Multiplex qPCR assay for simultaneous quantification of CYP51-S524T and SdhC-H152R substitutions in European populations of Zymoseptoria tritici

Demethylation inhibitor (DMI) and succinate dehydrogenase inhibitor (SDHI) fungicides are currently relied upon for the control of septoria tritici blotch (STB) in European wheat fields. However, multiple mutations have occurred over time in the genes encoding the targeted proteins that have led to a practical loss of fungicide efficacies. Among the different amino acid substitutions in Zymoseptoria tritici associated with resistance to these fungicides, S524T in CYP51 (DMI target) and H152R in SdhC (SDHI target) are regarded as conferring the highest resistance factors to DMI and SDHI, respectively. To facilitate further studies on the monitoring and selection of these substitutions in Z. tritici populations, a multiplex allele-specific quantitative PCR (qPCR) assay allowing for estimation of both allele frequencies in bulk DNA matrices was developed. The assay was then used on complex DNA samples originating from a spore trap network set up in Belgium, Denmark, Sweden, and Ireland in 2017 and 2018, as well as on leaf samples with symptoms. The S524T allele was present in all field samples and its proportion was significantly higher in Ireland than in Belgium, whereas the proportion of H152R was only sporadically present in both countries. The frequency of S524T varied greatly in the airborne inoculum of all four countries; however, the H152R allele was never detected in the airborne inoculum. The method developed in this study can be readily adopted by other laboratories and used for multiple applications including resistance monitoring in field populations of Z. tritici.     

The physiology of triploid fish: current knowledge and comparisons with diploid fish

This paper is aimed at underlining the limited knowledge available about the physiology of triploid fishes compared with diploids. Whereas many aspects (induction, detection, growth, resistance to diseases, etc.) in the production and rearing of triploid fishes have widely been developed and described in the literature, other numerous questions of ecophysiology remain in abeyance, and the study of triploid cells physiology is still in its infancy. Triploid fishes can be considered as models worthwhile for physiological investigations not only for the economical stake in relation to the development of triploid fishes rearing, but also for the cytological and molecular features of their tissues and organs. The functional implications of these features have been poorly studied although they are potential areas of applied and/or fundamental studies.     

Effects of hypoxia on respiratory physiology of turbot, Scophthalmus maximus

Variations in respiratory and acid-base status were studied in turbot (Scophthalmus maximus) during progressive severe hypoxia followed by recovery under normoxic conditions. The first behavioural strategy of turbot under hypoxia was an increase in amplitude and frequency of ventilation. Consequently, standard O₂ consumption remained unchanged over a broad range of O₂ tensions, until a low critical level of 30 mmHg. The hyperventilation induced a moderate blood alkalosis, compensated by a lactic acidosis. The fact that blood pH did not decrease below control values could be explained by the retention in white muscle of most of the lactate produced and by a high capacity for H⁺ excretion. During the recovery period, the marked increase in O₂ uptake corresponding to an oxygen debt repayment, was partly related to the lactate elimination. When total energy contributions of aerobic and anaerobic processes were assessed in terms of ATP, the anaerobic contribution, estimated at the deepest hypoxia level, was higher than 20% of the total energy budget and appeared to totally compensate for the decline in aerobic metabolism. Moreover, the high value of O₂ tension in arterial blood in normoxia and during recovery from hypoxia showed high diffusing capacity of gills in turbot. Our results explain the high tolerance of turbot for O₂ deficient waters.     

Detection of Fusarium langsethiae on wheat in Belgium

In 2010, the populations of Fusarium sp. and Microdochium sp. were monitored in Belgium and 16 strains were identified as Fusarium langsethiae on wheat in Belgium. The other species identified from the sampling were F. poae, F. tritinctum, F. graminearum, F. avenaceum and Microdochium nivale. The pathogenicity potential of the F. langsethiae strains was assessed via an in vitro coleoptile growth rate test on wheat seedlings and compared with strains of F. poae, F. tritinctum, F. graminearum and F. avenaceum known to cause Fusarium head blight. The results showed the ability of F. langsethiae to cause retardation in the wheat coleoptile growth rate, but at a lower rate than F. graminearum, F. avenaceum, F. poae and F. tricinctum. A test for mycotoxin production in vitro showed the ability of the four strains tested to produce T-2 and HT-2 toxins at a rate of up to 290 mg kg^?1. This is the first report on the potential pathogenicity of F. langsethiae on wheat in Belgium, a species known to produce T-2 and HT-2 toxins, which are highly toxic for humans and animals.     

Relationships of climate and cell features in stems and roots of black spruce and balsam fir

? The anatomical differences of mature black spruces and balsam firs were examined at stem and root level in order to characterize their wood properties at cellular level and link these differences to climate.

