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Using pyrosequencing and quantitative PCR to analyze microbial communities

Husen ZHANG

Frontiers of Environmental Science & Engineering 2011, Volume 5, Issue 1,   Pages 21-27 doi: 10.1007/s11783-011-0303-9

Abstract: New high-throughput technologies continue to emerge for studying complex microbial communities. In particular, massively parallel pyrosequencing enables very high numbers of sequences, providing a more complete view of community structures and a more accurate inference of the functions than has been possible just a few years ago. In parallel, quantitative real-time polymerase chain reaction (QPCR) allows quantitative monitoring of specific community members over time, space, or different environmental conditions. In this review, the principles of these two methods and their complementary applications in studying microbial ecology in bioenvironmental systems are discussed. The parallel sequencing of amplicon libraries and using barcodes to differentiate multiple samples in a pyrosequencing run are explained. The best procedures and chemistries for QPCR amplifications are also described and advantages of applying automation to increase accuracy are addressed. Three examples in which pyrosequencing and QPCR were used together to define and quantify members of microbial communities are provided: in the human large intestine, in a methanogenic digester whose sludge was made more bioavailable by a high-voltage pretreatment, and on the biofilm anode of a microbial electrolytic cell. The key findings in these systems and how both methods were used in concert to achieve those findings are highlighted.

Keywords： polymerase chain reaction (PCR)     microbial communities     pyrosequencing     gut     microbial fuel cell     sludge    

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Measurement and Control Technology for the Array of Low Cost PCR Microreactors

Li Xianming ,Zhang Yulin ,Li Jiantian

Strategic Study of CAE 2007, Volume 9, Issue 7,   Pages 76-81

Abstract:

In this paper a new style of polymerase chain reaction(PCR)equipmentIt is the concrete way to reduce the cost of PCR chips and improve the reliability of the chips.The measurement and control scheme for the low-cost PCR chips is designed.The uncertainty of indirect to the temperature of PCR liquid in low-cost PCR chips is also assessed.Its performance in temperature cycling meets the requirements of PCR.

Keywords： microreactors     measurement     control     polymerase chain reaction    

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Erratum to: Using pyrosequencing and quantitative PCR to analyze microbial communities

Husen ZHANG

Frontiers of Environmental Science & Engineering 2011, Volume 5, Issue 3,   Pages 488-488 doi: 10.1007/s11783-011-0342-2

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Safety evaluation of microbial pesticide (HaNPV) based on PCR method

Miao Zhao, Shufei Li, Qinghong Zhou, Dianming Zhou, Ning He, Zhiyong Qian

Frontiers of Chemical Science and Engineering 2019, Volume 13, Issue 2,   Pages 377-384 doi: 10.1007/s11705-018-1777-9

Abstract: Microbial pesticides can prevent and control diseases and pests of crops, and has become one of the important measures to ensure food and environmental safety. However, the potential harm of microbial pesticides to humans and animals is a serious concern at home and abroad. In this paper, we have investigated the infectivity and pathogenicity of a representative of viral microbial pesticides, helicoverpa armigera nuclear polyhedrosis virus (HaNPV), by specific and highly sensitive polymerase chain reaction technology. The results show that HaNPV can be gradually cleared in a short time after getting into blood of experimental rats, and does not infect other tissues or organs of animals; also indicate that the test subjects are not infectious to experimental rats after intravenous injection of HaNPV. Our method has good specificity and repeatability, and could provide an important reference for establishment of infectivity and pathogenicity detection methods for viral microbial pesticides in future.

Keywords： microbial pesticides     HaNPV     acute intravenous injection     infectivity     pathogenicity    

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A pooling strategy of a PCR-based assay to detect Angiostrongylus cantonensis in snail intermediate host

Fu-Rong WEI MS, Shan LV PhD, He-Xiang LIU, Ling HU, Yi ZHANG MS,

Frontiers of Medicine 2010, Volume 4, Issue 2,   Pages 204-207 doi: 10.1007/s11684-010-0043-4

Abstract: increase the efficiency in detecting and screening pathogen infections by polymerase chain reaction (PCRdiagnosis of in Two settings of specimens were prepared, divided into portions and detected by multiplex PCR

