Challenging loop-mediated isothermal amplification(LAMP) technique for molecular detection of Toxoplasma gondii

Objective:To compare analytical sensitivity and specificity of a newly described DNA amplification technique.LAMP and nested PCR assay targeting the RE and Bl genes for the detection of Toxoplasma gondii(T.gondii) DNA.Methods:The analytical sensitivity of LAMP and ncstcd-PCR was obtained against 10-fold serial dilutions of T.gondii DNA ranging from 1 ng to 0.01 fg.DNA samples of other parasites and human chromosomal DNA were used to determine the specificity of molecular assays.Results:After testing LAMP and nesled-PCR in duplicate,the detection limit of RE-LAMP.B1-LAMP,RE-nested PCR and B1-nested PCR assays was one fg.100 fg,1 pg and 10 pg of T.gondii DNA respectively.All the LAMP assays and nested PCRs were 100% specific.The RE-LAMP assay revealed the most sensitivity for the detection of T.gondii DNA.Conclusions:The obtained results demonstrate that the LAMP technique has a greater sensitivity for detection of T.gondii.Furthermore,these findings indicate that primers based on the RE are more suitable than those based on the B1 gene.However,the B1-LAMP assay has potential as a diagnostic tool for detection of T.gondii.

Using uncertainty and sensitivity analysis for finding the best rainfall-runoff model in mountainous watersheds(Case study: the Navrood watershed in Iran)

In watersheds that have not sufficient meteorological and hydrometric data for simulating rainfall-runoff events, using geomorphologic and geomorphoclimatic characteristics of watershed is a conventional method for the simulation. A number of rainfall-runoff models utilize these characteristics such as Nash-IUH, Clark-IUH, Geomorphologic Instantaneous Unit Hydrograph(GIUH), Geomorphoclimatic Instantaneous Unit Hydrograph(GcIUH), GIUH-based Nash(GIUH-Nash) and GcIUH-based Clark(GcIUH-Clark). But all these models are not appropriate for mountainous watersheds. Therefore, the objective of this study is to select the best of them for the simulation. The procedure of this study is: a) selecting appropriate rainfall-runoff events for calibration and validation of six hybrid models, b) distinguishing the best model based on different performance criteria(Percentage Error in Volume(PEV); Percentage Error in Peak(PEP); Percentage Error in Time to Peak(PETP); Root Mean Square Error(RMSE) and Nash-Sutcliffe model efficiency coefficient(ENS)), c) Sensitivity analysis for determination of the most effective parameter at each model, d) Uncertainty determination of different parameters in each model and confirmation of the obtained results by application of the performance criteria. For application of this procedure, the Navrood watershed in the north of Iran as a mountainous watershed has been considered. The results showed that the ClarkIUH and GcIUH-Clark are suitable models for simulation of flood hydrographs, while other models cannot simulate flood hydrographs appropriately. The sensitivity analysis shows that the most sensitive parameters are the infiltration constant rate and time of concentration in the Clark-IUH model. Also, the most sensitive parameters include the infiltration constant rate and storage coefficient in the GcIUHClark model. The Clark-IUH and GcIUH-Clark models are more sensitive to their parameters. The Latin Hypercube Sampling(LHS) on Monte Carlo(MC) simulation method was used for evaluation of uncertainty of data in rainfall-runoff models. In this method 500 sets of data values are produced and then the peak discharge of flood hydrographs for each produced data set is simulated with rainfall-runoff models. The uncertainty of data changes the value of simulated peak discharge of flood hydrograph. The uncertainty analysis shows that the observed peak discharges of different rainfall-runoff events are within the range of values of simulated by the six hybrid rainfall-runoff models and IUH that inputs of these models were the produced data sets. The range of the produced peak discharge of flood hydrographs by the Clark-IUH and GcIUH-Clark models is wider than those of other models.