? Anatomical variability of these species was evaluated in relation to climate data gathered from 2001 to 2004 during the cell enlargement (CE) and wall thickening and lignification (WTL) phases. Lumen area, single cell wall thickness and total tracheid radial diameter were analyzed and regrouped into earlywood and latewood.

? Results from a principal component analysis (PCA) indicated that both first eigenvectors account for 82% and 90% of total variance for CE and WTL respectively. These component factors revealed that precipitation, humidity and number of days with precipitation significantly influence the lumen area (p = 0.0168) and radial cell diameter (p = 0.0222) in earlywood. Significant differences were registered between species and tree parts (stem and root) for the lumen area, radial cell diameter and cell wall thickness in both earlywood and latewood.

? In our study, black spruce exhibited smaller tracheid size in both stem and roots compared to balsam fir. Furthermore, the lower amount of tracheids produced during the growing season and higher proportion of latewood ensure a higher wood density of black spruce. The influence of temperature on earlywood formation is significant, whereas no influence was observed on latewood.

     

The prediction of bread wheat quality: joint use of the phenotypic information brought by technological tests and the genetic information brought by HMW and LMW glutenin subunits

Attempts have been made to predict Chopin alveograph or French bread-making tests, using tree-based models and PLS regressions. Data came from three sets of trials, involving 130, 214 and 103 different genotypes, which were described for HMW-GS, LMW-GS and small-scale tests currently used in breeding programs. Segmentation trees and PLS regressions indicated that HMW-GS and LMW-GS were not sufficient to explain alone the variability of bread wheat quality. This could be partly due to “allele × environment” and “locus × locus” interactions. For HMW-GS, Glu-B1 was the predominant locus for alveograph and French bread-baking, and some differences in the alleles hierarchy were demonstrated according to the end-use parameter considered. For LMW-GS, Glu-B3 seemed to be preponderant, with alleles b′, c and g being favourable and allele c′ unfavourable. Joint use of the information brought by glutenin subunits and technological tests did not enable to predict satisfactorily, neither the different parameters of French bread-baking, nor the extensibility L of alveograph. Only the prediction of the strength W proved reliable, and robust PLS equations were proposed for this alveograph parameter. These prediction equations could be of interest to select for high values of W in the mid generations of breeding.     

Analysis of Accumulation Patterns of <Emphasis Type="Italic">Barley yellow dwarf virus-PAV</Emphasis> (BYDV-PAV) in Two Resistant Wheat Lines

Barley yellow dwarf (BYD) is one of the main viral diseases of small-grain cereals. This disease, reported on numerous plant species of the Poaceae family, is caused by a complex of eight viral species including the species Barley Yellow Dwarf Virus-PAV (BYDV-PAV), frequently found in western Europe. Resistance sources against BYDV-PAV are scarce and only identified in perennial Triticineae. Some BYDV-resistant wheat lines have been obtained by introgressing these resistances into bread wheat germplasms. Genetic and biological characterization of the resulting lines has been undertaken. However, little information on the resistant behaviour of these lines during the early stages of the infection process is available. To evaluate the resistance of two genetically distinct resistant lines (Zhong ZH and TC14), 1740 young plantlets, belonging to susceptible reference hosts (barley cv. Express and wheat cv. Sunstar), Zhong ZH or TC14 wheat lines, were inoculated in controlled conditions with French BYDV-PAV isolates. The infection process was monitored during the first 21 days after inoculation (DAI) using a semi-quantitative ELISA. A standardized protocol including five successive samplings of leaves from all inoculated plants and the collection of plant roots at the end of the monitored period was carried out. This protocol enabled an assessment of the infection percentage and the evolution of the viral load in plants from the 7th DAI to the 21st DAI. Statistical analyses of the BYDV infection kinetics using raw ELISA data, a model of the time-dependent variation of the percentage of infected plants and the area under concentration progress curves (AUCPC) demonstrated that Zhong ZH and TC14 lines (1) reduce the development rate of the BYD disease during the first days of infection, (2) decrease the infection efficiency of BYDV-PAV isolates, in the leaves, from 98.7% for susceptible plant genotypes to 81.9% and 71.7% for Zhong ZH and TC14, respectively, (3) reduce the virus load in the leaves of infected plants and (4) are not spared from BYDV infection, as 95.1% of Zhong ZH and 90.2% of TC14 inoculated plants accumulated viral particles in roots and/or in leaves at 21 DAI. These results confirm the BYDV-partial resistant behaviour of both Zhong ZH and TC14 lines. The development rate of the disease was the single parameter that allowed the distinction between the two resistant sources present in the tested lines.     

