A Lab-on-a-Chip Approach for the Detection of the Quarantine Potato Cyst Nematode Globodera pallida.

The potato cyst nematode (PCN), Globodera pallida, has acquired significant importance throughout Europe due to its widespread prevalence and negative effects on potato production. Thus, rapid and reliable diagnosis of PCN is critical during surveillance programs and for the implementation of control measures. The development of innovative technologies to overcome the limitations of current methodologies in achieving early detection is needed. Lab-on-a-chip devices can swiftly and accurately detect the presence of certain nucleotide sequences with high sensitivity and convert the presence of biological components into an understandable electrical signal by combining biosensors with microfluidics-based biochemical analysis. In this study, a specific DNA-probe sequence and PCR primers were designed to be used in a magnetoresistive biosensing platform to amplify the internal transcribed spacer region of the ribosomal DNA of G. pallida. Magnetic nanoparticles were used as the labelling agents of asymmetric PCR product through biotin−streptavidin interaction. Upon target hybridization to sensor immobilized oligo probes, the fringe field created by the magnetic nanoparticles produces a variation in the sensor's electrical resistance. The detection signal corresponds to the concentration of target molecules present in the sample. The results demonstrate the suitability of the magnetic biosensor to detect PCR target product and the specificity of the probe, which consistently distinguishes G. pallida (DV/V > 1%) from other cyst nematodes (DV/V < 1%), even when DNA mixtures were tested at different concentrations. This shows the magnetic biosensor's potential as a bioanalytical device for field applications and border phytosanitary inspections.

A study to evaluate the effect of ultrasound treatment on nodules in multiple sclerosis patients.

Purpose/aim: Ultrasound has demonstrated anti-inflammatory and pain-relief benefits in several conditions such as cellulite or trauma events. We assessed the efficacy of ultrasound therapy on nodules associated with first-line treatments in multiple sclerosis patients. MATERIALS AND METHODS: Twenty-two multiple sclerosis patients were enrolled during 2013 and randomized to two groups: in the control group patients were treated only with a conventional gel prescribed for cellulite and nodules, while in the experimental group the gel was combined with ultrasound therapy. Patients were treated during 10 weeks and followed up for 10 additional weeks. Three nodules were assessed for each patient, measuring size, pain and redness at 0, 10 and 20 weeks. RESULTS: We found a significant decrease in both groups in size, pain and redness across the three visits (p < 0.0001 for size, p = 0.01 and p < 0.0001 for pain, and p = 0.0002 and p < 0.0001 for redness, respectively for the difference at visit 2 and 3 with respect to visit 1). More interestingly, we observed a greater reduction in pain and redness in the ultrasound-treated group, but the difference was only statistically significant at 10 weeks (p = 0.01 for both pain and redness). On the third visit, no differences between control and experimental groups were detected, both achieving the same levels in measured variables. CONCLUSIONS: Both treatments are useful to improve skin reaction after first-line treatments, but ultrasound in combination with gel achieves a faster reduction in pain and redness, suggesting that ultrasound treatment might be a good analgesic for nodule management in multiple sclerosis patients.

FTA-LAMP based biosensor for a rapid in-field detection of Globodera pallida-the pale potato cyst nematode.

The combination of a sensitive and specific magnetoresistive sensing device with an easy DNA extraction method and a rapid isothermal amplification is presented here targeting the on-site detection of Globodera pallida, a potato endoparasitic nematode. FTA-cards were used for DNA extraction, LAMP was the method developed for DNA amplification and a nanoparticle functionalized magnetic-biosensor was used for the detection. The combinatorial effect of these three emerging technologies has the capacity to detect G. pallida with a detection limit of one juvenile, even when mixed with other related species. This combined system is far more interesting than what a single technology can provide. Magnetic biosensors can be combined with any DNA extraction protocol and LAMP forming a new solution to target G. pallida. The probe designed in this study consistently distinguished G. pallida (∆Vac binding/Vac sensor above 1%) from other cyst nematodes (∆Vac binding/Vac sensor below 1%). It was confirmed that DNA either extracted with FTA-cards or Lab extraction Kit was of enough quantity and quality to detect G. pallida whenever present (alone or in mixed samples), ensuring probe specificity and sensitivity. This work provides insights for a new strategy to construct advanced devices for pathogens in-field diagnostics. LAMP runs separately but can be easily integrated into a single device.

Assessment of Agricultural Practices for Controlling Fusarium and Mycotoxins Contamination on Maize Grains: Exploratory Study in Maize Farms.

Maize is a significant crop to the global economy and a key component of food and feed, although grains and whole plants can often be contaminated with mycotoxins resulting in a general exposure of the population and animals. To investigate strategies for mycotoxins control at the grain production level, a pilot study and exploratory research were conducted in 2019 and 2020 to compare levels of mycotoxins in grains of plants treated with two fertilizers, F-BAC and Nefusoil, under real agricultural environment. The 1650 grains selected from the 33 samples were assessed for the presence of both Fusarium species and mycotoxins. Only fumonisins and deoxynivalenol were detected. Fumonisin B1 ranged from 0 to 2808.4 µg/Kg, and fumonisin B2 from 0 to 1041.9 µg/Kg, while deoxynivalenol variated from 0 to 465.8 µg/Kg. Nefusoil showed to be promising in regard to fumonisin control. Concerning the control of fungal contamination rate and the diversity of Fusarium species, no significant differences were found between the two treatments in any of the years. However, a tendency for was observed Nefusoil of lower values, probably due to the guaranteed less stressful conditions to the Fusarium spp. present in the soil, which do not stimulate their fumonisins production.

