家养及大型养殖场鸡肠道微生物菌群及四环素耐药菌多样性的比较研究（英文）

本研究分析比较了农家家养鸡与大型养殖场鸡肠道菌群组成及抗四环素抗性基因在肠道菌群中的分布情况。通过454焦磷酸法对细菌16SrRNA V3区进行测序来分析肠道菌群的组成;用平板琼脂法筛选四环素耐药菌株,对其16SrRNA基因全长测序,并与RDP(Ribosomal Database Project)数据库比对进行菌种鉴定;聚合酶链反应(polymerase chain reaction,PCR)检测常见四环素耐药基因。家养鸡粪便中菌群香农多样性指数为5.321±0.590,养殖场鸡为4.398±0.440,前者显著大于后者(Mann-Whitney U test,P=0.008)。从家养鸡和养殖场鸡粪便中随机分离到69株和65株四环素耐药菌株,后者四环素耐药菌的种类多于前者。家养鸡较养殖场鸡的肠道菌群更具多样性,而抗生素抗性基因在养殖场鸡的肠道菌群中分布更广泛。结果表明,不同饲养方式对鸡的肠道菌群有影响,对抗生素抗性基因的分布也有一定影响。     

一对紫绀型先天性心脏病患儿肠道菌群动态变化及阴沟肠杆菌耐药性研究

[目的]观察一对紫绀型先天性心脏病双胞胎新生儿在接受手术及抗生素治疗前后的肠道菌群的动态变化,探究患儿肠道内阴沟肠杆菌在治疗期间耐药性如何发生改变。[方法]通过采集患儿在不同治疗阶段的粪便样本,进行培养组学和16S rRNA基因测序分析。同时,从双胞胎哥哥不同治疗阶段的粪便样本中分离得到10株阴沟肠杆菌进行体外药敏试验。[结果]培养组学结果显示使用抗生素导致术前粪便样本丰富度降低,仅可分离获得肠球菌属、不动杆菌属和肠杆菌属;长期医院环境暴露后,能分离出多种条件致病菌属,包括肠杆菌属、克雷伯菌属、不动杆菌属和假单胞菌属;母乳喂养4个月后,粪便菌群丰富度增加、构成发生改变,可分离获得乳杆菌属和双歧杆菌属等有益细菌。16S rRNA基因测序结果显示,在多种因素影响下不同治疗时间点α多样性指数和相对丰度最高的菌属不同。细菌耐药实验结果显示,治疗期间阴沟肠杆菌对哌拉西林逐渐产生耐药性。[结论]接受抗生素治疗、院内环境暴露和母乳喂养共同影响患儿治疗和康复期间的肠道菌群组成;在不同治疗阶段仅使用一种抗生素,也会导致阴沟肠杆菌对不同药物的耐药性发生改变。     

不同年龄大熊猫肠道可培养芽孢杆菌的多样性及部分功能特性分析

【背景】大熊猫的肠道内微生物类群丰富,种群结构与宿主的年龄、生存环境、季节变化等因素有关,其中年龄是影响肠道菌群组成的重要因素之一。【目的】以不同年龄阶段的大熊猫粪便为研究对象,旨在了解不同年龄大熊猫肠道内芽孢杆菌的多样性,探究大熊猫肠道芽孢杆菌种类与年龄段之间的关系,并为优良益生菌剂的开发提供菌种资源。【方法】用稀释涂布法分离大熊猫粪便中的芽孢杆菌,对分离的菌株进行BOXA1R-PCR、16S rRNA基因系统发育及主成分分析,揭示大熊猫肠道中可培养芽孢杆菌的多样性;采用对峙生长法和药敏纸片琼脂扩散法分别检测菌株的抗菌能力和药敏性。【结果】从大熊猫粪便中共分离出90株芽孢杆菌,基于BOXA1R-PCR分析菌株的遗传多样性并从中选取41株代表菌株,经16Sr RNA基因测序分析后,结果显示归属于枯草芽孢杆菌(Bacillus subtilis)、萎缩芽孢杆菌(Bacillus atrophaeus)、贝莱斯芽孢杆菌(Bacillus velezensis)、甲基营养型芽孢杆菌(Bacillusmethylotrophicus)、解淀粉芽孢杆菌(Bacillusamyloliquefaciens)和短小芽孢杆菌(Bacillus pumilus)6个种;主成分分析结果表明大熊猫肠道芽孢杆菌种群组成与年龄存在一定的相关性。所有的供试菌株都具有纤维素降解潜力,大部分菌株对病原菌具有不同程度的抑制作用;除对青霉素具有耐药性外,供试菌株对常见的抗生素耐受性低。【结论】年龄是影响大熊猫肠道芽孢杆菌分布的重要因素,成年大熊猫肠道芽孢杆菌种类多样性最丰富,这为大熊猫益生菌制剂的开发提供了菌种资源。     