Pacific-Ciguatoxin-2 and Brevetoxin-1 Induce the Sensitization of Sensory Receptors Mediating Pain and Pruritus in Sensory Neurons

Ciguatera fish poisoning (CFP) and neurotoxic shellfish poisoning syndromes are induced by the consumption of seafood contaminated by ciguatoxins and brevetoxins. Both toxins cause sensory symptoms such as paresthesia, cold dysesthesia and painful disorders. An intense pruritus, which may become chronic, occurs also in CFP. No curative treatment is available and the pathophysiology is not fully elucidated. Here we conducted single-cell calcium video-imaging experiments in sensory neurons from newborn rats to study in vitro the ability of Pacific-ciguatoxin-2 (P-CTX-2) and brevetoxin-1 (PbTx-1) to sensitize receptors and ion channels, (i.e., to increase the percentage of responding cells and/or the response amplitude to their pharmacological agonists). In addition, we studied the neurotrophin release in sensory neurons co-cultured with keratinocytes after exposure to P-CTX-2. Our results show that P-CTX-2 induced the sensitization of TRPA1, TRPV4, PAR2, MrgprC, MrgprA and TTX-r NaV channels in sensory neurons. P-CTX-2 increased the release of nerve growth factor and brain-derived neurotrophic factor in the co-culture supernatant, suggesting that those neurotrophins could contribute to the sensitization of the aforementioned receptors and channels. Our results suggest the potential role of sensitization of sensory receptors/ion channels in the induction or persistence of sensory disturbances in CFP syndrome.     

Effect of the management of seed flows and mode of propagation on the genetic diversity in an Andean farming system: the case of oca (<Emphasis Type="Italic">Oxalis tuberosa</Emphasis> Mol.)

The seed system is a major component of traditional management of crop genetic diversity in developing countries. Seed flows are an important part of this system. They have been poorly studied for minor Andean crops, especially those that are propagated vegetatively. We examine the seed exchanges of Oxalis tuberosa Mol. (oca), a vegetatively propagated crop capable of sexual reproduction. We studied the seed exchanges of four rural communities in Candelaria district (Cochabamba department, Bolivia) at the international and local levels, emphasizing the spread of new sexually-produced genotypes through these exchanges. Interviews with 44 farmers generated socioeconomic, agronomic, crop diversity and seed exchange information, and data on the potential incorporation of new sexually-produced genotypes in the crop germplasm. We interviewed merchants to evaluate the input and output of genetic diversity in the communities studied. Results showed a positive effect of the farmers’ wealth on the diversity cultivated and on seed exchanges. Most seed exchanges occurred at market, creating a distinction between cash and self-consumption landraces. Cash landraces were intensively exchanged; self-consumption landraces were isolated at the farmer level and prone to genetic drift and complete loss. Merchants exported seeds of cash landraces across Bolivia and into Peru and Argentina. New sexually produced genotypes are less incorporated into cash landraces than in self-consumed landraces. However, new genotypes incorporated into cash landraces are diffused faster and better, being more intensively exchanged. We propose conservation strategies that can be applied to other vegetatively propagated and minor Andean crops.     

Sensitivity and permissivity of Cyprinus carpio to cyprinid herpesvirus 3 during the early stages of its development: importance of the epidermal mucus as an innate immune barrier

Cyprinid herpesvirus 3 (CyHV-3) causes a lethal disease in common and koi carp (Cyprinus carpio). The present study investigated the ability of CyHV-3 to infect common carp during the early stages of its development (from embryos to fingerlings) after inoculation by immersion in water containing the virus. Fish were inoculated at different times after hatching with a pathogenic recombinant CyHV-3 strain expressing luciferase. The sensitivity and permissivity of carp to CyHV-3 were investigated using in vivo bioluminescence imaging. The susceptibility of carp to CyHV-3 disease was investigated by measuring the survival rate. Carp were sensitive and permissive to CyHV-3 infection and susceptible to CyHV-3 disease at all stages of development, but the sensitivity of the two early developmental stages (embryo and larval stages) was limited compared to later stages. The lower sensitivity observed for the early developmental stages was due to stronger inhibition of viral entry into the host by epidermal mucus. In addition, independent of the developmental stage at which inoculation was performed, the localization of light emission suggested that the skin is the portal of CyHV-3 entry. Taken together, the results of the present study demonstrate that carp are sensitive and permissive to CyHV-3 at all stages of development and confirm that the skin is the major portal of entry after inoculation by immersion in infectious water. The results also stress the role of epidermal mucus as an innate immune barrier against pathogens even and especially at the early stages of development.