Validation of Bacteroidales-based microbial source tracking markers for pig fecal pollution and their application in two rivers of North China

• Pig feces is the predominant excrement produced by animal husbandry in China. • The PF, Pig-1-Bac^TaqMan, and Pig-2-Bac^TaqMan MST assays showed better performance. • The pig-specific MST assays can contribute to managing the pig fecal pollution. In China, pig feces is the predominant source of excrement produced by animal husbandry. Improper use or direct discharge of pig feces can result in contamination of natural water systems. Microbial source tracking (MST) technology can identify the sources of fecal pollution in environmental water, and contribute to the management of pig fecal pollution by local environmental protection agencies. However, the accuracy of such assays can be context-dependent, and they have not been comprehensively evaluated under Chinese conditions. We aimed to compare the performance of five previously reported pig-specific MST assays (PF, Pig-Bac1^SYBR, Pig-Bac2^SYBR, Pig-1-Bac^TaqMan, and Pig-2-Bac^TaqMan, which are based on Bacteroidales 16S rRNA gene markers) and apply them in two rivers of North China. We collected a total of 173 fecal samples from pigs, cows, goats, chickens, humans, and horses across China. The PF assay optimized in this study showed outstanding qualitative performance and achieved 100% specificity and sensitivity. However, the two SYBR green qPCR assays (Pig-Bac1^SYBR and Pig-Bac2^SYBR) cross-reacted with most non-pig fecal samples. In contrast, both the Pig-1-Bac^TaqMan and Pig-2-Bac^TaqMan assays gave 100% specificity and sensitivity. Of these, the Pig-2-Bac^TaqMan assay showed higher reproducibility. Our results regarding the specificity of these pig-specific MST assays differ from those reported in Thailand, Japan, and America. Using the PF and Pig-2-Bac^TaqMan assays, a field test comparing the levels of pig fecal pollution in rivers near a pig farm before and after comprehensive environmental pollution governance indicated that pig fecal pollution was effectively controlled at this location.     

Fecal source tracking by antibiotic resistance analysis on a watershed exhibiting low resistance

The ongoing development of microbial source tracking has made it possible to identify contamination sources with varying accuracy, depending on the method used. The purpose of this study was to test the efficiency of the antibiotic resistance analysis (ARA) method under low resistance by tracking the fecal sources at Turkey Creek, Oklahoma exhibiting this condition. The resistance patterns of 772 water-isolates, tested with nine antibiotics, were analyzed by discriminant analysis (DA) utilizing a five-source library containing 2250 isolates. The library passed various representativeness tests; however, two of the pulled-sample tests suggested insufficient sampling. The resubstitution test of the library individual sources showed significant isolate misclassification with an average rate of correct classification (ARCC) of 58%. These misclassifications were explained by low antibiotic resistance (Wilcoxon test P < 0.0001). Seasonal DA of stream E. coli isolates for the pooled sources human/livestock/deer indicated that in fall, the human source dominated (P < 0.0001) at a rate of 56%, and that human and livestock respective contributions in winter (35 and 39%), spring (43 and 40%), and summer (37 and 35%) were similar. Deer scored lower (17–28%) than human and livestock at every season. The DA was revised using results from a misclassification analysis to provide a perspective of the effect caused by low antibiotic resistance and a more realistic determination of the fecal source rates at Turkey Creek. The revision increased livestock rates by 13–14% (0.04 ≤ P ≤ 0.06), and decreased human and deer by 6–7%. Negative misclassification into livestock was significant (0.04 ≤ P ≤ 0.06). Low antibiotic resistance showed the greatest effect in this category.     

The role of lipids in fermentative propionate production from the co-fermentation of lipid and food waste

