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Background Volatile fatty acid intoxication (acidosis), a common process failure recorded in anaerobic reactors, leads to drastic losses in methane production. Unfortunately, little is known about the microbial mechanisms underlining acidosis and the potential to recover the process. In this study, triplicate mesophilic anaerobic reactors of 100 L were exposed to acidosis resulting from an excessive feeding with sugar beet pulp and were compared to a steady-state reactor. Results Stable operational conditions at the beginning of the experiment initially led to similar microbial populations in the four reactors, as revealed by 16S rRNA gene T-RFLP and high-throughput amplicon sequencing. Bacteroidetes and Firmicutes were the two dominant phyla, and although they were represented by a high number of operational taxonomic units, only a few were dominant. Once the environment became deterministic (selective pressure from an increased substrate feeding), microbial populations started to diverge between the overfed reactors. Interestingly, most of bacteria and archaea showed redundant functional adaptation to the changing environmental conditions. However, the dominant Bacteroidales were resistant to high volatile fatty acids content and low pH. The severe acidosis did not eradicate archaea and a clear shift in archaeal populations from acetotrophic to hydrogenotrophic methanogenesis occurred in the overfed reactors. After 11 days of severe acidosis (pH 5.2 ± 0.4), the process was quickly recovered (restoration of the biogas production with methane content above 50 %) in the overfed reactors, by adjusting the pH to around 7 using NaOH and NaHCO3. Conclusions In this study we show that once the replicate reactors are confronted with sub-optimal conditions, their microbial populations start to evolve differentially. Furthermore the alterations of commonly used microbial parameters to monitor the process, such as richness, evenness and diversity indices were unsuccessful to predict the process failure. At the same time, we tentatively propose the replacement of the dominant Methanosaeta sp. in this case by Methanoculleus sp., to be a potential warning indicator of acidosis. Electronic supplementary material The online version of this article (doi:10.1186/s13068-015-0309-9) contains supplementary material, which is available to authorized users.

The progress of Indian peanut clump virus (Hyderabad isolate; IPCV-H) and its vector Polymyxa graminis in various monocotyledonous crops and groundnut was studied during the 1994, 1995 and 1996 rainy seasons in a naturally infested field in India. The roles of rainfall and temperature in the dynamics of infection by both the virus and its vector were analysed by exposing young seedlings for short periods in the field. Of the host crops studied, wheat, followed by barley, showed the highest virus incidence, although P. graminis was rarely observed in roots of wheat and was not detected in those of barley. The roots of maize, pearl millet and sorghum plants infected by P. graminis showed intense colonization by sporosori. IPCV accumulated in systemically infected maize plants; the sorghum and pearl millet cultivars studied showed a transient presence of IPCV-H. Rice was seldom infected by the virus and P. graminis was not detected in its roots. Groundnut was a systemic host for the virus, although during these experiments no P. graminis was found in its roots. Groundnut appeared to be susceptible to infection, mostly in the early stages of crop development, and the rate of IPCV-H transmission in groundnut seeds was highest (13%) for plants infected when young. The seed-transmission rate quickly decreased in plants showing symptoms 1 month after sowing. Time of infection had little influence on groundnut pod yield, which was always reduced by >60% in infected plants. There was some evidence that the quantity and distribution of rainfall influenced the incidences of IPCV-H and P. graminis: high rainfall resulted in high incidences of the virus and P. graminis, and a weekly rainfall of 14 mm was sufficient for P. graminis to initiate infection. Temperatures prevailing during the rainy season ranged from 23 to 30degreesC and were found to be conducive to natural virus transmission. These results suggest measures to be explored for controlling peanut clump disease.

A region of the nuclear ribosomal DNA containing the internal transcribed spacer 1 (ITS1), the 5.8S DNA and the internal transcribed spacer 2 (ITS2) was sequenced in 12 Polymyxa graminis and P. betae isolates, with particular emphasis on P. graminis from peanut clump-infested areas of the Indian subcontinent and West-Africa. Four different sequences were obtained from the seven isolates on sorghum or pearl millet, which differed from four sequences previously published for Polymyxa species and obtained for P. graminis isolates on barley, oat and wheat originating from temperate areas in Europe and America (two distinct sequences), for isolates on rice from Colombia and for P. betae isolates on sugar beet from several origins. The sequence variations concerned mainly the composition and length of ITS1 and ITS2 regions. Phylogenetic trees built with the distinct sequences currently known for Polymyxa spp. using parsimony, maximum likeihood and neighbour-joining methods separated P. betae from P. graminis, Within P. graminis, the hierarchy of the clustering partially matched the host range and geographical origin of the isolates. These results confirm the great diversity within P. graminis that has already been observed in host range and temperature requirements studies, and provide new arguments for considering several taxa within the species. On the basis of the ecological requirements and rDNA sequences of distinct P. graminis isolates, five special forms are proposed: P. graminis f. sp. temperata, P. graminis f. sp. tepida, P. graminis f. sp. subtropicalis, P. graminis f. sp. tropicalis and P. graminis f. sp. colombiana.