甘肃省武威市胃癌患者肠道菌群分布

目的 通过比较甘肃省武威市胃癌患者和健康对照人群肠道菌群的分布,探讨胃癌患者肠道菌群的变化与胃癌发生发展的关系,并寻找可能作为该地区胃癌患者的潜在生物标志物.方法 收集24例胃癌患者和24例健康对照人群的粪便样本,提取DNA,采用16S rRNA基因高通量测序进行肠道菌群分析.结果 分析2组研究对象肠道菌群的Alpha...     

不同养殖场林麝肠道微生物组成和功能的差异

肠道疾病是养殖林麝(Moschus berezovskii)常见疾病。动物肠道微生物伴随宿主进化并与胃肠道构成了复杂的微生态系统。为探究不同饲养环境对圈养林麝肠道微生物组成和功能的影响,本研究对采自国内5个不同养殖场的215份粪便样品进行了16S rRNA基因高通量测序,对比分析不同养殖场林麝肠道微生物组成、多样性和功能的差异。结果显示,厚壁菌门和拟杆菌门是未喂食复合益生菌的祁连县养殖场林麝肠道菌群的绝对优势菌门,而喂食复合益生菌的甘肃两当县和陕西凤县的4家养殖场林麝肠道菌群的绝对优势菌门为厚壁菌门和变形菌门。不同养殖场林麝肠道菌群组成、优势菌门、优势菌属、潜在致病菌、代谢及疾病相关功能均有显著差异。祁连县养殖场林麝肠道微生物的α多样性和疾病相关功能表达量显著低于其他养殖场,并以肠型2为主,其主导菌为厚壁菌门、UCG-005和拟杆菌属;两当县和凤县的4家养殖场林麝肠道菌群潜在致病菌相对丰度较低。本研究推测食物组成差异可能是导致不同养殖场林麝肠道微生物差异的主要因素,复合益生菌的使用可能是导致α多样性和潜在致病菌下降的重要因素。该结果可为林麝的人工养殖和有效管理提供科学依据,也对人工饲养...     

高通量 16S rRNA 标签测序法比较人与不同动物肠道微生物组多样性

收集人、大猪、小猪、大鼠、小鼠以及鸡五个不同个体的粪便样品 , 每个个体取两个平行样 , 提取总 DNA;PCR 扩增 , 获得 16S rRNA V6 标签片段 ; Illumina 测序 ; 经 BIPES 以及 QIIME 分析并比较菌群结构及多样性。研究结果发现 , 不同物种之间肠道菌群差异较大。五类物种肠道菌群均以厚壁菌门、拟杆菌门、以及变形菌门为主 , 但鸡的厚壁菌门显著减少 , 而变形菌门显著增加。从α多样性角度来看 , 猪肠道菌群种属丰富度及 Shannon 指数均显著高于人及鸡肠道菌群。从β多样性角度 , 尽管不同人之间的肠道菌群相似度较低 , 但不同物种之间相比较 , 小猪与大鼠肠道菌群与人相似性高于小鼠和鸡肠道菌群。与人类相比 , 小猪的肠道微生物组最相近 , 而鸡的肠道菌群相似度最低。     

