Progress in experimental models to investigate the in vivo and in vitro antidiabetic activity of drugs

Diabetes mellitus is one of the world's most prevalent and complex metabolic disorders,and it is a rapidly growing global public health issue.It is characterized by hyperglycemia,a condition involving a high blood glucose level brought on by deficiencies in insulin secretion,decreased activity of insulin,or both.Prolonged effects of diabetes include cardiovascular problems,retinopathy,neuropathy,nephropathy,and vascular alterations in both macro-and micro-blood vessels.In vivo and in vitro models have always been important for investigating and characterizing disease pathogenesis,identifying targets,and reviewing novel treatment options and medications.Fully understanding these models is crucial for the researchers so this review summarizes the different experimental in vivo and in vitro model options used to study diabetes and its consequences.The most popular in vivo studies involves the small animal models,such as rodent models,chemically induced diabetogens like streptozotocin and alloxan,and the possibility of deleting or overexpressing a specific gene by knockout and transgenic technologies on these animals.Other models include virally induced models,diet/nutrition induced diabetic animals,surgically induced models or pancreatectomy models,and non-obese models.Large animals or non-rodent models like porcine(pig),canine(dog),nonhuman primate,and Zebrafish models are also outlined.The in vitro models discussed are murine and human beta-cell lines and pancreatic islets,human stem cells,and organoid cultures.The other enzymatic in vitro tests to assess diabetes include assay of amylase inhibition and inhibition ofα-glucosidase activity.

The antitumor effect of bromophenol derivatives in vitro and Leathesia nana extract in vivo

To investigate the antitumor effect of bromophenol derivatives in vitro and Leathesia nana extract in vivo, six bromophenol derivatives 6-（2,3-dibromo-4,5-dihydroxybenzyl）-2,3-dibromo-4,5-dihydroxy benzyl methyl ether （1）, （＋）-3-（2,3-dibromo-4,5-dihydroxyphenyl）-4-bromo-5,6-dihydroxy-1,3- dihydroisobenzofuran （2）, 3-bromo-4-（2,3-dibromo-4,5-dihydroxybenzyl）-5-methoxymethyl-pyrocatechol （3）, 2,2＇,3,3＇-tetrabromo-4,4＇,5,5＇-tetrahydroxy-diphenylmethane （4）, bis（2,3-dibromo-4,5-dihydroxybenzyl） ether （5）, 2,2＇,3-tribromo-3＇,4,4＇,5-tetrahydroxy-6＇-ethyloxymethyldiphenylmethane （6） were isolated from brown alga Leathesia nana, and their cytotoxicity were tested by MTF assays in human cancer cell lines A549, BGC-823, MCF-7, B16-BL6, HT-1080, A2780, Be17402 and HCT-8. Their inhibitory activity against protein tyrosine kinase （PTK） with over-expression of c-kit was analyzed also by ELISA. The antitumor activity of ethanolic extraction of Leathesia nana （EELN） was evaluated on S180-bearing mice. All compounds showed very potent cytotoxicity against all of the eight cancer cell lines with IC50 below 10 pg/mL. In PTK inhibition study, all bromophenol derivatives showed moderate inhibitory activity and compounds 2, 5 and 6 showed significant bioactivity with the inhibition ratio of 77.5%, 80.1% and 71.4% respectively. Pharmacological studies reveal that EELN could inhibit the growth of Sarcoma 180 tumor and increase the indices of thymus and spleen to improve the immune system remarkably in vivo. Results indicated that the bromophenol derivatives and EELN can be used as potent antitumor agents for PTK over-expression of c-kit and considered in a new therapeutic strategy for treatment of cancer.