Fluorescent Pseudomonas species are characterized by the production of pyoverdin-type siderophores for Fe3+ acquisition in iron-limited environments. Since it produces a structurally specific pyoverdin, Pseudomonas putida strain BTP1 could represent a valuable tool in an attempt to correlate the structural features of these compounds with some specificity in their two main properties i.e. affinity for iron and recognition rate by other Pseudomonas strains. An uncommonly high affinity for iron of the pyoverdin synthetized by P. putida BTP1 was observed by comparing both the apparent stability constant and the decomplexation kinetic of its ferric complex with those of ferripyoverdins from other strains. On another hand, results from growth stimulation experiments and labeled ferripyoverdin uptake assays highlighted the very low recognition rate of BTP1 isopyoverdins by membrane receptors of foreign strains. By contrast, P. putida BTP1 was able to utilize a broad spectrum of structurally unrelated exogenous pyoverdins by means of multiple receptors that are likely constitutively expressed in its outer membrane. The unusual traits of its pyoverdin-mediated iron acquisition system should contribute to enlarge the ecological competence of Pseudomonas putida BTP1 in terms of colonization and persistence in the rhizosphere.

This study investigated the use of an electronic nose for on-line anaerobic reactor state monitoring at the pilot-scale level and then upscaling to the full-scale level. E-nose indicator was compared to classical state indicators such as pH, alkalinity, volatile fatty acids concentration and to other gas phase compounds. Multivariate statistical process control method, based on principal component analysis and the Hotelling’s T 2 statistics was used to derive an indicator representative of the reactor state. At the pilot-scale level, the e-nose indicator was relevant and could distinguish 3 process states: steady-state, transient and collapsing process. At the full-scale level, the e-nose indicator could provide the warning of the major disturbance whereas two slight disturbances were not detected and it gave one major false alarm. This work showed that gas phase relation with anaerobic process should be deeper investigated, as an e-nose could indicate the reactor state, focusing on the gas phase.

The host range of Polymyxa graminis isolates originating from peanut clump-infested areas in India (Andhra Pradesh and Rajasthan), Pakistan and Senegal was studied on monocotyledonous and dicotyledonous cultivated species, using known quantities of sporosori as inoculum. Profuse multiplication occurred only on some graminaceous species, but the various isolates showed different host specificity. All the isolates produced high infection on sorghum and pearl millet, and all but one isolate from Rajasthan infected maize. Wheat, rye and barley were susceptible to some of the tested isolates. The isolates from Rajasthan and Pakistan produced moderate to severe infection on at least one of these species. On rice, groundnut and sugar beet, only traces of infection by some isolates were detected, whereas no infection was observed on mustard and sunflower. Differences of susceptibility in Pennisetum spp. and Sorghum spp. were demonstrated. The variations in host specificity among isolates from peanut clump-infested areas may result from an adaptation of P. graminis populations to various biotopes. The implications of these results for the management of peanut clump disease are discussed. A comparison of the host ranges of isolates of P. graminis and P. betae from temperate areas demonstrated that distinct types of Polymyxa might be identified based on their relative ability to multiply on susceptible species. Nevertheless, overlapping in the host ranges among the different Polymyxa types, characterised by distinct ecological and genomic features, raises doubts about the host range as a classification criterion for the Polymyxa genus.

Hybridomas that secreted antibodies for aflatoxin B1 were selected using two immunization protocols referred to as A and B. Protocol A is a standard immunization method and resulted in the selection of only two clones that produced monoclonal antibodies against aflatoxin B1. In protocol B a unique immunization schedule which resulted in the generation of 10 hybridomas is described. Of the 10, one antibody was highly specific to B1, four antibodies reacted equally strongly with B1, G1 and weakly with B2. Another four reacted strongly with B1 and weakly with B2 and G1. One clone reacted equally strongly with B1, G1 and B2. Interestingly all the 10 antibodies showed little or no cross-reaction with G2.

