Quantification of multi-antibiotic resistant opportunistic pathogenic bacteria in bioaerosols in and around a pharmaceutical wastewater treatment plant

Pharmaceutical wastewater treatment plants(WWTPs) are thought to be a "seedbed" and reservoirs for multi-antibiotic resistant pathogenic bacteria which can be transmitted to the air environment through aeration. We quantified airborne multi-antibiotic resistance in a full-scale plant to treat antibiotics-producing wastewater by collecting bioaerosol samples from December2014 to July 2015. Gram-negative opportunistic pathogenic bacteria(GNOPB) were isolated, and antibiotic susceptibility tests against 18 commonly used antibiotics, including 11 β-lactam antibiotics, 3 aminoglycosides, 2 fluoroquinolones, 1 furan and 1 sulfonamide, were conducted.More than 45% of airborne bacteria isolated from the pharmaceutical WWTP were resistant to three or more antibiotics, and some opportunistic pathogenic strains were resistant to 16 antibiotics, whereas 45.3% and 50.3% of the strains isolated from residential community and municipal WWTP showed resistance to three or more antibiotics. The calculation of the multiple antibiotic resistance(MAR) index demonstrated that the air environment in the pharmaceutical WWTP was highly impacted by antibiotic resistance, while the residential community and municipal WWTP was less impacted by antibiotic resistance. In addition, we determined that the dominant genera of opportunistic pathogenic bacteria isolated from all bioaerosol samples were Acinetobacter, Alcaligenes, Citrobacter, Enterobacter, Escherichia, Klebsiella, Pantoea, Pseudomonas and Sphingomonas. Collectively, these results indicate the proliferations and spread of antibiotic resistance through bioaerosols in WWTP treating cephalosporin-producing wastewater, which imposed a potential health risk for the staff and residents in the neighborhood, calling for administrative measures to minimize the air-transmission hazard.     

Community analysis of ammonia-oxidizing bacteria in activated sludge of eight wastewater treatment systems

We investigated the communities of ammonia-oxidizing bacteria(AOB) in activated sludge collected from eight wastewater treatment systems using polymerase chain reaction(PCR) followed by terminal restriction fragment length polymorphism(T-RFLP),cloning,and sequencing of the α-subunit of the ammonia monooxygenase gene(amoA).The T-RFLP fingerprint analyses showed that different wastewater treatment systems harbored distinct AOB communities.However,there was no remarkable difference among the AOB TRFLP profiles from different parts of the same system.The T-RFLP fingerprints showed that a full-scale wastewater treatment plant(WWTP) contained a larger number of dominant AOB species than a pilot-scale reactor.The source of influent affected the AOB community,and the WWTPs treating domestic wastewater contained a higher AOB diversity than those receiving mixed domestic and industrial wastewater.However,the AOB community structure was little affected by the treatment process in this study.Phylogenetic analysis of the cloned amoA genes clearly indicated that all the dominant AOB in the systems was closely related to Nitrosomonas spp.not to Nitrosospira spp.Members of the Nitrosomonas oligotropha and Nitrosomonas communis clusters were found in all samples,while members of Nitrosomonas europaea cluster occurred in some systems.     

Characteristics of microbial community involved in early biofilms formation under the influence of wastewater treatment plant effluent

Effluents from wastewater treatment plants (WWTPs) containing microorganisms and residual nutrients can influence the biofilm formation. Although the process and mechanism of bacterial biofilm formation have been well characterized, little is known about the characteristics and interaction of bacteria, archaea and eukaryotes in the early colonization, especially under the influence of WWTP effluent. The aim of this study was to characterize the important bacterial, archaeal and eukaryotic species in the early stage of biofilm formation downstream of the WWTP outlet. Water and biofilm samples were collected 24 and 48 hr after the deposition of bio-cords in the stream. Illumina Miseq sequencing of the 16S and 18S rDNA showed that, among the three domains, the bacterial biofilm community had the largest alpha and beta diversity. The early bacterial colonizers appeared to be “biofilm-specific”, with only a few dominant operational taxonomic units (OTUs) shared between the biofilm and the ambient water environment. Alpha-proteobacteria and Ciliophora tended to dominate the bacterial and eukaryotic communities, respectively, of the early biofilm already at 24 hr, whereas archaea played only a minor role during the early stage of colonization. The network analysis showed that the three domains of microbial community connected highly during the early colonization and it might be a characteristic of the microbial communities in the biofilm formation process where co-occurrence relationships could drive coexistence and diversity maintenance within the microbial communities.     

