scholarly journals In Vivo Selection of Reduced Susceptibility to Carbapenems in Acinetobacter baumannii Related to ISAba1-Mediated Overexpression of the Natural blaOXA-66 Oxacillinase Gene

2009 ◽  
Vol 53 (6) ◽  
pp. 2657-2659 ◽  
Author(s):  
Samy Figueiredo ◽  
Laurent Poirel ◽  
Jacques Croize ◽  
Christine Recule ◽  
Patrice Nordmann
Keyword(s):  
Acinetobacter Baumannii ◽  
Reverse Transcription ◽  
Resistance Pattern ◽  
Pcr Analysis ◽  
In Vivo Selection ◽  
Reverse Transcription Pcr ◽  
Naturally Occurring ◽  
Carbapenemase Gene ◽  
Selection Of

ABSTRACT Two clonally related Acinetobacter baumannii isolates, A1 and A2, were obtained from the same patient. Isolate A2, selected after an imipenem-containing treatment, showed reduced susceptibility to carbapenems. This resistance pattern was related to insertion of the ISAba1 element upstream of the naturally occurring bla OXA-66 carbapenemase gene as demonstrated by sequencing, reverse transcription-PCR analysis, and inactivation of the bla OXA-66 gene.

scholarly journals In Vivo Analysis of Aspergillus fumigatus Developmental Gene Expression Determined by Real-Time Reverse Transcription-PCR

2008 ◽  
Vol 76 (8) ◽  
pp. 3632-3639 ◽  
Author(s):  
Fabrice N. Gravelat ◽  
Thomas Doedt ◽  
Lisa Y. Chiang ◽  
Hong Liu ◽  
Scott G. Filler ◽  
...  
Keyword(s):  
Gene Expression ◽  
Aspergillus Fumigatus ◽  
Real Time ◽  
Reverse Transcription ◽  
Invasive Pulmonary Aspergillosis ◽  
Pcr Analysis ◽  
Invasive Infection ◽  
Reverse Transcription Pcr

ABSTRACT Very little is known about the developmental stages of Aspergillus fumigatus during invasive aspergillosis. We performed real-time reverse transcription-PCR analysis on lung samples from mice with invasive pulmonary aspergillosis to determine the expression of A. fumigatus genes that are expressed at specific stages of development. In established infection, A. fumigatus exhibited mRNA expression of genes specific to developmentally competent hyphae, such as stuA. In contrast, mRNA of genes expressed by conidia and precompetent hyphae was not detected. Many genes required for mycotoxin synthesis, including aspHS, gliP, mitF, and metAP, are known to be expressed by developmentally competent hyphae in vitro. Interestingly, each of these genes was expressed at significantly higher levels during invasive infection than in vitro. The expression of gliP mRNA in vitro was found to be highly dependent on culture conditions. Furthermore, gliP expression was found to be dependent on the transcription factor StuA both in vitro and in vivo. Therefore, developmentally competent hyphae predominate during established invasive infection, and many mycotoxin genes are expressed at high levels in vivo. These results highlight the importance of the evaluation of putative virulence factors expressed by competent hyphae and analysis of gene expression levels during invasive infection rather than in vitro alone.

scholarly journals MicroRNA-182-5p relieves murine allergic rhinitis via TLR4/NF-κB pathway

Open Medicine ◽  
2020 ◽  
Vol 15 (1) ◽  
pp. 1202-1212
Author(s):  
Aichun Zhang ◽  
Yangzi Jin
Keyword(s):  
Allergic Rhinitis ◽  
Inflammatory Cells ◽  
Lavage Fluid ◽  
Specific Ige ◽  
Inflammatory Factors ◽  
Pcr Analysis ◽  
Reverse Transcription Pcr ◽  
Pathway Activation ◽  
Nasal Lavage Fluid

AbstractAllergic rhinitis (AR) is one of the most common chronic diseases. This study examined whether microRNA (miR)-182-5p plays a role in AR by regulating toll-like receptor 4 (TLR4). First, data demonstrated that TLR4 was a target of miR-182-5p. Subsequently, AR mouse model was established to explore the role of miR-182-5p and TLR4 in AR in vivo. Initially, quantitative reverse transcription-PCR (qRT-PCR) analysis indicated that miR-182-5p was downregulated, while TLR4 expression was upregulated in AR mice. Then we found that miR-182-5p mimic reduced the frequency of sneezing and nose rubbing of the AR mice. In addition, miR-182-5p mimic significantly increased ovalbumin (OVA)-specific IgE and leukotriene C4 expression levels in nasal lavage fluid (NLF) and serum of AR mice. miR-182-5p mimic decreased the number of inflammatory cells in NLF of AR mice. It also reduced the levels of inflammatory factors in the serum of AR mice, such as interleukin (IL)-4, IL-5, IL-13, IL-17 and tumor necrosis factor (TNF)-α, while increasing the release of IFN-γ and IL-2. Finally, miR-182-5p mimic inhibited NF-κB signaling pathway activation in AR mice. However, all effects of miR-182-5p mimic on AR mice were reversed by TLR4-plasmid. In conclusion, miR-182-5p/TLR4 axis may represent a novel therapeutic target for AR.

scholarly journals Reverse Transcription-PCR Analysis of Bottled and Natural Mineral Waters for the Presence of Noroviruses

