Escherichia coli Producing CTX-M and DHA-1 Beta-lactamases in Farm Animals in Chongqing City of China

Aim of this study was to investigate the genotypic distribution of extended-spectrum β-lactamases (ESBLs) and AmpC enzymes among Escherichia coli isolated from farm animals in Chongqing City. 326 E.coli isolates were obtained from piglets,cattle and chickens.In these isolates,19 ESBLs and 6 AmpC-producing E.coli isolates were detected by preliminary screening and phenotypic identification, and polymerase chain reaction and sequence analysis methods were used to detect ESBLs and AmpCβ-lactamases genes.The results showed that the frequencies of TEM,CTX-M,DHA-1 and CMY-2 genes were 2.15%,5.21%,0.61%and 0.61%,respectively,in E.coli of animal origin in the Chongqing area,and 5 E.coli isolates were found to carry two kinds of genotype.The DHA-1 gene was detected for the first time in a veterinary clinic in China;its GenBank accession number is FJ386455.This study indicated that TEM and CTX-M genes were the predominant genotypes in Chongqing,and the majority of ESBLs and AmpC-producing E.coli isolates were multi-resistant to antibiotics,so they deserve particular attention in veterinary clinics.

Identification and antibiotic susceptibility evaluation of Mycoplasma synoviae isolated from chickens in central China

Mycoplasma synoviae(M.synoviae)infections have become an increasingly serious concern in China because they cause huge economic losses to the poultry industry.Antibiotic treatment is one of control strategies that can be used to contain clinical outbreaks in M.synoviae-free flocks,especially because the bacteria can be transmitted through eggs.To understand M.synoviae infection status in farms of central China and the antibiotic susceptibility of the circulating strains in vivo and in vitro,485 samples were collected from five provinces from 2019 to 2021.Fifty-two strains were isolated and identified.Determination of the minimum inhibitory concentration(MIC)of eight antibiotics(tylvalosin,tiamulin,tilmicosin,lincomycin,enrofloxacin,chlortetracycline,doxycycline and tylosin)for isolates showed that tylvalosin,doxycycline and tiamulin were effective against 52 clinical isolates(MIC values≤0.0625-0.25μg/mL,≤0.0625-1μg/mL,and 0.25-2μg/mL,respectively).Tilmicosin,enrofloxacin and lincomycin had high MICo values(>32μg/mL).An artificial M.synoviae infection model was established in chickens for evaluation of the short-term therapeutic effect of these antibiotics.After 5 days of medication,doxycycline(200 mg/L)showed a superior ability to inhibit M.synoviae compared with other groups,as did tylvalosin(200 mg/L).Furthermore,the therapeutic efficacy of tylvalosin(0.4μg/mL)on intra-embryo-injected M.synoviae was higher than that of tiamulin at the same dose.A combination of MiC values determined in vitro and therapeutic effects observed in vivo revealed that tylvalosin and doxycycline had the best therapeutic effects.Tylvalosin also showed better inhibitory effects on the vertical transmis-sion of M.synoviae than tiamulin.

The Effect of Glycyl-Glutamine Dipeptide Concentration on Enzyme Activity, Cell Proliferation and Apoptosis of Jejunal Tissues from Weaned Piglets

An experiment was conducted in a singly factorial design to study the effect of glycyl-glutamine dipeptide on enzyme activity, cell proliferation and apoptosis of jejunal tissues from weaned piglets at different glycyl-glutamine concentration levels of 2, 4, 10, 20, and 30 mmol L-1, respectively. The glutaminase activity, diamine oxidase （DAO） activity, cell peoliferation, apoptosis, and perotein metabolism were measured by the tissue culture method in vitro using jejunal tissues. The immunohistochemical method was used to study the cell proliferation and apoptosis of jejunal tissues. The results showed that compared to the blank control, the percentage and MOD value of BrdU-positicve cells incubated with glycyl-glutamine dipeptide solution were significantly （P0.05） increased. Accordingly, the percentage and MOD value of caspase-3-positive cells from tissue incubated with glycyl-glutamine dipeptide were notably lower （P0.05） than that from the control treatment. The glycyl-glutamine dipeptide increased the glutaminase activity, DAO activity and protein content of jejunal tissues, as the dipeptide concentration was on the rise （P0.05）. These results indicated that glycyl-glutamine dipeptide affected the jejunum development and adaptation of weaned piglets, and the function might be fulfilled by enhancing the glutamine-related enzyme activity, thereby increasing the consumption of glutamine, and then improving the jejunal cell proliferation and suppressing cell apoptosis. The effects of glycyl-glutamine dipeptide relied in a dose-dependent manner, and the maximum effect was achieved at 20-30 mmol L-1 glycyl-glutamine dipeptide.