Keywords： Angiostrongylus cantonensis     Pomacea canaliculata     multiplex polymerase chain reaction     diagnosis     pooling    

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Research and application on high-end equipment of biological detection and molecular diagnosis

Peng Niancai,Li Lei,Li Zheng,Miao Baogang,Li Ming,Zhao Yao,Jiang Zhuangde

Strategic Study of CAE 2013, Volume 15, Issue 1,   Pages 57-62

Abstract: social development priorities of the real-time fluorescence quantitative polymerase chain reaction (PCR

Keywords： biological detection     molecular diagnosis     real-time quantitative PCR     ATP luminescence detection    

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Quantitative Analysis of the Effectiveness of Antigen- and Polymerase Chain Reaction-Based Combination Strategies for Containing COVID-19 Transmission in a Simulated Community Article

Qiangru Huang, Yanxia Sun, Mengmeng Jia, Ting Zhang, Fangyuan Chen, Mingyue Jiang, Qing Wang, Luzhao Feng, Weizhong Yang

Engineering 2023, Volume 28, Issue 9,   Pages 237-245 doi: 10.1016/j.eng.2023.01.004

Abstract: Antigen testing and its combination strategies were more efficient than polymerase chain reaction (PCRstrategies is the short turnaround time, which is also a critical factor to be optimized to improve PCR

Keywords： COVID-19     Antigen testing     PCR testing     Testing strategies    

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Full length cDNA cloning and expression of luteinizing hormone (LH) and which serum concentration was measured in half smooth tongue sole Cynoglossus semilaevis Günther

Liu Xuezhou,Shi Bao,Wang Shanshan,Xu Yongjiang,Li Xiaoxiao

Strategic Study of CAE 2014, Volume 16, Issue 9,   Pages 50-60

Abstract: Real-time PCR analysis showed that LH mRNA was expressed in all twelve tissues including brain，pituitaryQuantitative real- time PCR was used for analyzing the expression of female C.

Keywords： Cynoglossus semilaevis Günther     LH     cDNA cloning     Quantitative real-time PCR     RIA    

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Abundance and distribution of ammonia-oxidizing archaea in Tibetan and Yunnan plateau agricultural soils of China

Kun DING,Xianghua WEN,Liang CHEN,Daishi HUANG,Fan FEI,Yuyang LI

Frontiers of Environmental Science & Engineering 2014, Volume 8, Issue 5,   Pages 693-702 doi: 10.1007/s11783-014-0635-3

Abstract: Quantitative PCR (q-PCR) and clone library targeting on gene were used to quantify the abundances of

Keywords： ammonia-oxidizing archaea     ammonia-oxidizing bacteria     quantitative PCR     clone library     plateau    

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Interaction and independence on methane oxidation of landfill cover soil among three impact factors: water, oxygen and ammonium

Pinjing HE, Na YANG, Wenjuan FANG, Fan Lü, Liming SHAO

Frontiers of Environmental Science & Engineering 2011, Volume 5, Issue 2,   Pages 175-185 doi: 10.1007/s11783-011-0320-8

Abstract: To understand the influence patterns and interactions of three important environmental factors, i.e. soil water content, oxygen concentration, and ammonium addition, on methane oxidation, the soils from landfill cover layers were incubated under full factorial parameter settings. In addition to the methane oxidation rate, the quantities and community structures of methanotrophs were analyzed to determine the methane oxidation capacity of the soils. Canonical correspondence analysis was utilized to distinguish the important impact factors. Water content was found to be the most important factor influencing the methane oxidation rate and Type II methanotrophs, and the optimum value was 15% (w/w), which induced methane oxidation rates 10- and 6- times greater than those observed at 5% (w/w) and 20% (w/w), respectively. Ambient oxygen conditions were more suitable for methane oxidation than 3% oxygen. The addition of of ammonium induced different effects on methane oxidation capacity when conducted at low or high water content. With regard to the methanotrophs, Type II was sensitive to the changes of water content, while Type I was influenced by oxygen content. Furthermore, the methanotrophic acidophile, , was detected in soils with a pH of 4.9, which extended their known living environments.

