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Niche Differentiation of Phenol-Degrading Microorganisms in UASB Granular Sludge as Revealed by fluorescence in situ hybridization

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Engineering 2022年第2期8卷 61-66页

作者： Kengo Kubota Kei Igarashi Masayoshi Yamada Yasuyuki Takemura Yu-You Li Hideki Harada Department of Civil and Environmental Engineering Tohoku UniversitySendai 980-8579Japan Department of Urban Environmental Design and Engineering National Institute of TechnologyKagoshima CollegeKirishima 899-5193Japan New Industry Creation Hatchery Center Tohoku UniversitySendai 980-8579Japan

A microbial community structure of granules harvested from an anaerobic sludge blanket reactor treating phenolic wastewater was investigated using fluorescence in situ hybridization(FISH)and clone library construction.Clones of Syntrophorhabdaceae and Cryptanaerobacter were observed to be responsible for phenol degradation.For accurate taxonomic assignment of Cryptanaerobacter clones,phylogenetic analysis using nearly full-length 16S ribosomal RNA(rRNA)gene sequences was necessary.Three oligonucleotide probes were designed to detect the following three taxonomic groups:Syntrophorhabdaceae,Cryptanaerobacter,and Syntrophus.FISH analysis of thin sections of anaerobic granules showed a random distribution of bacteria and archaea.However,a well-defined distribution of Syntrophorhabdaceae,Cryptanaerobacter,and Syntrophus was observed.Cryptanaerobacter and Syntrophus were found on the outer layer of the granules and were closely associated with each other,while Syntrophorhabdaceae was located in the deeper part of the granules.Such specific distribution of the bacteria is most likely due to their metabolic association and affinity for the substrate.Phenol degradation in the granular sludge was observed to be carried out in the following way.First,Cryptanaerobacter converts phenol to benzoate,which is then degraded by Syntrophus into acetate.This syntrophic degradation of phenol occurs near the surface of the granule,where the phenol concen-tration is high.In the deeper part of the granule,where the phenol concentration is lower,Syntrophorhabdaceae degrades phenol into acetate.We observed that Syntrophorhabdaceae is less likely to produce benzoate as an intermediate to feed the neighboring organisms,which contradicts the theo-ries presented by previous studies.

关键词： Cryptanaerobacter fluorescence in situ hybridization Anaerobic phenol degradation Syntrophorhabdaceae Syntrophus UASB granular sludge

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fluorescence in situ hybridization polymorphism usinng 45S and 5S rDNA in different cultivars of sweet potato

Fluorescence in situ hybridization polymorphism usinng 45S a...

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江苏省遗传学会2016年学术年会

作者： Jianying Sun Yonghua Han Zongyun Li Institute of Integrative Plant Biology School of Life ScienceJiangsu Normal University

To detect if polymorphism could be observed in the hybridization patterns of different sweet potato cultivars,twenty sweet potato cultivars were analysed with two-colour fluorescence in situ hybridization（FISH） using the 45 S rDNA and 5 S rDNA.The number、region and intensity of rDNA loci on the metaphase chromosmes and interphase nucleus of all materials were observed.The average number of chromosmes in these cultivars is 90,but not always be.19 to 22 45 S rDNA signals should be examined,varies among cultivars.There were 3 regions of 45 S rDNA hybridization signals located in metaphase chromosmes.The 45 S rDNA signals were detected on the ends of chromosomes with high intensity brightness,weaker signals on the satellites and the weakest on the pericentromeric.The intensity of 45 S rDNA differs in different cultivars.When comparing the most strong signals,5 cultivars on the chromosome long arm,2 cultivars covering the entire chromosome and 2 cultivars on the short arm.The number of 5 S rDNA was consistent among these cultivars,all of them have 6 signals with different intensity.The location of 5 S rDNA is usually on the centromeric and the middle of chromosome arms.The former generally have a stronger signal,the latter have the weaker.When both of the 45 S and 5 S rDNA are present,their signals will always be weak,and they usually located near the pericentromere and in the middle of chromosme arm.In this study,a low level of polymorphism has been observed in the FISH pattern of various types of sweet potato cultivars.Thesse results may provide additional information on the influence of different origin on sweet potato genetics stocks.