Antibacterial Activity of 9-Octadecanoic Acid-Hexadecanoic Acid-Tetrahydrofuran-3,4-Diyl Ester from Neem Oil

The 9-octadecanoic acid-hexadecanoic acid-tetrahydrofuran-3,4-diyl ester from neem oil was investigated for antibacterial activity against three bacterial strains viz., Staphylococcus aureus ATCC No. 25923, Escherichia coli ATCC No. 44102 and Salmonella sp. ATCC No. 50 041 in vitro. The minimum inhibitory concentration （MIC） and minimum bactericidal concentration （MBC） values of the 9-octadecanoic acid-hexadecanoic acid-tetrahydrofuran-3,4-diyl ester were determined by using the broth microdilution dilution （BMD） method at different concentrations ranging from 20 to 0.625 mg mL-1. Its time-inhibition curve against E. coli was also tested and showed that the MIC values for the bacterial strains S. aureus, E. coli and Salmonella sp. were 20, 5 and 10 mg mL-1, respectively. Its MBC values were 20, 20 and 10 mg mL-1, respectively. The antibacterial activity of 9-octadecanoic acid-hexadecanoic acid-tetrahydrofuran-3,4-diyl ester against three strain tested showed the relationship with time and concentration.

The prolonged effect of glucagon-like peptide 2 pretreatment on growth performance and intestinal development of weaned piglets

Background： Glucagon-like peptide 2 （GLP-2） is a potent epithelium-specific intestinal growth factor. The aim of this study was to demonstrate the prolonged effect of GLP-2 on the growth performance of weaned piglets. Forty piglets weaned at the age of 28 d with an average BW of 6.8 ＋ 0.4 kg were assigned to four treatments： （i） non- challenged control; （ii） LPS-challenged control; （iii） LPS ＋ low GLP-2; and （iv） LPS ＋ high GLP-2. Piglets in groups （i）, （ii）, and （iv） were s.c. injected with PBS supplemented with human [Gly2]GLP-21-34 at doses of 0, 2 and 10 nmol/kg BW per day for seven consecutive days. BW, gain：feed ratio （G：F）, and plasma GLP-2 levels were determined on d 0 7, and 14 after weaning. Piglets were challenged with i.p. administration of Escherichia coil lipopolysaccharide （LPS） at a dose of 100 pg/kg on d 14 to induce intestinal damage. Twenty-four hours later, intestinal tract samples were collected to assess intestinal morphology and quantify enzyme activity. Results： Plasma GLP-2 levels decreased after weaning, but in the high GLP-2 group, plasma GLP-2 was maintained on d 7 and even increased to a level higher than the preweaning level on d ]4 （P 〈 0.05）. High GLP-2 treatment significantly increased the duodenal, jejunal and ileal weight, as well as the gross weight of the small intestine （SI）, and the SI weight index （P 〈 0.05）. LPS caused villous atrophy and disrupted intestinal morphology in the duodenum, jejunum and ileum. GLP-2 also significantly increased the villus height and the villus height/crypt depth ratio （VCR） of the duodenum, jejunum, and ileum （P 〈 0.05）. Histological examination revealed that in GLP-2-treated groups, the integrity of the villus was maintained, and the villus was protected against LPS-induced damage. GLP-2 significantly increased the activity of alkaline phosphatase （AKP）, y-glutamyltranspeptidase （y-G-i-）, and pancreatic lipase in the duodenum and jejunum （P 〈 0.05）. GLP-2 treatment also significantly increased the average daily gain （ADG） and G：F of piglets at 0 to 7, 7 to 14, as well as 0 to14 d （P 〈 0.05）, resulting in a significant increase of final 8W in high GLP-2 pigs （P = 0.016）. Conclusions： Exogenous GLP-2 improved the growth of weaned piglets and protected them against LPS-induced intestinal damage. These effects may be due to the ability of GLP-2 to promote the secretion of endogenous GLP-2 to stimulate the small intestinal development.

