畜禽养殖环境中抗生素抗性基因污染的研究进展

畜禽养殖中抗生素不当使用导致的抗生素抗性基因(ARGs)污染问题日益严重,引起了国际社会的广泛关注。养殖环境中的ARGs不但可通过吸附、解吸、迁移等途径发生跨介质扩散,也可通过水平基因转移(HGT)使耐药性跨菌属传播,甚至通过食物链传递给人类,对公众健康造成潜在威胁。然而,目前缺少以大健康视角总结分析畜禽养殖环境ARGs赋存特征、关键环境过程和阻控技术的研究,导致对ARGs传播风险的评估和污染阻控对策的制定难以有效开展。本文在分析不同国家、畜种、环境介质中ARGs污染水平差异的基础上,阐述了ARGs在畜禽养殖环境中的关键环境过程和主要影响因素,重点论述了当前畜禽养殖业ARGs的污染阻控技术,最后结合大健康的方法理念,指出了目前研究存在的不足和亟需加强的方面,尤其强调了明确ARGs时空演变规律和环境过程机制、开发绿色高效的污染阻控技术的重要性与紧迫性,以期为畜禽养殖环境中ARGs健康风险评估和污染阻控技术的开发提供理论依据。     

多重耐药菌在人类、动物和环境的耐药和传播机制

抗生素等抗菌药物的滥用在全球范围内造成了多重耐药菌的传播。多重耐药菌(Multidrug resistant organisms,MDRO)以及耐药基因(Antibiotic resistance genes,ARGs)可在人类、动物和环境之间进行传播,尤其是ARGs可以通过水平转移的方式在同种属或者不同种属的菌群之间进行传播,使得细菌耐药问题日益严重,耐药机制趋于复杂,疾病治疗更加困难,对人类公众健康造成严重的威胁。因此抗生素等抗菌药物的使用应加以规范。     

肺炎克雷伯菌基因组可移动遗传元件的研究进展

肺炎克雷伯菌是目前临床上最主要的耐药致病菌之一,对人类健康造成了很大威胁。近年来,细菌耐药成为治疗肺炎克雷伯菌感染的主要难题,尤其是高毒力、高耐药性肺炎克雷伯菌的出现对临床工作造成了巨大挑战,而研究表明其耐药基因和毒力基因主要由可移动遗传元件携带而传播。因此,为了更好地认识及防控肺炎克雷伯菌感染,本文对肺炎克雷伯菌基因组中几种常见可移动元件(包括质粒、前噬菌体、插入序列等)及其与肺炎克雷伯菌耐药性和致病性之间的关系进行了综述,并阐述了其在耐药基因和毒力基因传播过程中的作用机制。     

临床分离肺炎克雷伯菌Ⅰ类整合子的结构与功能

为了探索细菌多重耐药性的产生和播散的分子机制, 文章对2002～2007年间179株临床分离的肺炎克雷伯菌进行耐药性、I类整合子可变区基因盒结构以及基因盒携带的耐药性基因进行分段克隆和耐药性功能测定。结果显示：65.9%(118/179)的肺炎克雷伯菌表现出对至少两种以上的抗生素(主要为β-内酰胺类、氨基糖苷类和喹诺酮类抗菌药物)的耐药性; 36.3%(65/179)的菌株检出单条或者双条I类整合子基因盒条带; 对整合子阳性组与阴性组的耐药率进行比较发现, 除氨基糖苷类、喹诺酮类和复方新诺明等药物的耐药性存在显著性差异(P<0.01)外, 其余药物的差异不显著; 共发现15种耐药基因构成形式的整合子基因盒, 其中以dfrA17-aadA5最为多见, 实验证明整合子可由接合转移耐药性质粒携带; 对整合子基因盒(dhfr17-orfF-aadA2)分段克隆的耐药性功能研究发现, 3个克隆重组子(pET28a-dhfr17、pET28a-dhfr17-orfF和pET28a-dhfr17-orfF-aadA2)对复方新诺明的抗性(MIC值)均为256 µg/mL, 重组子pET28a-dhfr17-orfF与重组子pET28a-dhfr17对链霉素的抗性无明显区别, 和受体菌一样MIC值均为8 µg/mL, 而pET28a-dhfr17-orfF-aadA2对链霉素的抗性则明显提高, MIC值为256 µg/mL。结果表明, I类整合子在肺炎克雷伯菌中较常见, 携带氨基糖苷类和甲氧苄啶类的耐药基因盒在数量上占优势, 且整合子携带的耐药基因具有耐药性功能, 位于可水平转移耐药性质粒的耐药性基因相关的整合子对病原菌耐药性播散具有重要意义。 目的基因     