关键词： fluorescence in situ hybridization sweet potato rDNA

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fluorescence in situ hybridization COMBINED WITH IMMUNOFLUORESCENT STAINING FOR RAPID DETECTION OF Nmyc AMPLIFICATION IN NEUROBLASTOMA

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Chinese Journal of Cancer Research 2004年第4期16卷 260-264页

作者：王伟 Marianne Ifversen 赵春亭汪洪毅赵洪国 Department of Hematology Affiliated Hospital Qingdao University Medical College Qingdao 266003 China Bloodbank 2034 Laboratory Center Rigshospitalet Biegdamsvej 9 2100 Copenhagen Denmark

Objective: To establish a method to improve the detection of disseminated tumor cells in bone marrow and peripheral blood samples of neuroblastoma patients and analysis of cytogenetic aberration. Methods: Immunofluorescent staining was performed using a cocktail of primary monoclonal neuroblastoma antibodies (14.G2a, 5.1H11). fluorescence in situ hybridization was applied with fluorescent probes specific for Nmyc genes afterwards. A novel computer assisted scanning system for automatic search, image analysis and repositioning of these positive cells was developed. Fifty-six bone marrow and peripheral blood samples from 7 patients were evaluated by this method. Results: fluorescence in situ hybridization can be combined with immunofluorescent staining in detecting Nmyc amplification in neuroblastoma patients. fluorescence in situ hybridization results correlated well with data obtained by conventional cytogenetic procedures. Conclusion: The technique described allows search of tumor cells in the bone marrow as well as detection of Nmyc amplification in interphase nuclei.

关键词： Neuroblastoma fluorescence in situ hybridization Nmyc gene

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Application of fluorescence in situ hybridization in the detection of bladder transitional-cell carcinoma: A multi-center clinical study based on Chinese population

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Asian Journal of Urology 2019年第1期6卷 114-121页

作者： Liqun Zhou Kaiwei Yang Xuesong Li Yi Ding Dawei Mu Hanzhong Li Yong Yan Jinyi Li Dongwen Wang Wei Li Yulong Cong Jiangping Gao Kewei Ma Yajun Xiao Sheng Zhang Hongyi Jiang Weilie Hu Qiang Wei Xunbo Jin Zhichen Guan Qingyong Liu Danfeng Xu Xin Gao Yongguang Jiang Weimin Gan Guang Sun Qing Wang Yanhui Liu Jianquan Hou Liping Xie Xishuang Song Fengshuo Jin Jiafu Feng Ming Cai Zhaozhao Liang Jie Zhang Dingwei Ye Lin Qi Lulin Ma Jianzhong Shou Yuping Dai Jianyong Shao Ye Tian Shizhe Hong Tao Xu Chuize Kong Zefeng Kang Yuexin Liu Xun Qu Benkang Shi Shaobin Zheng Yi Lin Shujie Xia Dong Wei Jianbo Wu Weiling Fu Zhiping Wang Jianbo Liang Department of Urology Peking University First HospitalBeijingChina Institute of Urology Peking UniversityBeijingChina National Urological Cancer Center BeijingChina Air Force General Hospital PLABeijingChina Department of Urology Peking Union Medical College HospitalBeijingChina Department of Urology Beijing Chao-Yang HospitalBeijingChina Department of Urology Armed-Police General HospitalBeijingChina Department of Urology Shanxi Medical University First HospitalTaiyuanChina Department of Urology Hebei Medical University Second HospitalShijiazhuangChina Department of Urology The General Hospital of the People’s Liberation ArmyBeijingChina Department of Urology First Affiliated Hospital of Jilin UniversityChangchunChina Department of Urology Union HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina Department of Pathology The First Affiliated Hospital of Fujian Medical UniversityFuzhouChina Department of Urology The Second Xiangya Hospital of Central South UniversityChangshaChina Department of Urology General Hospital of Guangzhou Military Command of PLAGuangzhouChina Department of Urology West China School of MedicineChengduChina Department of Urology Shangdong Province-owned HospitalJinanChina Department of Urology Peking University Shenzhen HospitalShenzhenChina Department of Urology The Central Hospital of Jinan CityJinanChina Department of Urology Shanghai Changzheng HospitalShanghaiChina Department of Urology The Third Affiliated Hospital of Sun Yat-sen UniversityGuangzhouChina Department of Urology Beijing Anzhen Hospital of Capital Medical UniversityBeijingChina Department of Urology The Second Affiliated Hospital of Xi’an Jiaotong UniversityXi’anChina Department of Urology The Second Hospital of Tianjin Medical UniversityTianjinChina Department of Urology People’s Hospital of Xinjiang Uygur Autonomous RegionWulumuqiChina Department of Urology Guangdong General HospitalGuangzhouChina Department of Urology The First Affil