Quantification of viable bacteria in wastewater treatment plants by using propidium monoazide combined with quantitative PCR (PMA-qPCR)

The detection of viable bacteria in wastewater treatment plants(WWTPs) is very important for public health, as WWTPs are a medium with a high potential for waterborne disease transmission. The aim of this study was to use propidium monoazide(PMA) combined with the quantitative polymerase chain reaction(PMA-qPCR) to selectively detect and quantify viable bacteria cells in full-scale WWTPs in China. PMA was added to the concentrated WWTP samples at a final concentration of 100 μmol/L and the samples were incubated in the dark for 5 min, and then lighted for 4 min prior to DNA extraction and qPCR with specific primers for Escherichia coli and Enterococci, respectively. The results showed that PMA treatment removed more than 99% of DNA from non-viable cells in all the WWTP samples, while matrices in sludge samples markedly reduced the effectiveness of PMA treatment. Compared to qPCR, PMA-qPCR results were similar and highly linearly correlated to those obtained by culture assay, indicating that DNA from non-viable cells present in WWTP samples can be eliminated by PMA treatment, and that PMA-qPCR is a reliable method for detection of viable bacteria in environmental samples. This study demonstrated that PMA-qPCR is a rapid and selective detection method for viable bacteria in WWTP samples, and that WWTPs have an obvious function in removing both viable and non-viable bacteria. The results proved that PMA-qPCR is a promising detection method that has a high potential for application as a complementary method to the standard culture-based method in the future.     

Bacterial diversity in Fez tanneries and Morocco’s Binlamdoune River, using 16S RNA gene based fingerprinting

The response of bacteria to various carbohydrates in the deep-sea sediments and the Antarctic soils was investigated using cellulose, chitin, and olive oil. It was found that the carbohydrates significantly increased the corresponding specific ectoenzyme activity (β- glucosidase, β-N-acetylglucosaminidase, lipase) in the samples from deep-sea sediments. In the case of Antarctic soil samples, the cellulose or olive oil amendments had minor or no effect on β-glucosidase or lipase activity, except the chitin which stimulated β- N-acetylglucosaminidase production. The responses of the bacteria in the deep-sea sediment sample WP02-3 and the Antarctic soil sample CC-TY2 towards the chitin amendment were further analyzed. Chitin amendments were shown to stimulate the ectoenzyme activity in all the tested sediments and the soils. The bacterial response before and after the carbohydrates amendments were compared by denaturing gradient gel electrophoresis and quantitative competitive polymerase chain reaction. Significant changes were found in the structure and density of the bacterial community in the deep sea sediments as compared to the Antarctic soil sample, where the effects were relatively lower. There was no change in the bacterial population in both studied samples in response to carbohydrates amendments. These data indicate that the bacterial communities in the oligotrophic deep-sea sediments are more dynamic than that in the Antarctic soils as they respond to the nutrient sources efficiently by regulation of ectoenzyme activity and/or changing community structure.     

Reduction and characterization of bioaerosols in a wastewater treatment station via ventilation

Bioaerosols from wastewater treatment processes are a significant subgroup of atmospheric aerosols. In the present study,airborne microorganisms generated from a wastewater treatment station(WWTS) that uses an oxidation ditch process were diminished by ventilation.Conventional sampling and detection methods combined with cloning/sequencing techniques were applied to determine the groups,concentrations,size distributions,and species diversity of airborne microorganisms before and after ventilation. There were 3021 ± 537 CFU/m3 of airborne bacteria and 926 ± 132 CFU/m3 of airborne fungi present in the WWTS bioaerosol.Results showed that the ventilation reduced airborne microorganisms significantly compared to the air in the WWTS. Over 60% of airborne bacteria and airborne fungi could be reduced after4 hr of air exchange. The highest removal(92.1% for airborne bacteria and 89.1% for fungi) was achieved for 0.65–1.1 μm sized particles. The bioaerosol particles over 4.7 μm were also reduced effectively. Large particles tended to be lost by gravitational settling and small particles were generally carried away,which led to the relatively easy reduction of bioaerosol particles0.65–1.1 μm and over 4.7 μm in size. An obvious variation occurred in the structure of the bacterial communities when ventilation was applied to control the airborne microorganisms in enclosed spaces.     