市售酸奶中乳酸菌的鉴定与耐药性

[目的]检测市售酸奶中乳酸菌的种类及其耐药情况.[方法]收集10种来自5个不同厂家的酸奶,通过细菌基因组重复基因外回文序列-PCR (repetitive extragenic palindromic-PCR,rep-PCR)结合16S rRNA同源性分析的方法对分离的乳酸菌进行基因分型和菌种鉴定.利用药敏纸片扩散法(K-B法)对分离的乳酸菌进行针对7种抗生素的药敏实验,用PCR特异性扩增结合测序的方法检测每个样品中不同基因型菌株的耐药基因(包括红霉素耐药基因erm A、erm B和四环素耐药基因tetM、tetK、tet S、tetQ、tetO、tetL、tetW).[结果]10种市售酸奶中分离到100株乳酸菌.其中,德氏乳杆菌保加利亚亚种(Lactobacillus delbrueckii ssp.bulgaricus)23株,干酪乳杆菌(Lactobacillus casei)26株,嗜热链球菌(Streptococcus thermophilus)30株,嗜酸乳杆菌(Lactobacillus acidophilus)5株,植物乳杆菌(Lactobacillus plantarum)6株,副干酪乳杆菌(Lactobacillus paracasei)10株.药敏实验发现所有100株乳酸菌均对链霉素和庆大霉素耐药,42株对万古霉素耐药,没有菌株对头孢氨苄,四环素,红霉素以及土霉素耐药.在28株经过16S rRNA测序的乳酸菌中检测到5种不同的耐药基因,在8株乳酸菌中检测到erm B基因,4株检测到tetK基因,2株菌检测到tetL基因,4株菌检测到tet M基因,2株菌检测到tet O基因,没有检测到erm A,tet S,tet Q,tet W基因.28株乳酸菌中有15株(53.57%)检测到耐药基因,其中有4株L delbrueckii ssp.bulgaricus检测到2-3种不同的耐药基因.[结论]本研究在市售酸奶中除了检测到商品标签上标注的L.delbrueckii ssp.bulgaricus和S.thermophilus以外,还检测到商标上没有标注的乳酸菌;作为常用发酵剂的德氏乳杆菌保加利亚亚种和嗜热链球菌更容易检测到耐药基因;分离得到的乳酸菌均对红霉素和四环素敏感却检测到相应的耐药基因,再一次证明了没有耐药表型的菌株也可能携带耐药基因.     

携带armA基因的铜绿假单胞菌耐药性及基因周边环境

目的了解多重耐药(MDR)铜绿假单胞菌armA基因与可移动遗传元件的携带情况及其相关性;分析armA基因的周边环境,探讨armA基因转移的可能机制。方法收集MDR铜绿假单胞菌98株,琼脂稀释法测定MIC,PCR方法检测16S rRNA甲基化酶基因armA、I型整合子、可移动元件IS26及重要耐药基因侧翼基因环境,测序并拼接PCR产物明确耐药基因座位排列,并对armA基因进行周边序列分析。结果 98株MDR铜绿假单胞菌检出5株armA基因PCR扩增阳性,携带armA基因的菌株对庆大霉素和阿米卡星全耐药;检出20株携带I型整合子,17株携带可移动元件IS26;armA基因扩增阳性的菌株均携带I型整合子和IS26;序列测序显示armA定位于Tn1548相关区域,位于插入序列ISCR1的下游,该序列含多种移动元件。结论大连市氨基糖苷类高水平耐药基因armA广泛分布在MDR铜绿假单胞菌中,均对庆大霉素和阿米卡星高度耐药;该基因定位在转座子Tn1548的质粒上,提示16S rRNA甲基化酶基因armA的广泛播散可能是可移动元件ISCR1-armA-IS26结构参与其中。     

