Assessment of remdesivir and its nucleoside metabolite in beagle dogs and healthy humans by liquid chromatography coupled with triple quadrupole mass spectrometry.

The aim of this study was to assess the pharmacokinetics of the existing remdesivir intravenous formulation (100 mg dose) against the newly developed oral formulation (20 mg dose) for remdesivir and its active nucleoside metabolite (GS-441524) in beagle dogs followed by healthy human volunteers. A quantification method for remdesivir and its active nucleoside metabolite (GS-441524) in beagle dog and human plasma has been developed and validated using liquid chromatography coupled to triple quadrupole mass spectrometry detection. The analytical methods for beagle dogs and human differ in the calibration curve range, plasma matrix, processing volume, reconstitution volume and injection volume; however all other parameters were same in both methods. A simple protein precipitation extraction was carried out using acetonitrile containing the internal standard remdesivir D5. Remdesivir and GS-441524 were separated on an Endurus C-18P, 100 × 4.6 mm, 3 µm column and detected using a mass spectrometer with electrospray ionization in positive ion mode. The ion transitions used were m/z 603.1 → m/z 200.0 for remdesivir, m/z 292.0 → m/z 202.2 for GS-441524 and m/z 608.2 → m/z 205.1 for remdesivir D5. The calibration curve results were linear in beagle dog plasma (2.0-2,000.8 ng/ml range for remdesivir and 2.0-1,500.4 ng/ml for GS-441524) and human plasma (30.0-4,503.9 ng/ml range for remdesivir and 2.0-200.4 ng/ml for GS-441524). The recovery was >90% in beagle dog and human plasma. These methods were successfully used to determine the pharmacokinetic parameters of the intravenous injection and subcutaneous tablets dosage forms in beagle dogs and healthy humans.

Effect of particle size on gastric emptying of enteric-coated granules in fasted beagle dogs: Relationship with interdigestive migrating motor complex.

This study investigates the particle size threshold at which the interdigestive migrating motor complex (IMMC) becomes active in gastric emptying for fasted beagle dogs. Enteric-coated granules containing cetirizine dihydrochloride (CET) were prepared in three particle sizes, 200, 660, and 1,200 µm (D50). To mark IMMC timing and water movement from the stomach, enteric-coated aspirin tablets and acetaminophen solution were used. To six fasted beagle dogs with 50 mL of acetaminophen solution was administered each granule size as a multiple-unit and a single enteric-coated aspirin tablet (3-period crossover study). No significant difference in pharmacokinetic parameters of CET after oral administration of different particle sizes was observed. However, the appearance time of CET in plasma with smaller granules (200 and 660 µm) was significantly faster than that of salicylic acid (a major metabolite of aspirin) in all dogs. In the case of the largest granules (1,200 µm), no significant time difference was observed in the appearance of both compounds in plasma. Furthermore, in two dogs, both compounds appeared at the same time, implying IMMC-regulated gastric emptying for the largest CET granules. These results support a particle size threshold between 660 and 1,200 µm for gastric emptying without IMMC action in fasted beagle dogs.

Inflammatory Reprogramming Mediates Changes in Three-Dimensional Strain Capacity and Cardiac Function in Beagle Dogs with Doxorubicin-Related Cardiomyopathy.