The infection potential of sporosori of Polymyxa graminis involved in the transmission of the Indian peanut clump virus (IPCV) was assessed by culturing bait plants exposed to various concentrations of sporosorus suspensions and then determination of the numbers of plants that became infected. Storage of air-dried inoculum at temperatures above 30 degrees C resulted in an increase in the infection potential compared to that of sporosori stored at 15 degrees C or 20 degrees C. In contrast, when the sporosori were stored at 20 degrees C or freeze-dried, their infection potential was low. These results confirm the adaptation of P. graminis isolates associated with IPCV transmission to the tropical environment. The implication of storage temperature for the epidemiology of Indian peanut clump virus and for the assessment of the infection potential of the vector in the soil is discussed.

Wheat and barley crops were shown to be susceptible to Indian peanut clump virus (IPCV) under field conditions. In wheat, the Hyderabad isolate of IPCV (IPCV-H) induced symptoms resembling the rosette caused by soil-borne wheat mosaic virus, and these were apparent only three weeks after emergence. Early-infected plants were severely stunted and dark green, with chlorotic streaks on the youngest leaves, which turned necrotic as the plants aged; most of these plants died. Late-infected plants were also stunted and were conspicuous in the field because of their dark green appearance as a result of delayed maturity. The virus was detected by ELISA and nucleic acid hybridization in all plants with symptoms. These plants usually produced fewer tillers than healthy ones. Spikes were malformed, often did not emerge from the flag leaf, and they contained few shrivelled seeds. Grain yield was decreased, on average, by 58%. In barley, IPCV-H caused severe stunting and general leaf chlorosis. As the plants aged, the leaves became necrotic and the few infected plants that reached maturity produced small spikes. IPCV-H antigens were detected by ELISA in every wheat seed from infected plants and the virus was transmitted through wheat seed at a frequency of 0.5-1.3%. Storage at 4 degrees C for more than a year did not affect seed transmission frequency. The virus was detected in leaves and roots of seed-transmitted plants. Seed transmission was not detected in barley. The Durgapura isolate (IPCV-D) was detected in wheat crops (cv. RR-21) at 3 different locations in Rajasthan State, India. Infected plants showed reduced growth without any overt symptoms.

The temperature requirements of three single cystosorus strains of Polymyxa sp. from India were studied at 15–18, 19–22, 23–26 and 27–30 °C (night-day temperature), and compared with the temperature requirements of three strains of P. graminis from Belgium, Canada and France and two strains of P. betae from Belgium and Turkey. Sorghum was used as the host-plant for the Indian strains; the strains of P. graminis and P. betae from temperate areas were cultivated on barley and sugar beet, respectively. The cystosori germination and the development of plasmodia, zoosporangia and cystosori of Polymyxa sp. from India were optimal at 27–30 °C. Infection progression was slower at 23–26 °C than at 27–30 °C. At 19–22 °C, infection was insignificant. No infection occurred below 19 °C. In contrast, the infection of barley with P. graminis strains from temperate areas was optimal at 15–18 °C, but at 19–22 °C the progression appeared inconsistent and infection stayed low. Above 22 °C, infection was insignificant. P. betae strains showed consistent infection in the range of 15–18 °C to 27–30 °C. Plasmodia formation and cystosori detection of the Belgian strain were slightly advanced at 23–26 °C compared to 19–22 °C but clearly restrained at 27–30 °C. Fungus development of the P. betae strain from Turkey was almost as high at 27–30 °C as at the lower temperatures. These results strengthen the case for distinguishing between Polymyxa sp. from India and P. graminis or P. betae from temperate areas.

Biogas production from energy crops by anaerobic digestion is becoming increasingly important. The amount of biogas that can be produced per unit of biomass is referred to as the biomethane potential (BMP). For energy crops, the BMP varies among varieties and with crop state during the vegetation period. Traditional ways of analytical BMP determination are based on fermentation trials and require a minimum of 30 days. Here, we present a faster method for BMP retrievals using near infrared spectroscopy and partial least square regression (PLSR). PLSR prediction models were developed based on two different sets of spectral reflectance data: (i) laboratory spectra of silage samples and (ii) airborne imaging spectra (HyMap) of maize canopies under field (in situ) conditions. Biomass was sampled from 35 plots covering different maize varieties and the BMP was determined as BMP per mass (BMPFM, Nm3 biogas/t fresh matter (Nm3/t FM)) and BMP per area (BMParea, Nm3 biogas/ha (Nm3/ha)). We found that BMPFM significantly differs among maize varieties; it could be well retrieved from silage samples in the laboratory approach (Rcv2 = 0.82, n = 35), especially at levels >190 Nm3/t. In the in situ approach PLSR prediction quality declined (Rcv2 = 0.50, n = 20). BMParea, on the other hand, was found to be strongly correlated with total biomass, but could not be satisfactorily predicted using airborne HyMap imaging data and PLSR.