Objective:To evaluate the diagnostic value of fluorescence in situ hybridization(FISH)in bladder cancer.Methods:We enrolled healthy volunteers and patients who were clinically suspected to have bladder cancer and conducted FISH tests and cytology examinations from August 2007 to December 2008.Receiver operating characteristic(ROC)curve analysis was performed and the area under curve(AUC)values were calculated for both the FISH and urine cytology tests.Results:A cohort of 988 healthy volunteers was enrolled to establish a reference range for the normal population.A total of 4807 patients with hematuria were prospectively,randomly enrolled for the simultaneous analysis of urine cytology,FISH testing,and a final diagnosis as determined by the pathologic findings of a biopsy or a surgically-excised specimen.Overall,the sensitivity of FISH in detecting transitional-cell carcinoma was 82.7%,while that of cytology was 33.4%(p<0.001).The sensitivity values of FISH for non-muscle invasive and muscle invasive bladder transitional-cell carcinoma were 81.7%and 89.6%,respectively(p=0.004).The sensitivity values of FISH for low and high grade bladder cancer were 82.6%and 90.1%,respectively(p=0.002).Conclusion:FISH is significantly more sensitive than voided urine cytology for detecting bladder cancer in patients evaluated for gross hematuria at all cancer grades and stages.Higher sensitivity using FISH was obtained in high grade and muscle invasive tumors.

关键词： Bladder transitionalcell carcinoma fluorescence in situ hybridization Detection Grade Stage

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fluorescence in situ hybridization rapidly detects three different pathogenic bacteria in urinary tract infection samples

Fluorescence in situ hybridization rapidly detects three dif...

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第二届中国临床微生物学大会暨微生物学与免疫学三江论坛

作者： Qing Wu Yan Li Ming Wang Xiao P.Pan Yong F.Tang Department of Clinical Laboratory Renmin Hospital of Wuhan University

Objective The detection of pathogenic bacteria in urine is an important criterion for diagnosing urinary tract infections（UTIs）.By using fluorescence in situ hybridization（FISH） with rRNA-targeted,fluorescently labeled oligonucleotide probes,bacterial pathogens present in urine samples were identified within 3-4 h.In this study,three probes that are specific for Escherichia coli,Enterococcus faecalis and Staphylococcus aureus were designed based on the conserved 16 SRNA sequences,whereas probe Eub338 broadly recognizes all bacteria.Results and Discussion The samples for these experiments included 76 E.coli infected samples,32 E.faecalis infected samples and 9 S.aureus infected samples.Compared to conventional methods of bacterial identification,the FISH method produced positive results for ＞ 90%of the samples tested.Fig.1.Fluorescent microscopy（magnification,×100） of microorganisms from urine samples after FISH.Part A:FISH with probes specific for Escherichia coli（green signal）;Part B:FISH with probes specific for Enterococcus faecalli（red signal）;Part C:FISH with probes specific for Staphylococcus aureus（green signal）.Conclusions FISH has the potential to become an extremely useful diagnostic tool for UTIs because it has a quick turnaround time and high accuracy.Acknowledgements This work has been supported by "the Fundamental Research Funds for the Central Universities"

