Combining maxRatio analysis with real-time PCR and its potential application for the prediction of Meloidogyne incognita in field samples.

Diagnosing and quantifying plant-parasitic nematodes is critical for efficient nematode management. Several studies have been performed intending to demonstrate nematode quantification via real-time quantitative PCR. However, most of the studies used dilution of DNA templates to make standard curves, while few studies used samples with different nematode numbers to make the standard curve, resulting in a high standard error. The objective of the present study was to develop a high quality standard curve using samples containing different numbers of the root-knot nematode Meloidogyne incognita and evaluate the results of real time qPCR with maxRatio analysis. The results showed that a high quality standard curve was obtained with different nematode numbers using specific primers and cycle threshold (Ct)-PCR (R²=0.9962, P<0.001, n=9). With the maxRatio analysis, the fractional cycle number (FCN)-PCR cycle curve and adjusted FCN (FCNadj)-PCR cycle curve had similar patterns as those of the Ct-PCR cycle curve. For quantification of nematodes in field soil samples, qPCR estimations with a FCNadj-PCR cycle standard curve was very close to microscope counting of second-stage juveniles (R²=0.9064, P<0.001, n=10), qPCR estimations with a FCN-PCR cycle standard curve was comparably good (R²=0.8509, P<0.001, n=10), and the biases with a Ct-PCR cycle standard curve were large (R²=0.7154, P<0.001, n=10). Moreover, we found that the concentration of Triton X-100 had less of an effect on FCN as compared to Ct, with delta FCN 0.52, and delta Ct 3.94 at 0.8% Triton. The present study suggests, that combined with maxRatio methods, real time qPCR could be a practical approach for quantifying M. incognita in field samples.

[Nitrous Oxide Production in Response to Oxygen in a Solar Greenhouse Vegetable Soil].

To explore the sources of peak nitrous oxide (N2O) flushes in solar greenhouse vegetable field, an experiment was conducted with two conventional vegetable soils under different initial volume fractions of oxygen (O2) (0%, 1%, 3%, 5%, and 10%). A robotized incubation system was employed to analyze the gas kinetics[O2, N2O, nitric oxide (NO), nitrogen (N2), and carbon dioxide (CO2)] every 6 or 8 h and calculate the N2O/(NO+N2O+N2) index. Sodium chlorate (NaClO3) was used to inhibit the oxidation of NO2- to further explore the relationship between N2O and nitrite (NO2-). A parallel off-line incubation in triplicates was conducted under similar conditions to measure the dynamic changes in inorganic nitrogen content[ammonia (NH4+), nitrate (NO3-), and NO2-]. The results showed that N2O production under anaerobic condition was significantly higher than that under aerobic condition. The peak value of N2O in the soil collected from a straw-added plot (DIS) was significantly higher than that in the soil from non-straw added plot (DI) (P<0.01) when the volume fraction of oxygen was ≤ 1%. Oxygen can directly affect N2O production by delaying or inhibiting N2O reduction, with significant increase in N2O production rate under oxygen-depleted condition. However, the N2production rate decreased significantly with increase in initial oxygen volume fraction (P<0.01). When the initial volume fraction of oxygen was between 1% and 5%, a continuous accumulation of NO2- was observed during the incubation period, resulting in the significantly higher N2O/(NO+N2O+N2) index than that in either anaerobic or 10% of oxygen treatments. Furthermore, a linear correlation was observed between NO2- and N2O at 5% and 10% of oxygen with the addition of NaClO3 (R2 ≥ 0.85). Incomplete denitrification and nitrifier denitrification from NO2- induction co-occurred in the range of 1% and 5% volume fractions of oxygen, significantly increasing the soil N2O production and N2O/(NO+N2O+N2) index. In addition, N2O production under anaerobic condition was significantly higher than that under aerobic condition (P<0.01).

[Occurrence, Distribution and Ecological Risk of Aantibiotics in Surface Water of the Gonghu Bay, Taihu Lake].

In order to assess the contamination degree and ecological risk of antibiotics in Gonghu Bay, Taihu Lake, the contamination characteristics of four kinds of antibiotics (tetracycline, quinolones, sulfonamides and macrolides) in surface water of the Gonghu Bay were analyzed using high performance liquid chromatography and tandem mass spectrometry (HPLC-MS/MS). The results showed that 13 of the 16 target antibiotics were detected in the Gonghu Bay. The mass concentrations of the 13 antibiotics ranged 0.005-4.720 µg·L-1, and OTC was of the highest concentration. The detection level of antibiotics was also different between the northern and the southern parts of Gonghu Bay, and the mass concentration of tetracycline in the northern part was higher than that in the southern part. Moreover, the concentrations of quinolones and sulfonamides in the southern part were larger than those in the northern part. The potential ecological risks of the detected antibiotics were assessed via using the risk quotients (RQs) approach. The preliminary results showed that 5 kinds of antibiotics: OTC, NOR, OFL, CIP and ENR were at high risk, accounting for 38.5% and 4 kinds of antibiotics TC, CTC, ROX and SMX were at medium risk, accounting for 30.8% in the antibiotics detected in Gonghu Bay, while the others (SMR, SMP, SQX, TMP) were at low risk. Ecological risk assessment revealed that some risks were present in antibiotics in the Gonghu Bay. The tetracyclines and quinolones were particularly prominent, which should be drawn enough attention, and corresponding preventive measures should be taken as well.