Characterization and phylogenetics of a new species of genus Lactobacillus from the activated sludge in biohydrogen production

Anaerobic process of biohydrogen production was developed. There is a great deal of Lactobacillus bacteria in the activated sludge of biohydrogen reactor. The isolation and identification of different anaerobic bacteria in the reactor is important for fermented biohydrogen production process by anaerobic digesting organic wastewater. Considering with the physiological and biochemical traits, morphological characteristics and 16SrDNA sequence, the isolated Rennanqilyfl3 is a new species in Lactobacillus genus. And the temporary nomenclature of the species is Lactobacillus Strain Rennanqilyfl3 sp. nov.     

The molecular biological characterization of a strain of biohydrogen-producing anaerobe in Clostridium Genus

The anaerobic process of biohydrogen production was developed recently. The isolation and identification of biohydrogen producing anaerobic bacteria with high evolution rate and yield is an important foundation of the fermented biohydrogen production process through which anaerobic bacteria digest organic wastewater. By considering physiological and biochemical traits, morphological characteristics and a 16S rDNA sequence, the isolated Rennanqilyf33 is shown to be a new species.     

发酵产氢细菌分离培养的厌氧实验操作技术

厌氧产氢细菌的分离纯化与培养是发酵法生物制氢技术的基础课题之一,现有厌氧细菌的分离制备技术和培养基分别存在着操作繁琐和成份复杂问题．通过厌氧培养技术比较和细菌生长试验,确定了“煮沸吹氮去氧,固液交替分离”的厌氧产氢细菌分离纯化培养程序,改进了Hungate技术和建立了新的宽体窄颈培养瓶平板技术,通过培养基组成成份的增减和细菌增长、种类和数量的研究确立了厌氧发酵产氢细菌的分离和富集培养基．利用这些厌氧培养技术,已成功分离培养了550余厌氧菌株系．     

复合菌群降解玉米秸秆协同产氢特性

为加快生物制氢工业化进程,利用玉米秸秆这类来源广泛、储量巨大和价格低廉的可再生生物质纤维素资源作为发酵产氢的原料,从连续流发酵产氢反应器(ZL92114474.1)中新分离筛选出一株高效纤维素降解产氢细菌Clostridium sp.X9(NCBI注册号:EU434651)和一株高效产乙醇发酵产氢细菌Ethanoigenens harbinenseB2(NCBI注册号:EU639425),通过构建高效降解纤维素发酵产氢复合菌群进行同步降解玉米秸秆发酵产氢.结果表明,对玉米秸秆进行酸化汽爆预处理后可以显著提高复合菌群的产氢能力.复合菌群X9和B2比单一菌种具有更理想的降解玉米秸秆发酵产氢的能力,两菌种间存在协同产氢效应.复合菌群X9和B2降解玉米秸秆发酵产氢获得的最大产氢率和玉米秸秆降解率分别为8.7mmol/g和74%.液相代谢末端产物主要为乙醇、乙酸和丁酸.这说明复合菌群X9和B2在以木质纤维素为发酵底物的工业化生物制氢领域中具有很好的应用发展前景.     

Length polymorphisms for intergenic spacer regions of 16S-23S rDNA in members of the new hydrogen-producing bacteria

A method based on PCR amplification of the 16S rRNA gene (rDNA)-23S rDNA intergenic spacer regions (ISR) was developed for the identification of species within the novel group hydrogen-producing anaerobes. The sizes of the PCR products varied from 1264 to 398 bp. Strain of isolate Rennanqilyf 3 was characterized as having products of 1262,398,638,437 and 436 bp. The isolate Rennanqilyf 1 had product of 1264 bp. The isolate Rennanqilyf 13 had products of 1261,579 and 485 bp. Of the 3 species of the novel group hydrogen-producing anaerobes examined, no one was indistinguishable. Two environmental isolates were identified as hydrogen-producing bacteria, which were new species in present taxon. Rennanqilyf 3 could not be associated with any Clostridium sp. studied. Rennanqilyf 1 could be classified into Clostridium genus. The combination between 16S rDNA equencing and length polymorphisms of IRS in 16S-23S rDNA is a better method for determining species of the hydrogen-producing bacteria.     

