Effect Of Galla Chinensis Combined With Chebula Fructus Extract On Diarrhea Induced By Bovine Proteus Mirabilis In Mice

Diarrhea is a common disease among newborn calves.With the continuous scaling up of farming,its incidence and mortality rates have remained high,causing serious losses to the livestock industry.Proteus mirabilis is one of the main pathogenic bacteria causing calf diarrhea and is also an important zoonotic pathogen that poses a serious threat to public health.Antibiotics are commonly used in clinical treatment of bacterial calf diarrhea,which has led to the emergence of antibiotic resistance and issues related to drug residues in agricultural products.In order to better control this disease,"reducing antibiotic use and replacing antibiotics"has become the mainstream research direction.Traditional Chinese medicine,with its good antibacterial activity,has gradually become one of the important alternative drug development options for antibiotics.In this experiment,a strain of Proteus mirabilis was isolated and identified from clinical samples of calf diarrhea.20 kinds of Chinese herbs with antibacterial activity were selected to screen for an effective combination of traditional Chinese medicine through in vitro antibacterial tests.A mouse diarrhea model was established to evaluate the anti-diarrheal effect of the effective combination of traditional Chinese medicine.The following results were obtained:1.Bacterial isolation and identification were performed on samples of calf diarrhea.Through morphological observation,biochemical identification,and molecular biology identification,the pathogen was identified as Proteus mirabilis and named HY1.Pathogenicity testing in mice showed that the strain was highly pathogenic and could cause intestinal lesions in mice.2.The antimicrobial susceptibility test of the Proteus mirabilis strain was conducted using the microdilution method.Among the 20 kinds of Chinese herbs tested,Chebula Fructus,Menthae Haplocalycis Herba,Coptis Rhizoma,Artemisia Argyi Folium,Lonicera Japonicae Flos,Anemarrhena Rhizoma and Galla Chinensis showed good antibacterial effects.The herb with the lowest Minimum Inhibitory Concentration(MIC)value,Chebula Fructus,was paired with each of the other 6 herbs,and their combined antibacterial effects were tested using the checkerboard method and time-kill curve method.Among these combinations,the Chebula Fructus and Galla chinensis combination(CF+GC)exhibited the best results,achieving complete bactericidal activity within 8 hours.3.56 mice were randomly divided into 7 groups:CF+GC high-dose group,medium-dose group,low-dose group,positive drug group,single-drug group,model group,and control group.A mouse diarrhea model was established by intraperitoneal injection of the HY1 strain of Proteus mirabilis(0.1 m L/10 g bacterial solution/weight,6×10~8CFU/m L).The mice were treated continuously for 5 days,and physiological indicators of each group were observed.Compared with the control group,the model group showed decreased body weight,reduced food and water intake,increased fecal score,increased intestinal transit rate(p<0.001),disrupted intestinal villi integrity,and increased organ indexes(liver and spleen:p<0.001;heart,lung,and kidney:p<0.01;thymus:p<0.05).The blood routine test showed significantly increased WBC,LYM,MON,NEU,LY%,MO%,and NE%(p<0.001).Compared with the model group,the CF+GC group showed a trend towards normal values for the above abnormal indicators with increasing treatment days.4.The expression of inflammatory factors in mouse blood and ileum was detected by ELISA,the expression of inflammatory and mucosal repair factors in the jejunum was detected by RT-q PCR,and the expression of TGF-β1 in the duodenum was detected by immunohistochemistry.The results showed that the expression of IL-1β,IL-6,and TNF-αin the blood and ileum of the model group mice was significantly increased(p<0.001),and the expression of inflammatory factors in the blood and ileum of the treatment group mice was reduced,with the high-dose group showing the most significant effect.Compared with the model group,the expression of EGF,EGFR,and TGF-β1 in the jejunum of the treatment group mice was significantly increased(p<0.001),and the expression of TGF-β1 in the jejunum increased in a dose-dependent manner(high-dose and medium-dose groups:p<0.001;low-dose and positive drug groups:p<0.05).