Sustained release donepezil loaded PLGA microspheres for injection:Preparation,in vitro and in vivo study

The purpose of this study was to develop a PLGA microspheres-based donepezil(DP)formulation which was expected to sustain release of DP for one week with high encapsulation efficiency(EE).DP derived from donepezil hydrochloride was encapsulated in PLGA microspheres by the O/W emulsion-solvent evaporation method.The optimized formulation which avoided the crushing of microspheres during the preparation process was characterized in terms of particle size,morphology,drug loading and EE,physical state of DP in the matrix and in vitro and in vivo release behavior.DP microspheres were prepared successfully with average diameter of 30m,drug loading of 15.92±0.31%and EE up to 78.79±2.56%.Scanning electron microscope image showed it has integrated spherical shape with no drug crystal and porous on its surface.Differential scanning calorimetry and X-ray diffraction results suggested DP was in amorphous state or molecularly dispersed in microspheres.The Tg of PLGA was increased with the addition of DP.The release profile in vitro was characterized with slow but continuous release that lasted for about one week and fitted well with first-order model,which suggested the diffusion governing release mechanism.After single-dose administration of DP microspheres via subcutaneous injection in rats,the plasma concentration of DP reached peak concentration at 0.50 d,and then declined gradually,but was still detectable at 15 d.A good correlation between in vitro and in vivo data was obtained.The results suggest the potential use of DP microspheres for treatment of Alzheimer’s disease over long periods.

Effects of Size and Surface Charge of Polymeric Nanoparticles on in Vitro and in Vivo Applications

Biodegradable polymeric materials are the most common carriers for use in drug delivery systems. With this trend, newer drug delivery systems using targeted and controlled release polymeric nanoparticles (NPs) are being developed to manipulate their navigation in complex in vivo environment. However, a clear understanding of the interactions between biological systems and these nanoparticulates is still unexplored. Different studies have been performed to correlate the physicochemical properties of polymeric NPs with the biological responses. Size and surface charge are the two fundamental physicochemical properties that provide a key direction to design an effective NP formulation. In this critical review, our goal is to provide a brief overview on the influences of size and surface charge of different polymeric NPs in vitro and to highlight the challenges involved with in vivo trials.

Gan Shen Fu Fang ameliorates liver fibrosis in vitro and in vivo by inhibiting the inflammatory response and extracellular signalregulated kinase phosphorylation

BACKGROUND Liver fibrosis is a common health problem worldwide and there is still a lack of effective medicines.The Chinese herbal medicine,Gan Shen Fu Fang(GSFF)is composed of salvianolic acid B and diammonium glycyrrhizinate.In this study,we observed the effects of GSFF on liver fibrosis in vivo and in vitro in an attempt to provide some hope for the treatment.AIM To observe the effects of GSFF on liver fibrosis in vivo and in vitro and investigate the mechanism from the perspective of the inflammatory response and extracellular signal-regulated kinase(ERK)phosphorylation.METHODS Common bile duct-ligated rats were used for in vivo experiments.Hepatic stellate cells-T6(HSC-T6)cells were used for in vitro experiments.Hematoxylin and eosin staining and Masson staining,biochemical assays,hydroxyproline(Hyp)assays,enzyme-linked immunoasorbent assay and western blotting were performed to evaluate the degree of liver fibrosis,liver function,the inflammatory response and ERK phosphorylation.The CCK8 assay,immunofluorescence and western blotting were applied to test the effect of GSFF on HSC-T6 cell activation and determine whether GSFF had an effect on ERK phosphorylation in HSC-T6 cells.RESULTS GSFF improved liver function and inhibited liver fibrosis in common bile ductligated rats after 3 wk of treatment,as demonstrated by histological changes,hydroxyproline assays and collagen I concentrations.GSFF alleviated inflammatory cell infiltration and reduced the synthesis of pro-inflammatory cytokines[tumor necrosis factor-α(TNF-α)and interlukin-1β]and NF-κB.In addition,GSFF decreased ERK phosphorylation.In vitro,GSFF inhibited the viability of HSC-T6 cells with and without transforming growth factorβ1(TGF-β1)stimulation and decreased the synthesis of collagen I.GSFF had the greatest effect at a concentration of 0.5μmol/L.GSFF inhibited the expression ofα-smooth muscle actin(α-SMA),a marker of HSC activation,in HSC-T6 cells.Consistent with the in vivo results,GSFF also inhibited the phosphorylation of ERK and downregulated the expression of NF-κB.CONCLUSION GSFF inhibited liver fibrosis progression in vivo and HSC-T6 cell activation in vitro.These effects may be related to an alleviated inflammatory response and downregulated ERK phosphorylation.

