ObjectiveTo investigate the effects of different fermentation methods and times on the fungal flora and chemical composition of Aurantii Fructus， in order to obtain the optimal fermentation conditions and flora structure， and to ensure the stability and controllability of the fermented varieties. MethodsScanning electron microscopy was used to observe and analyze the colony characteristics on the surface of Aurantii Fructus under different fermentation conditions. Internal transcribed spacer 2（ITS2） high-throughput sequencing， combined with fungal community diversity analysis and fungal community structure analysis， were used to obtain the fungal flora microbial categories of Aurantii Fructus under the conditions of traditional pressure-shelf fermentation and non-pressure-shelf natural fermentation for 7， 14， 21 d（numbered Y1-Y3 for the former， and numbered F1-F3 for the latter）， respectively. At the same time， the chemical components in the fermentation process were detected by ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS/MS）， combined with principal component analysis（PCA）， partial least squares-discriminant analysis（PLS-DA） and compound retention time， parent ions， characteristic fragment ions and other information， the differential compounds between the different fermentation samples were screened and identified. ResultsThe analysis of fungal community diversity showed that the dominant flora did not change at different fermentation time points in the traditional pressure-shelf fermentation method， while in the non-pressure-shelf natural fermentation method， there was a significant difference with the fermentation process， and at the genus level， the dominant genus of samples Y1， Y2， Y3 and F2 was Aspergillus， while the dominant genera of samples F1 and F3 were both Rhizopus. This indicated that the microbial growth environment provided by the traditional fermentation method was more stable， and the microbial community structure was more stable， which was more conducive to the stable and controllable fermentation process and fermented products. A total of 155 compounds were identified by compositional analysis， including 70 flavonoids， 38 coumarins， 10 alkaloids， 34 organic acids and 3 other compounds. After fermentation， two new components of ribalinine and pranferin were produced. Different fermentation conditions also brought about differences in chemical composition， multivariate statistical analysis obtained 26 differential compounds under two different fermentation methods， mainly including flavonoids， organic acids and coumarins. Comprehensively， the microbial community structure of samples fermented by the traditional pressure-shelf method of Aurantii Fructus for 14 d was stable， the species richness was high and the overall content of differential compounds was high， which was the optimal processing condition. ConclusionCompared with non-pressure-shelf natural fermentation， the traditional method has obvious advantages in terms of the stability of the microbial community structure and the content of chemical compounds， and the optimal condition is 14 days of fermentation. This study is helpful to promote the quality stability and fermentation bioavailability of fermented products of Aurantii Fructus， as well as to provide an experimental basis for the further improvement of the quality control methods of this variety.

ObjectiveTo investigate the effects of different fermentation methods and times on the fungal flora and chemical composition of Aurantii Fructus， in order to obtain the optimal fermentation conditions and flora structure， and to ensure the stability and controllability of the fermented varieties. MethodsScanning electron microscopy was used to observe and analyze the colony characteristics on the surface of Aurantii Fructus under different fermentation conditions. Internal transcribed spacer 2（ITS2） high-throughput sequencing， combined with fungal community diversity analysis and fungal community structure analysis， were used to obtain the fungal flora microbial categories of Aurantii Fructus under the conditions of traditional pressure-shelf fermentation and non-pressure-shelf natural fermentation for 7， 14， 21 d（numbered Y1-Y3 for the former， and numbered F1-F3 for the latter）， respectively. At the same time， the chemical components in the fermentation process were detected by ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry（UPLC-Q-TOF-MS/MS）， combined with principal component analysis（PCA）， partial least squares-discriminant analysis（PLS-DA） and compound retention time， parent ions， characteristic fragment ions and other information， the differential compounds between the different fermentation samples were screened and identified. ResultsThe analysis of fungal community diversity showed that the dominant flora did not change at different fermentation time points in the traditional pressure-shelf fermentation method， while in the non-pressure-shelf natural fermentation method， there was a significant difference with the fermentation process， and at the genus level， the dominant genus of samples Y1， Y2， Y3 and F2 was Aspergillus， while the dominant genera of samples F1 and F3 were both Rhizopus. This indicated that the microbial growth environment provided by the traditional fermentation method was more stable， and the microbial community structure was more stable， which was more conducive to the stable and controllable fermentation process and fermented products. A total of 155 compounds were identified by compositional analysis， including 70 flavonoids， 38 coumarins， 10 alkaloids， 34 organic acids and 3 other compounds. After fermentation， two new components of ribalinine and pranferin were produced. Different fermentation conditions also brought about differences in chemical composition， multivariate statistical analysis obtained 26 differential compounds under two different fermentation methods， mainly including flavonoids， organic acids and coumarins. Comprehensively， the microbial community structure of samples fermented by the traditional pressure-shelf method of Aurantii Fructus for 14 d was stable， the species richness was high and the overall content of differential compounds was high， which was the optimal processing condition. ConclusionCompared with non-pressure-shelf natural fermentation， the traditional method has obvious advantages in terms of the stability of the microbial community structure and the content of chemical compounds， and the optimal condition is 14 days of fermentation. This study is helpful to promote the quality stability and fermentation bioavailability of fermented products of Aurantii Fructus， as well as to provide an experimental basis for the further improvement of the quality control methods of this variety.