● Lipid can promote PA production on a target from food waste. ● PA productivity reached 6.23 g/(L∙d) from co-fermentation of lipid and food waste. ● Lipid promoted the hydrolysis and utilization of protein in food waste. ● Prevotella , Veillonella and norank _f _Propioni bacteriaceae were enriched. ● Main pathway of PA production was the succinate pathway. Food waste (FW) is a promising renewable low-cost biomass substrate for enhancing the economic feasibility of fermentative propionate production. Although lipids, a common component of food waste, can be used as a carbon source to enhance the production of volatile fatty acids (VFAs) during co-fermentation, few studies have evaluated the potential for directional propionate production from the co-fermentation of lipids and FW. In this study, co-fermentation experiments were conducted using different combinations of lipids and FW for VFA production. The contributions of lipids and FW to propionate production, hydrolysis of substrates, and microbial composition during co-fermentation were evaluated. The results revealed that lipids shifted the fermentation type of FW from butyric to propionic acid fermentation. Based on the estimated propionate production kinetic parameters, the maximum propionate productivity increased significantly with an increase in lipid content, reaching 6.23 g propionate/(L∙d) at a lipid content of 50%. Propionate-producing bacteria Prevotella, Veillonella, and norank_f_Propionibacteriaceae were enriched in the presence of lipids, and the succinate pathway was identified as a prominent fermentation route for propionate production. Moreover, the Kyoto Encyclopedia of Genes and Genomes functional annotation revealed that the expression of functional genes associated with amino acid metabolism was enhanced by the presence of lipids. Collectively, these findings will contribute to gaining a better understanding of targeted propionate production from FW.     

镁铁复合絮凝剂的表征及应用研究

以FeSO4、MgO和H2SO4为原料,制备了新型镁铁复合絮凝剂(PFMS),考察了PFMS的结构和Fe(Ⅲ)分布形态,为了考察自制PFMS、自制聚合硫酸铁(PFS)与工业PFS产品的絮凝性能差别,选择高岭土模拟浊度废水、直接耐晒黑G模拟染料废水和焦化废水生化出水,分别进行了去除浊度、色度和COD的絮凝实验.结果表明,Mg2+未参与聚合过程,但引入Mg2+提高了PFMS的稳定性.PFMS对浊度去除效果略高于自制PFS和工业PFS,余浊<5NTU;在碱性条件下,PFMS的脱色率、COD去除率高于自制PFS和工业PFS.     

Impact of microbial communities from tropical soils on the mobilization of trace metals during dissolution of cinnabar ore

Biodissolution experiments on cinnabar ore(mercury sulphide and other sulphide minerals,such as pyrite) were performed with microorganisms extracted directly from soil. These experiments were carried out in closed systems under aerobic and anaerobic conditions with 2 different soils sampled in French Guyana. The two main objectives of this study were(1) to quantify the ability of microorganisms to mobilize metals(Fe, Al, Hg) during the dissolution of cinnabar ore, and(2) to identify the links between the type and chemical properties of soils, environmental parameters such as season and the strategies developed by indigenous microorganisms extracted from tropical natural soils to mobilize metals.Results indicate that microbial communities extracted directly from various soils are able to(1) survive in the presence of cinnabar ore, as indicated by consumption of carbon sources and,(2) leach Hg from cinnabar in oxic and anoxic dissolution experiments via the acidification of the medium and the production of low molecular mass organic acids(LMMOAs). The dissolution rate of cinnabar in aerobic conditions with microbial communities ranged from 4.8 × 10~(-4) to 2.6 × 10~(-3) μmol/m~2/day and was independent of the metabolites released by the microorganisms. In addition, these results suggest an indirect action by the microorganisms in the cinnabar dissolution. Additionally, because iron is a key element in the dynamics of Hg, microbes were stimulated by the presence of this metal,and microbes released LMMOAs that leached iron from iron-bearing minerals, such as pyrite and oxy-hydroxide of iron, in the mixed cinnabar ore.     

新型重金属絮凝剂巯基乙酰化羟甲基聚丙烯酰胺的优化制备

为提高含铜废水的处理效果及简化处理流程,以聚丙烯酰胺(PAM)、甲醛、氢氧化钠、巯基乙酸(TGA)为原料,先经羟甲基化反应制备中间产物羟甲基聚丙烯酰胺(MPAM),再通过酰胺化反应将巯基接枝到MPAM分子链上,制备出新型重金属絮凝剂巯基乙酰化羟甲基聚丙烯酰胺(MAMPAM).以水样中Cu(Ⅱ)的去除率为考察目标,采用Plackett-Burman实验、最陡爬坡实验和响应面法中CCD实验优化MAMPAM的制备条件.结果表明,MAMPAM最优制备条件为:MPAM浓度0.31%、MPAM与TGA物质的量比为1∶3.2、反应介质p H值为4.76、反应温度为25℃、反应时间为2 h,在此条件下制备的MAMPAM对Cu(Ⅱ)的去除率为95.30%,与模型的理论预测值94.47%接近,相对偏差仅为0.83%,模型合理可靠.红外分析表明MPAM分子链上成功接上了巯基.MAMPAM对不同初始浓度的含Cu(Ⅱ)水样具有很好的去除效果,Cu(Ⅱ)去除率均能达到90%以上.MAMPAM有望成为一种有效的含铜废水处理剂,具有一定的应用前景.     