Pyoverdin type siderophores produced by six fluorescent Pseudomonas strains isolated from different rhizospheres were purified and characterized. The purified ferri-pyoverdins were tested for their ability to promote the growth of other strains grown under iron deficiency conditions. Only the one obtained from Pseudomonas putida BTP1 did not act as a growth promoter. The structure of the BTP1 siderophore was elucidated by spectroscopic methods and degradation studies. It turned out that it contains a chromophore which differs from the one typical for pyoverdins insofar as it carries the carboxyl group in 3- rather than in 1-position ((3S)-5-amino-1,2-dihydro-8,9-dihydroxy-3 H -pyrimido[1,2 a]quinoline-3-carboxylic acid). The amino group of the chromophore is substituted with the 5-carboxyl group of ʟ-glutamic acid and its carboxyl group with the N -terminus of the peptide ʟ -A sp -ʟ -A la-ʟ -Asp - ᴅ-N5- Ac - N5- OH - Orn - ʟ - Ser - ʟ - ᴄ - N5- OH - Orn . This isopyoverdin fits into the biogenetic scheme which postulates ferribactins as the precursors of pyoverdins

This study aimed at analysing the utilization of an electronic nose (e-nose) to serve as a specific monitoring tool for anaerobic digestion process, especially for detecting organic overload. An array of non specific metal oxide semiconductor gas sensors were used to detect process faults due to organic overload events in twelve 1.8-L anaerobic semi-continuous reactors. Three different load strategies were followed: (1) a cautious organic load (1.3 gVS L−1 day−1); (2) an increasing load strategy (1.3–5.3 gVS L−1 day−1) and (3) a cautious organic load with load pulses of up to 12 gVS L−1 day−1. A first monitoring campaign was conducted with three different substrates: sucrose, maize oil and a mix of sucrose/oil during 60 days. The second campaign was run with dry sugar beet pulp for 45 days. Hotelling’s T 2 value and upper control limit to a reference set of digesters fed with a cautious OLR (1.3 gVS L−1 day−1) was used as indirect state variable of the reactors. Overload situations were identified by the e-nose apparatus with Hotelling’s T 2 values at least four times higher in magnitude than the upper control limit of 23.7. These results confirmed that the e-nose technology appeared promising for online detection of process imbalances in the domain of anaerobic digestion.

The 3'-end region of many virus isolates has been shown to possess conserved sequences in addition to the presence of numerous genomic and subgenomic RNAs. Utilizing these sequences, a broad-spectrum reverse transcription-polymerase chain reaction protocol has been developed to detect all the known Indian peanut clump virus and Peanut clump virus isolates, that cause peanut clump diseases in West Africa and India. The primers were targeted at the highly conserved 3'-untranslated regions of the PCV RNA-1 and RNA-2. The conservation was confirmed by sequencing these untranslated regions of RNA-1 for six isolates and RNA-2 for one isolate. The conserved structure of the RNA-1 and RNA-2 was observed and the importance of this region for the virus survival was confirmed. The primers were also designed for virus quantitation using a Taqman((R))-based real-time RT-PCR. The use of RT-PCR and real-time quantitative RT-PCR improved the sensitivity of PCV detection compared to ELISA. RT-PCR also led to the detection of IPCV and PCV on two new natural hosts: Oldenlandia aspera and Vigna subterranea. Real-time RT-PCR is considered to be an ideal tool for identifying resistant sources to both IPCV and PCV.

Fusarium head blight (FHB) is among the major causes of reduced quality in winter wheat and its products. In addition, the causal fungi produce a variety of toxins. A relatively high FHB infection rate in winter wheat was observed in 2007 and 2008 in Luxembourg. A fusariotoxin survey was carried out in 17 different geographical locations. Three groups of Fusarium mycotoxins (trichothecenes A and B and zearalenone) were analysed by a multi-detection HPLC-MS/MS method. Fusarium strains were also investigated by morphological and molecular methods. In addition, questionnaires relating to cultural practices were sent to the farmers managing the 17 fields investigated. FHB prevalence ranged from 0.3 to 65.8% (mean: 8.5%) in 2007 and from 0 to 24.5% (mean: 8.3%) in 2008. Results of morphological and molecular identification showed that the most common species isolated from diseased wheat spikes was F. graminearum (33.1%), followed by F. avenaceum (20.3%) and F. poae (17.8%). The chemical analysis revealed that 75% of the investigated fields were contaminated by deoxynivalenol (DON, range 0-8111 microg/kg). The preceding crop was highly and significantly correlated to the number of grains infected and had a significant impact on disease prevalence (p = 0.025 and 0.017, respectively, Fisher's F-test). A trend was found for maize as the preceding crop (p = 0.084, Tukey's test) to predict the amount of DON in the fields. This is the first report on the occurrence of DON and ZON in naturally infected wheat grains sampled from Luxembourg.