Objective To investigate the prevalence of Schistosoma japonicum infections in wild rodents in schistosomiasis-endemic areas of China, so as to provide insights into formulation of technical guidelines for monitoring of and the precise control strategy for S. japonicum infections in wild rodents during the elimination phase. Methods Two administrative villages where schistosomiasis was historically highly prevalent were selected each from Dongzhi County, Anhui Province, and Duchang County, Jiangxi Province as study villages. Wild rodents were captured from study villages with baited traps or cages at night in June and September, 2021. The number of rodents captured was recorded, and the rodent species was characterized based on morphologi-cal characteristics. Liver tissues were sampled from captured rodents for macroscopical observation of the presence of egg granu- lomas, and S. japonicum infection was detected simultaneously using liver tissue homogenate microscopy, examinations of mesenteric tissues for parasites, and modified Kato-Katz thick smear technique (Kato-Katz technique). A positive S. japonicum infection was defined as detection of S. japonicum eggs or adult worms by any of these methods. The rate of wild rodent capture and prevalence of S. japonicum infections in wild rodents were compared in different study villages and at different time periods, and the detection of S. japonicum infections in wild rodents was compared by different assays. Results The overall rate of wild ro- dent capture was 8.28% (237/2 861) in Dongzhi County, and the wild rodent capture rates were 9.24% (133/1 439) and 7.31% (104/1 422) in two study villages (χ² = 3.503, P = 0.061), and were 8.59% (121/1 409) and 7.99% (116/1 452) in June and September, 2021, respectively (χ² = 0.337, P = 0.561). The overall rate of wild rodent capture was 3.72% (77/2 072) in Duchang County, and the wild rodent capture rates were 6.91% (67/970) and 0.91% (10/1 102) in two study villages (χ² = 51.901, P < 0.001), and were 4.13% (39/945) and 3.37% (38/1 127) in June and September, 2021, respectively (χ² = 0.815, P = 0.365). Rattus norvegicus was the predominant rodent species captured in both counties, accounting for 70.04% (166/237) of all captured wild rodents in Dongzhi County and 88.31% (68/77) in Duchang County. No S. japonicum infection was detected in wild rodents captured in Duchang County. Nevertheless, the overall prevalence of S. japonicum infections was 51.05% (121/237) in wild rodents captured in Dongzhi County, with prevalence rates of 50.38% (67/133) and 51.92% (54/104) in two study villages (χ² = 0.098, P = 0.755), and 54.31% (63/116) and 47.93% (58/121) in September and June, 2021, respectively (χ² = 0.964, P = 0.326). Of 237 wild rodents captured in Dongzhi County, there were 140 (59.07%) rodents with visible hepatic egg granulomas, 117 (49.47%) tested positive for S. japonicum eggs by liver tissue homogenate microscopy, 34 (14.35%) tested positive for S. japonicum eggs with Kato-Katz technique; however, no adult S. japonicum worms were detected in mesenteric tissues. In addition, hepatic egg granulomas were found in all wild rodents tested positive for S. japonicum eggs with liver tissue homogenate microscopy. Conclusions The rate of wild rodent capture and prevalence of S. japonicum infection in wild rodents vary greatly in schistosomiasis-endemic areas of China, and the prevalence of S. japonicum infection is slightly higher in wild rodents captured in autumn than in summer. Liver tissue is recommended as the preferred sample for surveillance of S. japonicum infection in wild rodents, and a combination of macroscopical observation of hepatic egg granulomas and liver tissue homogenate microscopy may be a standard method for surveillance of S. japonicum infection in wild rodents.

