a replicated 3 × 3 Latin square design with three dietary treatments:(1) low-NFC diet(NFC = 35.6%, L-NFC),(2) high-NFC diet(NFC = 40.1%,H-NFC), and(3) alfalfa hay diet(NFC = 38.9%, AH).Results: Intake of DM was lower for cows fed H-NFC compared to L-NFC and AH, while the milk yield was higher in AH than in H-NFC and L-NFC(P < 0.01). The feed efficiency(milk yield/DM intake, 1.15 vs. 1.08, P < 0.01) were greater for cows fed H-NFC than L-NFC. The contents of milk protein and lactose were not different among the groups(P > 0.11), but milk fat content was higher for cows fed H-NFC and L-NFC compared to AH(P < 0.01). The rumen ammonia nitrogen concentration and the concentrations of urea nitrogen in blood and milk were lower for cows fed H-NFC and AH compared to L-NFC(P < 0.05). The concentrations of rumen propionate and total volatile fatty acids were different among groups(P < 0.05) with higher concentration for cows fed AH compared to H-NFC and L-NFC, and acetate concentration tended to be different among groups(P = 0.06).Conclusions: From the results obtained in this study, it was inferred that the increased NFC content in a diet containing corn stover can improve the feed efficiency and benefit the nitrogen conversion.

Development of a PCR Diagnostic Kit for Cryptosporidium andersoni in Dairy Cow

Cryptosporidiosis is an important zoonosis caused by the Cryptosporidium species. To develop a PCR diagnostic kit for molecular detection and differential diagnosis of Cryptosporidium spp., a portion of ITS-1 sequence of Cryptosporidium. andersoni was chosen as the target DNA for designing the species-specific primers （ZRQF/ZR）. The kit components were determined after the PCR amplification conditions were serially optimized. A series of tests were conducted in the specificity, sensitivity, stability, reproducibility, and stored period of the kit, respectively. The results showed that only C. andersoni were amplified specific band of about 500 bp, while Cryptosporidium. parvum, Cryptosporidium. baileyi, Eimeria sp of dairy cow, Toxoplasma gondii, Eimeria sp of pig, Ascaris suum, Cyclospora sp, and E. coli could not be amplified. 254 oocysts of C. andersoni was the lowest number that could be detected using the kit. The kit worked well after being stored at room temperature, 4 and -20℃ for nine months. Fecal specimens, which were collected from a total of 243 calves on four different dairy farms in Guangdong Province, China, and one dairy farm in Henan Province, China, were examined using the kit; the positive rate of the kit was 2-13% higher than that of the routine methods. The results indicated that the kit can detect fecal samples faster, more sensitively, and conveniently, and can provide a useful tool for the identification and differentiation of C. andersoni from the other Cryptosporidium species; it also has implications for further studies on molecular epidemiology and differential diagnostics of cryptosporidiosis in animals.