Effects of selected pharmaceuticals on riverine biofilm communities

2005 ◽  
Vol 51 (8) ◽  
pp. 655-669 ◽  
Author(s):  
John R Lawrence ◽  
George D.W Swerhone ◽  
Leonard I Wassenaar ◽  
Thomas R Neu
Keyword(s):  
In Situ Hybridization ◽  
Algal Biomass ◽  
Lectin Binding ◽  
Stable Carbon Isotope ◽  
Bacterial Biomass ◽  
Relative Increase ◽  
Carbon Utilization ◽  
The Impact ◽  
Isotope Analyses

Although pharmaceutical and therapeutic products are widely found in the natural environment, there is limited understanding of their ecological effects. Here we used rotating annular bioreactors to assess the impact of 10 µg·L–1of the selected pharmaceuticals ibuprofen, carbamazepine, furosemide, and caffeine on riverine biofilms. After 8 weeks of development, community structure was assessed using in situ microscopic analyses, fluor-conjugated lectin binding, standard plate counts, fluorescent in situ hybridization, carbon utilization spectra, and stable carbon isotope analyses. The biofilm communities varied markedly in architecture although only caffeine treated biofilms were significantly thicker. Cyanobacteria were suppressed by all 4 compounds, whereas the nitrogen containing caffeine, furosemide, and carbamazepine increased algal biomass. Ibuprofen and carbamazepine reduced bacterial biomass, while caffeine and furosemide increased it. Exopolymer content and composition of the biofilms was also influenced. Significant positive and negative effects were observed in carbon utilization spectra. In situ hybridization analyses indicated all treatments significantly decreased the gamma-proteobacterial populations and increased beta-proteobacteria. Ibuprofen in particular increased the alpha-proteobacteria, beta-proteobacteria, cytophaga-flavobacteria, and SRB385 probe positive populations. Caffeine and carbamazepine additions resulted in significant increases in the high GC354c and low GC69a probe positive cells. Live–dead analyses of the biofilms indicated that all treatments influenced the ratio of live-to-dead cells with controls having a ratio of 2.4, carbamazepine and ibuprofen being 3.2 and 3.5, respectively, and furosemide and caffeine being 1.9 and 1.7, respectively. Stable isotope analyses of the biofilms indicated δ13C values shifted to more negative values relative to control biofilms. This shift may be consistent with proportional loss of cyanobacteria and relative increase in algal biomass rather than incorporation of pharmaceutical carbon into microbial biofilm. Thus, at 10 ug·L–1levels pharmaceuticals exhibit both nutrient-like and toxic effects on riverine microbial communities.Key words: carbamazepine, ibuprofen, furosemide, caffeine, laser microscopy, digital imaging, in situ hybridization, stable isotopes.

Fluorescence in situ hybridization for identification and visualization of microorganisms in infected heart valve tissue as addition to standard diagnostic tests improves diagnosis of endocarditis

2019 ◽  
Vol 29 (5) ◽  
pp. 678-684
Author(s):  
Simone Eichinger ◽  
Judith Kikhney ◽  
Annette Moter ◽  
Alexandra Wießner ◽  
Walter B Eichinger
Keyword(s):  
In Situ Hybridization ◽  
Fluorescence In Situ Hybridization ◽  
Diagnostic Tests ◽  
Diagnostic Testing ◽  
Bacterial Biofilm ◽  
Causative Organism ◽  
Histopathological Diagnosis ◽  
Duke Criteria ◽  
The Impact