不同程度颅脑疾病患者肠道菌群分析

目的 检测不同程度颅脑疾病患者肠道菌群分布情况,探讨患者肠道菌群分布与疾病的关系,为该类患者的治疗提供参考。方法 通过MiSeq PE2x300 bp高通量测序技术对21例重度颅脑损伤患者(G1组,GCS≤8分)和14例轻度颅脑损伤患者(G2组,GCS>8分)粪便样本中细菌的16S rRNA基因V3-V4可变区进行定性分析,并对两组患者肠道菌群进行生物学分类比较、多样性分析和组间物种差异分析。结果 两组患者肠道菌群多样性差异无统计学意义(均P>0.05)。G1组患者肠道厌氧球菌属(Anaerococcus)、埃希菌-志贺菌属(Escherichia-Shigella)、链球菌属(Streptococcus)、肠球菌属(Enterococcus)、棒状杆菌属(Corynebacterium)数量高于G2组(均P<0.05)。结论 颅脑疾病患者肠道菌群紊乱,其肠道菌群丰度及多样性显著下降,有害菌数量升高,有益菌数量下降。     

圈养大熊猫肠道乳酸菌分离及益生特性

【背景】大熊猫数量稀少、繁殖困难,在其生长过程中极易感染消化系统疾病甚至死亡。【目的】分析圈养大熊猫肠道可培养乳酸菌的群落结构与功能,筛选具有益生特性的乳酸菌菌株,为大熊猫消化系统疾病的预防和肠道微生物研究提供理论参考及菌种资源。【方法】采用3种培养基分离大熊猫粪便中的乳酸菌,通过革兰氏染色镜检和过氧化氢试验对分离的菌株进行初步鉴定,基于BOXA1R-PCR图谱遗传多样性选取代表菌株进行16S rRNA基因测序分析并进行主成分分析(principal component analysis, PCA),同时分析乳酸菌菌株安全性和益生特性。【结果】通过初步鉴定共分离获得58株乳酸菌,根据BOXA1R-PCR结果挑选20株菌进行测序,结果显示20株菌分属于明串珠菌属(Leuconostoc)、肠球菌属(Enterococcus)、魏斯氏菌属(Weissella)和链球菌属(Streptococcus)这4个属,主成分分析结果表明不同年龄段的大熊猫肠道乳酸菌群落结构存在差异。20株乳酸菌均不溶血,17株乳酸菌对11种抗生素均敏感;11株菌耐pH值2.0酸性条件,14株菌对0.3%的胆盐具有良好...     

Abrupt temporal fluctuations in the chicken fecal microbiota are explained by its gastrointestinal origin

One of the main challenges in understanding the composition of fecal microbiota is that it can consist of microbial mixtures originating from different gastrointestinal (GI) segments. Here, we addressed this challenge for broiler chicken feces using a direct 16S rRNA gene-sequencing approach combined with multivariate statistical analyses. Broiler feces were chosen because of easy sampling and the importance for pathogen transmission to the human food chain. Feces were sampled daily for 16 days from chickens with and without a feed structure-induced stimulation of the gastric barrier function. Overall, we found four dominant microbial phylogroups in the feces. Two of the phylogroups were related to clostridia, one to lactobacilli, and one to Escherichia/Shigella. The relative composition of these phylogroups showed apparent stochastic temporal fluctuations in feces. Analyses of dissected chickens at the end of the experiment, however, showed that the two clostridial phylogroups were correlated to the microbiota in the cecum/colon and the small intestine, while the upper gut (crop and gizzard) microbiota was correlated to the lactobacillus phylogroup. In addition, chickens with a stimulated gizzard also showed less of the proximate GI dominating bacterial group in the feces, supporting the importance of the gastric barrier function. In conclusion, our results suggest that GI origin is a main determinant for the chicken fecal microbiota composition. This knowledge will be important for future understanding of factors affecting shedding of both harmful and beneficial gastrointestinal bacteria through feces.     

禽畜养殖粪便中多重抗生素抗性细菌研究

通过对新乡地区8家养猪场和11家养鸡场饲喂抗生素情况的调研,发现头孢氨苄、阿莫西林、卡那霉素、庆大霉素等4种抗生素是该地区被普遍使用的兽药抗生素。通过多点取样法和微生物培养技术对3家养鸡场和3家养猪场不同养殖时期的粪便进行单一抗生素和多重抗生素抗性细菌的检测,结果表明养鸡场堆置1周的粪便中抗头孢氨苄的细菌比例最高,达到65.90%,对所研究的3种和4种抗生素同时抗性的比例高达8.60%—12.51%和9.73%,明显高于饲喂中药的对照养鸡场样本检测结果(0.02%—2.73%和0.12%)。养猪场堆置1周的粪便中检测到抗头孢氨苄的细菌比例也是最高,达到49.12%上,但养猪场粪便中多重抗生素抗性细菌的比例明显低于养鸡场。同时研究发现,在两种养殖场中,幼龄期粪便中检测到的多重抗性细菌比例明显高于成熟期粪便,这可能与养殖过程中鸡、猪在幼龄期由于防病和促生长等因素而同时大剂量使用多种抗生素有关。     