Morphological Characteristics of the Dorsal Skin of Two Hynobiids and Their Adaptive Role in Aquatic and Terrestrial Habitats

The morphological characteristics of the dorsal skin of trunk in two species of hynobiid salamanders, Batrachuperus pinchonii and Hynobius chinensis were examined by light microscopy. The basic structures of the skin in the two species are similar and consist of two layers： epidermis and dermis. The epidermis consists of stratum corneum, stratum intermedium and stratum germinativum, while the dermis is composed of a stratum spongiosum and stratum compactum. However, some species-specific variation has been identified （e.g., the distribution of capillary vessels and gland cells, and the thickness of skin）. H. chinensis is a terrestrial species and only lives in water during breeding period, but B. pinchonii is aquatic and remains aquatic throughout its lifetime. The differences in the distribution of capillary vessels and gland cells are related to their different habitats, and show a morphological adaptation.

Antibacterial Activity of Crude Extracts of Prinsepia utilis Royle in vitro

The purpose of this study is to isolate the active fraction from the crude extract of Prinsepia utilis Royle and to determine its antibacterial activity against the standard and drug-resistant bacteria strains in vitro. The Prinsepia utilis Royle was extracted in three portions by using petroleum ether, ethanol and water by the continuation recirculate extraction. The antibacterial activity of the extracts were evaluated by the determination of Minimum Inhibitory Concentration （MIC） and Minimum Bactericidal Concentration （MBC） values. The results showed that the ethanol extract and aqueous extract of petroleum ether showed the weakest inhibition. MIC values of the aqueous Prinsepia utilis Royle were found that they had significant effects against bacteria, while ethanol extracts against three standard bacteria （S. aureus ATCC25923, E. coli 44102, Salmonella 50041） were 25, 25, 50 mg·mL^-1 and 50, 100, 100 mg·mL^-1, MIC values of three drug-resistant bacteria （S. aureus GL17, E. coli EYAC08-56, Salmonella STQD2G.） were 50, 50, 50 mg·mL^-1 and 100, 100, 200 mg·mL^-1, respectively. MBC values of two extracts against three standard strains were 25, 50, 100mg·mL^-1 and 50, 100, 200 mg·mL^-1, MBC values of three drug-resistant bacteria were 100, 100, 100mg·mL^-1 and 200, 200, 200 mg·mL^-1, respectively.

Targeted myostatin loss-of-function mutation increases type Ⅱ muscle fibers in Meishan pigs

