雌二醇短期暴露对非洲爪蟾性腺和输卵管形态及性二态基因表达影响的初步研究

环境雌激素对两栖动物的生殖内分泌干扰作用已引起关注,目前的研究主要集中在长期暴露对性腺分化和发育的影响。以雌二醇(E2,10 nmol·L-1)为模式化合物,以非洲爪蟾为实验动物,6月龄的幼蛙暴露6 d,研究雌激素对性腺和输卵管形态以及性二态性基因表达的影响。结果发现,6 d的E2暴露对精巢和卵巢的形态没有明显影响,但从组织学结构来看对卵和精子的发育有微弱的促进作用。E2能够明显地促进了雌性输卵管的发育,表现为输卵管变粗变弯曲。E2暴露显著抑制精巢和卵巢中性激素合成相关基因cyp11a1、3βHSD、Srd5α、ARO的表达,对cyp17的表达无影响,使脑中LHβ和FSHβ的表达下调;对性别分化和性腺发育相关基因Dax、AR、ERα、TFⅢA、TBPL2的表达均无影响,但能够促进卵巢中Sox3的表达;同时促进脑中Emi2的表达,抑制脑中Dmrt1的表达。因此,输卵管可作为敏感的形态指标,性腺中Srd5α、cyp11a1、3βHSD、ARO、Sox3和脑中LHβ、FSHβ、Dmrt1、Emi2可作为分子指标。以上结果可为后续建立评价化学品对两栖动物雌激素效应的短期试验方法提供参考。     

裂片石莼和土霉素复合压力下弧菌群落及耐药基因变化特性研究

近年来,水产养殖病源弧菌污染及其耐药性问题备受关注,本研究首次采用宏基因组测序技术分析了海水养殖沉积物在大型海藻裂片石莼（12 g·L^-1）与不同浓度（0、10、100、500 μg·L^-1）土霉素作用3个月后,弧菌群落结构及其耐药基因变化特性.结果表明,经裂片石莼诱导培养后,沉积物中弧菌属相对丰度降低了0.22%,但对弧菌多样性和弧菌耐药基因含量提升明显.裂片石莼和土霉素复合压力下,100 μg·L^-1土霉素对弧菌群落多样性及弧菌耐药基因影响最大,而500 μg·L^-1对其影响最小.沉积物弧菌属共检测出21种耐药基因,多为BL1_fox、BL2be_per、BL2_veb等β-内酰胺类耐药基因.裂片石莼能减少耐药基因tetPB富集,而进一步添加土霉素能诱导产生VatB、BcrA耐药基因.此外,弧菌耐药基因迁移与转化还受弧菌群落结构变化影响.     

Short-term inhalation exposure evaluations of airborne antibiotic resistance genes in environments

Antibiotic resistance is a sword of Damocles that hangs over humans. In regards to airborne antibiotic resistance genes (AARGs), critical knowledge gaps still exist in the identification of hotspots and quantification of exposure levels in different environments. Here, we have studied the profiles of AARGs, mobile genetic elements (MGEs) and bacterial communities in various atmospheric environments by high throughput qPCR and 16S rRNA gene sequencing. We propose a new AARGs exposure dose calculation that uses short-term inhalation (STI). Swine farms and hospitals were high-risk areas where AARGs standardised abundance was more abundant than suburbs and urban areas. Additionally, resistance gene abundance in swine farm worker sputum was higher than that in healthy individuals in other environments. The correlation between AARGs with MGEs and bacteria was strong in suburbs but weak in livestock farms and hospitals. STI exposure analysis revealed that occupational intake of AARGs (via PM₁₀) in swine farms and hospitals were 110 and 29 times higher than in suburbs, were 1.5 × 10⁴, 5.6 × 10⁴ and 5.1 × 10² copies, i.e., 61.9%, 75.1% and 10.7% of the overall daily inhalation intake, respectively. Our study comprehensively compares environmental differences in AARGs to identify high-risk areas, and forwardly proposes the STI exposure dose of AARGs to guide risk assessment.     

Biofilm-overproducing Bacillus subtilis B12ΔYwcc decreases Cd uptake in Chinese cabbage through increasing Cd-immobilizing related gene abundance and root surface colonization