Genome editing for healthy crops: traits, tools and impacts.

Crop cultivars in commercial use have often been selected because they show high levels of resistance to pathogens. However, widespread cultivation of these crops for many years in the environments favorable to a pathogen requires durable forms of resistance to maintain "healthy crops". Breeding of new varieties tolerant/resistant to biotic stresses by incorporating genetic components related to durable resistance, developing new breeding methods and new active molecules, and improving the Integrated Pest Management strategies have been of great value, but their effectiveness is being challenged by the newly emerging diseases and the rapid change of pathogens due to climatic changes. Genome editing has provided new tools and methods to characterize defense-related genes in crops and improve crop resilience to disease pathogens providing improved food security and future sustainable agricultural systems. In this review, we discuss the principal traits, tools and impacts of utilizing genome editing techniques for achieving of durable resilience and a "healthy plants" concept.

Development and Validation of a Loop-Mediated Isothermal Amplification Diagnostic Method to Detect the Quarantine Potato Pale Cyst Nematode, Globodera pallida.

The potato cyst nematode (PCN) Globodera pallida has acquired significant importance throughout Europe due to its nefarious effects on potato production. Rapid and reliable diagnosis of PCN is critical during the surveillance programs and for the implementation of control measures. Molecular DNA-based methods are available, but they require expensive laboratory facilities, equipment and trained technicians. Moreover, there is an additional need of time for sample shipment and testing. In this work, we have developed a new and simple assay which reliably discriminates G. pallida from other cyst nematodes in less than 40 min. This assay may be applied either on cysts or juveniles with the ability to detect a single juvenile of G. pallida in a sample of at least 40 juveniles of the non-target species G. rostochiensis. This test should be a tool to improve the performance of the laboratory and has the potential to be performed on-site.

Integrative Morphometric and Molecular Approach to Update the Impact and Distribution of Potato Cyst Nematodes Globodera rostochiensis and Globodera pallida (Tylenchida: Heteroderidae) in Algeria.

Morphological and molecular studies were conducted to characterize the specific identity of 36 isolates of potato cyst nematodes (PCNs) recovered from soil samples collected in several potato producing areas of Algeria. Morphometric data revealed that 44% of isolates contained Globodera pallida alone, 28% contained Globodera rostochiensis alone and 28% mixtures of the two species. Morphometric values of cysts and second-stage juveniles were generally distributed with slight differences in the expected ranges for both Globodera species. Inter- and intraspecific morphometric variability in nematode isolates was noted. Molecular analysis using conventional multiplex PCR with species-specific primers and TaqMan real-time PCR confirmed the morphological identification. In addition, the distribution of both potato cyst nematode species throughout various parts of the country was investigated. In the central areas, the isolates of G. pallida alone dominate, whereas isolates of G. rostochiensis alone are more frequent in the southern areas. In the eastern regions, mixed isolates are more representative. Most isolates examined in the western areas are mixtures of the two species or G. rostochiensis alone. Comparatively, G. pallida remains the most widely distributed species in its geographic range. This study confirms the presence of two PCN species, G. pallida and G. rostochiensis, in Algeria and provides additional information on their biogeographic distribution.

Occurrence of Fusarium spp. in Maize Grain Harvested in Portugal and Accumulation of Related Mycotoxins during Storage.

Maize is an important worldwide commodity susceptible to fungal contamination in the field, at harvest, and during storage. This work aimed to determine the occurrence of Fusarium spp. in maize grains produced in the Tagus Valley region of Portugal and the levels of related mycotoxins in the 2018 harvest and during their storage for six months in barrels, mimicking silos conditions. Continuous monitoring of temperature, CO2, and relative humidity levels were done, as well as the concentration of mycotoxins were evaluated and correlated with the presence of Fusarium spp. F. verticillioides was identified as the predominant Fusarium species. Zearalenone, deoxynivalenol and toxin T2 were not found at harvest and after storage. Maize grains showed some variability in the levels of fumonisins (Fum B1 and Fum B2). At the harvest, fumonisin B1 ranged from 1297 to 2037 µg/kg, and fumonisin B2 ranged from 411 to 618 µg/kg. Fumonisins showed a tendency to increase (20 to 40%) during six months of storage. Although a correlation between the levels of fumonisins and the monitoring parameters was not established, CO2 levels may be used to predict fungal activity during storage. The composition of the fungal population during storage may predict the incidence of mycotoxins.

Potato Cyst Nematodes: Geographical Distribution, Phylogenetic Relationships and Integrated Pest Management Outcomes in Portugal.

The identification and phylogenetic relationships of potato cyst nematodes (PCN) were studied to assess the potential value of geographical distribution information for integrated pest management of potato production in Portugal. This research focused on PCN species, Globodera pallida and Globodera rostochiensis. From 2013 until 2019, 748 soil samples from the rhizosphere of different potato cultivars were surveyed in the Portuguese mainland to detect and identify both species and track their location. PCN are widespread invasive species throughout Portugal. In fact, during the survey period an incidence of 22.5% was estimated for the tested samples. The patterns of infestation vary among regions, increasing from south to north, where PCN were first detected. Currently, both species are present in all potato producing regions of the country, with a greater incidence of G. pallida. Phytosanitary control measures are influencing to the observed results. The use of potato cultivars resistant to G. rostochiensis led to a decrease of this species but had no influence on G. pallida detections, which continues its reproduction freely since there are no effective resistant cultivars for this species. The relationship between the presence, infestation rate, spread and geographical distribution of PCN is discussed in terms of behavioral responses of the potato cultivars and the implications for developing new integrated crop protection measures.