新型抗生素抗性基因传播元件ISCR及其生态风险

抗生素抗性基因(antibiotic resistance genes, ARGs)作为一种新型的环境污染物,成为多个学科关注的焦点.其在不同环境介质中的扩散和传播具有极大的环境危害性,对人类健康造成严重威胁.插入序列共同区(insertion sequence common region, ISCR),是一种新发现的抗性基因传播元件,因其特殊的遗传结构,能够通过滚环复制及同源重组等机制移动邻近的任何DNA序列,是ARGs在不同DNA分子或不同种属细菌间水平传播的高效媒介.目前世界上发现了27种ISCR元件.大量间接证据表明,ISCR可能与许多耐药基因的移动和扩散有关,特别是多重耐药性(multiple drug resistance, MDR)形成与传播.因此,ISCR很可能是抗生素抗性基因在环境中扩散传播的关键因子.本文就ARGs水平传播、ISCR结构特征、ISCR种类及其相关ARGs及其研究方法等进行综述,并揭示ISCR元件可能的生态风险,提出了今后的研究重点,以期为今后深入开展相关研究打下基础.     

气溶胶中抗生素抗性基因研究进展

抗生素的不合理使用导致细菌耐药问题日趋严峻,给人类健康造成巨大威胁。学者们对抗生素抗性菌和抗生素抗性基因(antibiotic resistance genes, ARGs)在多种环境介质中的环境行为开展了大量研究。气溶胶作为ARGs的潜在储存库,是抗生素抗性基因在环境中的重要传播途径之一。目前缺乏对其来源、传播、人类接触和健康风险系统性的梳理。本文针对人类生活功能场所、养殖场、城市污水处理厂和医院等4类气溶胶研究的典型场所,重点综述了上述4类典型场所中气溶胶ARGs的来源、传播途径及对人体的暴露和对健康的危害,为气溶胶中ARGs的预防和控制提供参考。     

土壤噬菌体及其介导的抗生素抗性基因水平转移研究进展

土壤中抗生素耐药性的扩散对全球的公共卫生和食品安全造成威胁,严重挑战人类感染类疾病的预防与治疗。噬菌体介导的抗生素抗性基因(ARGs)的水平转移是环境中抗性基因扩散的重要机制。但是,噬菌体对土壤环境中抗性基因传播的贡献尚未见报道。本文综述了土壤环境中噬菌体的分布特征与影响因子,总结了纯化和富集土壤噬菌体的主要研究方法;同时阐述了土壤环境中噬菌体介导抗性基因水平转移的作用机制等相关研究进展,并提出了土壤噬菌体研究领域尚未解决的一些科学问题。本综述将有助于进一步深入理解噬菌体在抗性基因水平传播中的重要生态角色,为制定相关管理政策以减缓抗生素抗性基因污染问题提供基础。     

环境中抗生素抗性基因与Ⅰ型整合子的研究进展

抗生素抗性基因(Antibiotic resistance genes,ARGs)作为一种新型污染物在不同环境中广泛分布、来源复杂,对生态环境和人类健康造成了很大的潜在风险。同时,Ⅰ型整合子(Int Ⅰ)介导的ARGs水平转移是环境中微生物产生耐药性的重要途径,Ⅰ型整合子整合酶基因(intI1)与ARGs丰度在环境中表现出了较高的正相关性,Int Ⅰ可以作为标记物在一定程度上反映ARGs在环境中的迁移转化规律和人类活动影响程度。本文介绍ARGs与Int Ⅰ在环境中的来源与分布,总结Int Ⅰ介导的ARGs迁移转化机制以及相关研究方法,并展望未来的研究发展趋势。     