Myostatin(MSTN) is a negative regulator of skeletal muscle growth and development. The skeletal muscle in MSTN^(-/-)mice is significantly hypertrophied, with muscle fiber type II increasing significantly while muscle fiber type I decreasing.However, it is still not clear how the skeletal muscle types change in MSTN^(-/-)pigs, and how the mechanism for MSTN regulates fiber types, especially in large animals like pigs. This study conducted a comprehensive analysis of the composition of skeletal muscle fibers in MSTN^(-/-)pigs produced in our laboratory. It was observed that, compared with wild-type(WT) pigs, both the total mass of skeletal muscle and type IIb muscle fibers increased significantly(P<0.01),while the type I and type IIa muscle fibers decreased significantly(P<0.01), in MSTN^(-/-)Meishan pigs. In addition, to explore the influence of MSTN on muscle fiber type and its regulation mechanism in the embryonic stage, this study selected a few genes(Myf5, Mef2 d, MyoD and Six1) associated with muscle fiber type and validated their expression by quantitative RT-PCR. Herein, it was found that Myh7, Myh2, Myh4 and Myh1 can be detected in the skeletal muscle of pigs at 65 days of gestation(dg). Compared with WT pigs, in MSTN^(-/-)Meishan pigs, Myh7 decreased significantly(P<0.01), while Myh4(P<0.001) and Myh1(P<0.05) increased significantly. Meanwhile, the increased expression of Myf5(P<0.05), Mef2 d(P<0.01) and Six1(P<0.05) in MSTN^(-/-)Meishan pigs suggested that MSTN should regulate the directional development of muscle fiber types in the early stage of embryonic development. Thus, at the embryonic stage, the type II muscle fibers began to increase in MSTN^(-/-)pigs. These results can provide valuable information not only for pig meat quality improvement, but also for the study of human skeletal muscle development and disease treatment.

Preliminary Isolation and Antibacterial Activity of the Ethyl Acetate Extract of <i>Prinsepia utilis</i>Royle <i>in Vitro</i>

Seven fractions were isolated from the ethyl acetate extract of Prinsepia utilis Royle by silica gel column chromatography. The antibacterial activities determined by disc diffusion method in vitro, indicated that the first and fourth fractions showed better antibacterial activity than the other fractions, while the sixth and seventh fractions did not showed any antibacterial activity. The diameters of the inhibition zone of first and fourth fractions were greater than 10 mm against three standard strains (Staphylococcus aureus ATCC25923, Escherichia coli ATCC44102 and Salmonella sp. CMCC(B)50041) at the concentration of 20 mg/ml. The first fraction was then repeatedly recrystallized in acetone to yield a white snowflake-like compound A, the inhibition zones of which were 14.03 mm, 11.54 mm and 12 mm, respectively. The Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were measured by broth dilution method at the concentrations ranging from 20 to 0.313 mg/mL. The MIC and MBC values of the first, fourth fractions and compound A were lower than that of oregano oil (positive control) against S. aureus ATCC25923.

ArcA Gene Expression Level of Actinobacillus pleuropneumoniae in Anaerobic Culture

In the present study,mRNA levels of the ArcA gene of Actinobacillus pleuropneumoniae serotype1 were measured during response to stress by growing under anaerobic conditions.Expression levels were measured by semi-quantitative RT-PCR using the housekeeping gene recF as an internal standard.The expression of ArcA was undetectable until about 3 hours under standard culture conditions,but it was promptly and highly expressed in anaerobic culture.The results are consistent with ArcA being a potential virulence gene of Actinobacillus pleuropneumoniae,and likely playing an important role in pathogenesis caused by this organism.

Dramatic Changes of Matrix Metalloproteinases-7 and Lysozyme in the Ulcerative Colitis of Mice Induced by Dextran Sulfate Sodium

Ulcerative colitis （UC） is a lifelong illness with profound emotional and social impacts, and could cause serious damage to large intestine, especially in colon. However, the pathogenesis of UC remained unclear. The present study attempts to find out the role of matrix metalloproteinases-7 （MMP-7） and lysozyme in the pathogenesis of UC through a mice model induced by dextran sulfate sodium （DSS）. The UC model was evaluated both by disease activity index （DAI） and the intestinal histopathology. The results show that there is a high correlation between the DAI score and the pathological changes of colon. Interleukin-6 （IL-6） serum levels and large intestinal fluids levels in UC mice are always higher than that of the control groups, which might be associated with the degree of the inflammation damage in the colon. The change tendency of the MMP-7 mRNA and protein expressions are both up-regulated firstly and then down-regulated from 1 to 5 d in the colon, but only the MMP-7 protein is up-regulated at 7 d again. The up-regulated MMP-7 levels in the early stage of UC may play a protective role through the activated defensins, while the down-regulated levels in the mid-later stage of UC may be connected with the severe lesions in the colon. However, the up-regulated MMP-7 levels in the later stage of UC in the colon may also contribute to the tissue repair or be served as a marker to CRC （colorectal cancer）. The distribution of lysozyme protein indicates that there may be Paneth-like cells in the colon. Both the changes of MMP-7 and lysozyme in the small intestine may play a protective role for the safe environment of the whole gut, especially to the colon of UC.