Molecular analysis of long-term biofilm formation on PVC and cast iron surfaces in drinking water distribution system

To understand the impacts of different plumbing materials on long-term biofilm formation in water supply system, we analyzed microbial community compositions in the bulk water and biofilms on faucets with two different materials-polyvinyl chloride （PVC） and cast iron, which have been frequently used for more thanlO years. Pyrosequencing was employed to describe both bacterial and eukaryotic microbial compositions. Bacterial communities in the bulk water and biofilm samples were significantly different from each other. Specific bacterial populations colonized on the surface of different materials. Hyphomicrobia and corrosion associated bacteria, such as Acidithiobacillus spp., Aquabacterium spp., Limnobacter thiooxidans, and Thiocapsa spp., were the most dominant bacteria identified in the PVC and cast iron biofilms, respectively, suggesting that bacterial colonization on the material surfaces was selective. Mycobacteria and Legionella spp. were common potential pathogenic bacteria occurred in the biofilm samples, but their abundance was different in the two biofilm bacterial communities. In contrast, the biofilm samples showed more similar eukaryotic communities than the bulk water. Notably, potential pathogenic fungi, i.e., Aspergillus spp. and Candida parapsilosis, occurred in similar abundance in both biofilms. These results indicated that microbial community, especially bacterial composition was remarkably affected by the different pipe materials （PVC and cast iron）.     

Bacterial diversity in soils around a lead and zinc mine

Five samples of soil collected from a lead and zinc mine were used to assess the effect of combined contamination of heavy metals on soil bacterial communities using a polyphasic approach including characterization of isolates by culture method, community level catabolic profiling in BIOLOG GN microplates, and genetic community fingerprinting by denaturing gradient gel electrophoresis of 16S rDNA fragments amplified by PCR from community DNA （PCR-DGGE）. The structure of the bacterial community was affected to a certain extent by heavy metals. The PCR-DGGE analysis of 16S rRNA genes showed that there were significant differences in the structure of the microbial community among the soil samples, which were related to the contamination levels. The number of bacteria and the number of denaturing gradient gel electrophoresis （DGGE） bands in the soils increased with increasing distance from the lead and zinc mine tailing, whereas the concentration of lead （Pb） and cadmium （Cd） was decreased. Heavily polluted soils could be characterized by a community that differs from those of lightly polluted soils in richness and structure of dominating bacterial populations. The clustering analysis of the DGGE profiles showed that the bacteria in all the five samples of soil belonged to three clusters. The data from the BIOLOG analysis also showed the same result. This study showed that heavy metal contamination decreased both the biomass and diversity of the bacterial community in soil.     

Bioaugmentation with a pyridine-degrading bacterium in a membrane bioreactor treating pharmaceutical wastewater

The bacterial strain Paracoccus denitrificans W12, which could utilize pyridine as its sole source of carbon and nitrogen, was added into a membrane bioreactor （MBR） to enhance the treatment of a pharmaceutical wastewater. The treatment efliciencies investigated showed that the removal of chemical oxygen demand, total nitrogen, and total phosphorus were similar between bioaugmented and non-bioaugmented MBRs, however, significant removal of pyridine was obtained in the bioaugmented reactor. When the hydraulic retention time was 60 hr and the influent concentration of pyridine was 250-500 mg/L, the mean effluent concentration of pyridine without adding W12 was 57.2 mg/L, while the pyridine was degraded to an average of 10.2 mg/L with addition of W12. The bacterial community structure of activated sludge during the bioaugmented treatment was analyzed using polymerase chain reaction-denaturing gradient gel electrophoresis （PCR-DGGE）. The results showed that the W12 inoculum reversed the decline of microbial community diversity, however, the similarity between bacterial community structure of the original sludge and that of the sludge after bioaugmentation decreased steadily during the wastewater treatment. Sequencing of the DNA recovered from DGGE gel indicated that sp., Sphingobium sp., Comamonas sp., and Hyphomicrobium sp. were the dominant organisms in time sequence in the bacterial community in the bioaugmented MBR. This implied that the bioaugmentation was affected by the adjustment of whole bacterial community structure in the inhospitable environment, rather than being due solely to the degradation performance of the bacterium added.     

Temporal and spatial changes of microbial community in an industrial effluent receiving area in Hangzhou Bay

Anthropogenic activities usually contaminate water environments, and have led to the eutrophication of many estuaries and shifts in microbial communities. In this study, the temporal and spatial changes of the microbial community in an industrial effluent receiving area in Hangzhou Bay were investigated by 454 pyrosequencing. The bacterial community showed higher richness and biodiversity than the archaeal community in all sediments. Proteobacteria dominated in the bacterial communities of all the samples; Marine_Group_Ⅰ and Methanomicrobia were the two dominant archaeal classes in the effluent receiving area. PCoA and AMOVA revealed strong seasonal but minor spatial changes in both bacterial and archaeal communities in the sediments. The seasonal changes of the bacterial community were less significant than those of the archaeal community, which mainly consisted of fluctuations in abundance of a large proportion of longstanding species rather than the appearance and disappearance of major archaeal species. Temperature was found to positively correlate with the dominant bacteria, Betaproteobacteria, and negatively correlate with the dominant archaea,Marine_Group_Ⅰ; and might be the primary driving force for the seasonal variation of the microbial community.     