高温(70℃)玉米秸秆水解液产氢行为及群落结构

考察了高温条件下(70℃)牛粪堆肥、土壤、厌氧污泥、腐烂秸秆种接种物利用玉米秸秆水解液的产氢行为。结果表明:牛粪堆肥接种时达到最大的产气量(1355.7mL/L)和氢气产量(608.4mL/L),随后依次为腐烂秸秆、厌氧污泥和土壤。修改的Gompertz方程可以较好描述产氢量随时间变化趋势(R2>0.99)。牛粪堆肥接种时达到最大的产氢潜力(676.0mL/L),而土壤接种时的迟滞时间最小(9.8h)。DGGE图谱显示:不同接种物对应不同的微生物群落结构。Bacillus thermozeamaize,Enterobacter sp.JDM-19和Thermoanaerobacterium polysaccharolyticum strain,KMTHCJT和可能分别是牛粪堆肥,厌氧污泥和腐烂秸秆接种条件下的关键产氢微生物。     

无机-有机复合絮凝剂PST处理印染废水效果研究

以模拟印染废水和实际印染废水为实验对象,检验了一种新合成有机-无机复合絮凝剂(PST)的絮凝效果。结果表明,PST具有良好的絮凝性能。在受试条件下,絮凝效果与絮凝剂投加量、染料浓度有较大相关性。废水的pH值应用范围较宽;搅拌时间对COD去除率和脱色率没有显著影响。采用PST处理广东省高明市某印染厂的印染废水后,COD去除率为75.16%,脱色率为76.18%,处理效果良好,处理成本明显低于目前市场上普遍使用的聚铝等絮凝剂。     

Investigation of physico-chemical properties and microbial community during poultry manure co-composting process

Co-composting of poultry manure and rubber wood sawdust was performed with the ratio of 2:1(V/V) for a period of 60 days. An investigation was carried out to study the extracellular enzymatic activities and structural degradation utilizing Fourier transform infrared spectroscopy(FT-IR), thermogravimetry and differential thermal analysis(TG/DTA)and scanning electron microscopy(SEM). The microbial succession was also determined by using denaturing gel gradient electrophoresis(DGGE). The compost was able to reach its highest temperature of 71°C at day 3 and stabilized between 30 and 40°C for 8 weeks.CMCase, FPase and β-glucosidase acted synergistically in order to degrade the cellulosic substrate. The xylanase activities increased gradually during the composting and reached the peak value of 11.637 U/g on day 35, followed by a sharp decline. Both Li P and Mn P activities reached their peak values on day 35 with 0.431 and 0.132 U/g respectively. The FT-IR spectra revealed an increase in aromaticity and a decrease in aliphatic compounds such as carbohydrates as decomposition proceeded. TGA/DTG data exhibited significant changes in weight loss in compost samples, indicating degradation of organic matter. SEM micrographs showed higher amounts of parenchyma exposed on the surface of rubber wood sawdust at day 60, showing significant degradation. DGGE and 16 S r DNA analyses showed that Burkholderia sp., Pandoraea sp., and Pseudomonas sp. were present throughout the composting process. Ornithinibacillus sp. and Castellaniella ginsengisoli were only found in the initial stage of the composting, while different strains of Burkholderia sp. also occurred in the later stage of composting.     

Shifts in microbial community structure and diversity in a MBR combined with worm reactors treating synthetic wastewater

The chemical oxygen demand (COD) and NH₃-N removal, membrane fouling, sludge characteristics and microbial community structure in a membrane bioreactor (MBR) coupled with worm reactors (SSBWR) were evaluated for 210 days. The obtained results were compared to those from a conventional MBR (C-MBR) operated in parallel. The results indicated that the combined MBR (S-MBR) achieved higher COD and NH₃-N removal efficiency, slower increase in membrane fouling, better sludge settleability and higher activities of the related enzymes in the activated sludge. Denaturing gradient gel electrophoresis was used to analyze the microbial community structures in the C-MBR and the S-MBR. The microbial community structure in the S-MBR was more diverse than that in the C-MBR. Additionally, the slow-growing microbes such as Saprospiraceae, Actinomyces, Frankia, Clostridium, Comamonas, Pseudomonas, Dechloromonas and Flavobacterium were enriched in the S-MBR, further accounting for the sludge reduction, membrane fouling alleviation and wastewater treatment.