MicroRNA regulates the toxicological mechanism of four mycotoxins in vivo and in vitro

Mycotoxins can cause body poisoning and induce carcinogenesis,often with a high mortality rate.Therefore,it is of great significance to seek new targets that indicate mycotoxin activity and to diagnose and intervene in mycotoxin-induced diseases in their early stages.MicroRNAs(miRNAs)are physiological regulators whose dysregulation is closely related to the development of diseases.They are thus important markers for the occurrence and development of diseases.In this review,consideration is given to the toxicological mechanisms associated with four major mycotoxins(ochratoxin A,aflatoxin B1,deoxynivalenol,and zearalenone).The roles that miRNAs play in these mechanisms and the interactions between them and their target genes are explained,and summarize the important role of histone modifications in their toxicity.As a result,the ways that miRNAs are regulated in the pathogenicity signaling pathways are revealed which highlights the roles played by miRNAs in preventing and controlling the harmful effects of the mycotoxins.It is hoped that this review will provide a theoretical basis for the prevention and control of the damage caused by these mycotoxins.

Effect of recombinant growth hormone on expression of growth hormone receptor, insulin-like growth factor mRNA and serum level of leptin in growing pigs

Sixteen Large White ?Landrace castrated male pigs were allotted into treatment and control group. The treatment group was injected intramuscularly with recombinant porcine growth hormone (rpGH, 4 mg·d-1) and the control group with vehicle for 28 days. Animals were slaugh-tered 4 h after final injection for liver, longissimus dorsi (LD) muscle and blood sampling. Serum concentration of insulin-like growth factor 1 (IGF-I) and leptin were determined by RIA. The total RNA was extracted from tissues to measure the abundance of growth hormone receptor (GHR), IGF-I mRNA by RT-PCR with 18S rRNA internal standard. Results showed that rpGH enhanced the average daily weight gain by 26.1% (P < 0.05), the serum IGF-I concentration by 70.94% (P < 0.01), decreased serum leptin by 34.8% (P < 0.01). The relative abundance of GHR and IGFmRNA in liver were increased by 24.45% (P < 0.05) and 45.30% (P < 0.01), respectively, but no difference of GHR (P > 0.05) and IGF-I mRNA (P > 0.05) in LD between GH treated and control group was found. These results suggest that rpGH can up-regulate hepatic GHR and IGF-I gene expression and improve animal growth. However the effect of rpGH on GHR and IGF-I gene ex-pression are tissue-specific.

Taurolidine improved protection against highly pathogenetic avian influenza H5N1 virus lethal-infection in mouse model by regulating the NF-κB signaling pathway

Taurolidine(TRD),a derivative of taurine,has anti-bacterial and anti-tumor effects by chemically reacting with cell-walls,endotoxins and exotoxins to inhibit the adhesion of microorganisms.However,its application in antiviral therapy is seldom reported.Here,we reported that TRD significantly inhibited the replication of influenza virus H5N1 in MDCK cells with the half-maximal inhibitory concentration(EC_(50))of 34.45μg/mL.Furthermore,the drug inhibited the amplification of the cytokine storm effect and improved the survival rate of mice lethal challenged with H5N1(protection rate was 86%).Moreover,TRD attenuated virus-induced lung damage and reduced virus titers in mice lungs.Administration of TRD reduced the number of neutrophils and increased the number of lymphocytes in the blood of H5N1 virus-infected mice.Importantly,the drug regulated the NF-κB signaling pathway by inhibiting the separation of NF-κB and IκBa,thereby reducing the expression of inflammatory factors.In conclusion,our findings suggested that TRD could act as a potential anti-influenza drug candidate in further clinical studies.