16S rDNA测序快速鉴定废水生物处理系统目标细菌

在废水生物处理系统中建立分子生物学技术快速鉴定有关微生物,具有十分重要的意义．以发酵法生物制氢系统的活性污泥分离培养的厌氧发酵细菌为研究对象,采用16S rDNA碱基测序分子生物学技术,通过生物信息学数据库NCBI将DNA序列输入、比对,可以直接鉴定从废水生物处理系统中分离培养的细菌进行种属科的系统发育学地位的判定,从而简化了细菌鉴定程序．通过16S rDNA测序技术,进行了分离产氢细菌的分子生物学鉴定,发现生物制氢厌氧活性污泥中所分离的细菌可能存在的菌属有：Lactobacillus,Clostridium,Klebsiella,Pmpionibacterium和可能新发现的1个新属Biohydrogenbacterium等5个菌属的菌种,其中新属Biohydrogenbacterium的菌种都以利用碳水化合物生产氢气为其主要特征．16S rDNA测序技术为废水生物处理系统的细菌学研究提供了方便、准确和经济的技术基础．     

牛奶废水厌氧水解酸化试验研究

利用CSTR反应器进行了牛奶废水厌氧酸化实验研究。结果表明,牛奶废水的厌氧水解酸化过程表现为丁酸型发酵;蛋白质水解为氨基酸是蛋白质类物质厌氧处理的限速步骤;长链脂肪酸C18：1(含一个双键的不饱和油酸)的积累是造成蛋白质厌氧水解酸化效率降底的主要原因之一;在牛奶废水的厌氧处理过程中用CSTR反应器作为生物产氢反应器是不适宜的。     

生物制氢细菌分离培养与分子鉴定技术进展

分离鉴定、培养产氢发酵细菌是提高利用高浓度有机废水制氢系统产氢能力的重要因素.首先对连续流发酵法生物制氢系统进行工程调控和生态位调整,达到产氢最佳的乙醇型发酵阶段后,设计HPB-LR培养基,用改良Hungater等3种厌氧培养技术分离培养产氢菌;采用16S rRNA/rDNA序列分析和16S-23S rDNA间隔区测序技术对分离出产氢细菌进行鉴定.     

厨余生质能源化再利用评估

本研究进行厨余厌氧酦酵生质能源应用之可行性评估,针对厨余转换生质能之产能、用能及可不可能等3大部分,作为评估之重要范畴,同时根据厨余特性、分类收集、反应器形式、操作控制技术、反应物再利用去化管道及经济评估等6大因子,作为后续能源应用可行性评估之重要准则.搜集国内外相关资料,汇整相关国家政策发展规划及技术可行性,以期提供推动低碳城市厨余生质能源化政策之建议方向.根据目前评估数据显示,利用生质能源于LNG、CNG车及氢气电动车等车辆之燃料,已有相关之应用实绩,未来可经由建置生质能源中心示范厂先期进行试运,再配合公部门(例如:公共运输工具与清洁车等)及未来施政之需求与考虑后,再行验证推动示范之可行性.     

硫酸盐还原过程中氢分压的平衡与调节

采用两相厌氧工艺系统的产酸相反应器处理硫酸盐废水,通过连续流试验考察硫酸盐还原过程中分子氢的产生和去路．试验结果表明：在硫酸盐还原过程中,分子氢的产生者是产酸菌(AB)、产氢产乙酸菌(HPA)和利用脂肪酸的硫酸盐还原菌(ESRB);分子氢的消费者主要是利用氢的硫酸盐还原菌(HSRB)、产酸菌、产氢产乙酸菌和硫酸盐还原菌之间生物链式的协同代谢关系是维持系统低氢分压的前提．同时,揭示出在微生物的协同作用下,种间氢转移的途径和反应体系中氢分压的平衡与调节规律．     

混合菌种非固定化技术制氢反应器产氢效能

为了研究混合菌种非固定化技术生物制氢反应器的产氢效能，实验采用厌氧Hungate技术和MPN法，从采用混合菌种非固定化技术的生物制氢反应器厌氧活性污染中分离到210株优势发酵菌株，其中18株为产氢细菌（HPB）。实验结果表明，主要决定反应器产氢效能的因素是反应器内HPB的数量和活性。采用混合菌种非固定化技术可以充分发挥HPB的产氢活性，但是由于反应器内HPB的数量和比例不高，大大制约了混合菌种非固定技术生物制氢反应器效能的充分发挥。针对这一结论，提出采用有自絮凝能力的高效产氢细菌进行快速启动和投加高效产氢菌株的方法提高生物制氢反应器的产氢效能。     