Formation of Superoxide Radical and Hydrogen Peroxide Enhanced by Trinitrotoluene in Rat Liver, Brain, Kidney, and Testicle in Vitro and Monkey Liver in Vivo

Trinitrotoluene (TNT) increased the formation of adrenochrome from adrenaline and the formation of formaldehyde from methanol in rat liver mitochondria and microsomes in vitro as well as in monkey liver mitrochondria and microsomes in vivo. The effects were more prominent at higher TNT concentrations. These findings indicate that TNT enhances the production of superoxide radicals (O_2^-) and hydrogen peroxide (H_2O_2). The production of O_2^- was more prominent in systems containing added TNT than in those containing added benzyl viologen. H_2O_2 production by mitochondria was more pronounced in the liver than in other organs, but its production by microsomes was more pronounced in the brain than in other organs. The results suggest that TNT undergoes cycling reduction which produces oxidative stress. 1989 Academic Press, Inc.

The Effect of a Dietary Supplement Spirulina and Bifidobacterium adolescentis on the Cholesterol-Lowering in Vitro and in Vivo

The survey＇s results showed a significant percentage of women and especially men having an above normal cholesterol. In order to help reduce excessive rate of blood cholesterol, we used a local strain of Spirulina （Tamanrasset, in south of Algeria） associated with probiotic bacteria （Bf adolescentis）. Experiments in vitro showed a significant degradation of total cholesterol by the combination of Spirulina and Bf adolescentis （74.5%） after 72 hours incubation at 37 ~C. A cholesterol is added to the standard diet mice in order to increase the total cholesterol for three lots. However, the rates of total cholesterol in mice receiving lower fermented milk with Bf adolescentis enriched by dry Spirulina. This shows that the decrease of cholesterol rate is closely related to the presence of Bf adolescentis and Spirulina. In vitro and in vivo results show, however, that it is possible to obtain a natural product （Spirulina） and a pseudo-strain lactic （Bf adolescentis） and to participate in the prevention of cardiovascular disease risk factor whose hand is cholesterol.

Can semipermeable membranes coating materials influence in vivo performance for paliperidone tri-layer ascending release osmotic pump tablet:In vitro evaluation and in vivo pharmacokinetics study

One purpose of this study was to develop a paliperidone(PAL)tri-layer ascending release pushepull osmotic pump(TA-PPOP)tablet which could meet the needs of clinical applications.And another purpose was to investigate whether different coating materials influenced in vivo performance of TA-PPOP.The ascending release mechanism of this trilayer delivery system on theory was elaborated.TA-PPOP was prepared by means of coating with cellulose acetate(CA)or ethyl cellulose(EC).Several important influence factors such as different core tablet compositions and different coating solution ingredients involved in the formulation procedure were investigated.The optimization of formulation and process was conducted by comparing different in vitro release behaviors of PAL.In vitro dissolution studies indicated that both the two formulations of different coating materials were able to deliver PAL at an ascending release rate during the whole 24 h test.The in vivo pharmacokinetics study showed that both self-made PPOP tablets with different coating had a good in vitro-in vivo correlation(IVIVC)and were bioequivalent with the brand product,which demonstrated no significant influence of the coating materials on the in vivo release acceleration of TA-PPOP.