关键词： Rapid detection Bacteria fluorescence in situ hybridization Urinary tract infection

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Clinical significance of chromosomal abnormalities detected by interphase fluorescence in situ hybridization in newly diagnosed multiple myeloma patients

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Chinese Medical Journal 2011年第19期124卷 2981-2985页

作者： HU Yu CHEN Lei SUN Chun-yan SHE Xiao-mei AI Li-sha QIN You Institute of Hematology Union Hospital Tongji Medical CollegeHuazhong University of Science and Technology Wuhan Hubei 430022 China

Background Chromosome 13q14 deletion （del13q14）, chromosome 1q21 gain （amp1q21） and chromosome 17p13 deletion （del17p13） are the most frequent chromosomal aberrations in multiple myeloma （MM）. They play an important role in prognosis. The aim of this study was to investigate the clinical significance of the chromosomal changes in Chinese MM patients.Methods Interphase fluorescence in situ hybridization （FISH） on bone marrow （BM） cells was performed in 72 enrolled MM patients. Relationships between chromosomal abnormalities and clinical features, response to therapies and prognosis were analyzed.Results As a result of interphase FISH, 77.8% （56/72） patients had chromosome changes. The incidences of each probe were RB1 51.4% （37/72）, D13S319 47.2% （34/72）, 1q21 45.8% （33/72） and p53 22.2% （12/72）. Osteolytic lesion,BM plasma cells index, serum calcium and serum M component were significantly correlated to del13q14. BM plasma cells and hemoglobin were correlated to amp1q21. Serum lactate dehydrogenase （LDH） was correlated with del17p13.Patients with del13q14 treated with bortezomib had a notably higher overall response rate than the patients treated with traditional chemotherapies （93% vs. 65%, P=0.048）. Patients carrying amp1q21 or/and del17p13 did not achieve satisfactory response to bortezomib. The median progression-free survival （PFS） for patients with amp1q21 was 5 months and patients without amp1q21 got 9-month PFS （P=0.001）. The median PFS for patients with del13q14 was 5 months （vs. 8 months, P=0.026）. The median PFS for patients with del17p13 was 3 months （vs. 8 months, P=0.002）.Patients with β2-microglobulin 〉5.5 mg/L also had a worse outcome, whose median PFS was 5 months （vs. 8 months,P=0.016）.Conclusions The prevalence of chromosomal abnormalities of MM patients was similar in Chinese and Caucasian people. Genetic changes were associated with patients＇ responses to therapies and prognosis.

关键词： multiply myeloma fluorescence in situ hybridization cytogenetics prognosis progression free survival

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Optimizing the hybridization chain reaction-fluorescence in situ hybridization(HCR-FISH)protocol for detection of microbes in sediments

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Marine Life Science & Technology 2021年第4期3卷 529-541页

作者： Zeyu Jia Yijing Dong Heng Xu Fengping Wang State Key Laboratory of Microbial Metabolism Joint International Research Laboratory of Metabolic and Developmental SciencesSchool of Life Sciences and BiotechnologyShanghai Jiao Tong UniversityShanghai 200240China School of Physics and Astronomy Shanghai Jiao Tong UniversityShanghai 200240China Institute of Natural Science Shanghai Jiao Tong UniversityShanghai 200240China School of Oceanography Shanghai Jiao Tong UniversityShanghai 200240China