蜂巢石强化 UASB 反应器处理垃圾渗滤液研究

向厌氧系统中投加载体材料是提升反应器稳定性及厌氧效率的有效手段.本试验将蜂巢石投加到上流式污泥床反应器(UASB)中,研究其对垃圾焚烧发电厂渗滤液厌氧消化效率的影响及厌氧污泥中产甲烷菌群落结构的变化.试验结果显示,投加蜂巢石缩短了UASB污泥驯化时间,与未投加蜂巢石的反应器相比,CODCr去除率提前12天稳定达到85％以上;污泥形貌观察及粒径分布测试表明,蜂巢石作为固定微生物的载体能够有效加速污泥颗粒化进程,运行至第15天,投加蜂巢石的反应器中粒径大于0.2 mm的污泥占比53.17％,高于未投加蜂巢石反应器的31.92％,形成了更加密实、更大的颗粒污泥.当有机负荷逐步提升至28.77 kgCODCr/(m3·d)时,投加蜂巢石的反应器CODCr去除率仍稳定达到97％,所能承受的最大有机负荷是未投加蜂巢石反应器的2倍以上.高通量测序表明,有机负荷提升后,投加蜂巢石反应器中产甲烷丝菌(Methanosaeta)代替产甲烷杆菌(Methanobacterium)成为优势种属.     

硫酸盐废水生物处理工艺研究进展

综述了国内外硫酸盐废水生物处理工艺的沿革和最新进展．20世纪90年代前国际上大多采用单相厌氧工艺处理硫酸盐废水,但常常运转失败．多数研究针对此问题,探讨硫酸盐还原菌(SRB)对产甲烷菌(MPB)和其他厌氧微生物的初级抑制作用和次级抑制作用机制以及改进措施．90年代以后,国内外开发出多种硫酸盐废水生物处理新工艺,重点介绍了单相吹脱工艺、硫酸盐还原与硫化物光合氧化联用工艺、硫酸盐还原与硫化物化学沉淀联用工艺、生物膜法工艺、两相厌氧工艺、两相厌氧与硫化物生物氧化联用工艺等．简述了每种工艺的要点和研究成果．同时,提出了进一步提高硫酸盐废水处理工艺效能的要点．     

Treatment of methanol wastewater with two-stage and two-phase anaerobic process

The two-stage and two-phase anaerobic process (TSTP) composed of hydrolytic acidification reactor,first-order and second-order external circulation anaerobic reactors (EC) was taken to treat methanol wastewater. Test results show that TSTP process is quick start-up in 51 d, and the maximum VFA of hydrolytic acidification reactor effluent reaches 876 mg/L. Under the condition of volume loading of 6.56 kgCOD/m3·d, COD removal rate of the first-order EC reactor is about 85%, and under the condition of volume loading of 1.02 kgCOD/m3·d, COD removal rate of the second-order EC reactor is about 50%. When the inflow COD of TSTP process is between 7000-11000 mg/L, its effluent COD is lower than 600 mg/L. In the biological conversion process of methanol into methane,the production of acetic acids as an intermediate product can be ignored and the direct production of methane from methanol is predominant.     

两种类型生物制氢反应器的运行及产氢特性

为探求反应器型式对发酵法生物制氢过程的影响,分别采用连续流搅拌槽式反应器(CSTR)和颗粒污泥膨胀床反应器(EGSB)接种厌氧活性污泥,从糖蜜废水中制取氢气.运行中控制温度为35℃,通过缩短水力停留时间(HRT)和增加进水COD质量浓度的方式逐渐提高容积负荷(OLR),分别对CSTR系统和EGSB系统的产氢速率、pH、液相末端产物及生物量进行研究.结果表明,两个系统中,产氢速率均随OLR提高而逐渐升高.CSTR的最佳产氢OLR为25~35 kg/(m3.d),而EGSB的最佳产氢OLR为70~80 kg/(m3.d);此时,CSTR系统的最大产氢速率为6.21 L/(L.h),EGSB系统的最大产氢速率可达18.0 L/(L.h).稳定运行期,EGSB系统的生物量为27.6 gVSS/L,而CSTR的生物量仅为7.8 gVSS/L,说明较高的生物量是生物制氢反应器稳定运行和高效产氢的关键.两个系统均可形成乙醇型发酵,说明发酵类型的形成不受反应器型式影响.与CSTR反应器相比,EGSB反应器具有更好的耐酸能力.     