THE SPECIFIC PHOTODYNAMIC EFFECTS OF MONOCLONAL ANTIBODIES CONJUGATED WITH HEMATOPORPHYRIN DERIVATIVE ON GASTRIC CANCER IN VITRO AND IN VIVO

Murine monoclonal antibody (MoAb) BB4.3, raised against the human gastric cancer cell line BGC823, was puriffied with Protein A-Sepharose CL-4B affinity chromatography and identified as IgG2a. It was then conjugated with a hematoporphyrin derivative (HPD) by using carbodiimide. The qualitative analysis of this conjugate showed that the amount of free HPD was negligible and there were no IgG aggregates among the conjugates. The conjugate retained both the antibody and photochemical activity of HPD.In vitro, the phototoxic effect of this HPD-BB4.3 conjugate on target cells was about 15 times higher than that of free HPD. The quality of selective photocytotoxicity was proven by the greater cytotoxi-city this conjugate showed than that of corresponding normal mouse IgG (NIgG) conjugated with HPD. It showed less cytotoxicity to colon cancer cell line B-80 (negative reaction to MoAb BB4.3) than to BGC825. Moreover, its cytotoxicity to BGC823 cells could be blocked specifically by excess BB4.3 antibody, but not by another MoAb 3G9, which combines with BGC823 at different binding sites from MoAb BB4.3.Nude mice inoculated with 2 × 10- BGC823 cells were given HPD-BB4.3, HPD, HPD-NIgG, HPD plus BB4.3 and PBS, respectively then exposed to light. Four out of six animals treated with the HPD-BB4.3 conjugate remained tumor-free for a long period. Although two developed tumors, there was a significant difference between the HPD-BB4.3-treated group and all the control groups in tumor induction time, tumor growth rate, and survival time (p<0.001). The HPD-BB4.3 conjugate inhibited the growth of established tumors by more than 40% in comparison with control groups (p<0.05).

Genetic Diversity of Intragenomic Rearrangement of Porcine Circovirus Type 2 in vitro and in vivo

We characterized the genome sequences of defective-interfering(DI) particle DNA of porcine circovirus type 2(PCV2) by sequencing and bioinformatics analyses. DI particles were both generated by serial passage of PCV2 in PK15 cells and obtained from sera of pigs with postweaning multisystemic wasting syndrome(PMWS). These subviral isolates ranged from 358 nt to 1 125 nt genomes. Investigating the complexity and diversity of PCV2 DI in vivo and in vitro can help elucidating the evolutionary history of PCV2.

Plant Regeneration of Ziziphus spina-christi by in vivo and in vitro Methods

Christ's thorn (Ziziphus spina-christi L. Desf.) is ecologically and economically important in Iran. Since it is a cross-pollinated plant with a wide range of genetic variability, an investigation was carried out to determine the best method for mass and clonal propagation. Three vegetative methods were examined, including 1) cutting, where shoots with a 22-25 cm length and three shoot diameters were treated with two culture media; 2) layering, where air layering and trench layering treatments consisted of three growth regulators at three levels of concentration; and 3) tissue culture with nodal segments bearing axillary buds that were removed from shoots of mature trees at different seasons. Several experiments were carried out to determine the best disinfectant chemical, the best culture method, the best season, the appropriate explant characteristics and media type. Rooting was successful only on the sandy beds for cuttings with more than (S.O mm diameter. For the trench layering method, only one specimen in one replicate rooted a very small root, while for the air layering method, only seedlings with growth regulator treatments rooted successfully. The best season for explant harvesting was determined as mid summer, and among the disinfecting treatments, Ca(OCI)2 at a concentration of 0 5 g kg-1 for 20 minutes was the best. There were no significant differences for shoot weight and length with a Murashige and Skoog (MS) medium with and without a hormone.