Background: The cardiotoxicity of doxorubicin (DOX) limits its use in cancer treatment. To address this limitation, we developed a novel animal model that uses beagle dogs to investigate DOX-induced cardiac disorders. Unfortunately, the lack of effective cardioprotection strategies against DOX-induced cardiotoxicity poses a significant challenge. To establish a canine model for low-mortality DOX-induced cardiac dysfunction and explore the relationship between inflammatory reprogramming and DOX-related cardiotoxicity. Methods: Twenty male beagle dogs aged two years were randomly assigned into the DOX (N = 10) and control (CON) (N = 10) groups. DOX was infused (1.5 mg/kg) every two weeks until doses cumulatively reached 12 mg/kg. Serum biomarkers and myocardial pathology were evaluated, while real-time fluorescence-based quantitative polymerase chain reaction (RTFQ-PCR), two- and three-dimensional echocardiography (2DE and RT3DE), functional enrichment, and matrix correlation were also performed. Results: In the DOX group, high-sensitive cardiac troponin T (hs cTnT) and N-terminal pro-brain natriuretic peptide (NT-proBNP) were significantly increased. Myocardial pathology indicated early to medium myocardial degeneration via a decreased cardiomyocyte cross-sectional area (CSA). Increased levels of inflammatory gene transcripts (interleukin 6 (IL6), tumor necrosis factor (TNF), transforming growth factor ß (TGF ß ), intercellular adhesion molecule 1 (ICAM1), interleukin 1 (IL1), interleukin 1 ß (IL1 ß ), and interleukin 8 (IL8)), of collagen metabolism and deposition regulatory genes (matrix metalloproteinase (MMP) family and tissue inhibitor of matrix metalloproteinase (TIMP) family), and the natriuretic peptide family (NPS) (natriuretic peptide A, B and C (NPPA, NPPB, and NPPC)) were observed. Strain abnormalities in the right ventricular longitudinal septal strain (RVLSS), right ventricular longitudinal free-wall strain (RVLFS), left ventricular global longitudinal strain (LVGLS), and left ventricular global circumferential strain (LVGCS) were detected at week 28 (vs. week 0 or CON group, p < 0.05, respectively). A significant decline in RVLSS and RVLFS occurred at week 16, which was earlier than in the corresponding left ventricular areas. A significant right ventricular ejection fraction (RVEF) decline was noted at week 16 (vs. week 0, 33.92 ± 3.59% vs. 38.58 ± 3.58%, p < 0.05), which was 12 weeks earlier than for the left ventricular ejection fraction (LVEF), which occurred at week 28 (vs. week 0, 49.02 ± 2.07% vs. 54.26 ± 4.38%, p < 0.01). The right ventricular strain and functional damages correlated stronger with inflammatory reprogramming (most R from 0.60 to 0.90) than the left ones (most R from 0.30 to 0.65), thereby indicating a more pronounced correlation. Conclusions: Inflammatory reprogramming mediated disorders of strain capacity and cardiac function predominantly in the right side of the heart in the newly established DOX-related cardiomyopathy beagle dog model.

Computer-aided design and 3D printing for a stable construction of segmental bone defect model in Beagles: a short term observation.

OBJECTIVE: Segmental bone defect animal studies require stable fixation which is a continuous experimental challenge. Large animal models are comparable to the human bone, but with obvious drawbacks of housing and costs. Our study aims to utilize CAD and 3D printing in the construction of a stable and reproducible segmental bone defect animal mode. METHODS: CAD-aided 3D printed surgical instruments were incorporated into the construction of the animal model through preoperative surgical emulation. 20 3D printed femurs were divided into either experimental group using 3D surgical instruments or control group. In Vitro surgical time and accuracy of fixation were analysed and compared between the two groups. A mature surgical plan using the surgical instruments was then utilized in the construction of 3 segmental bone defect Beagle models in vivo. The Beagles were postoperatively assessed through limb function and imaging at 1, 2 and 3 months postoperatively. RESULTS: In vitro experiments showed a significant reduction in surgical time from 40.6 ± 14.1 (23-68 min) to 26 ± 4.6 (19-36 min) (n = 10, p < 0.05) and the accuracy of intramedullary fixation placement increased from 71.6 ± 23.6 (33.3-100) % to 98.3 ± 5.37 (83-100) %, (n = 30, p < 0.05) with the use of CAD and 3D printed instruments. All Beagles were load-bearing within 1 week, and postoperative radiographs showed no evidence of implant failure. CONCLUSION: Incorporation of CAD and 3D printing significantly increases stability, while reducing the surgical time in the construction of the animal model, significantly affecting the success of the segmental bone defect model in Beagles.

Dog leukocyte antigen genotyping across class I and class II genes in beagle dogs as laboratory animals.

Dog leukocyte antigen (DLA) polymorphisms have been found to be associated with inter-individual variations in the risk, susceptibility, and severity of immune-related phenomena. While DLA class II genes have been extensively studied, less research has been performed on the polymorphisms of DLA class I genes, especially in beagle dogs commonly used as laboratory animals for safety evaluations in drug development. We genotyped four DLA class I genes and four DLA class II genes by locus-specific Sanger sequencing using 93 laboratory beagle dogs derived from two different strains: TOYO and Marshall. The results showed that, for DLA class I genes, 11, 4, 1, and 2 alleles, including a novel allele, were detected in DLA-88, DLA-12/88L, DLA-64, and DLA-79, while, for DLA class II genes, 1, 10, 6, and 7 alleles were detected in DLA-DRA, DLA-DRB1, DLA-DQA1, and DLA-DQB1, respectively. It was estimated that there were 14 DLA haplotypes, six of which had a frequency of ≥ 5%. Furthermore, when comparing the DLA diversity between TOYO and Marshall strains, the most common alleles and haplotypes differed between them. This is the first study to genotype all DLA loci and determine DLA haplotypes including all DLA class I and class II genes in dogs. Integrating information on the DLA diversity of laboratory beagle dogs should reinforce their benefit as an animal model for understanding various diseases associated with a specific DLA type.