A purification procedure was developed to separate Polymyxa graminisresting spores from sorghum root materials. The spores were used as im-munogen to produce a polyclonal antiserum. In a direct antigen coating enzyme-linked immunosorbent assay (DAC ELISA), the antiserum could detect one sporosorus per well of the ELISA plate. In spiked root samples, the procedure detected one sporosorus per mg of dried sorghum roots. The majority of isolates of P. graminis from Europe, North America, and India reacted strongly with the antiserum. Interestingly, P. graminis isolates from the state of Rajasthan (northern India), from Pakistan, and an isolate from Senegal (West Africa) reacted weakly with the antiserum. The cross-reactivity of the serum with P. betae isolates from Belgium and Turkey was about 40% of that observed for the homologous isolate. There was no reaction with common fungi infecting roots or with the obligate parasite Olpidium brassicae. However, two isolates of Spongospora sub-terranea gave an absorbance similar to that observed with the homologous antigen. The DAC ELISA procedure was successfully used to detect various stages in the life cycle of P. graminis and to detect infection that occurred under natural and controlled environments. A simple procedure to conjugate antibodies to fluorescein 5-isothiocyanate (FITC) is described. Resting spores could be detected in root sections by using FITC-labeled antibodies. The potential for application of the two serological techniques for studying the epidemiology of peanut clump disease and for the characterization of Polymyxa isolates from various geographical origins is discussed.

Peanut clump and sugarcane red leaf mottle diseases are caused by viruses of the genus Pecluvirus. Indian peanut clump virus occurs in the Indian sub-continent and Peanut clump virus in West Africa. A feature of these viruses is that they are both seed and soil transmitted. Both modes of transmission contribute to long-term persistence and field spread. Data on seed transmission in pearl millet, virus movement within the plant and virus diversity based on RNA-1 partial sequences are presented. This study emphasizes that pecluviruses are also viruses of cereals infecting sorghum and pearl millet, and highlights a correlation between the countries cultivating these two crops and the virus distribution. Ways of controlling pecluviruses and their vector, Polymyxa graminis, taking into account the virus dissemination routes, are proposed.

Zusammenfassung Die Abbauraten von Grassilage wurden bei 6 Rindern mit Hilfe von nylon bags in situ gemessen. Die Tiere erhielten eine Standardration zu 50% Heu und 50% kommerzielles Kraftfutter oder die zu untersuchende Grassilage, die teilweise durch Zuckerrubenschnitzel ersetzt wurde. Der Ersatz von Grassilage durch Zuckerrubenschnitzel hatte keinen Einflus auf die Abbaurate der organischen Substanz bzw. des Rohproteins aus Grassilage. Das Gleiche gilt fur die Futterung der Standardration. Dagegen ist die Abflusrate der unverdauten Futterpartikel im Pansen fur die Berechnung der tatsachlichen Abbaurate von erheblicher Bedeutung und beeinflust damit auch die Rationsgestaltung fur Milchvieh. Die Unterschiede hinsichtlich der Abbauraten zwischen den Tieren waren nicht reproduzierbar und mussen als zufallig bezeichnet werden.

Fluorescent Pseudomonas species are characterized by the production of pyoverdin-type siderophores for Fe3+ acquisition in iron-limited environments. Since it produces a structurally specific pyoverdin, Pseudomonas putida strain BTP1 could represent a valuable tool in an attempt to correlate the structural features of these compounds with some specificity in their two main properties i.e. affinity for iron and recognition rate by other Pseudomonas strains. An uncommonly high affinity for iron of the pyoverdin synthetized by P. putida BTP1 was observed by comparing both the apparent stability constant and the decomplexation kinetic of its ferric complex with those of ferripyoverdins from other strains. On another hand, results from growth stimulation experiments and labeled ferripyoverdin uptake assays highlighted the very low recognition rate of BTP1 isopyoverdins by membrane receptors of foreign strains. By contrast, P. putida BTP1 was able to utilize a broad spectrum of structurally unrelated exogenous pyoverdins by means of multiple receptors that are likely constitutively expressed in its outer membrane. The unusual traits of its pyoverdin-mediated iron acquisition system should contribute to enlarge the ecological competence of Pseudomonas putida BTP1 in terms of colonization and persistence in the rhizosphere.