环境微生物的抗生素抗性和抗性组

环境和医疗实践中广泛存在的细菌抗生素抗性已经成为食品安全和人类健康领域的主要威胁。近年来的研究表明,病原菌主要通过水平基因转移而不是基因突变获得抗性,大量的研究支持病原微生物抗生素抗性基因的环境来源。系统论述了环境微生物抗生素抗性起源、进化及病原菌抗性基因与环境抗生素抗性组相互之间的交叉传播和水平转移机制,介绍了近年来环境微生物抗生素抗性组生态和进化生物学研究的最新进展和方法学应用。     

环境中抗生素抗性基因与I型整合子的研究进展

抗生素抗性基因(Antibiotic resistance genes,ARGs)作为一种新型污染物在不同环境中广泛分布、来源复杂,对生态环境和人类健康造成了很大的潜在风险。同时,Ⅰ型整合子(Int Ⅰ)介导的ARGs水平转移是环境中微生物产生耐药性的重要途径,Ⅰ型整合子整合酶基因(intI1)与ARGs丰度在环境中表现出了较高的正相关性,Int Ⅰ可以作为标记物在一定程度上反映ARGs在环境中的迁移转化规律和人类活动影响程度。本文介绍ARGs与Int Ⅰ在环境中的来源与分布,总结Int Ⅰ介导的ARGs迁移转化机制以及相关研究方法,并展望未来的研究发展趋势。     

食品动物养殖环境中细菌耐药性研究进展

抗生素耐药性被世界卫生组织认为是21世纪人类面临的最大的公共卫生安全问题之一。近年来,抗生素耐药基因作为一种新型污染物而受到广泛关注。养殖场现已成为耐药基因的一个重要储库,耐药菌及耐药基因随着动物排泄物进入环境,从而加速了耐药基因在环境中的传播。畜禽养殖环境中耐药基因和耐药菌可能经食物链、空气等途径传至人类,给人类健康带来巨大威胁。文中结合最新文献,主要介绍了动物养殖场抗菌药物耐药菌和耐药基因的分布特点、耐药基因的持留和传播扩散、研究方法等方面的研究进展,为食品动物养殖环境的抗菌药物耐药性风险评估提供一定支持。     

Status of pathogens,antibiotic resistance genes and antibiotic residues in wastewater treatment systems

Pathogens are becoming nearly untreatable due to the rise in gaining new resistance against standard antibiotics. Coexistence of microbial pathogens, antibiotics and antibiotic resistant genes (ARGs) in wastewater treatment plants (WWTP) provide favourable conditions for the development of new antibiotic resistant bacteria (ARB); facilitate horizontal gene transfer among pathogens and may also serve as a hotspot for the spread of ARB and genes into the environment. In this study, the current status of wastewater treatment systems in the removal of pathogens, ARGs, and antibiotic residues are discussed. WWTP are efficient in removing pathogens and antibiotic residues to a greater extend during secondary and tertiary treatment processes. Recent studies, however, have shown high variations in the presence of pathogens including ARB as well as antibiotic resistance genes (ARG) in the final effluent. Prolonged sludge retention time (SRT) and hydraulic retention time (HRT) during secondary treatment will facilitate antibiotic removal by adsorption and biodegradation. However, the above conditions can also lead to the enhancement of antibiotic resistance process in microbes. Therefore, optimum conditions for the operation of conventional WWTP for the efficient removal of antibiotics are yet to be established. The removal of antibiotic residues can be accelerated by combining conventional activated sludge (CAS) process with an additional treatment technology involving dosing with ozone. The advanced biological treatment method using membrane bioreactors (MBR) in combination with coagulation reportedly has the best ARG removal efficiency, and removes both ARB and extracellular ARGs. While studies have predicted the fate for ARGs in wastewater treatment plants, the mechanisms of ARGs acquisition remains to be conclusively established. Thus, strategies to investigate the underlying mechanism of acquisition of ARGs within the WWTP are also provided in this review.     