[Determination of nine components of fried Ziziphi Spinosae Semen extract in beagle dog plasma by UPLC-MS/MS and its application in pharmacokinetic research].

This study established a convenient, rapid, and sensitive ultra-performance liquid chromatography tandem mass spectrometry(UPLC-MS/MS) method for simultaneous determination of magnoflorine,(R)-coclaurine, vicenin Ⅱ, isospinosin, spinosin, swertisin, N-nornuciferine, 6-feruloylspinosin, and jujuboside B in beagle dog plasma after oral administration of fried Ziziphi Spinosae Semen(FZSS) extract. The Waters HSS-T3 C_(18) column(2.1 mm×100 mm, 1.8 µm) was used. The methanol-aqueous solution(containing 0.01% formic acid) was adopted as the mobile phase for gradient elution. The nine components and two internal standards were completely separated within 8 min. The mass spectrometry detection was performed in multiple reaction monitoring(MRM) mode by positive and negative ion switching of electrospray ionization. The analytical method was validated in terms of specificity, selectivity, linear range, accuracy, precision, recovery, matrix effect, and stability. It could meet the requirement of pharmacokinetic research after oral administration of FZSS extract to beagle dogs. The results showed that the time to reach the peak concentration(T_(max)) of magnoflorine,(R)-coclaurine, vicenin Ⅱ, isospinosin, spinosin, 6-feruloylspinosin, and jujuboside B was 2.40-3.20 h, and the elimination halflife(t_(1/2)) was 2.08-6.79 h after a single-dose oral administration of FZSS to beagle dogs. The exposure of magnoflorine and spinosin was high, with a peak concentration(C_(max)) of 76.7 and 31.5 ng·mL~(-1) and an area under the curve(AUC_(0-∞)) of 581 and 315 ng·h·mL~(-1), respectively. The exposure of the remaining five compounds was lower, with a C_(max) of 0.81-13.0 ng·mL~(-1) and an AUC_(0-∞) of 6.00-106 ng·h·mL~(-1). This study provides a reference for the follow-up research of FZSS.

A novel magnetic compression technique for establishment of a vesicovaginal fistula model in Beagle dogs.

Vesicovaginal fistula lacks a standard, established animal model, making surgical innovations for this condition challenging. Herein, we aimed to non-surgically establish vesicovaginal fistula using the magnetic compression technique, and the feasibility of this method was explored using eight female Beagle dogs as model animals. In these dogs, cylindrical daughter and parent magnets were implanted into the bladder and vagina, respectively, after anesthesia, and the positions of these magnets were adjusted under X-ray supervision to make them attract each other, thus forming the structure of daughter magnet-bladder wall-vaginal wall-parent magnet. Operation time and collateral damage were recorded. The experimental animals were euthanized 2 weeks postoperatively, and the vesicovaginal fistula gross specimens were obtained. The size of the fistula was measured. Vesicovaginal fistula was observed by naked eye and under a light microscope. Magnet placement was successful in all dogs, and remained in the established position for the reminder of the experiment. The average operation time was 14.38 min ± 1.66 min (range, 12-17 min). The dogs were generally in good condition postoperatively and were voiding normally, with no complications like bleeding and urine retention. The magnets were removed from the vagina after euthanasia. The vesicovaginal fistula was successfully established according to gross observation, and the fistula diameters were 4.50-6.24 mm. Histological observation revealed that the bladder mucosa and vaginal mucosa were in close contact on the internal surface of the fistula. Taken together, magnetic compression technique is a simple and feasible method to establish an animal model of vesicovaginal fistula using Beagle dogs. This model can help clinicians study new surgical techniques and practice innovative approaches for treating vesicovaginal fistula.

Transcript abundance of hepatic drug-metabolizing enzymes in two dog breeds compared with 14 species including humans.

Drug-metabolizing enzymes are important in drug development and therapy, but have not been fully identified and characterized in many species, lines, and breeds. Liver transcriptomic data were analyzed for phase I cytochromes P450, flavin-containing monooxygenases, and carboxylesterases and phase II UDP-glucuronosyltransferases, sulfotransferases, and glutathione S-transferases. Comparisons with a variety of species (humans, rhesus macaques, African green monkeys, baboons, common marmosets, cattle, sheep, pigs, cats, dogs, rabbits, tree shrews, rats, mice, and chickens) revealed both general similarities and differences in the transcript abundances of drug-metabolizing enzymes. Similarly, Beagle and Shiba dogs were examined by next-generation sequencing (RNA-seq). Consequently, no substantial differences in transcript abundance were noted in different breeds of pigs and dogs and in different lines of mice and rats. Therefore, the expression profiles of hepatic drug-metabolizing enzyme transcripts appear to be similar in Shiba and Beagle dogs and pig breeds and the rat and mouse lines analyzed, although some differences were found in other species.

