Negative cross-resistance of benzimidazole-resistant strains of Botrytis cinerea,Fusarium nivale and Pseudocercosporella herpotrichoides to various pesticides

Among benzimidazole-resistant strains ofFusarium nivale andPseudocercosporella herpotrichoides negative cross-resistance to N-phenylcarbamates like barban or chlorpropham was more common than to diethofencarb. Such differences were also observed with N-methylcarbamate or organophosphorus insecticides and with triazine herbicides. Several compounds belonging to these various groups of pesticides were highly toxic against the most common benzimidazole-resistant isolates ofBotrytis cinerea. They were not active, however, against isolates recently found in French vineyards that were treated with a mixture of diethofencarb and carbendazim. Some diphenylether derivatives seemed to be effective against all the benzimidazole-resistant phenotypes of this fungus.     

Identification and characterization of <Emphasis Type="Italic">Colletotrichum</Emphasis> spp. associated with chili anthracnose in peninsular Malaysia

Anthracnose fruit rot caused by Colletotrichum spp. is a serious post-harvest disease of chili fruits (Capsicum spp.). One hundred-thirty isolates of Colletotrichum spp. were isolated from anthracnose of green and red cayenne pepper (Capsicum annuum) and bird’s eye chili (Capsicum frutescens). The isolates were morphologically identified as Colletotrichum acutatum sensu lato (62 isolates), Colletotrichum truncatum (54 isolates), and Colletotrichum gloeosporioides sensu lato (14 isolates). Molecular identification and phylogenetic analyses were based on internal transcribed spacer regions, β-tubulin, actin, and glyceraldehyde-3-phosphate dehydrogenase genes, and the isolates were re-identified as C. scovillei (58 isolates), C. truncatum (54 isolates), C. siamense (11 isolates), C. fioriniae (four isolates), and C. fructicola (3 isolates). Maximum likelihood trees using combined sequences showed that isolates of the same species grouped in the same main clade with the isolates used for comparison. Pathogenicity testing showed that the tested isolates from each species were pathogenic towards green and red Capsicum annuum and Capsicum frutescens upon treatment of wounded fruit, using both the mycelial plug and conidial suspension methods. Only five isolates of C. truncatum and seven isolates of C. scovillei were found to be pathogenic upon treatment of unwounded fruit. The occurrence of five Colletotrichum spp. (C. siamense, C. fructicola, C. scovillei, C. fioriniae, and C. truncatum) associated with chili anthracnose in Peninsular Malaysia indicates that correct species identification is important to formulate not only effective disease management, but also effective quarantine policy.     

Multi-resistance to thiophanate-methyl,diethofencarb, and procymidone among <Emphasis Type="Italic">Alternaria alternata</Emphasis> populations from tobacco plants,and the management of tobacco brown spot with azoxystrobin

In 2014 and 2015, a total of 151 tobacco brown spot (Alternaria alternata) isolates were collected from Guizhou Province in China to evaluate their resistance to the benzimidazole thiophanate-methyl, the carbamate diethofencarb, and the dicarboximide procymidone. Resistance to thiophanate-methyl and diethofencarb was observed in all isolates. Resistance to all the three fungicides, thiophanate-methyl, diethofencarb, and procymidone was detected at a frequency of 6.0%. The F167Y single mutation in the β-tubulin gene was found to be associated with resistance to thiophanate-methyl,but no mutation was found in the coiled-coil region of the histidine kinase-encoding gene OS1, a fungal gene for dicarboximide resistance. Procymidone applied at the rate of 20 mg l^?1 inhibited spot lesion formation on tobacco leaves with an efficacy of 51.7% for the low resistance (LR) isolates and 74.2% for the procymidone-sensitive isolates. Thiophanate-methyl applied at 100 mg l^?1, however, slightly promoted the expansion of disease lesions with an efficacy of ?7.7%. Azoxystrobin applied at 10 and 20 mg l^?1 provided efficacies of 91.1 and 100%, respectively, regardless of whether the isolates were thiophanate-methyl resistant or procymidone-LR. Further studies suggested that azoxystrobin exhibited excellent protective activity and good curative activity against A. alternata in plants. The baseline sensitivity to azoxystrobin was then determined. In the presence SHAM, the mean EC₅₀ values for conidial germination inhibition were 0.49?±?0.22 (Mean?±?SD) mg l^?1. Interestingly, no resistance was recovered through UV irradiation or Agrobacterium tumefaciens-mediated mutagenesis. This research indicated widespread resistance to thiophanate-methyl and diethofencarb, low frequency of (6.0%) resistance to procymidone in A. alternata populations from tobacco, and suggested that azoxystrobin could potentially constitute a good alternative for the management of tobacco brown spot disease.     