OBJECTIVE: To investigate the effects of combined platelet-rich plasma (PRP) and arterial supercharging technique on the survival rate and functional restoration of cross-body region skin flaps in rabbits. METHODS: Twelve healthy 6-month-old New Zealand White rabbits were randomly assigned to 4 groups ( n=3): sham group, PRP group, anastomosis group, and combined treatment group. An axial skin flap with an area of 12 cm×6 cm on the inner side of the hind limbs of all animals were prepared, with the saphenous artery as the main blood supply. Following the ligation of both the proximal and distal ends of the saphenous artery across all groups, the sham group received no further intervention, the PRP group was subjected to PRP injection, the anastomosis group underwent in situ end-to-end anastomosis of the distal saphenous artery, and the combined treatment group received both in situ distal saphenous artery anastomosis and PRP administration. Flap survival was evaluated and recorded on postoperative days 1, 3, and 7, with survival rates calculated accordingly. On day 7, flap tissue samples were harvested for HE staining to assess basal tissue morphology. Additionally, immunohistochemical staining was conducted to detect the expression of α-smooth muscle actin (α-SMA), vascular endothelial growth factor (VEGF), and CD31 in the flap tissues. RESULTS: At postoperative day 1, no significant difference in flap survival rates were observed among the 4 groups ( P>0.05). At day 3, the PRP group showed no significant difference compared to the sham group ( P>0.05); however, both the anastomosis and combined treatment groups exhibited significantly higher survival rates than the sham group ( P<0.05), the combined treatment group further demonstrated superior survival rates compared to both the PRP and anastomosis groups ( P<0.05). At day 7, the combined treatment group maintained significantly higher survival rates than all other groups ( P<0.05), while both the PRP and anastomosis groups exceeded the sham group ( P<0.05). HE staining at day 7 revealed persistent inflammatory cell infiltration, sheet-like erythrocyte deposition, and disordered collagen fibers in the sham group. The PRP group showed nascent microvessel formation and early collagen reorganization, whereas the anastomosis group displayed mature microvasculature with resolved interstitial edema. The combined treatment group exhibited differentiated microvessels with densely packed collagen bundles. Immunohistochemical analysis at day 7 demonstrated significantly larger relative area percentages of α-SMA, VEGF, and CD31 positive cells in the combined treatment group compared to all other groups ( P<0.05). Both the PRP and anastomosis groups also showed significantly higher values than the sham group ( P<0.05). CONCLUSION The combination of PRP and arterial supercharging techniques significantly enhances flap healing, potentially through mechanisms involving augmented angiogenesis and improved blood supply.
Animals ; Rabbits ; Platelet-Rich Plasma ; Surgical Flaps/blood supply* ; Graft Survival ; Anastomosis, Surgical/methods* ; Ischemia/surgery* ; Arteries/surgery* ; Skin/blood supply* ; Vascular Endothelial Growth Factor A/metabolism* ; Male ; Skin Transplantation/methods*

