scholarly journals Development of a Reverse Transcription Loop-Mediated Isothermal Amplification Method for the Rapid Detection of Subtype H7N9 Avian Influenza Virus

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Hongmei Bao ◽  
Yuhui Zhao ◽  
Yunhe Wang ◽  
Xiaolong Xu ◽  
Jianzhong Shi ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Reverse Transcription ◽  
Influenza A ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
H7n9 Virus ◽  
Loop Mediated Isothermal Amplification ◽  
H7n9 Influenza Virus ◽  
Virus Rna ◽  
H7n9 Influenza

A novel influenza A (H7N9) virus has emerged in China. To rapidly detect this virus from clinical samples, we developed a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for the detection of the H7N9 virus. The minimum detection limit of the RT-LAMP assay was 0.01 PFU H7N9 virus, making this method 100-fold more sensitive to the detection of the H7N9 virus than conventional RT-PCR. The H7N9 virus RT-LAMP assays can efficiently detect different sources of H7N9 influenza virus RNA (from chickens, pigeons, the environment, and humans). No cross-reactive amplification with the RNA of other subtype influenza viruses or of other avian respiratory viruses was observed. The assays can effectively detect H7N9 influenza virus RNA in drinking water, soil, cloacal swab, and tracheal swab samples that were collected from live poultry markets, as well as human H7N9 virus, in less than 30 min. These results suggest that the H7N9 virus RT-LAMP assays were efficient, practical, and rapid diagnostic methods for the epidemiological surveillance and diagnosis of influenza A (H7N9) virus from different resource samples.

scholarly journals Emergence of H7N9 Influenza A Virus Resistant to Neuraminidase Inhibitors in Nonhuman Primates

2015 ◽  
Vol 59 (8) ◽  
pp. 4962-4973 ◽  
Author(s):  
Yasushi Itoh ◽  
Shintaro Shichinohe ◽  
Misako Nakayama ◽  
Manabu Igarashi ◽  
Akihiro Ishii ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Influenza A ◽  
Cynomolgus Macaque ◽  
Influenza Virus Infection ◽  
Influenza Viruses ◽  
H7n9 Virus ◽  
Macaque Model ◽  
H7n9 Influenza Virus ◽  
Number Of Patients ◽  
H7n9 Influenza

ABSTRACTThe number of patients infected with H7N9 influenza virus has been increasing since 2013. We examined the efficacy of neuraminidase (NA) inhibitors and the efficacy of a vaccine against an H7N9 influenza virus, A/Anhui/1/2013 (H7N9), isolated from a patient in a cynomolgus macaque model. NA inhibitors (oseltamivir and peramivir) barely reduced the total virus amount because of the emergence of resistant variants with R289K or I219T in NA [residues 289 and 219 in N9 of A/Anhui/1/2013 (H7N9) correspond to 292 and 222 in N2, respectively] in three of the six treated macaques, whereas subcutaneous immunization of an inactivated vaccine derived from A/duck/Mongolia/119/2008 (H7N9) prevented propagation of A/Anhui/1/2013 (H7N9) in all vaccinated macaques. The percentage of macaques in which variant H7N9 viruses with low sensitivity to the NA inhibitors were detected was much higher than that of macaques in which variant H5N1 highly pathogenic influenza virus was detected after treatment with one of the NA inhibitors in our previous study. The virus with R289K in NA was reported in samples from human patients, whereas that with I219T in NA was identified for the first time in this study using macaques, though no variant H7N9 virus was reported in previous studies using mice. Therefore, the macaque model enables prediction of the frequency of emerging H7N9 virus resistant to NA inhibitorsin vivo. Since H7N9 strains resistant to NA inhibitors might easily emerge compared to other influenza viruses, monitoring of the emergence of variants is required during treatment of H7N9 influenza virus infection with NA inhibitors.

scholarly journals Rapid and Sensitive Detection of Tomato Brown Rugose Fruit Virus in Tomato and Pepper Seeds by Reverse Transcription Loop-Mediated Isothermal Amplification Assays (Real Time and Visual) and Comparison With RT-PCR End-Point and RT-qPCR Methods

2021 ◽  
Vol 12 ◽  
Author(s):  
Domenico Rizzo ◽  
Daniele Da Lio ◽  
Alessandra Panattoni ◽  
Chiara Salemi ◽  
Giovanni Cappellini ◽  
...  
Keyword(s):  
Real Time ◽  
Reverse Transcription ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
The European Union ◽  
Rt Pcr ◽  
Loop Mediated Isothermal Amplification ◽  
End Point ◽  
Specificity And Sensitivity ◽  
The Status

Tomato brown rugose fruit virus (ToBRFV) represents an emerging viral threat to the productivity of tomato and pepper protected cultivation worldwide. This virus has got the status of quarantine organism in the European Union (EU) countries. In particular, tomato and pepper seeds will need to be free of ToBRFV before entering the EU and before coming on the market. Thus, lab tests are needed. Here, we develop and validate a one-step reverse transcription LAMP platform for the detection of ToBRFV in tomato and pepper leaves, by real-time assay [reverse transcription loop-mediated isothermal amplification (RT-LAMP)] and visual screening (visual RT-LAMP). Moreover, these methods can also be applied successfully for ToBRFV detection in tomato and pepper seeds. The diagnostic specificity and sensitivity of both RT-LAMP and visual RT-LAMP are both 100%, with a detection limit of nearly 2.25 fg/μl, showing the same sensitivity as RT-qPCR Sybr Green, but 100 times more sensitive than end-point RT-PCR diagnostic methods. In artificially contaminated seeds, the proposed LAMP assays detected ToBRFV in 100% of contaminated seed lots, for up to 0.025–0.033% contamination rates in tomato and pepper, respectively. Our results demonstrate that the proposed LAMP assays are simple, inexpensive, and sensitive enough for the detection of ToBRFV, especially in seed health testing. Hence, these methods have great potential application in the routine detection of ToBRFV, both in seeds and plants, reducing the risk of epidemics.

Sign in / Sign up

Export Citation Format

Share Document