Stability of Cereal Crops to Drought and Saline Stress in Vivo and in Vitro

Complex research is devoted to basic non-specific stress-reactions caused by abiotic factors such as drought and salinity in vivo and in vitro. A comparative physiological, biochemical and cytogenetic analysis was performed and showed the peculiarities of growth and viability on various （cellular, tissular, organismic） levels of plants structural arrangement at stress conditions. Determined the parameters of the growth, ion balance, the content of free proline, superoxide dismutase activity and conducted the cytological studies. The commonness of cytological reactions of plant cells to abiotic stress was revealed. The considerable positive correlation relationships between growth of callus biomass and increases of primary roots number under abiotic stressess, between growth of callus biomass and capacity for survival of seedlings under osmotic stress were registered. Such correlation tells about comparability of stress tolerance valuation at different levels of plants structural arrangement. The considerable negative correlation between K~/Na~ ions relations and percent increase of free proline in calluses were showed. Physiological and biochemical indicators of abiotic stresses impact on plants cells and tissues, such as SOD activity and K^＋/Na^＋ ions correlation were noted. These indicators are effective as metabolic markers in the course of testing and selection of stress-resistant cereals in vivo and in vitro.

Cancer Chemopreventive Retinoids: Validation and Analysis of in Vivo and in Vitro Bioassay Results

Several natural and synthetic retinoids (vitamin-A derived analogies) were examined for their potential anti-cancer activity in both in vivo animal models and a novel in vitro human keratinocyte clonal growth bioassay system. The natural retinoids included all-trans-retinoic (RA), 13-cis-retinoic acid, 4-oxoretinoic acid, and retinol. Among the synthetic retinoids tested were all trans N-(4-hydroxy(phenyl)retinamide, 3-substituted oxoretinoic acids, and 13 cis-N-ethylretinamide. The animal models employed were: 1) vitamin A-deficient hamster tracheal organ assay (HTOC);2) the benzo(α)pyrene-induced squamous metaplasia in a hamster tracheal organ system (BP-HTOC);3) the mouse skin tumor promoter (TPA)-induced ornithine decarboxylase enzyme assay(ODC);4) the mouse skin papilloma (MPA) assay;and 5) a novel retinoid bioassay in which retinoids display IC50 values to inhibit clonal growth of NHK. All-trans-RA, 4-oxoretinoic acid and retinol were consistently more active than any of the synthetic derivatives in all bioassays tested. A statistical model was developed and significant positive correlations were found between: 1) ED50 values in the HTOC system and reduction in TPA-induced ODC enzyme activity;2) tumors per animal in the MPA bioassay and suppression of TPA-induced ODC activity;and 3) a positive correlation between suppression of tumors per animal in the MPA assay, and retinoid inhibition of keratinocyte clonal growth. Test retinoids, were tested for their capacity to inhibit the clonal growth of a squamous carcinoma cell line (SCC-25), which were found to be 2 - 3 logs less sensitive for each tested retinoid than the corresponding activity against NHK cells. Antineoplastic retinoid drugs were reviewed.

The Effects of Dietary Restriction and Aging on in Vivo and in Vitro Binding of Aflatoxin B_1 to Cellular DNA

Laboratory animals maintained on a reduced calorie but nutritionally adequate diet have extended life spans and lowered incidences of spontaneous and chemically induced cancers compared to ad libitum- fed counterparts. Many of the effects of dietary restriction on laboratory animals have been suggested to be related to a deceleration of the aging process. The inhibition of age-related changes in xenobiotic metabolizing enzyme activities by dietary restriction has previously been reported. Alterations of these enzyme activities may cause changes in metabolic activation of carcinogens and, therefore, carcinogen-DNA binding. DNA-repair capability has also been reported to be enhanced in diet-restricted rats. Using AFB1 as a model carcinogen, we have studied in vivo and in vitro hepatic AFB1 -DNA binding, demonstrating that dietary restriction (60% of ad libitum consumption) may decrease the metabolic activation of AFB1, and subsequently reduce AFB 1-DNA binding. Our preliminary results obtained from the AFB 1-DNA binding experiments in isolated hepatocytes suggest that the observed age-dependent reduction in AFB 1-DNA binding which may be attributed to a loss of metabolic activating capability was delayed in the diet-restricted rats.