Screening for Antifungal Potential of Plant Extracts of Fifteen Plant Species Against Four Pathogenic Fungi Species

The antifungal activity of fifteen wild plant species grown in the Al-Qassim region, Saudi Arabia, was investigated against the fungi species Fusarium solani, Botrytis cinerea, Alternaria alternata and Stemphylium botryosum at concentrations of 10, 20, 40 and 80?g/L. The inhibitory effect of test extracts varied among examined fungi, and Fusarium solani exhibited the least sensitivity compared to other fungi species. Antifungal activity of the tested extracts was proportional to the applied dose. Extracts of Lactuca virosa, Neurada procumbens, Ochradenus baccatus and Cyperus conglomerates showed relatively low inhibition effects on the fungi species. The most effective plants were Pulicaria undulata, Artemisia monosperma, Prosopis juliflora, Withania somnifera and Rumex vesicarius. At 80?g/L, extracts of these plant species reduced mycelial growth of Botrytis cinerea by 16.5–32.2%, of Fusarium solani by 11.1–27.9%, of Alternaria alternaae by 26.9–63.5% and of Stemphylium botryosum by 22–40%. The methanolic extract of the most effective plant species was further fractioned with hexane, methylene chloride and ethyl acetate. The obtained fractions varied in their effects on mycelial growth of the four tested fungi. Using the same fraction resulted in different inhibition effects on mycelial growth of all tested fungi. The antifungal activity of each crude extract tended to be distributed among its three fractions, probably because the bioactive components were also distributed among the fractions.     

Disease resistance in Vicia faba and Phaseolus vulgaris

Droplets of spore suspensions of each of four isolates ofBotrytis cinerea but not those of each of four isolates ofB. fabae proved to contain an antifungal compound 24 h after application on pod tissue ofVicia faba. Partial inhibition of germ-tube growth of three highly pathogenic isolates ofB. fabae was caused at 2.5 times the concentration of inhibitor needed to cause similar inhibition of each isolate ofB. cinerea and a weakly pathogenic isolate ofB. fabae. After extraction, concentration and chromatographic separation, 5–10 times more inhibitor was obtained from lesions in pods caused byB. cinerea than from those caused byB. fabae. However, the amounts of inhibitor extracted from whole leaves bearing either large lesions caused byB. fabae or small lesions caused byB. cinerea were almost the same. It is suggested that infection by either fungus induces inhibitor formation, but thatB. fabae metabolizes the inhibitor to an inactive form.No relation was found between amounts of an inhibitor produced in droplets of spore suspensions 24 h after application on pods of differential varieties ofPhaseolus vulgaris and the disease reactions caused by races ofColletotrichum lindemuthianum. Each race appeared to have a similar sensitivity to the inhibitor. Anatomical studies showed that only superficial growth of germ-tubes occurred in seed cavities in the first two days, after which penetration took place. Resistant or susceptible reactions were distinguished after 6 days in young pods, and even later in old pods. Before rejecting the hypothesis that the inhibitor may have a role in the mechanism of disease resistance, amounts of inhibitor in and around infection sites on leaves or stems should be measured. Apparent protection of leaf areas against infection was caused by prior inoculation with a race which was avirulent on the leaf. This phenomenon is consistent with the action of an inhibitor of the type found in pod tissues, but could be caused by reactions as yet unknown.     

Comparison among Japanese isolates of <Emphasis Type="Italic">Pseudomonas savastanoi</Emphasis> pv. <Emphasis Type="Italic">savastanoi</Emphasis>, causal agent of olive knot disease

Olive knot disease in Japan was first reported in Shizuoka Prefecture in 2014, and the causal agent was identified as Pseudomonas savastanoi pv. savastanoi. Subsequently, olive trees having knots were also found in Aichi and Kanagawa Prefectures in 2015, and the isolates from knots were also suspected to be P. savastanoi pv. savastanoi through preliminary examinations. Therefore, the Aichi and Kanagawa isolates were identified through comparison of isolates from three prefectures. Phylogenic analysis based on 16S rDNA and housekeeping genes (gyrB, rpoD, gltA and gap1) revealed that the isolates belonged to the same cluster as the pathotype strain, ICMP4352^PT. The iaaM, H and L genes, which are involved in promotion of symptoms, and the ina gene coding the ice nucleation protein, were detected by PCR from all the isolates. In rep-PCR (ERIC and REP) analyses, the isolates yielded DNA fragment-banding patterns that were nearly identical to that of ICMP4352^PT, but slight variations in banding patterns were observed among them. In a pathogenicity test, the isolates formed distinct knots on olive and pink jasmine. Phenotypic properties of the isolates were almost identical to those of ICMP4352^PT, with the exception of d-sorbitol utilization. Consequently, Aichi and Kanagawa isolates from olive were identified as P. savastanoi pv. savastanoi, and several genetic diversities in terms of rep-PCR were found in the Japanese population of P. savastanoi pv. savastanoi, indicating their heterogeneity.     