Feasibility of an innovative integrated process of simultaneous desulfurization and denitrification for high strength wastewater

An anaerobic expanding-bed reactor was adopted to investigate the feasibility of an innovative integrated process of simultaneous desulfurization and denitrification （SDD） for high strength wastewater. In the reactor, heterotrophic bacteria （including sulfate reducing bacterium and denitrifying bacteria） and autotrophic bacteria （ including Thiobacillus denitrificans） cooperated together by incubating and enriching functional bac- teria on different carriers in the anaerobic activated sludge. Synthetic wastewater with high concentrations of sulfate and nitrate was employed. The experimental resuhs showed that the removal efficiency of sulfate and nitrate was above 85% , elemental sulfur was observed while nitrate was absent in effluent. The balance of sulfur, nitrogen and electron was discussed respectively, which indicated that the integrated SDD process could be actualized. These resuhs might provide a guidance to further investigate the key factors affecting the integrated SDD process and to improve the efficiency of desulfurization and denitrification in wastewater treatment.     

处理难生物降解有机物的厌氧颗粒污泥形成的技术进展

为了加快以上流式厌氧污泥床(Upflow Anaerobic Sludge Blanket，简称UASB)为代表的无载体厌氧反应器处理含难生物降解有机物废水的启动速度，综述了影响厌氧颗粒污泥形成的因素．此外，为了高效、快速地降解废水中的难生物降解有机物，建议向厌氧反应器中投加优势茵，以进一步提高厌氧反应器降解废水中难生物降解有机物的效率和速度．     

氨三乙酸对废水厌氧消化过程中产甲烷的影响

在连续运行的槽式搅拌反应器(CSTR)中考察了氨三乙酸(NTA)对厌氧消化反应器运行效果的影响.在中温35℃条件下,有机负荷化学需氧量(COD)为1.0～3.4g/(L.d)时,NTA的添加可使CSTR反应器COD去除率增加,产气率提高20%～42%,反应器中厌氧污泥的辅酶F430和F420浓度分别提高了1.33倍和24%;采用荧光原位杂交技术(FISH)对古细菌和甲烷八叠球菌进行了检测,结果表明:NTA还促进了甲烷八叠球菌的生长.作为微量元素螯合剂,NTA增强了微量元素镍的生物可利用性,优化了产甲烷菌的营养条件,强化了产甲烷菌对微量元素的生物吸收,对厌氧消化反应具有促进作用.     

Fermentative Hydrogen Production by Pure Culture with a New H2-producing Anaerobe

As a new clean energy source, the utilization and demand for hydrogen fuel are rapidly increasing. The integrated process of wastewater treatment of DESAR and energy recovery was developed in the studies. A new hydrogen anaerobe was isolated from the activated sludge. The optimal glucose concentration and the optimal initial pH were 12.0 g/L and 5. 5 respectively. The optimum C/N of the growth and hydrogen production in Rennanqilyf3 was (3.0 ～3.5): 1. The integrated process between DESAR system and biohydrogen production will be an important progress on energy recovery of DESAR system.     

五氯酚在厌氧生物体系中的降解及反馈作用

为探讨厌氧系统降解五氯酚(PCP)能力及PCP对厌氧系统的毒性影响,采用UASB反应器进行了PCP模拟污水连续流试验,考察了不同PCP浓度及负荷条件下系统对PCP和COD的降解去除规律,揭示了PCP对系统产酸相和产甲烷相影响状况.结果表明,在进水PCP含量从无到开始加入、PCP浓度提高以及PCP负荷提高3个冲击变化阶段,厌氧系统对PCP和COD的去除率均表现出先迅速降低后缓慢升高的变化趋势,运行稳定后PCP和COD的去除率分别达91.2%和83.5%以上.PCP对厌氧系统中微生物种群产生影响,使系统产酸菌由丁酸型菌群为主逐渐演替为丙酸型菌群为主,PCP对系统产甲烷菌活性产生明显抑制,甲烷产量由PCP加入前的12.1L/d降低至加入后的2.5L/d.