One extraction tool for in vitro-in vivo extrapolation?SPME-based metabolomics of in vitro 2D,3D,and in vivo mouse melanoma models

Solid phase microextraction(SPME)in combination with high-resolution mass spectrometry was employed for the determination of metabolomic profile of mouse melanoma growth within in vitro 2D,in vitro 3D,and in vivo models.Such multi-model approach had never been investigated before.Due to the low-invasiveness of SPME,it was possible to perform time-course analysis,which allowed building time profile of biochemical reactions in the studied material.Such approach does not require the multiplication of samples as subsequent analyses are performed from the very same cell culture or from the same individual.SPME already reduces the number of animals required for experiment;therefore,it is with good concordance with the 3Rs rule(replacement,reduction,and refinement).Among tested models,the largest number of compounds was found within the in vitro 2D cell culture model,while in vivo and in vitro 3D models had the lowest number of detected compounds.These results may be connected with a higher metabolic rate,as well as lower integrity of the in vitro 2D model compared to the in vitro 3D model resulting in a lower number of compounds released into medium in the latter model.In terms of in vitro-in vivo extrapolation,the in vitro 2D model performed more similar to in vivo model compared to in vitro 3D model;however,it might have been due to the fact that only compounds secreted to medium were investigated.Thus,in further experiments to obtain full metabolome information,the intraspheroidal assessment or spheroid dissociation would be necessary.

Effect of temsirolimus on bladder cancer cells in vitro and in vivo

Objective To examine the effects of temsirolimus, an inhibitor of mammalian target of rapamycin,on bladder cancer cell lines T24 and BIU-87 in vitro and in vivo for purpose of evaluating the probability of mTOR targeted therapy for bladder cancer. Methods After

Adsorption,in vitro digestion and human gut microbiota regulation characteristics of three Poria cocos polysaccharides

Poria cocos(PC)is a famous traditional Chinese medicine(TCM)and a widely used healthcare ingredient,which has antiobesity,enhancing immunity and improving sleep effects.Traditionally,only water-soluble poria polysaccharide(WSP)is extracted and applied for clinical application,while insoluble polysaccharide(alkali-soluble poria polysaccharide,ASP)is discarded as herb residue.However,the whole PC has also been historically utilized as functional herbal food.Considering the beneficial role of dietary fiber and the traditional use of PC,ASP may also contribute substantially to the therapy function of PC.Compared to WSP,little attention has been paid to ASP and ASP modified product carboxymethyl poria polysaccharide(CMP)which has been used as an antitumor adjuvant drug.In this study,the oil,cholesterol,metal ions and polyphenols adsorption ability,in vitro simulated digestive and the gut microbiota fermentation characteristics of WSP,ASP and CMP were studied to evaluate the functional values of three P.cocos polysaccharides(PCPs).The results showed that all three PCPs had good adsorption capacity on cholesterol,polyphenols and metal ions(Cd^(2+)/Zn^(2+)/Mg^(2+)),among which ASP showed the highest capacity than WSP and CMP.The adsorption capacity of all three PCPs on heavy metal ions(Cd^(2+)/Zn^(2+))was stronger than that of non-heavy metal ions(Mg^(2+));The in vitro digestibility of all three PCPs was very low,but WSP was slightly higher than ASP and CMP;Moreover,the indigestible residue of all three PCPs could improve the richness and diversity of gut microbiota,among which ASP had the greatest influence.In general,ASP and CMP could significantly promote the proliferation of some probiotics and inhibit the growth of some harmful bacteria.The gut microbiota diversity of CMP was reduced,but the richness of probiotics,especially Parabacteroides distasonis was significantly enhanced compared with the ASP group,and the growth of harmful bacteria Klebsiella pneumoniae was inhibited after CMP treatment.The short-chain fatty acids(SCFAs)analysis results showed that all three PCPs could significantly promote the production of acetic acid,propionic acid and the total acid content compared with blank control group,and SCFAs producing activity was positively correlated with the proliferative capacity of probiotics.Taken together,the good adsorption characteristics and gut microbiota regulatory activity of ASP may lay foundation for its lipid-lowering and immune-improving function.Additionally,the probiotic effect of CMP and ASP indicated that except for only use the water extract of PC in clinic,CMP and ASP also can be used in healthcare to take full advantage of this valuable medicine.