Extraintestinal sporozoites of chicken Eimeria in chickens and turkeys

Oocysts were found in the feces of chickens (recipients) dosed orally with whole blood, liver, lung, or heart homogenates from chickens and turkeys (donors) inoculated 3 and 4 days previously with a mixture of 3.5 X 10(6) oocysts of chicken Eimeria. No oocysts were found in the feces of recipients given spleen homogenates from these same chickens and turkeys and none were found in the feces of recipients given similar material from uninoculated donors. Intracellular sporazoites were found in the peripheral blood of a turkey inoculated with chicken Eimeria. The results indicate that a small number of sporozoites are capable of invading and surviving for at least 4 days in the peripheral blood of chickens and turkeys.     

Occurrence of ceftriaxone-resistant commensal bacteria on a dairy farm and a poultry farm

Approximately 40 samples of animal feces, drinking water, feed, bedding, pine wood shavings, compost, and manure slurry were collected from two animal research farms (one dairy and one poultry) and analyzed for ceftriaxone-resistant bacteria. Our study revealed that the total percentage of aerobic bacteria with reduced susceptibility to ceftriaxone (minimal inhibitory concentration (MIC) > or = 16 micro g/mL) ranged from 0.9% to 10.8% in dairy feces and from 0.05% to 3.93% in chicken feces. The percentages of ceftriaxone-resistant bacteria (MIC > or = 64 micro g/mL) were in the range of 0.01% - 2.3% in dairy feces and 0.01% - 0.79% in chicken feces. Environmental samples contained a wide range of ceftriaxone-resistant bacterial populations. Among those environmental samples, fresh pine wood shavings used as chicken bedding contained the highest percentages (41.5%) of ceftriaxone-resistant bacteria, as determined by a plating method. A total of 105 ceftriaxone-resistant (MIC > or = 128 micro g/mL) bacterial isolates were isolated from the above samples and tested for resistance to nine antibiotics: ampicillin, ceftriaxone, streptomycin, kanamycin, gentamicin, chloramphenicol, tetracycline, ciprofloxacin, and nalidixic acid. The most prevalent resistance pattern (34.3%) among isolates included resistance to all nine antibiotics. Results from this study suggest that ceftriaxone-resistant bacteria exist in farm environments, and the ceftriaxone resistance was frequently associated with resistance to multiple antibiotics. Environmental sources such as pine wood shavings used as bedding can be a potential reservoir for transmitting the multidrug-resistant bacteria.     

Avian leukosis virus subgroup J infection influences the gut microbiota composition in Huiyang bearded chickens

Avian leukosis virus (ALV) poses a major threat to poultry. The chicken gut microbiota plays critical roles in host performance, health and immunity. However, the effect of viral infection on the microbiota of Chinese local chickens is not well understood. In this study, we performed high-throughput 16S rRNA gene sequencing and evaluated the gut microbiota profiles using faeces from ALV subgroup J (ALV-J)-infected and healthy Huiyang bearded chickens (Chinese local chickens). At the phylum level, ALV-J infection mainly increased the abundance of Bacteroidetes and Proteobacteria and decreased that of Firmicutes. An analysis at the order, family and genus levels showed that the abundance of Lactobacillales, Lactobacillaceae and Lactobacillus was the highest in normal chicken faeces, accounting for 89·07%, 86·47% and 86·46%, respectively, of phylotypes. Moreover, samples from ALV-J-infected chickens were enriched with Bacteroidales, Clostridiales, Bacteroidaceae, Ruminococcaceae, Lachnospiraceae and Bacteroides. Our findings highlight that ALV-J infection alters the gut microbiota and disrupts the host–microbial homeostasis in chickens, which may be involved in the pathogenesis of ALV-J infection.     