Biocompatibility of dental implants coated with hydroxyapatite using pulsed Er:YAG laser deposition.

We aimed to improve the biocompatibility and osteoinductive potential of Ti implants using a simulated intraoral hydroxyapatite (HAp) coating. We devised a novel surface treatment for aggressive induction of osteoblast adhesion and bone regeneration on the implant surface. A thin α-tricalcium phosphate (α-TCP) film was deposited on the implant surface using a pulsed Er:YAG laser. The coating was converted to HAp through artificial saliva immersion, which was confirmed using scanning electron microscopy (SEM) and X-ray diffraction (XRD). SEM showed needle-like HAp crystals on the Ti disks and sandblasted implant surfaces after immersion in artificial saliva for 96 h. Microcomputed tomography and histological evaluation 4 and 8 weeks after implantation into beagle dog mandibles showed that the HAp-coated implant was biocompatible and exhibited superior osteoinduction compared to that of sandblasted implants. Coating the implant surface with HAp using an Er:YAG laser has potential as a new method of the implant-surface debridement.

Establishment of a spontaneous ventilation anesthesia model for beagle dogs.

While spontaneous ventilation (SV) anesthesia is in use for clinical patients, there is still little systematic experimental research into its basic aspects. The rabbit SV model that we established previously has some limitations including the model being too small, differences in anesthetic drugs and anesthesia procedures, so we set out to establish an SV anesthesia model for beagle dogs.•Single lumen tracheal intubation was performed on beagles connecting a ventilator, and the anesthetic dosage was adjusted for spontaneous ventilation before surgery.•5 mL of 1 % lidocaine was applied as a local infiltration anesthesia at the surgical incision.•After thoracotomy, 5 mL of 1% lidocaine was sprayed onto the surface of the lungs and a T3-T7 intercostal nerve block (1:1 2 % lidocaine:0.75 % ropivacaine) was performed.

Investigation of circulating infectious agents in experimental Beagle dogs of a production colony and three research facilities in China from June 2021 to May 2022.

To understand the epizootiologic characteristics of pathogens and opportunistic infections in one Beagle dog production colony and three research facilities, viruses and mycoplasma were detected in 1777 samples collected from Beagle dogs in China by polymerase chain reaction/reverse transcription polymerase chain reaction, and bacteria were isolated and identified by 16S rRNA sequence analysis. In addition, genotyping of the major circulating viruses was carried out by amplification of gene fragments and homology analysis. Canine coronavirus (CCoV), Escherichia coli, canine parvovirus (CPV), Bordetella bronchiseptica, Clostridium perfringens, Mycoplasma cynos, Klebsiella pneumoniae, Streptococcus canis, canine astrovirus (CaAstV), canine kobuvirus (CaKV), Pseudomonas aeruginosa, Proteus mirabilis, Macrococcus canis, Pasteurella canis, canine bocavirus (CBoV) and canine adenovirus (CAdV) were detected in the samples. Single, double, triple and quadruple infections accounted for 6.6%, 1.4%, 1.2% and 0.96% of samples, respectively. CCoV strains in 81 samples included three genotypes, CCoV-I, CCoV-IIa and CCoV-IIb, by analysis of S gene. The rate of single infection of CCoV-I, CCoV-IIa or CCoV-IIb was 19%, 38% or 7.4% respectively. The double and triple infection rates of CCoV were 32.8% and 2.5% respectively. All CPV strains in 36 samples belonged to CPV-2c. There were three amino acid differences in the Fiber protein of CAdV-positive sample QD2022, compared with the reference strain Toronto A26/61 and the vaccine strain YCA-18. These results suggest that CCoV and CPV are primary infectious agents, and that these two viruses were often identified in mixed infections, or coinfections alongside mycoplasma or other bacteria. These results will provide the basis for improvements in prevention and control of naturally occurring infectious diseases in Beagle dog production colonies and research facilities.

Effects of combined application of fibroblast growth factor (FGF)-2 and carbonate apatite for tissue regeneration in a beagle dog model of one-wall periodontal defect.