Polyclonal antibodies were produced for Ochratoxin A (OA) by injecting OA-bovine serum albumin (BSA) conjugate subcutaneously at multiple sites into a New Zealand White inbred rabbit. Antiserum could be used at a dilution exceeding 1:100 000 in an indirect competitive enzyme-linked immunosorbent assay (ELISA), and detected OA concentrations up to 0.1 ng/mL. The 50% inhibition binding (I(50)) of OA was 5 ng/mL. Antibodies did not react with ochratoxin B, coumarin, 4-hydroxycoumarin, L-phenylalanine, and aflatoxin B1. OA contamination in chilies (Capsicum annum L.) collected from commercial markets and cold storage units was determined. The mean recoveries from OA-free chilies spiked with 1 to100 microg of OA per kg of chili sample were 90-110% with a standard deviation of

Tn5 mutagenesis of different fluorescent pseudomonads was achieved by conjugational transfer of the suicide vector pSUP 10141. Pyoverdine negative (Pvd-) mutants were detected by the absence of fluorescence on King's B medium and by their inability to grow in the presence of the iron chelator EDDHA [ethylenediamine di(o-hydroxyphenylacetic acid)]. In P. fluorescens ATCC 17400 and three rhizosphere isolates (one P. putida and two P. fluorescens), the percentage of Pvd- mutants ranged between 0 and 0.54%. In a P. chlororaphis rhizosphere isolate, this percentage was higher (4%). In these mutants both of the Tn5 antibiotic resistances (Km and Tc) were stable and the transposon could be detected by hybridization. In Pvd- mutants of P. fluorescens ATCC 17400, the transposon was found to be inserted twice in the chromosome while single insertions were detected in the DNA of other, randomly tested mutants. In P. aeruginosa PAO1, where 13.1% of the mutants were Pvd-, both antibiotic resistances were rapidly lost and accordingly no transposon insertion could be detected by hybridization. However, the Pvd- phenotype was generally stable in these mutants. The plasmid pNK862 containing a mini-Tn10 transposon was introduced by electroporation into P. aeruginosa PAO1 and Kmr mutants were recovered, 89% of which were Pvd- and confirmed to be P. aeruginosa by PCR amplification of the P. aeruginosa lipoprotein gene. The mini-Tn10 insertions were also found to be unstable in PAO1.

Rapeseed (Brassica napus L.) is the third most important crop after wheat and barley in the Grand Duchy of Luxembourg. Since 2005, clubroot symptoms in this crop have been reported by farmers in the Gutland Region. In February 2009, plants of the hybrid rapeseed cv. Exocet, with stunted growth, yellow leaves, and club-shaped roots, were sampled from a field in Oberkorn village near Differdange. Microscopic observations of the rapeseed root fragments revealed the presence of the three life stages characteristic of Plasmodiophora brassicae Woronin. Plasmodia and zoosporangia were observed in the root hairs and resting spores were present in root galls. Individual spores were 2 to 3 μm in diameter. Total DNA was extracted from the root galls with a FAST DNA Kit (MP Biomedicals, Irvine, CA). The internal transcribed spacer region (ITS) and 5.8S gene of the rDNA region were amplified with ITS5 and ITS4 primers as described by White et al. (2) and part of this region was sequenced. A BLASTn search in GenBank revealed that the sequence closely resembled (98% identity) sequences of P. brassicae (Genbank Accession No. EF195335) from an isolate of the pathogen from Switzerland. To confirm the presence of the pathogen, seeds of the susceptible ecotype cvi-0 of Arabidopsis thaliana were grown in a soil sample (1 liter) collected near the infected rapeseed plants. After 55 days of growth in a glasshouse at 15 to 20°C, the roots of 11 plants were analyzed. Two showed clear clubroot symptoms and four others exhibited small swellings. The remaining five plants were symptomless, but plasmodia and zoosporangia were found in root hair cells. Clubroot caused by P. brassicae has previously been described on B. napus and other crucifers (1). To our knowledge, this is the first report of clubroot disease caused by P. brassicae in Luxembourg. Because its presence has since been observed in new fields in the Gutland Region and because of the ability of the pathogen to survive for a long period in the soil, this disease could represent a severe threat for cropping of Brassicaceae in Luxembourg and neighboring countries. References: (1) I. R. Crute et al. Plant Breed. Abstr. 50:91, 1980. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic press: San Diego, 1990.