A review of the emergence of antibiotic resistance in bioaerosols and its monitoring methods

Despite significant public health concerns regarding infectious diseases in air environments, potentially harmful microbiological indicators, such as antibiotic resistance genes (ARGs) in bioaerosols, have not received significant attention. Traditionally, bioaerosol studies have focused on the characterization of microbial communities; however, a more serious problem has recently arisen due to the presence of ARGs in bioaerosols, leading to an increased prevalence of horizontal gene transfer (HGT). This constitutes a process by which bacteria transfer genes to other environmental media and consequently cause infectious disease. Antibiotic resistance in water and soil environments has been extensively investigated in the past few years by applying advanced molecular and biotechnological methods. However, ARGs in bioaerosols have not received much attention. In addition, ARG and HGT profiling in air environments is greatly limited in field studies due to the absence of suitable methodological approaches. Therefore, this study comprehensively describes recent findings from published studies and some of the appropriate molecular and biotechnological methods for monitoring antibiotic resistance in bioaerosols. In addition, this review discusses the main knowledge gaps regarding current methodological issues and future research directions.

     

抗生素耐药基因在畜禽粪便-土壤系统中的分布、扩散及检测方法

抗生素耐药基因作为一种新型的环境污染物已引起研究者的高度关注。畜禽养殖业长期将抗生素添加到饲料中,在促进动物生长、预防和治疗动物疾病等方面起了重要作用。这些抗生素大多数不能被动物完全吸收,在动物肠道中诱导出耐抗生素细菌和抗生素耐药基因,并随着粪便排出体外。畜禽粪便作为重要的抗生素、耐抗生素细菌和抗生素耐药基因储存库,通过堆粪、施肥等农业活动进入土壤环境中,可刺激土壤中耐抗生素细菌和抗生素耐药基因的富集。耐药基因借助于基因水平转移等方式在土壤介质中进一步传播扩散,甚至进入植物中随食物链传播,对生态环境和人类健康造成极大的威胁。为了正确评估抗生素耐药基因的生态风险,本文结合国内外相关研究,系统阐述了畜禽粪便-土壤系统中抗生素耐药基因的来源、分布及扩散机制,同时探讨了细菌耐药性的主要研究方法,指出堆肥化处理仍是目前去除抗生素耐药基因的主要手段,并对今后的研究方向进行展望。     

Research progress on distribution,migration, transformation of antibiotics and antibiotic resistance genes (ARGs) in aquatic environment

Antimicrobial and antibiotics resistance caused by misuse or overuse of antibiotics exposure is a growing and significant threat to global public health. The spread and horizontal transfer of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) by the selective pressure of antibiotics in an aquatic environment is a major public health issue. To develop a better understanding of potential ecological risks die to antibiotics and ARGs, this study mainly summarizes research progress about: (i) the occurrence, concentration, fate, and potential ecological effects of antibiotics and ARGs in various aquatic environments, (ii) the threat, spread, and horizontal gene transfer (HGT) of ARGs, and (iii) the relationship between antibiotics, ARGs, and ARB. Finally, this review also proposes future research direction on antibiotics and ARGs.     

Aquatic animals promote antibiotic resistance gene dissemination in water via conjugation: Role of different regions within the zebra fish intestinal tract,and impact on fish intestinal microbiota

The aqueous environment is one of many reservoirs of antibiotic resistance genes (ARGs). Fish, as important aquatic animals which possess ideal intestinal niches for bacteria to grow and multiply, may ingest antibiotic resistance bacteria from aqueous environment. The fish gut would be a suitable environment for conjugal gene transfer including those encoding antibiotic resistance. However, little is known in relation to the impact of ingested ARGs or antibiotic resistance bacteria (ARB) on gut microbiota. Here, we applied the cultivation method, qPCR, nuclear molecular genetic marker and 16S rDNA amplicon sequencing technologies to develop a plasmid‐mediated ARG transfer model of zebrafish. Furthermore, we aimed to investigate the dissemination of ARGs in microbial communities of zebrafish guts after donors carrying self‐transferring plasmids that encode ARGs were introduced in aquaria. On average, 15% of faecal bacteria obtained ARGs through RP4‐mediated conjugal transfer. The hindgut was the most important intestinal region supporting ARG dissemination, with concentrations of donor and transconjugant cells almost 25 times higher than those of other intestinal segments. Furthermore, in the hindgut where conjugal transfer occurred most actively, there was remarkable upregulation of the mRNA expression of the RP4 plasmid regulatory genes, trbBp and trfAp. Exogenous bacteria seem to alter bacterial communities by increasing Escherichia and Bacteroides species, while decreasing Aeromonas compared with control groups. We identified the composition of transconjugants and abundance of both cultivable and uncultivable bacteria (the latter accounted for 90.4%–97.2% of total transconjugants). Our study suggests that aquatic animal guts contribute to the spread of ARGs in water environments.     