Development and assessment of a real-time pcr assay for rapid and sensitive detection of a novel thermotolerant bacterium, Lactobacillus thermotolerans, in chicken feces

A new real-time PCR assay was successfully developed using a TaqMan fluorescence probe for specific detection and enumeration of a novel bacterium, Lactobacillus thermotolerans, in chicken feces. The specific primers and probe were designed based on the L. thermotolerans 16S rRNA gene sequences, and these sequences were compared to those of all available 16S rRNA genes in the GenBank database. The assay, targeting 16S rRNA gene, was evaluated using DNA from a pure culture of L. thermotolerans, DNA from the closely related bacteria Lactobacillus mucosae DSM 13345(T) and Lactobacillus fermentum JCM 1173(T), and DNA from other lactic acid bacteria in quantitative experiments. Serial dilutions of L. thermotolerans DNA were used as external standards for calibration. The minimum detection limit of this technique was 1.84 x 10(3) cells/ml of an L. thermotolerans pure culture. The assay was then applied to chicken feces in two different trials. In the first trial, the cell population was 10(4) cells/g feces on day 4 and 10(5) cells/g feces on days 11 to 18. However, cell populations of 10(6) to 10(7) cells/g feces were detected in the second trial. The total bacterial count, measured by 4',6-diamidino-2-phenylindole (DAPI) staining, was approximately 10(11) cells/g feces. These results suggest that in general, L. thermotolerans is a normal member of the chicken gut microbiota, although it is present at relatively low levels in the feces.     

Antimicrobial resistance in generic Escherichia coli isolates from wild small mammals living in swine farm, residential, landfill, and natural environments in southern Ontario, Canada

To assess the impacts of different types of human activity on the development of resistant bacteria in the feces of wild small mammals, we compared the prevalences and patterns of antimicrobial resistance and resistance genes in generic Escherichia coli and Salmonella enterica isolates from fecal samples collected from wild small mammals living in four environments: swine farms, residential areas, landfills, and natural habitats. Resistance to antimicrobials was observed in E. coli isolates from animals in all environments: 25/52 (48%) animals trapped at swine farms, 6/69 (9%) animals trapped in residential areas, 3/20 (15%) animals trapped at landfills, and 1/22 (5%) animals trapped in natural habitats. Animals trapped on farms were significantly more likely to carry E. coli isolates with resistance to tetracycline, ampicillin, sulfisoxazole, and streptomycin than animals trapped in residential areas. The resistance genes sul2, aadA, and tet(A) were significantly more likely to be detected in E. coli isolates from animals trapped on farms than from those trapped in residential areas. Three S. enterica serotypes (Give, Typhimurium, and Newport) were recovered from the feces of 4/302 (1%) wild small mammals. All Salmonella isolates were pansusceptible. Our results show that swine farm origin is significantly associated with the presence of resistant bacteria and resistance genes in wild small mammals in southern Ontario, Canada. However, resistant fecal bacteria were found in small mammals living in all environments studied, indicating that environmental exposure to antimicrobials, antimicrobial residues, resistant bacteria, or resistance genes is widespread.     

Beta-Lactamase Producing Escherichia coli Isolates in Imported and Locally Produced Chicken Meat from Ghana