The potential of entomopathogenic fungi to control the lettuce root aphid,<Emphasis Type="Italic">pemphigus bursarius</Emphasis>

The lettuce root aphid,Pemphigus bursarius, is a commercially important pest of lettuce and occupies a niche amenable to the action of entomopathogenic fungi. As the first stage of a biological control programme, a bioassay was developed against adult apterousP. bursarius. An isolate ofMetarhizium flavoviride was pathogenic toP. bursarius and sporulated abundantly upon aphid cadavers. However, two isolates ofVerticillium lecanii, pathogenic against glasshouse whitefly and aphids, were only weakly pathogenic toP. bursarius.     

Inhibitory effect of <Emphasis Type="Italic">Lycium europaeum</Emphasis> extracts on phytopathogenic soil-borne fungi and the reduction of late wilt in maize

The ability to control soil-borne pathogens in agriculture is highly conditioned by the restricted use of synthetic pesticides. Allelopathy, the antimicrobial activity of plant extracts, is a promising option against crop pathogens. Extracts from Lycium spp. such as L. barbarum, L. chinense and L. intricatum possess biological and therapeutic properties. Individual methanolic extracts from leaves and stems of the Mediterranean medicinal species L. europaeum collected in two locations of Tunisia were each evaluated in vitro against Verticillium dahliae (Vd), Sclerotinia sclerotiorum (Ss) and Harpophora maydis (Hm). The mycelial growth of the three fungi was significantly reduced by all the extracts at doses of 10 and 30 μl mL^?1 (equivalent to 1 and 3 mg plant tissue mL^?1). The sporulation of Hm was almost completely inhibited in all the amendments, but that of Vd was stimulated by one of the leaf extracts when 1 and 3 mg dried plant tissue mL^?1 were used. Sclerotia of Ss were formed in a smaller number, their total weight increasing at extract doses equivalent to 1 mg plant tissue mL^?1 and higher. In greenhouse, the pathogenicity of Hm was confirmed as early as 6 weeks after inoculation, since it caused significant decreases of weights in both roots and aboveground parts of maize. The detrimental effect of Hm on maize root weight in greenhouse was significantly counteracted by one of the leaf extracts added by watering. In total, 11 phenolic compounds were separated in the four extracts. The hydroxycinnamic acid family, including chlorogenic acid as a major compound, represented more than 50% of the total content in all the samples. Rutin was the most abundant flavonoid. The results of this work show the detrimental effect of L. europaeum extracts against the soil-borne pathogens Hm, Ss and Vd, and highlight their potential in crop protection if adequately developed into final products and used in combination with other tools.     

Characterization of <Emphasis Type="Italic">Myoviridae</Emphasis> and <Emphasis Type="Italic">Podoviridae</Emphasis> family bacteriophages of <Emphasis Type="Italic">Erwinia amylovora</Emphasis> from Hungary - potential of application in biological control of fire blight

The virulence spectrum of 300 isolates of Xanthomonas oryzae pv. oryzae (Xoo), representing 17 districts of Punjab, Pakistan was elucidated through inoculation on a set of six rice IRRI-differentials. The virulence level was assessed by using principal component and cluster analysis. Among six principal components (PCs), PC-1 exhibited 59.3 % of the total variance. The highly virulent isolates clusters on the positive side of the ordination away from the point of intersection of PC1 and PC2 and classifies the Xoo isolates from slow disease to the highest disease causing entities. The 300 isolates were categorized into 29 pathotypes (Pt1-29) wherein the highly virulent pathotype (Pt-1), comprises of 39 Xoo isolates were widespread in 12 districts. The majority of Xoo isolates were moderately to least virulent (21.7–43 %) and average disease progress curves confirmed the field reactions of these pathotype clusters for an efficient recognition of Xoo isolates. Interaction of the pathogen with differentials harboring different resistant genes was well investigated in the current study for future management approaches for which the surveillance of the new Xoo pathotypes may expedite the disease resistant rice breeding programme in the country.     