Introduction: There has been an increasing desire for the development of predictive periodontal regenerative therapy for severe periodontitis. In this study, we investigated the effect of the combined use of fibroblast growth factor-2 (FGF-2), a drug for periodontal regeneration approved in Japan, and carbonated apatite (CO3Ap), bioresorbable and osteoconductive scaffold, on periodontal regeneration in beagle dog model of one-wall periodontal defect (severe intraosseous defect) for 24 weeks in comparison with CO3Ap or vehicle alone. Methods: One-wall periodontal defects were created (mesiodistal width × depth: 4 × 4 mm) on the mesial portion of the mandibular first molar (M1) of beagle dogs on both side. Mixture of FGF-2 and CO3Ap, vehicle and CO3Ap, or vehicle alone were administered to the defects and designated as groups FGF-2+CO3Ap, CO3Ap, and control, respectively. To assess the periodontal regeneration, radiographic analysis over time for 24 weeks, and micro computed tomography (µCT) and histological evaluation at 6 and 24 weeks were performed. Results: For the regenerated tissue in the defect site, the mineral content of the FGF-2+CO3Ap group was higher than that of the CO3Ap group in the radiographic analysis at 6-24 weeks. In the context of new bone formation and replacement, the FGF-2+CO3Ap group exhibited significantly greater new bone volume and smaller CO3Ap volume than the CO3Ap group in the µCT analysis at 6 and 24 weeks. Furthermore, the density of the new bone in the FGF-2+CO3Ap group at 24 weeks was similar to those in the control and CO3Ap groups. Histological evaluation revealed that the length of the new periodontal ligament and cementum in the FGF-2+CO3Ap group was greater than that in the CO3Ap group at 6 weeks. We also examined the effect of the combined use of the FGF-2 and CO3Ap on the existing bone adjacent to the defect and demonstrated that the existing bone height and volume in the FGF-2+CO3Ap group remained significantly greater than those in the CO3Ap group. Conclusion: This study demonstrated that the combination of FGF-2 and CO3Ap was effective not only in enhancing new bone formation and replacing scaffold but also in maintaining the existing bone adjacent to the defect site in a beagle dog model of one-wall periodontal defect. Additionally, new periodontal tissues induced by FGF-2 and CO3Ap may follow a maturation process similar to that formed by spontaneous healing. This suggests that the combined use of FGF-2 and CO3Ap would promote periodontal regeneration in severe bony defects of periodontitis patient.

Comparison of methods for the effective evaluation of the energy content of poultry byproduct meal for beagles.

The objectives of this study were to compare the energy values of poultry byproduct meal (PBM) as feed for adult beagle dogs using the direct, difference, and regression methods to examine dogs' nitrogen metabolism, energy utilization, gaseous metabolism, and body health. Five groups of six 12 mo old female beagles with an average body weight of 9.67 ±â€…0.52 kg were tested in a 5 × 6 incomplete Latin square design, with six repetitions in each group. Five experimental diets were tested consisting of 100% PBM; three substitution diets containing either 15%, 30%, or 45% PBM (termed 15PBM, 30PBM, and 45PBM, respectively); and a basal diet (included 6.90% PBM). Each experimental period lasted for 10 d, comprising 4 d of dietary acclimation followed by 6 d of testing (including 3 d feeding period and 3 d fasting period), during which the heat production (HP) was determined and feces and urine were collected. Results showed that, in the feeding state, the nitrogen intake, urinary nitrogen, apparent nitrogen digestibility, retained nitrogen, andHP increased significantly (P < 0.05) as the PBM level increased. The net protein utilization, biological value of protein, and total apparent digestibility of amino acids did not differ between the 30PBM and 45PBM diets (P > 0.05). The O2 consumption and CO2 production of beagles during the fasting period were not influenced by the PBM level (P > 0.05). The digestible energy and metabolizable energy values of the PBM estimated by the regression method were 20.16 and 18.18 MJ/kg dry matter (DM), respectively, and did not differ from those determined by the direct method (P > 0.05). The fecal DM percentages and fecal PBM scores were significantly higher in the PBM diet than in the difference method groups (P < 0.05). The direct method group had a significantly higher fecal score (4.63) than the other groups (P < 0.05), The fecal score of the 45PBM diet (3.50) was significantly higher than the 30PBM diet (2.90; P < 0.05). In summary, the direct and difference methods of determining the effective energy value of PBM for beagles, produce significantly different results. Under the conditions of this test, the best proportion of PBM in beagle feed for optimum energy provision is 30%.

