Therapeutic Effect Evaluation Of Nanogel Enhancing Valnemulin Cows Hydrochloride For Milk Borne Staphylococcus Aureus In Dairy

Mastitis in dairy cattle is one of the diseases that have the greatest impact on the cost of dairy farming as it leads to a decrease in milk yield and quality,with the highest incidence of mastitis in dairy cattle caused by Staphylococcus aureus(S.aureus).The irrational use of antibiotics has led to increased resistance of S.aureus in clinical practice,making mastitis in dairy cows a chronic and recurrent infection,leading to a decrease in its cure rate year by year.Valnemulin hydrochloride(VH)is a broad-spectrum antibiotic with the advantages of high safety,low side effects and less resistance,but its short half-life and rapid metabolism greatly limit its application in clinical practice.Therefore,in this study,chitosan and gelatin were used to wrap VH and then cross-linked with Kynnipin to prepare VH nanogel with good slow release effect,to prolong its release time at the drug delivery site,to improve the antibacterial activity of Vonimycin hydrochloride against S.aureus,to enhance the therapeutic effect of VH on clinical-type dairy cows with mastitis,and then to promote the healthy and rapid development of dairy farming in the southern border area.The main findings of this study are as follows:1.Isolation and identification of dairy-derived S.aureus from a dairy farm in Alar,Xinjiang and study of its pathogenicityMilk samples were collected from affected cows,bacterial culture,biochemical identification and molecular biology identification,and 21 strains of S.aureus were isolated,with an isolation rate of39.6%,and 7 virulence genes were detected by relevant virulence gene tests.Some Staphylococcus aureus isolates could carry five virulence genes at the same time,among which more isolates carried the adhesion factor clfa and clfb genes,90.5%(19/21)and 95.2%(20/21),respectively.One S.aureus isolate with strong pathogenicity was screened by mouse pathogenicity test,and the minimum inhibition concentration(MIC)of VH against this S.aureus was 0.063 μg/m L.2.Preparation and characterization of VH nanogelsIn order to compensate for the fast metabolism of VH in animals,VH nanogels were prepared by changing the VH dosage form,and orthogonal tests were designed to evaluate the encapsulation efficiency(EE)and loading capacity(LC)as the evaluation indexes.The EE and LC were 67.5±2.1 %and 58.8±1.2 %,respectively,when the gelatin concentration was 75 mg/m L,the chitosan concentration was 20 mg/m L and the genipin concentration was 0.3 %.The prepared VH nanogels were light blue gel-like in appearance,and no flocculation or precipitation was observed.Scanning electron microscopy observed that VH formed nanoparticles with small particle size,uniform size and distribution under the inclusions of chitosan-gelatin and cross-linking of genipin.After freeze-drying the samples,it was observed that VH was wrapped by the network structure of gelatin,chitosan and genipin cross-linking,forming a dense three-dimensional network-like structure with homogeneous network voids.The average particle size of VH nanogel was 19.54±2.38 nm and the surface potential was 6.14 m V as determined by nano-laser particle size meter.(PXRD)results showed that the VH nanogels generated new heterocyclic ammonia compounds by cross-linking,and the VH drug molecules were completely encapsulated in the amorphous form in the nanogels cross-linked with chitosan/gelatin.In vitro release experiments showed good slow release in a simulated release environment of affected bovine milk(p H7.2).Safety tests showed that the VH nanogels were biocompatible.In vitro inhibition assays showed that the MIC value of VH nanogel against dairy-derived S aureus in cows was significantly better than that of the commercial VH drug at 0.063 μg/m L.3.Clinical efficacy validation of VH nanogelBased on the completion of the characterization and safety evaluation of VH nanogels,the in vitro pharmacodynamics and clinical animal therapeutic effects of VH nanogels were investigated.The results showed that VH nanogel was more effective than VH commercial drugs in the treatment of mastitis caused by S.aureus infection in dairy cows.