Cloned cDNA fragments representing different regions of the genome of the H serotype of Indian peanut clump furovirus (IPCV) were used as hybridization probes. Dot-blot assays using digoxigenin-labeled probes readily detected IPCV in extracts from infected plants with a sensitivity approaching that of using 32 P-labeled probes. The specificity of the probes depended on the part of the IPCV genome represented in the cDNA. Two probes representing the coat protein gene and adjacent gene in RNA-2 readily detected infection by H serotype IPCV, but reacted poorly with samples of plants infected with other serotypes. A probe corresponding to sequence near the 5'-end of RNA-1 reacted with samples of all IPCV serotypes, but not with those of peanut clump virus (PCV) from West Africa. A probe corresponding to the 3'-terminal 742 nucleotides of RNA-1 readily detected RNA of any IPCV serotype and of PCV. Assays using this probe, labeled with a nonradioactive reporter-digoxigenin, could detect IPCV in samples of seed tissue, as well as in tissue of several weed species and crop species such as wheat. The probe did not react with samples from plants infected by other furoviruses.

International audience; The goal of this study was to investigate how the microbial community structure establishes during the start-up phase of a full-scale farm anaerobic reactor inoculated with stale and cold cattle slurry. The 16S/18S high-throughput amplicon sequencing results showed an increase of the bacterial, archaeal and eukaryotic diversity, evenness and richness during the settlement of the mesophilic anaerobic conditions. When a steady performing digestion process was reached, the microbial diversity, evenness and richness decreased, indicating the establishment of a few dominant microbial populations, best adapted to biogas production. Interestingly, among the environmental parameters, the temperature, alkalinity, free-NH3, total solids and O2 content were found to be the main drivers of microbial dynamics. Interactions between eukaryotes, characterized by a high number of unknown organisms, and the bacterial and archaeal communities were also evidenced, suggesting that eukaryotes might play important roles in the anaerobic digestion process.

Projected climate changes will affect wheat crop production both in the main processes of plant growth and development but also in the occurrences and severities of plant diseases. We assessed the potential infection periods of wheat leaf rust (WLR) at two climatologically different sites in Luxembourg. A threshold-based model, taking hourly values of air temperatures, relative humidity and precipitation during night-time into account, was used for calculating favourable WLR infection days during three periods throughout the cropping season. Field experiments were conducted during the 2003–2013 period at the selected sites. Projected climate data, from a multi model ensemble of regional climate models (spatial resolution 25 km) as well as an additional projection with a higher spatial resolution of 1.3 km, were used for investigating the potential WLR infection periods for two future time spans. Results showed that the infections of WLR were satisfactorily simulated during the development of wheat at both sites for the 2003–2013 period. The probabilities of WLR detection were close to 1 and the critical success index ranged from 0.80 to 0.94 (perfect score = 1 for both). Moreover, the highest proportions of favourable days of WLR infection were simulated during spring and summer at both sites. Regional climate projections showed an increase in temperatures by 1.6 K for 2041–2050 and by 3.7 K for 2091–2100 compared to the reference period 1991–2000. Positive trends in favourable WLR infection conditions occur at both sites more conducive than in the reference period due to projected climatic conditions.

Adult pollen beetles, Meligethes spp. (Coleoptera: Nitidulidae), were sampled from 27 and 35 oilseed rape (Brassica napus subsp. oleifera) fields scattered over Luxembourg in 2008 and 2009, respectively. Ten well-developed and active individuals from each field were transferred into 40-ml glass vials (4 replicates) with their interior surface being coated with the pyrethroid deltamethrin at rates corresponding to 0%, 0.8%, 4%, 20%, 100%, or 500% of the recommended field application rate. The percentage of dead insects was recorded at 0 h, 1 h, 5 h, and 24 h, and increased with time to a maximum. The application rate required to obtain a mortality of 50% (LC50) after 24 h of exposure was estimated by logistic regression models. The overall median LC50 calculated for all locations and years was 1.15 ± 0.29% of the field rate. The median LC50 for 2008 was 0.79 ± 0.40% of the field rate and 1.80 ± 0.40% in 2009. In 2008, no population with an LC50 below the overall median LC50 was found in the north-eastern part of the country, whereas an approximately homogeneous spatial distribution of populations with LC50s below the overall median was found in 2009, suggesting a spread of less sensitive Meligethes spp. from southwest to northeast within the period of observation. Pollen beetles from four locations were additionally tested for their sensitivity towards bifenthrin and chlorpyri- fos-methyl. The period of time until 50% of the individuals were affected (LT50) decreased exponentially with rate and differed significantly among insecticides (P = 0.005), but not among locations (P = 0.753). The average LT50 at 4% of the field rate was 9.3 ± 2.2 h, 5.4 ± 0.3 h, and 2.9 ± 0.1 h for deltamethrin, bifenthrin, and the organophosphate chlorpyrifos-methyl, respectively.