SBR工艺城市污水处理厂微生物气溶胶逸散特征

在采用SBR工艺的某污水处理厂设置采样点,研究各污水处理工艺段微生物气溶胶的逸散特征.结果表明,各工艺段均有细菌气溶胶逸散,浓度为82～1 525 CFU·m~(-3),粗格栅、生化池和污泥脱水间为主要逸散源.各工艺段检测到的细菌气溶胶主要菌属为Cyanobacteria,其它丰度较高的菌属有Aeromonas、Peptostreptococcaceae、Moraxellaceae、Chroococcidiopsis、Sphingomonas、Arcobacter及Acinetobacter等,其中Aeromonas、Arcobacter、Acinetobacter及Sphingomonas为潜在致病菌.微生物气溶胶的浓度和丰度沿垂直方向和水平方向减少.适宜的温度和相对湿度利于微生物气溶胶在空气中保持活性(P 0. 01),风速则与微生物气溶胶的逸散呈负相关(P 0. 05).污水处理过程产生的微生物气溶胶的暴露风险较小(HQ 1),但是污染物的累积会增加人体的暴露风险.生物除臭反应器在处理臭味气体的同时还可以有效削减微生物气溶胶.     

高通量测序技术应用于污水处理厂细菌气溶胶群落结构分析

由于污水及其处理设施中细菌气溶胶的逸散作用,导致污水处理厂细菌气溶胶的群落结构较为复杂,因此,选择适宜的采样和分析方法对其多样性的认识和风险评估具有重要意义.本研究以某城市污水厂为研究对象,采用"总悬浮颗粒物采样器+高通量测序技术"分别对污水处理全过程进行细菌气溶胶的收集与分析;作为对照,采用"Andersen六级采样器+克隆文库技术"进行同步采样和分析.结果表明,前者对各采样点气溶胶中细菌的多样性、总体群落组成、优势群落的解析均明显优于后者,分析更简便快捷,数据更全面、更接近其真实的分布状态.本研究结果可为污水处理厂细菌气溶胶的采样和分析方法的选择提供科学依据.     

Exposure to Airborne Noroviruses and Other Bioaerosol Components at a Wastewater Treatment Plant in Denmark

Exposure to bioaerosols associated with wastewater treatment processes may represent an occupational health risk for workers at wastewater treatment plants (WWTPs). A high frequency of acute symptoms in the gastrointestinal tract among the wastewater workers at a Danish WWTP has been reported. The objective of the study was therefore to examine the exposure of the workers to aerosolised microorganisms. Sampling of inhalable endotoxin, bacteria, moulds and viruses was performed on one occasion using personal samplers. Noroviruses (NoVs) and endotoxin were detected at concentrations that could pose an occupational health risk and possibly contribute to the increased frequency of gastrointestinal illness among the workers and should therefore be investigated further. In addition, positive correlations between exposure to endotoxin, bacteria, moulds and NoVs were found and indicate that the exposure to bioaerosols may be related to work tasks. This is the first study directly showing an occupational exposure to airborne NoVs by its detection in airborne dust.     

焦化废水活性污泥细菌菌群结构分析

焦化废水是一种高毒难降解的有机废水,以细菌菌群为主的好氧活性污泥决定焦化废水的处理效率,处理焦化废水的活性污泥细菌群落结构鲜见报道.利用454测序技术分析实际焦化废水污泥中的细菌菌群结构和多样性.结果表明,热图聚类分析和主成分分析说明不同焦化废水活性污泥细菌菌群多样性存在差异;焦化废水活性污泥中的细菌门类群主要为Proteobacteria、Planctomycetes、Acidobacteria、Candidatus Saccharibacteria、Bacteroidetes、Cyanobacteria、Actinobacteria、Chloroflexi、Firmicutes、Thaumarchaeota、Ignavibacteriae和Verrucomicrobia,其中Proteobacteria门占主导地位,丰度为36.00%~76.98%;主要属为Thiobacillus、Thauera、Comamonas、Caldimonas、Steroidobacter、Nitrosomonas、Phycisphaera和Gp4,大多数主要属与芳香烃的降解和硝化反硝化过程有关.这些结果为焦化废水污染物的去除机制提供了理论基础.     