Transfection and expression of exogenous gene in laying hens oviduct in vitro and in vivo

To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken ovalbumin gene. Jellyfish green fluorescence protein (EGFP) reporter gene and bacterial LacZ reporter gene were respectively inserted into the downstream of the 5'-regulatory region. The recombinants were named as pOVEGFP and pOVLacZ. Two transfer systems, in vitro and in vivo, were used to verify the function of the vector. In vitro, the plasmid DNA pOVEGFP and pEGFP-N1 were transfected respectively by the polyethyle-neimine procedure into the primary chicken oviduct epithelium (PCOE) and fibroblasts cells isolated from laying hens. In vivo, the recombinant vector pOVLacZ was injected into egg-laying hens via wing vein and the tissues were collected for RT-PCR analysis. The results showed that expression of pEGFP-Nl was achieved at low level in oviduct epithelial cells and at high level in fibroblasts, but that the recombinant vector was not expressed in both cells. RT-PCR analysis showed that the LacZ gene was transcribed in the oviduct, but not in the heart, liver, kidney and spleen of the injected hens. Accordingly, the β-galactosidase activity was only detected in the oviduct magnum (116.7 mU/ml) and eggs (16.47 mU/ml). These results indicated that the cloned regulation regions of chicken ovalbumin gene could drive exogenous gene expression specifically in the oviducts of hens. In vivo gene injection via wing vein may serve as a rapid production system of recombinant proteins in chicken eggs. In addition, the cultured primary oviduct cells from laying hens were not efficient temporary expression systems for analyzing the function of regulating elements of ovalbumin gene.

Studies on Antimicrobial Resistance Transfer In vitro and Existent Selectivity of Avian Antimicrobial-Resistant Enterobacteriaccae In vivo

Increasing antimicrobial resistance （AR） has become a severe problem of public health in the world, whereas control of the AR of bacteria will be based on investigation of the AR mechanism. Furthermore, understanding the existent selectivity of AR organisms from animals can prevent the emergence and diffusion of AR effectively. PCR amplifications of gyrA and parC genes have been performed for detecting fluoroquinolones-resistance （FR） genes. A conjugational transfer test has been carried out using a donor which is resistant to tetracycline （TE）, ampicillin （AMP）, sulfamethoxazole-trimethoprim （SXT）, and a recipient which is sensitive to TE, AMP, and SXT. The AR strains have been passed 20 passages. Two groups of chicken inoculated multi-AR Escherichia coli （E. coli） and multi-AR Salmonella, respectively, are mix-fed. The result shows that amino acid codons of Ser-83 and Asp-87 are mutations from gyrA and there are no mutations from parC genes in all the FR strains. Resistance to TE, AM, and SXT can transfer among E. coli and the conjugal transfer frequency of TE is 3×10^-7. AR can inherit in 20 passages at least. The multi-AR E. coli and Salmonella can be isolated from all chickens three days after inoculation but CIP-resistant strains decrease during the time run out and disappear at 23 days after inoculation. The results indicate that the mutations of gene gyrA are correlative with the FR phenotype. AR genes that are not connected to the chromosome can transfer horizontally and vertically. AR bacteria can diffuse quickly and eliminate naturally from the host if the chicken is not under the pressure of this antibiotic.