The use of antibiotics in food animals is of public health concern, because resistant zoonotic pathogens can be transmitted to humans. Furthermore, global trade with food may rapidly spread multi-resistant pathogens between countries and even continents. The purpose of the study was to investigate whether imported chicken meat and meat from locally reared chicken are potential sources for human exposure to multi resistant Escherichia coli isolates. 188 samples from imported and locally produced chicken meat were sampled and analyzed. 153 bacteria isolates were successfully cultured and identified as E. coli using MALDI-ToF. Of these 109 isolates were from meat whereas the remaining 44 were isolated from the cloaca of locally reared live chickens. Antimicrobial susceptibility test was done on the identified E. coli isolates. Additionally, beta-lactamases production (ESBL and/or AmpC) were phenotypically confirmed on all isolates showing resistance to cefpodoxime. Beta-lactamase producing (BLP) E. coli meat isolates were further genotyped. Antimicrobial resistance to four antibiotic markers with highest resistance was detected more frequently in isolates from local chickens compared to imported chickens (tetracycline 88.9% vs. 57.5%, sulphonamide 75.0% vs. 46.6%, ampicillin 69.4% vs. 61.6% and trimethoprim 66.7% vs. 38.4%). Beta-lactamase production was found in 29 E. coli meat isolates, with 56.9% of them being multiple drug resistant (≥ 3). The predominant phylogroup identified was B1 followed by A and D, with similar distribution among the isolates from meat of locally reared chickens and imported chickens. Beta-lactamase producing genotype bla _CTX-M-15 (50%; 10/20) was the most frequently drug resistant gene detected. More BLP E. coli isolates were found in imported chicken meat compared to locally reared chickens, demonstrating that these isolates may be spreading through food trade. In conclusion, both imported and locally produced chicken meats are potential sources for human exposure to BLP E. coli.     

Synthesis of low molecular weight alginic acid nanoparticles through persulfate treatment as effective drug delivery system to manage drug resistant bacteria

The purpose of this study was to prepare low molecular weight alginic acid (LMWA) nanoparticles by cation-induced, controlled gelification of depolymerized alginic acid for effective drug delivery to drug resistant bacteria. The depolymerization reaction was performed using potassium persulfate oxidation at an optimized condition. The optimized conditions for depolymerization were anticipated to be 37°C, pH 4, 2 days reaction time, and a 0.075 M concentration of potassium persulphate containing 0.001 M silver nitrate in the final reaction mixture. Gel permeation chromatography showed depolymerized alginic acid had an average molecular weight of 20.95 ± 0.49 kDa. Depolymerized alginic acid was also characterized for its structural integrity by X-ray diffraction, nuclear magnetic resonance, and Fourier transform spectroscopy. Depolymerized alginic acid was used to prepare low molecular weight nanoparticles with a particle size of 54 ± 0.41 nm, and a zetapotential of −32.2 ± 3.91 mV. The nanoparicles were then subjected to tetracycline loading. In vitro drug loading and drug release efficiencies after 100 h were determined to be 66.56 ± 1.88 and 61.8 ± 0.141%, respectively. Finally, the minimal inhibitory concentration and a putative mode of action for the tetracycline nanoparticles were determined using tetracycline resistant bacteria, Escherichia coli XL-1.     

Characterization of intestinal microbiota and response to dietary virginiamycin supplementation in the broiler chicken

The inclusion of antibiotic growth promoters, such as virginiamycin, at subtherapeutic levels in poultry feeds has a positive effect on health and growth characteristics, possibly due to beneficial effects on the host gastrointestinal microbiota. To improve our understanding of the chicken gastrointestinal microbiota and the effect of virginiamycin on its composition, we characterized the bacteria found in five different gastrointestinal tract locations (duodenal loop, mid-jejunum, proximal ileum, ileocecal junction, and cecum) in 47-day-old chickens that were fed diets excluding or including virginiamycin throughout the production cycle. Ten libraries (five gastrointestinal tract locations from two groups of birds) of approximately 555-bp chaperonin 60 PCR products were prepared, and 10,932 cloned sequences were analyzed. A total of 370 distinct cpn60 sequences were identified, which ranged in frequency of recovery from 1 to 2,872. The small intestinal libraries were dominated by sequences from the Lactobacillales (90% of sequences), while the cecum libraries were more diverse and included members of the Clostridiales (68%), Lactobacillales (25%), and Bacteroidetes (6%). To assess the effects of virginiamycin on the gastrointestinal microbiota, 15 bacterial targets were enumerated using quantitative, real-time PCR. Virginiamycin was associated with increased abundance of many of the targets in the proximal gastrointestinal tract (duodenal loop to proximal ileum), with fewer targets affected in the distal regions (ileocecal junction and cecum). These findings provide improved profiling of the composition of the chicken intestinal microbiota and indicate that microbial responses to virginiamycin are most significant in the proximal small intestine.