The scale of the global pet food market is expanding daily, but there are relatively few evaluations that have been performed on many of the common ingredients in pet food recently. This study compares direct, difference, and regression methods of evaluating the effects of the energy values of poultry byproduct meal (PBM), which is commonly used in beagle foods. Beagles were fed five diets containing different proportions of PBM, and the effects on diet quality were compared. The direct evaluation method enabled relatively accurate assessment of the energy value of PBM but could cause diarrhea in the test dogs. Different proportions of PBM significantly affected the results, and the difference method identified feed with 30% poultry meal as optimal. The regression method effectively determined the digestible energy and metabolizable energy values of the various tested levels of PBM. The difference method performed best in evaluating the energy values of beagle feed, and a diet consisting of 30% PBM was determined to be optimal.

Different Diet Energy Levels Alter Body Condition, Glucolipid Metabolism, Fecal Microbiota and Metabolites in Adult Beagle Dogs.

Diet energy is a key component of pet food, but it is usually ignored during pet food development and pet owners also have limited knowledge of its importance. This study aimed to explore the effect of diet energy on the body condition, glucolipid metabolism, fecal microbiota and metabolites of adult beagles and analyze the relation between diet and host and gut microbiota. Eighteen healthy adult neutered male beagles were selected and randomly divided into three groups. Diets were formulated with three metabolizable energy (ME) levels: the low-energy (Le) group consumed a diet of 13.88 MJ/kg ME; the medium-energy (Me) group consumed a diet of 15.04 MJ/kg ME; and the high-energy (He) group consumed a diet of 17.05 MJ/kg ME. Moreover, the protein content of all these three diets was 29%. The experiment lasted 10 weeks, with a two-week acclimation period and an eight-week test phase. Body weight, body condition score (BCS), muscle condition score (MCS) and body fat index (BFI) decreased in the Le group, and the changes in these factors in the Le group were significantly higher than in the other groups (p < 0.05). The serum glucose and lipid levels of the Le and He groups changed over time (p < 0.05), but those of the Me group were stable (p > 0.05). The fecal pH of the Le and He groups decreased at the end of the trial (p < 0.05) and we found that the profiles of short-chain fatty acids (SCFAs) and bile acids (BAs) changed greatly, especially secondary BAs (p < 0.05). As SCFAs and secondary BAs are metabolites of the gut microbiota, the fecal microbiota was also measured. Fecal 16S rRNA gene sequencing found that the Me group had higher α-diversity indices (p < 0.05). The Me group had notably higher levels of gut probiotics, such as Faecalibacterium prausnitzii, Bacteroides plebeius and Blautia producta (p < 0.05). The diet-host-fecal microbiota interactions were determined by network analysis, and fecal metabolites may help to determine the best physical condition of dogs, assisting pet food development. Overall, feeding dogs low- or high-energy diets was harmful for glucostasis and promoted the relative abundance of pathogenic bacteria in the gut, while a medium-energy diet maintained an ideal body condition. We concluded that dogs that are fed a low-energy diet for an extended period may become lean and lose muscle mass, but diets with low energy levels and 29% protein may not supply enough protein for dogs losing weight.

Abrupt Dietary Change and Gradual Dietary Transition Impact Diarrheal Symptoms, Fecal Fermentation Characteristics, Microbiota, and Metabolic Profile in Healthy Puppies.

Dietary changes are inevitable for pets, yet little is known about the impact of different dietary change methods on the gastrointestinal response. The current comparative study evaluated the effects of different dietary changes on the diarrheal symptoms, fecal fermentation characteristics, microbiota, and metabolic profile of healthy puppies. A total of 13 beagle puppies were randomly divided into two groups; puppies in the abrupt change (AC) group were given 260 g of a chicken- and duck-based extruded diet (CD)daily for the one-week transition period, whereas puppies in the gradual transition (GT) group were fed according to a gradual transition ratio of a salmon-based extruded diet (SA) and a CD diets with a difference of 40 g per day for seven consecutive days. Serum samples were collected on D7, and fecal samples were collected on D0 and D7. The results indicated that GT reduced the incidence of diarrhea in puppies throughout the trial period. Dietary change methods had no influence on serum inflammatory factors or fecal SCFAs, but isovaleric acid was significantly reduced after GT. Meanwhile, 16S rRNA sequencing showed that the fecal microbiota was changed after different dietary changes. Compared with the bacterial changes after AC, the relative abundances of beneficial bacteria (i.e., Turicibacter and Faecalibacterium) in feces were increased after GT in puppies. Additionally, both GT and AC caused changes in amino acid metabolism, while AC also altered lipid metabolism. AC increased fecal histamine and spermine concentrations, but decreased concentrations of metabolites such as 5-hydroxyindoleacetic acid and serotonin. Our findings indicated that GT most likely reduced the diarrhea rate in puppies by modulating the composition and metabolism of the gut microbiota.