水环境中耐药菌污染与危害研究进展

细菌在抗菌药选择性压力下产生耐药性并可传代,通过质粒和整合子等可移动基因元件将耐药基因在相同或不同种属中广泛传播,导致细菌多重耐药,并可通过多种途径进入水体,水环境日益成为庞大的耐药基因库,为致病菌及条件致病菌提供获得大量耐药基因的机会,若多重耐药菌再次侵入人体,可能引发严重的公共卫生问题。     

Metagenomic analysis reveals the prevalence and persistence of antibiotic- and heavy metal-resistance genes in wastewater treatment plant

The increased antibiotic resistance among microorganisms has resulted into growing interest for investigating the wastewater treatment plants (WWTPs) as they are reported to be the major source in the dissemination of antibiotic resistance genes (ARGs) and heavy metal resistance genes (HMRGs) in the environment. In this study, we investigated the prevalence and persistence of ARGs and HMRGs as well as bacterial diversity and mobile genetic elements (MGEs) in influent and effluent at the WWTP in Gwangju, South Korea, using high-throughput sequencing based metagenomic approach. A good number of broad-spectrum of resistance genes (both ARG and HMRG) were prevalent and likely persistent, although large portion of them were successfully removed at the wastewater treatment process. The relative abundance of ARGs and MGEs was higher in effluent as compared to that of influent. Our results suggest that the resistance genes with high abundance and bacteria harbouring ARGs and MGEs are likely to persist more through the treatment process. On analyzing the microbial community, the phylum Proteobacteria, especially potentially pathogenic species belonging to the genus Acinetobacter, dominated in WWTP. Overall, our study demonstrates that many ARGs and HMRGs may persist the treatment processes in WWTPs and their association to MGEs may contribute to the dissemination of resistance genes among microorganisms in the environment.     

Shifts in microbial community,pathogenicity-related genes and antibiotic resistance genes during dairy manure piled up

The uncomposted faeces of dairy cow are usually stacked on cow breeding farms, dried under natural conditions and then used as cow bedding material or they may be continuously piled up. However, no information is available to evaluate variations in the human and animal pathogen genes and antibiotic resistance during the accumulation of fresh faeces of dairy cow to manure. Here, we present the metagenomic analysis of fresh faeces and manure from a dairy farm in Ning Xia, showing a unique enrichment of human and animal pathogen genes and antibiotic resistance genes (ARGs) in manure. We found that manure accumulation could significantly increase the diversity and abundance of the pathogenic constituents. Furthermore, pathogens from manure could spread to the plant environment and enphytotic pathogens could affect the yield and quality of crops during the use of manure as a fertilizer. Levels of virulence genes and ARGs increased with the enrichment of microbes and pathogens when faeces accumulated to manure. Accumulated manure was also the transfer station of ARGs to enrich the ARGs in the environment, indicating the ubiquitous presence of environmental antibiotic resistance genes. Our results demonstrate that manure accumulation and usage without effective manure management is an unreasonable approach that could enrich pathogenic microorganisms and ARGs in the environment. The manure metagenome structure allows us to appreciate the overall influence and interaction of animal waste on water, soil and other areas impacted by faecal accumulation and the factors that influence pathogen occurrence in products from dairy cows.     

环境中抗生素抗性基因的水平传播扩散

抗生素抗性基因作为一类新型环境污染物,其在不同环境介质中的传播扩散可能比抗生素本身的环境危害更大,其中,水平基因转移是抗生素抗性基因传播的重要方式,是造成抗性基因环境污染日益严重的原因之一.本文系统阐述了抗生素抗性基因在环境中发生水平转移的主要分子传播元件及其影响因素,这对于正确揭示抗性基因的分子传播机制具有重要意义.结合多重抗药性的传播扩散机制,探讨了行之有效的遏制抗生素抗性基因传播扩散的方法和途径,并针对目前的污染现状,对今后有关抗生素抗性基因水平转移的研究重点进行了展望.