Biofilm-producing bacteria can decrease Cd uptake in vegetables, but mechanisms underlying this effect are poorly characterized. In this study, two mutant strains B12ΔYwcc and B12ΔSlrR were constructed from a biofilm-producing Bacillus subtilis strain B12. Then, the impacts of strain B12 and its high biofilm-producing mutant strain B12ΔYwcc and low biofilm-producing mutant strain B12ΔSlrR on Cd availability and uptake in Chinese cabbage and the related mechanisms were investigated in the Cd-polluted soil. Strain B12 and its mutants B12ΔYwcc and B12ΔSlrR increased the dry biomasses of edible tissues by 54%–130% compared with the controls. Strain B12 and its mutant B12ΔYwcc reduced the soil available Cd content by 36%–50% and root and edible tissue Cd contents by 23%–50% compared with the controls. Furthermore, the mutant strain B12ΔYwcc reduced the edible tissue Cd content by 40% and increased the polysaccharide content by 23%, invertase activity by 139%, and gene copies of the cumA by 4.5-fold, epsA by 7.1-fold, and cadA by 4.3-fold, which were involved in Cd adsorption in the rhizosphere soils, respectively, compared with strain B12. The polysaccharide content and cumA, epsA, and cadA gene copy numbers showed significantly reverse correlations with the available Cd content. Notably, the mutant strain B12ΔYwcc showed better ability to colonize the vegetable root surface than strain B12. These findings demonstrated that the biofilm-overproducing mutant strain B12ΔYwcc increased the polysaccharide production and Cd-immobilizing related cumA, epsA, and cadA gene copies, resulting in lower Cd availability and accumulation in Chinese cabbage in the Cd-polluted soil.     

高效石油降解菌修复石油污染土壤与强化机制分析

利用定向驯化高效石油降解菌系对石油污染土壤进行为期120 d原位修复,考察生物强化修复效果、土壤理化性质和酶活性的变化,结合宏基因组测序及生物信息学分析揭示其强化机制.结果表明,与空白对照组(Ctrl)相比,生物修复组(Exp-BT)总石油烃降解率显著提升,增幅达81.23%; 高效石油降解菌生物强化修复期间土壤pH变化稳定,体系氧化能力提高,电导率处于适宜农业活动范围内; 脂肪酶和脱氢酶在修复期间保持较高活性; 另对初始污染土壤样本(B0)、驯化所得高效石油降解菌系样本(GZ)和生物修复后土壤样本(BT)的分析显示,门水平上变形菌门与放线菌门相对丰度增加17.1%,属水平上Nocardioides、Achromobacter、Gordonia和Rhodococcus等丰度明显上升,COG和KEGG物种与功能贡献度分析证明以上菌属对石油烃降解有重要贡献; 修复后土壤中发现高丰度的石油烃相关代谢酶及5个降解基因:alkM、tamA、rubB、ladA 和alkB,分析表明外源石油烃降解菌群的引入增强了微生物相关酶的代谢活性与相应功能基因的表达.     

历史抗生素胁迫改变磺胺甲唑和甲氧苄啶对活性污泥的影响:ARGs及其潜在宿主

抗生素共暴露对污水处理厂抗生素抗性基因（ARGs）及微生物群落聚集过程有着重要影响.然而,历史抗生素接触胁迫差异（遗留效应）是否能决定微生物和ARGs对抗生素复合污染的响应尚不清楚.通过选择高浓度（30 mg ·L^-1）磺胺甲唑（SMX）和甲氧苄啶（TMP）作为历史接触胁迫条件,探究SMX和TMP复合污染对ARGs、细菌群落及其交互作用的短期影响.基于高通量荧光定量PCR,共检测出13种ARGs,它们的绝对丰度介于2.21~5.42 copies ·μL^-1（对数值,以DNA计,下同）,其中sul2、ermB、mefA和tetM-01是样品中最主要的亚型,绝对丰度在2.95~5.40 copies ·μL^-1之间;SMX和TMP复合污染会引起ARGs和可移动基因元件（MGEs）的富集,但其对各亚型的影响不同,且SMX的历史遗留效应高于TMP;不同接触史的复合污染下,ARGs间的共现与互斥模式均存在;MGEs （尤其是intI-1）与磺胺类（sul1、sul2）、四环素类［tet （32）］和大环内酯-林可酰胺-链霉菌素类抗性基因（ermB）均呈显著正相关性;基于微生物全尺度分类发现各类群微生物群落结构对复合污染的响应不同,且条件丰富菌属（CAT）得到了显著富集;陶厄氏菌属（Thauera）、假黄单胞菌属（Pseudoxanthomonas）和副球菌属（Paracoccus）是优势的抗性菌属;网络分析共发现31个ARGs的潜在宿主,以条件稀有菌属（CRT）为主,尤其是Candidatus_Alysiosphaera和纺锤状菌属（Fusibacter）与多种ARGs呈现正相关,是多种ARGs的潜在公共细菌宿主;部分稀有菌属［RT,变异杆菌属（Variibacter）、气单胞菌属（Aeromonas）和管道杆菌属（Cloacibacterium）等］是转座子IS 613的潜在宿主,在ARGs的增殖传播中发挥重要作用.综上,研究揭示了污水处理系统中抗生素历史胁迫对ARGs赋存特征和及其宿主细胞的遗留效应,可为削减抗生素复合污染引起的ARGs污染提供新思路与理论依据.     