Abstract OBJECTIVES In infective endocarditis (IE), identification of the causative organism and consecutive treatment are crucial for patient survival. Although the macroscopic aspect resembles infected tissue, standard diagnostic tests often fail to allow one to identify bacteria. Fluorescence in situ hybridization (FISH) is a molecular, culture-independent technique that allows one to identify and visualize microorganisms within tissue and to recognize their morphology, number and activity. We analysed the diagnostic benefit of FISH/polymerase chain reaction (PCR) by comparing its results to those of standard diagnostic tests. METHODS From September 2015 to April 2018, 128 patients underwent first-time or redo valve surgery to treat IE. Patients were designated according to the modified Duke criteria as definite (n = 61), possible (n = 34) or rejected (n = 33) IE. Tissue specimens obtained intraoperatively were analysed using FISH/PCR in addition to undergoing standard diagnostic testing and PCR alone. RESULTS We used blood cultures to detect microorganisms in 67/128 patients; valve cultures, in 34/128; PCR, in 67/128; histopathological diagnosis showed IE in 72/128 cases. We were able to detect microorganisms in 103/128 cases using FISH/PCR, with 55/61 in definite IE. Furthermore, we were able to identify 26 cases of bacterial biofilm using FISH/PCR, despite antibiotic treatment of 61 in the definite, 13 in the possible and 1 in the rejected group, including 8/33 patients in the rejected group with active bacteria. In all cases, the patient’s therapy was altered. CONCLUSIONS FISH/PCR was used to identify microorganisms in cases in which standard diagnostic tests failed to provide sufficient results for various reasons. Furthermore, FISH/PCR enabled us to identify bacterial biofilms and to differentiate between active versus degraded bacteria, thus indicating the impact of treatment. Therefore, we suggest FISH/PCR as an additional diagnostic tool in IE alongside standard diagnostic tests.

scholarly journals Investigation of an Acetate-Fed Denitrifying Microbial Community by Stable Isotope Probing, Full-Cycle rRNA Analysis, and Fluorescent In Situ Hybridization-Microautoradiography

2005 ◽  
Vol 71 (12) ◽  
pp. 8683-8691 ◽  
Author(s):  
Maneesha P. Ginige ◽  
Jürg Keller ◽  
Linda L. Blackall
Keyword(s):  
In Situ Hybridization ◽  
Stable Isotope ◽  
Activated Sludge ◽  
Fluorescent In Situ Hybridization ◽  
Batch Reactor ◽  
External Source ◽  
Stable Isotope Probing ◽  
Rrna Gene ◽  
The Impact

ABSTRACT The acetate-utilizing microbial consortium in a full-scale activated sludge process was investigated without prior enrichment using stable isotope probing (SIP). [13C]acetate was used in SIP to label the DNA of the denitrifiers. The [13C]DNA fraction that was extracted was subjected to a full-cycle rRNA analysis. The dominant 16S rRNA gene phylotypes in the 13C library were closely related to the bacterial families Comamonadaceae and Rhodocyclaceae in the class Betaproteobacteria. Seven oligonucleotide probes for use in fluorescent in situ hybridization (FISH) were designed to specifically target these clones. Application of these probes to the sludge of a continuously fed denitrifying sequencing batch reactor (CFDSBR) operated for 16 days revealed that there was a significant positive correlation between the CFDSBR denitrification rate and the relative abundance of all probe-targeted bacteria in the CFDSBR community. FISH-microautoradiography demonstrated that the DEN581 and DEN124 probe-targeted cells that dominated the CFDSBR were capable of taking up [14C]acetate under anoxic conditions. Initially, DEN444 and DEN1454 probe-targeted bacteria also dominated the CFDSBR biomass, but eventually DEN581 and DEN124 probe-targeted bacteria were the dominant bacterial groups. All probe-targeted bacteria assessed in this study were denitrifiers capable of utilizing acetate as a source of carbon. The rapid increase in the number of organisms positively correlated with the immediate increase in denitrification rates observed by plant operators when acetate is used as an external source of carbon to enhance denitrification. We suggest that the impact of bacteria on activated sludge subjected to intermittent acetate supplementation should be assessed prior to the widespread use of acetate in the wastewater industry to enhance denitrification.

scholarly journals Significance of p53-Binding Protein 1 Nuclear Foci in Cervical Squamous Intraepithelial Lesions: Association With High-Risk Human Papillomavirus Infection and P16INK4a Expression

2020 ◽  
Vol 27 (1) ◽  
pp. 107327481990117
Author(s):  
Sayaka Kawashita ◽  
Katsuya Matsuda ◽  
Hisayoshi Kondo ◽  
Yuriko Kitajima ◽  
Yuri Hasegawa ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Human Papillomavirus ◽  
High Risk ◽  
Binding Protein ◽  
Histological Grade ◽  
Neoplastic Progression ◽  
Cervical Lesions ◽  
Nuclear Foci ◽  
The Impact