A Validated Chiral LC-MS/MS Method for the Enantioselective Determination of (S)-(+)- and (R)-(-)-Ibuprofen in Dog Plasma: Its Application to a Pharmacokinetic Study.

The purpose of this study was to develop a method for simultaneously separating ibuprofen enantiomers using electrospray ionization (ESI) liquid chromatography with tandem mass spectrometry (LC-MS/MS). LC-MS/MS was operated with negative ionization and multiple reaction monitoring modes; transitions were monitored at m/z of 205.1 > 160.9 for ibuprofen enantiomers, 208.1 > 163.9 for (S)-(+)-ibuprofen-d3 [internal standard 1 (IS1)], and 253.1 > 208.9 for (S)-(+)-ketoprofen (IS2), respectively. In a one-step liquid-liquid extraction, 10 µL plasma was extracted with ethyl acetate:methyl tertiary-butyl ether of 7:3. Enantiomer chromatographic separation was carried out with an isocratic mobile phase consisting of 0.008% formic acid in water-methanol (v/v) at a flow rate of 0.4 mL/min on a CHIRALCEL® OJ-3R column (150 × 4.6 mm, 3 µm). This method was fully validated for each enantiomer and results were in compliance with the regulatory guidelines of the U.S. Food and Drug Administration and the Korea Ministry of Food and Drug Safety. The validated assay was executed for nonclinical pharmacokinetic studies after oral and intravenous administration of racemic ibuprofen and dexibuprofen in beagle dogs.

Effects of black soldier fly larvae as protein or fat sources on apparent nutrient digestibility, fecal microbiota, and metabolic profiles in beagle dogs.

Black soldier fly (Hermetia illucens) larvae (BSFL) act as a biological system converting organic waste into protein and fat with great potential application as pet food. To evaluate the feasibility of BSFL as a protein and fat source, 20 healthy beagle dogs were fed three dietary treatments for 65 days, including (1) a basal diet group (CON group), (2) a basal diet that replaced 20% chicken meal with defatted black soldier fly larvae protein group (DBP group), and (3) a basal diet that replaced 8% mixed oil with black soldier fly larvae fat group (BF group). This study demonstrated that the serum biochemical parameters among the three groups were within the normal range. No difference (p > 0.05) was observed in body weight, body condition score, or antioxidant capacity among the three groups. The mean IFN-γ level in the BF group was lower than that in the CON group, but there was no significant difference (p > 0.05). Compared with the CON group, the DBP group had decreasing (p < 0.05) apparent crude protein and organic matter digestibility. Furthermore, the DBP group had decreasing (p < 0.05) fecal propionate, butyrate, total short-chain fatty acids (SCFAs), isobutyrate, isovalerate, and total branched-chain fatty acids (BCFAs) and increased (p < 0.05) fecal pH. Nevertheless, there was no difference (p > 0.05) in SCFAs or BCFAs between the CON and BF groups. The fecal microbiota revealed that Lachnoclostridium, Clostridioides, Blautia, and Enterococcus were significantly enriched in the DBP group, and Terrisporobacter and Ralstonia were significantly enriched in the BF group. The fecal metabolome showed that the DBP group significantly influenced 18 metabolic pathways. Integrating biological and statistical correlation analysis on differential fecal microbiota and metabolites between the CON and DBP groups found that Lachnoclostridium, Clostridioides, and Enterococcus were positively associated with biotin. In addition, Lachnoclostridium, Clostridioides, Blautia, and Enterococcus were positively associated with niacinamide, phenylalanine acid, fumaric acid, and citrulline and negatively associated with cadavrine, putrescine, saccharopine, and butyrate. In all, 20% DBP restrained the apparent CP and OM digestibility, thereby affecting hindgut microbial metabolism. In contrast, 8% BF in the dog diet showed no adverse effects on body condition, apparent nutrient digestibility, fecal microbiota, or metabolic profiles. Our findings are conducive to opening a new avenue for the exploitation of DBP and BF as protein and fat resources in dog food.

Differential miRNA expression profiles in the bone marrow of Beagle dogs at different stages of Toxocara canis infection.