Keywords： quantitative polymerase chain reaction (PCR)     denaturing gradient gel electrophoresis (DGGE)     principal    

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Assessment of antibiotic resistance genes in dialysis water treatment processes

Xuan Zhu, Chengsong Ye, Yuxin Wang, Lihua Chen, Lin Feng

Frontiers of Environmental Science & Engineering 2019, Volume 13, Issue 3, doi: 10.1007/s11783-019-1136-1

Abstract:

• Quantitative global ARGs profile in dialysis water was investigated.

• Totally 35 ARGs were found in the dialysis treatment train.

• 29 ARGs (highest) were found in carbon filtration effluent.

• erm and mtrD-02 occurred in the final effluent.

• The effluent was associated with health risks even after RO treatment.

Keywords： Dialysis water     Treatment process     Antibiotic resistance gene     High-throughput quantitative PCR     Horizontal    

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Seasonal microbial community shift in a saline sewage treatment plant

Qingmei YAN, Xuxiang ZHANG, Tong ZHANG, Herbert H P FANG

Frontiers of Environmental Science & Engineering 2011, Volume 5, Issue 1,   Pages 40-47 doi: 10.1007/s11783-011-0304-8

Abstract: Activated sludge was monthly sampled from a saline sewage treatment plant of Hong Kong (China) during June 2007 to May 2008 to analyze the microbial community shift along with environmental variations using denaturing gradient gel electrophoresis of polymerase chain reaction amplified 16S rDNA fragments. Environmental changes resulted into a seasonal microbial community shift characterized by alterations in species number and abundance in the sewage treatment plant. Correspondence analysis and cluster analysis on community structure profile showed that the 12 monthly samples fell into four groups, which is in accordance with season changing in Hong Kong. Canonical correspondence analysis revealed that and posed more significant effects on community structure than total phosphorus and total nitrogen, respectively. Compared with sludge retention time, influent total phosphorus had an inverse effect on the community structure shift, and chemical oxygen demand and showed a similar effects. Results of this study may contribute to the development of new knowledge involving the microbial community shift in sewage treatment plants.

Keywords： sewage treatment plants     polymerase chain reaction (PCR)     denaturing gradient gel electrophoresis (DGGE    

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662 A/G gene variation in human tumor necrosis factor receptor superfamily, member 9 (TNFRSF9)

QU Yanchun, YANG Ze, SUN Liang, JI Linong

Frontiers of Medicine 2008, Volume 2, Issue 3,   Pages 283-285 doi: 10.1007/s11684-008-0053-7

Abstract: diabetes mellitus by D-HPLC mutation screening method and confirmed our results by direct sequencing and PCR-RFLP

Keywords： D-HPLC mutation     development     autoimmune     PCR-RFLP     candidate    

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Nucleic acid-based diagnostics for infectious diseases in public health affairs

Albert Cheung-Hoi Yu, Greg Vatcher, Xin Yue, Yan Dong, Mao Hua Li, Patrick H. K. Tam, Parker Y.L. Tsang, April K.Y. Wong, Michael H.K. Hui, Bin Yang, Hao Tang, Lok-Ting Lau

Frontiers of Medicine 2012, Volume 6, Issue 2,   Pages 173-186 doi: 10.1007/s11684-012-0195-5

Abstract:

Infectious diseases, mostly caused by bacteria and viruses but also a result of fungal and parasitic infection, have been one of the most important public health concerns throughout human history. The first step in combating these pathogens is to get a timely and accurate diagnosis at an affordable cost. Many kinds of diagnostics have been developed, such as pathogen culture, biochemical tests and serological tests, to help detect and fight against the causative agents of diseases. However, these diagnostic tests are generally unsatisfactory because they are not particularly sensitive and specific and are unable to deliver speedy results. Nucleic acid-based diagnostics, detecting pathogens through the identification of their genomic sequences, have shown promise to overcome the above limitations and become more widely adopted in clinical tests. Here we review some of the most popular nucleic acid-based diagnostics and focus on their adaptability and applicability to routine clinical usage. We also compare and contrast the characteristics of different types of nucleic acid-based diagnostics.