某集约化肉鸡饲养场PM2.5中抗生素抗性基因的分布特征

集约化家禽饲养场是抗生素抗性基因(ARGs)的重要来源,而PM_(2.5)作为ARGs可能向人体暴露的重要途径还未得到很好地研究.本文采集了集约化肉鸡饲养场舍内PM_(2.5)和粪便以及舍外PM_(2.5)样品,利用荧光定量PCR(q PCR)进行一类整合子(int I1)、总细菌(16S r DNA)和6类共19种ARGs丰度的检测.结果显示,除blaGES-1和blaSHV-1之外,其余17种ARGs在6类样品中均有检出.磺胺类、四环素类、大环内酯类和氨基糖苷类抗性基因在舍内粪便中丰度较高,达到1. 04×109～3. 27×1010copies·g-1,粪便是饲养场PM_(2.5)中ARGs的主要来源.舍内PM_(2.5)中以磺胺类和大环内酯类抗性基因丰度较高,分别为(8. 9±1. 9)×107copies·m-3和(5. 6±3. 1)×107copies·m-3,且舍内PM_(2.5)中ARGs丰度明显高于舍外. PM_(2.5)质量浓度与16S r DNA、int I1和ARGs丰度呈显著正相关,表明集约化饲养场中PM_(2.5)是空气传播细菌、ARGs和int I1的储存库和传播者. 6类样品中int I1丰度均高于ARGs,同时int I1和ARGs的共存关系表现出了多药耐药性的威胁,易对饲养人员和家禽健康及周边空气环境造成危害.     

Contribution of the Kodama and 4S pathways to the dibenzothiophene biodegradation in different coastal wetlands under different C/N ratios

Dibenzothiophene (DBT) degradation mechanisms and the transformation of pathways during the incubation of three types of coastal sediments with C/N ratios ranging from 1 to 9 were investigated. The DBT degradation efficiencies were clearly improved with increasing C/N ratio in reed wetland sediments, tidal wetlands sediments and estuary wetland sediments. The quantitative response relationships between DBT degradation rates and related functional genes demonstrate that the Kodama pathway-related gene groups were dominant factors at low C/N ratios, while the 4S-related gene groups mainly determined the degradation rate when the C/N ratio was up to 5. Network analysis also shows that the pathway shifts from the Kodama pathway to the 4S pathway occurred through changes in the connections between functional genomes and rates. Furthermore, there were competition and collaboration between the Kodama and 4S pathways. The 4S pathway-related bacteria were more active in estuary wetland sediments compared with reed wetland sediments and tidal wetland sediments. The higher degradation efficiency in estuary wetland sediments may indicate the greater participation of the 4S pathway in the DBT biodegradation reaction. And the effects of ring cleavage of Kodama pathway caused more complete metabolizing of DBT.     

ClO2消毒工艺对污水处理厂出水超级耐药基因的影响

为了研究二氧化氯（chlorine dioxide,ClO₂）消毒工艺对污水处理厂出水中超级耐药基因（super antibiotic resistance genes,SARGs）的去除效果,对污水处理厂出水消毒前后的水样进行了全年采集,并采用微孔滤膜正压过滤法及核酸吸附柱-洗脱法分别富集水中细菌和胞外核酸后,利用荧光定量PCR技术对其中的9种SARGs进行定量检测.结果表明,无论是胞内还是胞外核酸,均有NDM-1、MCR-1 和MEC-A被检测出;同时,ClO₂消毒后上述3种SARGs的胞内相对总浓度明显上升（P<0.05）,且ClO₂消毒对胞内SARGs相对浓度的影响与季节有关,其中春季、夏季和秋季均发生上升,且春季升高最为明显,达2倍,而冬季出水在消毒前后均未测出胞内SARGs;胞外SARGs则在ClO₂消毒前后没有明显浓度变化.因此,ClO₂消毒不能有效去除污水处理厂出水中胞内和胞外SARGs污染.     

上海沙田湖养殖区及周边水体中氟喹诺酮类抗性基因的分布特征及其与环境因子关系

集约化水产养殖区常被认为是水体抗生素和抗性基因的主要潜在来源,然而对其该类水体中抗生素和抗性基因污染情况的研究仍相对匮乏.本文以上海市沙田湖养殖区及周边水体为调查对象,利用宏基因组学高通量测序技术,于2020年9月对水体中的氟喹诺酮类抗生素(fluoroquinolones,FQs)和氟喹诺酮类抗性基因(fluoroquinolones antibiotic resistance genes,FQs-ARGs)的种类和丰度进行了调查,并利用多元统计分析方法研究了水体理化因子和FQs对FQs-ARGs的影响.结果表明,沙田湖养殖区及周边水体共检测出5类耐药机制、共46种FQs-ARGs,其中各水体共有基因为44种.抗性基因mfd和patA在养殖水体和周边水体中均为优势抗性基因,抗生素抗性基因变异或突变为优势耐药机制.养殖水体和周边河湖水体、不同养殖品种水体之间FQs-ARGs的平均丰度无显著差异(P＞0.05).偏冗余分析(pRDA)表明,除了诺氟沙星和环丙沙星2种抗生素外,水体电导率(Spc)、总磷(TP)、总氮(TN)、氨氮(NH4+-N)、溶解氧(DO)、pH和叶绿素a(Chl-a)等环境因子是影响水体中FQs-ARGs组成与分布的关键性因子,且非抗生素因子的总贡献率远高于抗生素.