不同好氧颗粒污泥中微生物群落结构特点

为了探讨活性污泥好氧颗粒化过程对微生物种群的影响、不同底物及不同颗粒化方法培养的好氧颗粒污泥中微生物群落结构的差异,以接种污泥、模拟废水好氧颗粒污泥和分别投加粉末活性炭和硅藻土的实际生活污水好氧颗粒污泥为研究对象,利用PCR-DGGE对比分析了接种污泥和好氧颗粒污泥中的微生物群落结构.结果表明:活性污泥好氧颗粒化过程会减少微生物种群多样性,影响颗粒污泥稳定性的细菌被淘汰,而聚磷菌、反硝化菌、难降解有机物降解菌等污水处理功能微生物都在颗粒化过程中得到保留.活性污泥好氧颗粒化过程中能够实现亚硝化细菌(AOB)一定程度的富集.与接种活性污泥相比,好氧颗粒污泥中AOB的多样性指数与均匀性指数均有提高.好氧颗粒污泥中的优势菌群主要分布于变形菌门(Proteobacteria)、拟杆菌门(Bacteroidetes)和未培养菌(uncultured bacterium).其中AOB均属于β-Proteobacteria的亚硝化单胞菌属(Nitrosomonas).     

细菌对城市污水中小球藻生长和油脂积累的影响

利用城市污水培养微藻,可在实现污水无害化处理的同时,培养微藻回收生物质能源.污水为微藻的培养提供氮、磷等营养组分和所需水源,同时污水中的细菌可分解污水中的有机物产生CO_2,为微藻提供生长所需碳源.菌藻混合培养既可以收获藻类,又可以净化污水,由于城市污水含有大量的原生菌类,且微藻与细菌之间存在着互生、拮抗等复杂的相互关系,因此,需要筛选出既能够适应于城市污水又能促进微藻生长和油脂积累的优势菌种.本文从不同来源的13种细菌中筛选出2种能够显著促进蛋白核小球藻(Chlorella pyrenoidosa)生长和油脂积累细菌,并分析了微藻培养结束后城市污水的菌群结构.结果表明:污水中光合细菌初始吸光度D600为0.01,W4菌初始吸光度D_(600)为0.02时,对小球藻的干重和油脂产量促进作用最显著,油脂产量分别可达0.114 g·L~(-1)、0.113 g·L~(-1),油脂产量比空白对照组分别提高了22.58%、21.50%.通过对生成的脂肪酸甲酯进行气相色谱分析,结果显示光合细菌和W4菌的添加并未改变小球藻脂肪酸成分,但提升了单不饱和脂肪酸的含量,有利于提升所得生物柴油的品位.培养结束后污水的菌群结构分析显示投加细菌会降低污水中菌群的丰富度和多样性,初步判断是投加的菌在藻液中能够成为优势菌群,且实验组中丛毛单胞菌属(Comamonas)和假单胞菌属(Pseudomonas)的丰度大于对照组.     

北京大气降水中细菌气溶胶的多样性研究

利用16S rRNA基因测序分类学技术,分别以北京市区2011及2012年不同月份的降水样品中细菌的基因组DNA为模板,通过克隆、测序构建基因组文库,研究了北京市大气降水中细菌的群落结构组成及多样性变化.系统发育分析结果表明,变形菌门(Proteobacteria) (α-,β-,γ-)是北京市降水样品中细菌的优势菌群(75.6%~100%),另外包括拟杆菌门(Bacteroidetes)、放线菌门(Actinobacteria)、异常球菌门(Deinococcus-Thermus)、蓝藻门(Cyanobacteria)、硝化螺菌门(Nitrospira)、厚壁菌门(Firmicutes)共7个主要门类的细菌,以及未定菌(TM7).多样性指数分析结果显示,不同的降水样品,细菌群落结构组成及多样性均存在着差异性,冬季12月份雪水样品细菌群落结构多样性明显高于其他季节的样品,细菌群落多样性(Shannon, H)特点是,冬季>秋季>夏季.     

Evaluation on the performance of triple oxidation ditch system treating municipal wastewater

ＥｖａｌｕａｔｉｏｎｏｎｔｈｅｐｅｒｆｏｒｍａｎｃｅｏｆｔｒｉｐｌｅｏｘｉｄａｔｉｏｎｄｉｔｃｈｓｙｓｔｅｍｔｒｅａｔｉｎｇｍｕｎｉｃｉｐａｌｗａｓｔｅｗａｔｅｒＺｈｏｕＬｕ，ＱｉａｎＹｉＮａｔｉｏｎａｌＫｅｙＪｏｉｎｔＬａｂｏｒａｔｏｒｙｏｎＥｎｖ...