Effective annotation of in vivo drug metabolites using liquid chromatography-mass spectrometry (LC-MS) remains a formidable challenge. Herein, a metabolic reaction-based molecular networking (MRMN) strategy is introduced, which enables the "one-pot" discovery of prototype drugs and their metabolites. MRMN constructs networks by matching metabolic reactions and evaluating MS² spectral similarity, incorporating innovations and improvements in feature degradation of MS² spectra, exclusion of endogenous interference, and recognition of redundant nodes. A minimum 75% correlation between structural similarity and MS² similarity of neighboring metabolites was ensured, mitigating false negatives due to spectral feature degradation. At least 79% of nodes, 49% of edges, and 97% of subnetworks were reduced by an exclusion strategy of endogenous ions compared to the Global Natural Products Social Molecular Networking (GNPS) platform. Furthermore, an approach of redundant ions identification was refined, achieving a 10%-40% recognition rate across different samples. The effectiveness of MRMN was validated through a single compound, plant extract, and mixtures of multiple plant extracts. Notably, MRMN is freely accessible online at https://yaolab.network, broadening its applications.

Objective To analyze the distribution of carbapenem-resistant strains and the transmission mode of resistance genes in the bacteria strains from urinary system of the individuals outside hospitals,and further explore the impacts of the antibiotics with different antibacterial mechanisms on the conjugation transfer of resistant plasmids.Methods The imipenem-resistant strains from 991 non-re-petitive urine samples of healthy individuals were screened for determining the carbapenem-resistance genotypes.Using the multilocus sequence typing(MLST)method,the main carbapenem-resistant strains were classified to determine the phylogenetic relationship be-tween different sequence types(STs)and reveal the transmission mode of resistance genes in the individuals outside hospitals.The conjugation experiment was used to analyze the effects of different sub-inhibitory concentrations of ampicillin,ciprofloxacin,and genta-micin on the conjugation efficiency of resistant plasmids in the bacteria of donor and/or recipient at different times.Results A total of 18 non-repetitive carbapenem-resistant strains(1.82％,18/991)were detected in healthy individuals outside hospitals,among which Escherichia coli(E.coli)(44.44％,8/18)was the main strain with main genotype being blaKPC-2(7/8).MLST showed that the 7 strains of E.coli with blaKPC-2 genotype belonged to ST-10,ST-101,ST-131,ST-405,ST-410,ST-1193,and ST-2562,respectively.Homologous clustering analysis showed that the type of the 7 E.coli strains exhibited high diversity.The range of joint efficiency was(1.59±0.20)× 10-4 to(4.01±1.31)× 10-4.The highest conjugation efficiency was observed in the donor bacteria treated with 1/8 of MIC of antibiotics for 9 hours,and the conjugation efficiency most significantly increased compared to the group without antibiotic treat-ment.Three antibiotics with different mechanisms of action could all improve conjugation efficiency,among which ampicillin exhibited the most significant improvement.Conclusion In this study,the genotype of carbapenem-resistant Escherichia coli(CREC)was mainly blaKPC-2 with a main mode of horizontal transmission.Antibiotics with sub-inhibitory concentrations could promote the conjugation transfer of the resistant plasmid.There may be significant differences in the effects of antibiotics with different mechanisms of action and different antibiotic stress conditions on the conjugation efficiency.

Objective:To investigate the clinicopathological and molecular genetic characteristics of Crohn′s disease (CD).Methods:A retrospective analysis was conducted on 52 CD patients who underwent surgical resection at the First Affiliated Hospital of Nanjing Medical University between January 2014 and June 2023. Clinical presentations and histopathological features were assessed. Whole-genome sequencing was performed on 17 of the samples, followed by sequencing and pathway enrichment analyses. Immunohistochemistry was used to assess the expression of frequently mutated genes.Results:Among the 52 patients, 34 were males and 18 were females, male-to-female ratio was 1.9∶1.0, with a median age of 45 years at surgery and 35 years at diagnosis. According to the Montreal classification, A3 (51.9%,27/52), B2 (61.5%, 32/52), and L3 (50.0%,26/52) subtypes were the most predominant. Abdominal pain and diarrhea were the common symptoms. Histopathological features seen in all 52 patients included transmural inflammation, disruption of cryptal architecture, lymphoplasmacytic infiltration, varying degrees of submucosal fibrosis and thickening, increased enteric nerve fibers and neuronal proliferation. Mucosal defects, fissure ulcers, abscesses, pseudopolyps, and adenomatous proliferation were also observed in 51 (98.1%), 38 (73.1%), 28 (53.8%), 45 (86.5%), and 28 (53.8%) cases, respectively. Thirty-one (59.6%) cases had non-caseating granulomas, and 3 (5.8%) cases had intestinal mucosal glandular epithelial dysplasia. Molecular analysis showed that 12/17 CD patients exhibited mutations in at least one mucin family gene (MUC2, MUC3A, MUC4, MUC6, MUC12, MUC17), and MUC4 was the most frequently mutated in 7/17 of cases. Immunohistochemical stains showed reduced MUC4 expression in epithelial cells, with increased MUC4 expression in the epithelial surface, particularly around areas of inflammatory cell aggregation; and minimal expression in the lower half of the epithelium.Conclusions:CD exhibits diverse clinical and pathological features, necessitating a comprehensive multidimensional analysis for diagnosis. Mutations and expression alterations in mucin family genes, particularly MUC4, may play crucial roles in the pathogenesis of CD.