fluorescence in situ hybridization(FISH)is a canonical tool commonly used in environmental microbiology research to visualize targeted cells.However,the problems of low signal intensity and false-positive signals impede its widespread application.Alternatively,the signal intensity can be amplified by incorporating hybridization Chain Reaction(HCR)with FISH,while the specificity can be improved through protocol modification and proper counterstaining.Here we optimized the HCR-FISH protocol for studying microbes in environmental samples,particularly marine sediments.Firstly,five sets of HCR initiator/amplifier pairs were tested on the laboratory-cultured bacterium Escherichia coli and the archaeon Methano-coccoides methylutens,and two sets displayed high hybridization efficiency and specificity.Secondly,we tried to find the best combination of sample pretreatment methods and HCR-FISH protocol for environmental sample analysis with the aim of producing less false positive signals.Various detachment methods,extraction methods and formulas of hybridization buffer were tested using sediment samples.Thirdly,an image processing method was developed to enhance the DAPI signal of microbial cells against that of abiotic particles,providing a reliable reference for FISH imaging.In summary,our optimized HCR-FISH protocol showed promise to serve as an addendum to traditional FISH for research on environmental microbes.

关键词： fluorescence in situ hybridization hybridization chain reaction HCR-FISH Microbial detection Sediment

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Development of taxonomic rRNA-targeted probes of two harmful algae: Prorocentrum minimum and Karenia mikimotoi by fluorescence in situ hybridization

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Acta Oceanologica Sinica 2013年第2期32卷 66-75页

作者： CHEN Guofu WANG Quanfu ZHANG Chunyun ZHANG Baoyu WANG Guangce LU Douding XU Zhong YAN Peishen Harbin Institute of Technology (Weihai) The First Institute of Oceanography State Ocenic Administration Institute of Oceanology Chinese Academy of Sciences The Second Institute of Oceanography State Ocenic Administration

Harmful algal blooms recently have been under the spotlight throughout the world, because of their nega- tive impact on the marine environment, aquaculture, fisheries as well as public health. The development of methods for rapid and precise identification and quantification of causative species is essential for the warning and monitoring of blooms, among which the techniques based on taxonomic probes are the most favored. In this study, two harmful algae, i.e., Prorocentrum minimum and Karenia mikimotoi were tak- en into consideration. The partial large subunit rDNA （D1-D2） of both species were firstly PCR-amplified, cloned and sequenced. The obtained sequences were then introduced to carry out alignment analysis for gene specific regions. Three respective candidate probes for each species were designed and used to screen the optimal probe by performing fluorescence in situ hybridization （FISH） tests. The results showed that the probes Pmin0443 and Kmik0602 displayed the best hybridization for P. minimum and K.. mikimotoi, respectively. Both the specific （taxonomic）（Pmin0443 and Kmik0602） and the control probes （UniC0512 and UniR0499） were used for cross-reactivity tests with other microalgae in our laboratory. The probes Pmin0443 and Kmik0602 are specific and could be served as taxonomic probes introduced into the tech- niques targeting rRNA, such as FISH, sandwich hybridization, and DNA-microarray assay of P minimum and K. mikimotoi in the future. Finally, FISH analyses with both probes were performed on the simulated field samples. The probes could hybridize exclusively with the target cells well, and no significant differ- ence （p 〉0.05） was observed in the cell densities of the samples determined by FISH and light microscopy （LM）. All suggest that the probes are specific and could be introduced into FISH for the monitoring of both harmful algae.

关键词： Prorocentrum minimum Karenia mikimotoi fluorescence in situ hybridization taxonomic probe

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Association of chromosome 7 aneuploidy measured by fluorescence in situ hybridization assay with muscular invasion in bladder cancer

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Cancer Communications 2020年第4期40卷 167-180页

作者： Xiayao Diao Jinhua Cai Junjiong Zheng Jianqiu Kong Shaoxu Wu Hao Yu Hao Huang Weibin Xie Xiong Chen Chengran Huang Lifang Huang Haide Qin Jian Huang Tianxin Lin Department of Urology Sun Yat-sen Memorial HospitalSun Yat-senUniversityGuangzhouGuangdong510120P.R.China Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation Sun Yat-Sen Memorial HospitalSun Yat-Sen UniversityGuangzhouGuangdong510120P.R.China Department of Neurology Sun Yat-sen Memorial HospitalSun Yat-sen UniversityGuangzhouGuangdong510120P.R.China State Key Laboratory of Oncology in South China GuangzhouGuangdong510120P.R.China