The anaerobic digestion process leads to the production of biomethane, a versatile renewable energy vector. The dynamics and interactions between specific microbial groups are currently considered as key research subjects towards the improvement of the anaerobic digestion (AD) process. Indeed, deeper knowledge of the ecology of AD, the dynamics of the microbial populations and their structure could provide valuable information regarding unexplained and unpredictable failures or malfunctioning of the anaerobic digestion process. The aim of this work was to characterize the microbial communities involved in the AD process, and to study their responses due to the change of operational parameters such as an increase of the organic loading rate, the reactor type (completely stirred tank reactor vs anaerobic baffled reactor), or the start-up phase of a farm reactor with a shift from psychrophilic to mesophilic temperature range. While we observed the installation of similar microbial populations between replicated reactors under stable conditions, best adapted to biogas production, the microbial communities started to diverge once the operational parameters changed. Indeed, due to deterministic environment, most of bacteria and archaea showed redundant functional adaptation to the changing environmental conditions. However, some dominant bacterial populations were also resistant in terms of presence and abundance to the environmental change. The specific microbial communities established in our studied reactors showed also discrepancies in terms of biogas yields. Furthermore, correlations between the bacterial, archaeal and eukaryotic communities were pointed out, indicating the putative influence of eukaryotes on the anaerobic digestion process and the establishment of the other microbes having crucial functions during the anaerobic biomass digestion. Interestingly, shifts inside the anaerobic microbial community due to the gradual change of operational parameters, were detected prior to any biogas production inhibition, giving the opportunity for the development of potential early microbial indicators for assessing the AD process status and improving the microbial management of anaerobic reactors, Le processus de digestion anaérobie conduit à la production de biométhane, un vecteur flexible d’énergie renouvelable. L’amélioration du rendement de ce processus est souvent évoquée comme dépendante de la compréhension approfondie de la structure et de la dynamique des communautés microbiennes qui y sont impliquées. L’objectif de la thèse a été de caractériser les communautés microbiennes impliquées dans le processus de digestion anaérobie et de déterminer l’influence de facteurs opérationnels sur leurs dynamiques. Nous nous sommes en particulier intéressés à l’augmentation du taux de charge organique, le type de digesteurs anaérobies (réacteur continu perpétuellement mélangé vs réacteur anaérobie à chicane), mais aussi à la phase de démarrage d’un digesteur de ferme avec une montée en température. En absence de conditions contraignantes, nous avons observé l’installation de populations méthanogènes les mieux adaptées à la production de biogaz dans les réacteurs étudiés et la mise en place de communautés microbiennes similaires entre réacteurs réplicats. Cependant, des changements au niveau opérationnel ont conduit au développement de communautés divergentes en termes de structure. En effet, en présence d’un environnement déterministe, la plupart des bactéries et archées impliquées en digestion anaérobie ont montré une redondance fonctionnelle à la perturbation. Toutefois, certaines populations bactériennes dominantes ont également pu montrer des phénomènes de résistance, en termes de présence et d’abondance, à l’évolution des conditions environnementales. Au cours de nos études, les différentes communautés s’installant dans les digesteurs étudiés ont également montré des aptitudes variables pour la production de biogaz. De plus, des corrélations entre les communautés bactériennes, archées et eucaryotes ont aussi été démontrées, soulignant le rôle non négligeable des eucaryotes dans le processus de digestion anaérobie et l’installation de communautés microbiennes dominantes et spécifiques à la production de biogaz. Ainsi, les changements au sein de la communauté microbienne résultant de la modification progressive de facteurs opérationnels, et ce bien avant l’apparition des premiers symptômes d’inhibition de la production de biogaz, pourraient permettre le développement d’indicateurs microbiens de l’état du processus de digestion anaérobie et donc la mise en place d’une gestion microbiologique raisonnée des digesteurs anaérobies