The present jaw relationship record tray is designed in an upper and lower integrated type,which can not accurately ob-tain important information of maxillary occlusal plane,the position about 1 mm below the upper lip and the support degree of the upper lip during clinical work.Therefore,the base-plate and occlusal-rim composite solid(diagnostic denture)is not accurate and the accu-rate jaw relationship can not be tranfered in the following steps.This paper introduces the design,clinical application process and key points of a split-type jaw relationship record tray.It can accurately obtain important information such as the occlusal plane,the position about 1mm below the upper lip,and the degree of support of the upper lip.It provides reference for the accurate recording and transfer of the jaw relationship.It also lays a foundation for the direct fabrication of full denture based on the information obtained by the split-type jaw relationship record tray after the digital impression obtained by intraoral scanning.

More than 300 million people world-wide suffer from asthma,and the incidence is in-creasing year by year.As one of the most common chronic diseases,asthma is an immune-mediated inflammatory disease with complex triggering mechanisms and strong heterogeneity.With the in-depth study of physiological and pathological mech-anisms,therapeutic small molecule and hormone drugs have been introduced to control and treat most patients,but about 5％-10％of patients still suffer from various subtypes of difficult to control and treat asthma,that is,severe asthma.In the past decade,with the rapid development of bio-pharmaceutical research,protein and antibody have become the key drugs for the treatment of se-vere asthma with high efficacy,high specificity and high safety.However,biological drugs are usually administered by injection,they cannot be noninva-sive and directly delivered into the lung to quickly absorb and take effect.Therefore,there is an ur-gent need for the introduction of inhaled biologics with quick effectiveness,convenience,economy and safety in clinical.The review summarizes the existing small molecule,hormone and biological therapy drugs,and summarizes the development of inhalable biological agents of asthma,and ana-lyzes the future prospects of the inhalable biologi-cal drugs,which is designed to deepen the percep-tion of the direction of the inhalable biological drugs research,and update the information of the field,in order to provide reference for the develop-ment of more inhalable biologics.

More than 300 million people worldwide suffer from asthma, and the incidence is increasing year by year. As one of the most common chronic diseases, asthma is an immune-mediated inflammatory disease with complex triggering mechanisms and strong heterogeneity. With the in-depth study of physiological and pathological mechanisms, therapeutic small molecule and hormone drugs have been introduced to control and treat most patients, but about 5% - 10% of patients still suffer from various subtypes of difficult to control and treat asthma, that is, severe asthma. In the past decade, with the rapid development of bio-pharmaceutical research, protein and antibody have become the key drugs for the treatment of severe asthma with high efficacy, high specificity and high safety. However, biological drugs are usually administered by injection, they cannot be noninvasive and directly delivered into the lung to quickly absorb and take effect. Therefore, there is an urgent need for the introduction of inhaled biologics with quick effectiveness, convenience, economy and safety in clinical. The review summarizes the existing small molecule, hormone and biological therapy drugs, and summarizes the development of inhalable biological agents of asthma, and analyzes the future prospects of the inhalable biological drugs, which is designed to deepen the perception of the direction of the inhalable biological drugs research, and update the information of the field, in order to provide reference for the development of more inhalable biologics.