2003 ◽  
Vol 69 (11) ◽  
pp. 6541-6549 ◽  
Author(s):  
Gilbert Thierry Lamothe ◽  
Thierry Putallaz ◽  
Han Joosten ◽  
Joey D. Marugg
Keyword(s):  
Reverse Transcription ◽  
Membrane Filtration ◽  
Limit Of Detection ◽  
Mineral Waters ◽  
Pcr Analysis ◽  
Rna Sequences ◽  
Natural Mineral ◽  
Reverse Transcription Pcr ◽  
Viral Particles ◽  
Nonspecific Amplification

ABSTRACT A seminested reverse transcription-PCR method coupled to membrane filtration was optimized to investigate the presence of norovirus (NV) RNA sequences in bottled and natural mineral waters. The recovery of viral particles by filtration varied between 28 and 45%, while the limit of detection of the overall method ranged from 6 to 95 viral particles. The assay was broadly reactive, as shown by the successful detection of 27 different viral strains representing 12 common genotypes of NVs. A total of 718 bottled and natural mineral water samples were investigated, including 640 samples of finished, spring, and line products (mostly 1 to 1.5 liters), collected from 36 different water brands of various types and from diverse geographic origins over a 2-year period. In addition, 78 samples of larger volume (10 and 400 to 500 liters) and environmental swabs were investigated. From the 1,436 analyses that were performed for the detection of NVs belonging to genogroups I and II, 34 samples (2.44%) were presumptively positive by seminested RT-PCR. However, confirmation by DNA sequence analysis revealed that all presumptive positive results were either due to nonspecific amplification or to cross-contamination. In conclusion, these results do not provide any evidence for the presence of NV genome sequences in bottled waters.

scholarly journals Selection of Valid Reference Genes for Reverse Transcription Quantitative PCR Analysis inHeliconius numata(Lepidoptera: Nymphalidae)

2016 ◽  
Vol 16 (1) ◽  
pp. 50 ◽  
Author(s):  
Florence Piron Prunier ◽  
Mathieu Chouteau ◽  
Annabel Whibley ◽  
Mathieu Joron ◽  
Violaine Llaurens
Keyword(s):  
Quantitative Pcr ◽  
Reverse Transcription ◽  
Reference Genes ◽  
Pcr Analysis ◽  
Reverse Transcription Quantitative Pcr ◽  
Selection Of

scholarly journals YY1-modulated Long Non-coding RNA SNHG12 Predicts and Promotes Gastric Cancer Metastasis via Activating miR-218-5p/YWHAZ-dependent β-catenin: A Bed to Bench Approach

2020 ◽  
Author(s):  
Tian Qi Zhang ◽  
Qingqiang Dai ◽  
Maneesh Kumarsing Beeharry ◽  
Zhenqiang Wang ◽  
Liping Su ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Transcription Factor ◽  
Peritoneal Carcinomatosis ◽  
Signaling Pathway ◽  
Cell Lines ◽  
Reverse Transcription ◽  
Reverse Transcription Pcr ◽  
Reporter Assays

Abstract Background: Gastric Cancer (GC) is one of the leading causes of cancer-related deaths and mortality. Long non-coding RNAs (lncRNAs) such as SNHG12 play important roles in the pathogenesis and progression of cancers. However, the role and significanve of SNHG12 in the metastasis of GC has not yet been thoroughly investigated.Methods: The SNHG12 expression pattern was detected in GC tissue samples from our faculty and cell lines using quantitative reverse transcription PCR. In vivo and in vitro gain and loss assays were conducted to observe the effects of SNHG12 regulation on GC cell metastasis potential. The underlying mechanisms of SNHG12 regulation on EMT and metastatic potential of GC cells were further determined by quantitative reverse transcription PCR, western blotting, dual luciferase reporter assays, co-immunoprecipitation, immunoprecipitation, RIP assays, TOPFlash/FOPFlash reporter assays and Ch-IP assays.Results: SNHG12 was upregulated in GC tissues and cell lines. The expression levels of SNHG12 in GC samples was significantly related to tumor invasion depth, TNM staging and lymph node metastasis, and was associated with poorer DFS and OS in the GC patients. SNHG12 was significantly highly expressed in peritoneal metastatic tissues from GC patients and mice subjects, suggesting a possible role of SNHG12 in peritoneal carcinomatosis from GC. Further in vivo and in vitro gain and loss assays indicated that SNHG12 promoted GC metastasis and EMT. Based on hypothetical bioinformatic analysis findings, our mechanistic analyses revealed that miR-218-5p was a direct target of SNHG12 and suggested that both SNHG12 and miR-218-5p could collectively regulate YWHAZ, forming the SNHG12/ miR-218-5p/YWHAZ axis, hereby decreasing the ubiquitination of β-catenin, thus activating the β-catenin signaling pathway and facilitating metastasis and EMT. Further analysis also revealed that the transcription factor YY1 could negatively modulate SNHG12 transcription.Conclusions: Our findings demonstrate that SNHG12 is be a potential prognostic marker and therapeutic target for GC. Negatively modulated by transcription factor YYI, SNHG12 promotes GC metastasis and EMT by regulating the miR-218-5p/YWHAZ axis and hence activating the β-catenin signaling pathway. Furthermore, we discovered high SNHG12 expression could be related to peritoneal carcinomatosis from GC but this requires further validation.

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