Keywords： nucleic acid-based diagnostics     infectious disease     PCR     NASBA     LAMP     microarray     LOAC     public health affairs    

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Simultaneous detection and characterization of toxigenic：Clostridium difficile directly from clinical stool specimens

Hanjiang Lai, Chen Huang, Jian Cai, Julian Ye, Jun She, Yi Zheng, Liqian Wang, Yelin Wei, Weijia Fang, Xianjun Wang, Yi-Wei Tang, Yun Luo, Dazhi Jin

Frontiers of Medicine 2018, Volume 12, Issue 2,   Pages 196-205 doi: 10.1007/s11684-017-0560-5

Abstract:

We employed a multiplex polymerase chain reaction (PCR) coupled with capillary electrophoresis (mPCR-CEstrains, 44 (97.8%) were determined as non-ribotype 027 by the mPCR-CE, which was fully agreed with PCR

Keywords： Clostridium difficile     multiplex PCR     capillary electrophoresis     detection     characterization    

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Title Author Date Type Operation

Using pyrosequencing and quantitative PCR to analyze microbial communities

Husen ZHANG

Journal Article

Measurement and Control Technology for the Array of Low Cost PCR Microreactors

Li Xianming ,Zhang Yulin ,Li Jiantian

Journal Article

Erratum to: Using pyrosequencing and quantitative PCR to analyze microbial communities

Husen ZHANG

Journal Article

Safety evaluation of microbial pesticide (HaNPV) based on PCR method

Miao Zhao, Shufei Li, Qinghong Zhou, Dianming Zhou, Ning He, Zhiyong Qian

Journal Article

A pooling strategy of a PCR-based assay to detect Angiostrongylus cantonensis in snail intermediate host

Fu-Rong WEI MS, Shan LV PhD, He-Xiang LIU, Ling HU, Yi ZHANG MS,

Journal Article

Research and application on high-end equipment of biological detection and molecular diagnosis

Peng Niancai,Li Lei,Li Zheng,Miao Baogang,Li Ming,Zhao Yao,Jiang Zhuangde

Journal Article

Quantitative Analysis of the Effectiveness of Antigen- and Polymerase Chain Reaction-Based Combination Strategies for Containing COVID-19 Transmission in a Simulated Community

Qiangru Huang, Yanxia Sun, Mengmeng Jia, Ting Zhang, Fangyuan Chen, Mingyue Jiang, Qing Wang, Luzhao Feng, Weizhong Yang

Journal Article

Full length cDNA cloning and expression of luteinizing hormone (LH) and which serum concentration was measured in half smooth tongue sole Cynoglossus semilaevis Günther

Liu Xuezhou,Shi Bao,Wang Shanshan,Xu Yongjiang,Li Xiaoxiao

Journal Article

Abundance and distribution of ammonia-oxidizing archaea in Tibetan and Yunnan plateau agricultural soils of China

Kun DING,Xianghua WEN,Liang CHEN,Daishi HUANG,Fan FEI,Yuyang LI

Journal Article

Interaction and independence on methane oxidation of landfill cover soil among three impact factors: water, oxygen and ammonium

Pinjing HE, Na YANG, Wenjuan FANG, Fan Lü, Liming SHAO

Journal Article

Assessment of antibiotic resistance genes in dialysis water treatment processes

Xuan Zhu, Chengsong Ye, Yuxin Wang, Lihua Chen, Lin Feng

Journal Article

Seasonal microbial community shift in a saline sewage treatment plant

Qingmei YAN, Xuxiang ZHANG, Tong ZHANG, Herbert H P FANG

Journal Article

662 A/G gene variation in human tumor necrosis factor receptor superfamily, member 9 (TNFRSF9)

QU Yanchun, YANG Ze, SUN Liang, JI Linong

Journal Article

Nucleic acid-based diagnostics for infectious diseases in public health affairs

Albert Cheung-Hoi Yu, Greg Vatcher, Xin Yue, Yan Dong, Mao Hua Li, Patrick H. K. Tam, Parker Y.L. Tsang, April K.Y. Wong, Michael H.K. Hui, Bin Yang, Hao Tang, Lok-Ting Lau

Journal Article

Simultaneous detection and characterization of toxigenic：Clostridium difficile directly from clinical stool specimens

Hanjiang Lai, Chen Huang, Jian Cai, Julian Ye, Jun She, Yi Zheng, Liqian Wang, Yelin Wei, Weijia Fang, Xianjun Wang, Yi-Wei Tang, Yun Luo, Dazhi Jin

Journal Article