Background:The preoperative prediction of muscular invasion status is important for adequately treating bladder cancer(BC)but nevertheless,there are some existing dilemmas in the current preoperative diagnostic accuracy of BC with muscular invasion.Here,we investigated the potential association between the fluorescence in situ hybridization(FISH)assay and muscular invasion among patients with BC.A cytogenetic-clinical nomogram for the individualized preoperative differentiation of muscle-invasive BC(MIBC)from non-muscle-invasive BC(NMIBC)is also proposed.Methods:All eligible BC patients were preoperatively tested using a FISH assay,which included 4 sites(chromosome-specific centromeric probe[CSP]3,7,and 17,and gene locus-specific probe[GLP]-p16 locus).The correlation between the FISH assay and BC muscular invasion was evaluated using the Chi-square tests.In the training set,univariate and multivariate logistic regression analyses were used to develop a cytogenetic-clinical nomogram for preoperative muscular invasion prediction.Then,we assessed the performance of the nomogram in the training set with respect to its discriminatory accuracy and calibration for predicting muscular invasion,and clinica usefulness,which were then validated in the validation set.Moreover,model comparison was set to evaluate the discrimination and clinical usefulness between the nomogram and the individual variables incorporated in the nomogram.Results:Muscular invasion was more prevalent in BC patients with positive CSP3,CSP7 and CSP17 status(OR[95%CI],2.724[1.555 to 4.774],P<0.001;3.406[1.912 to 6.068],P<0.001 and 2.483[1.436 to 4.292],P=0.001,respectively).Radiologydetermined tumor size,radiology-determined clinical tumor stage and CSP7 status were identified as independent risk factors of BC muscular invasion by the multivariate regression analysis in the training set.Then,a cytogenetic-clinical nomogram incorporating these three independent risk factors was constructed and was observed to have satisfactory discrimination in the trai

关键词： chromosome aneuploidy fluorescence in situ hybridization muscular invasion nomogram preoperative prediction urinary bladder neoplasms

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Comparison of fluorescence in situ hybridization and Immunohistochemistry for Assessment of HER-2 Status in Breast Cancer Patients

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Journal of Huazhong University of Science and Technology(Medical Sciences) 2009年第3期29卷 354-358页

作者：王琳王晓蓓聂秀马玲 Department of Clinical Laboratory Union Hospital Tongji Medical College Huazhong University of Science and Technology

The accurate assessment ofa proto-oncogene, human epidermal growth factor receptor-2 gene （HER-2）, is extremely important for the therapy and prognosis of breast cancer. Currently, immunohistochemistry （IHC） is the method widely used for the detection of HER-2 protein. fluorescence in situ hybridization （FISH） has been suggested to be a golden standard assay for HER-2 amplification. This study examined the expression and amplification of HER-2 in paraffin-embedded sections of breast cancer tissues, and compared the two methods on the measurement of HER-2 status. HER-2 gene and protein were determined in breast cancer samples from 52 Chinese women by FISH and IHC respectively. The findings indicated that the HER-2 gene amplification was found in 18 cases （34.6%） by FISH and the HER-2 protein over-expression （score 3＋） in 15 cases （28.8%） by IHC. hnmunohistochemically, 28.6% of the cases scored as 2＋ and 93.3% of the cases scored as 3＋ were HER-2-positive by FISH. There was a significant correlation between the HER-2 gene amplification and HER-2 protein over-expression in breast cancer （P〈0.005）. No correlation was noted between the HER-2 gene amplification and any of the clinicopathological parameters examined, including age, menopausal status, menarche age, tumor size, histological tumor type, histological grade, lymph node status, and the expression of ER and PR. It was concluded that the detection of HER-2 gene amplification in breast cancer by FISH is valuable and can compare with HER-2 protein detection by IHC.

关键词： HER-2 fluorescence in situ hybridization immunohistochemistry breast cancer

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