BACKGROUND: Toxocara canis is distributed worldwide, posing a serious threat to both human and dog health; however, the pathogenesis of T. canis infection in dogs remains unclear. In this study, the changes in microRNA (miRNA) expression profiles in the bone marrow of Beagle dogs were investigated by RNA-seq and bioinformatics analysis. RESULTS: Thirty-nine differentially expressed (DE) miRNAs (DEmiRNAs) were identified in this study. Among these, four DEmiRNAs were identified at 24 h post-infection (hpi) and all were up-regulated; eight DEmiRNAs were identified with two up-regulated miRNAs and six down-regulated miRNAs at 96 hpi; 27 DEmiRNAs were identified with 13 up-regulated miRNAs and 14 down-regulated miRNAs at 36 days post-infection (dpi). Among these DEmiRNAs, cfa-miR-193b participates in the immune response by regulating the target gene cd22 at 24 hpi. The novel_328 could participate in the inflammatory and immune responses through regulating the target genes tgfb1 and tespa1, enhancing the immune response of the host and inhibiting the infection of T. canis at 96 hpi. In addition, cfa-miR-331 and novel_129 were associated with immune response and self-protection mechanisms at 36 dpi. 20 pathways were significantly enriched by KEGG pathway analysis, most of which were related to inflammatory response, immune response and cell differentiation, such as Cell adhesion molecules (CAMs), ECM-receptor interaction and Focal adhesion. CONCLUSIONS: These findings suggested that miRNAs of Beagle dog bone marrow play important roles in the pathogenesis of T. canis infection in dogs and provided useful resources to better understand the interaction between T. canis and the hosts.

Effects of Softening Dry Food with Water on Stress Response, Intestinal Microbiome, and Metabolic Profile in Beagle Dogs.

Softening dry food with water is believed to be more beneficial to the intestinal health and nutrients absorption of dogs by some owners, but there appears to be little scientific basis for this belief. Thus, this study aimed to compare feeding dry food (DF) and water-softened dry food (SDF) on stress response, intestinal microbiome, and metabolic profile in dogs. Twenty healthy 5-month-old beagle dogs were selected and divided into two groups according to their gender and body weight using a completely randomized block design. Both groups were fed the same basal diet, with one group fed DF and the other fed SDF. The trial lasted for 21 days. The apparent total tract digestibility (ATTD) of nutrients, inflammatory cytokines, stress hormones, heat shock protein-70 (HSP-70), fecal microbiota, short-chain fatty acids (SCFAs), branch-chain fatty acids (BCFAs), and metabolomics were measured. Results showed that there was no significant difference in body weight, ATTD, and SCFAs between the DF and SDF groups (p > 0.05), whereas feeding with SDF caused a significant increase in serum cortisol level (p < 0.05) and tended to have higher interleukin-2 (p = 0.062) and HSP-70 (p = 0.097) levels. Fecal 16S rRNA gene sequencing found that the SDF group had higher alpha diversity indices (p < 0.05). Furthermore, the SDF group had higher levels of Streptococcus, Enterococcus, and Escherichia_Shigella, and lower levels of Faecalibacterium (p < 0.05). Serum and fecal metabolomics further showed that feeding with SDF significantly influenced the purine metabolism, riboflavin metabolism, and arginine and proline metabolism (p < 0.05). Overall, feeding with SDF caused higher cortisol level and generated effects of higher intestinal microbial diversity in dogs, but it caused an increase in some pathogenic bacteria, which may result in intestinal microbiome disturbance and metabolic disorder in dogs. In conclusion, feeding with SDF did not provide digestive benefits but caused some stress and posed a potential threat to the intestinal health of dogs. Thus, SDF is not recommended in the feeding of dogs.

Lung Lipidomic Alterations in Beagle Dogs Infected with Toxocara canis.

Toxocariasis, mainly caused by Toxocara canis, and to a lesser extent, Toxocara cati, is a neglected parasitic zoonosis. The mechanisms that underlie the changes in lipid metabolism of T. canis infection in Beagle dogs' lungs remain unclear. Lipidomics is a rapidly emerging approach that enables the global profiling of lipid composition by mass spectrometry. In this study, we performed a non-targeted lipidomic analysis of the lungs of Beagle dogs infected with the roundworm T. canis using liquid chromatography-tandem mass spectrometry (LC-MS/MS). A total of 1197 lipid species were identified, of which 63, 88, and 157 lipid species were significantly altered at 24 h post-infection (hpi), 96 hpi, and 36 days post-infection (dpi), respectively. This global lipidomic profiling identified infection-specific lipid signatures for lung toxocariasis, and represented a comprehensive comparison between the lipid composition of dogs' lungs in the presence and absence of T. canis infection. The potential roles of the identified lipid species in the pathogenesis of T. canis are discussed, which has important implications for better understanding the interaction mechanism between T. canis and the host lung.