As p53-binding protein 1 (53BP1) localizes to the sites of DNA double-strand breaks and rapidly forms nuclear foci (NF), and its presence may be an indicator of endogenous genomic instability (GIN). We previously showed that 53BP1 NF in cervical cells increase with neoplastic progression, indicating the significance of 53BP1 expression for the estimation of malignant potential during cervical carcinogenesis. This study aimed to further elucidate the impact of 53BP1 expression as a biomarker for cervical squamous intraepithelial lesion (SIL). A total of 81 tissue samples, including 17 of normal cervical epithelium, 22 of cervical intraepithelial neoplasia (CIN) 1, 21 of CIN2, and 21 of CIN3, from patients positive for high-risk human papillomavirus (HR-HPV) were used for double-label immunofluorescence of 53BP1 and Ki-67/p16INK4a expression and HR-HPV in situ hybridization. We analyzed associations between 53BP1 expression type with parameters such as CIN grade, HR-HPV infection status, p16INK4a expression, and CIN prognosis. Expression type of 53BP1 was significantly associated with histological grade of CIN and HR-HPV in situ hybridization signal pattern ( P < .0001). There was a significant correlation between 53BP1 and p16INK4a expression levels ( r = .73, P < .0001). However, there was no association between 53BP1 expression type and CIN prognosis. We propose that 53BP1 expression type is a valuable biomarker for SIL, which can help estimate the grade and GIN of cervical lesions reflecting replication stress caused by the integration of HR-HPV to the host genome.

The impact of 2018 ASCO-CAP HER2 testing guidelines on breast cancer HER2 results. An audit of 2132 consecutive cases evaluated by immunohistochemistry and in situ hybridization

2020 ◽  
Vol 33 (9) ◽  
pp. 1783-1790
Author(s):  
Gelareh Farshid ◽  
Deepak Dhatrak ◽  
Amardeep Gilhotra ◽  
Barbara Koszyca ◽  
James Nolan
Keyword(s):  
Breast Cancer ◽  
In Situ Hybridization ◽  
Her2 Testing ◽  
The Impact

Evaluation of the impact of bioaugmentation and biostimulation by in situ hybridization and microelectrode

2003 ◽  
Vol 37 (9) ◽  
pp. 2206-2216 ◽  
Author(s):  
Hisashi Satoh ◽  
Satoshi Okabe ◽  
Yuki Yamaguchi ◽  
Yoshimasa Watanabe
Keyword(s):  
In Situ Hybridization ◽  
The Impact

scholarly journals Elucidating the Burden of HIV in Tissues Using Multiplexed Immunofluorescence and In Situ Hybridization: Methods for the Single-Cell Phenotypic Characterization of Cells Harboring HIV In Situ

2018 ◽  
Vol 66 (6) ◽  
pp. 427-446 ◽  
Author(s):  
Joshua J. Vasquez ◽  
Rajaa Hussien ◽  
Brandon Aguilar-Rodriguez ◽  
Henrik Junger ◽  
Dejan Dobi ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Phenotypic Characterization ◽  
Major Barrier ◽  
Formalin Fixed Paraffin ◽  
Cellular Source ◽  
Formalin Fixed Paraffin Embedded ◽  
Infected Cells ◽  
Hiv Cure Research ◽  
The Impact

Persistent tissue reservoirs of HIV present a major barrier to cure. Defining subsets of infected cells in tissues is a major focus of HIV cure research. Herein, we describe a novel multiplexed in situ hybridization (ISH) (RNAscope) protocol to detect HIV-DNA (vDNA) and HIV-RNA (vRNA) in formalin-fixed paraffin-embedded (FFPE) human tissues in combination with immunofluorescence (IF) phenotyping of the infected cells. We show that multiplexed IF and ISH (mIFISH) is suitable for quantitative assessment of HIV vRNA and vDNA and that multiparameter IF phenotyping allows precise identification of the cellular source of the ISH signal. We also provide semi-quantitative data on the impact of various tissue fixatives on the detectability of vDNA and vRNA with RNAscope technology. Finally, we describe methods to quantitate the ISH signal on whole-slide digital images and validation of the quantitative ISH data with quantitative real-time PCR for vRNA. It is our hope that this approach will provide insight into the biology of HIV tissue reservoirs and to inform strategies aimed at curing HIV.

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