Studies on the selective fungitoxicity of aliphatic amines

A comparison of the uptake of simple (C₁–C₄) aliphatic amines byPenicillium digitatum revealed that the fungitoxicity of (?)sec-butylamine [(?)SBA] was not due to its accumulation by hyphae since (+)SBA was accumulated to the same extent and methylamine, which showed negligible antifungal activity, accumulated to twice the level of SBA. Amines with a secondary alkyl structure were resistant to fungal metabolism, whereas primary amines were degraded to a significant extent byP. digitatum during a 4 h incubation period.(?)SBA accumulated in the fresh hyphae ofP. digitatum to a level 24 times higher than that in the culture medium containing 1 μmol ml^?1; most of the SBA effluxed from the hyphae when transferred to fresh culture medium minus SBA. SBA did not accumulate when hyphae were incubated in N₂ or in the presence of respiratory inhibitors. The absorption and accumulation of SBA is characteristic of active transport.Penicillium species and biotypes that are sensitive to SBA did not accumulate more (?)SBA than resistant fungi. SBA-resistant biotypes ofP. digitatum accumulated SBA to twice the level of SBA-sensitive biotypes, but did not accumulate pyruvate in the hyphae, which is characteristic of SBA-altered metabolism.No evidence was found to implicate exclusion or metabolic detoxification as mechanisms of SBA resistance. More probably, resistance involves cytoplasmic sequestration of SBA or low affinity of the biochemical target, pyruvic dehydrogenase.     

Special issue “Deepen knowledge in plant pathology for innovative agroecology”

This study aimed to determine the genetic variability of isolates of rotting of pineapple fruitlet core in Brazil on the states of Paraiba, Pernambuco and Rio Grande do Norte, based on phylogenetic analysis of the RPB2 gene, morphocultural markers and aggressiveness of the isolates. The phylogenetic reconstruction of maximum parsimony and bayesian inference of the isolates were performed. Twenty-seven phylogenetic lineages were characterized with morphocultural markers on potato dextrose agar and synthetic nutrient-poor agar. The aggressiveness of these lineages were characterized in leaves and fruits of pineapple ‘Pérola’ cultivar. The Phylogenetic reconstruction showed close relationship between isolates of this study and phylogenetic lineages of F. guttiforme, F. ananatum and F. oxysporum by analysis of RPB2 gene. Phylogenetic lineages of this study shared significant morphocultural markers similar to those described for this species. Overall, the lineages related with Fusarium oxysporum Species Complex were more aggressive to the fruits of the Fusarium lineages related with Fusarium fujikuroi Species Complex. It is possible that F. ananatum and F. guttiforme or other lineages closely related to these species or F. oxysporum are present in the fields causing pineapple rotting fruitlet core in fruits of pineapple ‘Pérola’ in Brazil. The used markers determined high genetic variability in isolates analyzed in leaves and pineapple fruit ‘Pérola’ in the states of Paraíba, Pernambuco and Rio Grande do Norte of Brazil and the pathogenic lineages analyzed were better adapted to the fruits than to the detached leaves in pineapple ‘Pérola’ analyzed.     

Korean <Emphasis Type="Italic">Brassica oleracea</Emphasis> germplasm offers a novel source of qualitative resistance to blackleg disease

Blackleg disease, caused by the hemibiotrophic fungal pathogen Leptosphaeria maculans, is one of the most devastating disease of Brassica species worldwide. To date, a total of 20 race-specific blackleg resistance (R) genes have been reported and all of those loci are located in either the A or B genomes of various Brassica species. The B. oleracea genome (CC) shares a high ancestral synteny with the A genome of B. rapa, suggesting the presence of qualitative (race specific) resistance to blackleg disease is also possible in B. oleracea germplasm. In the present study the C genome of Korean B. oleracea germplasm was screened for the presence of blackleg R genes. Thirty-two inbred cabbage lines with unknown resistance profiles, along with five control B. napus lines with well-characterised race-specific R genes, were assessed for cotyledon resistance against two L. maculans isolates with known and highly-contrasting avirulence gene (Avr) profiles. Two cabbage accessions were identified which produced a strong resistance when challenged with either isolate, demonstrating the presence of effective blackleg R genes in the cabbage C genome. Additionally, 16 microsatellite markers linked to seven different R genes of the B. napus A genome were converted into markers for their homologous regions on the B. oleracea C genome. These markers were used to screen all B. oleracea lines to assess if the novel C genome R genes were syntenous to known R gene-homologous regions of the A genome. The resistant cabbage lines offer C genome R genes for the protection of B. oleracea varieties against incursion of blackleg disease, as well as novel additional resistance sources for introgression into B. napus and B. carinata breeding material.     

Fungicide sensitivity,growth rate,aggressiveness and frost hardiness of <Emphasis Type="Italic">Monilinia fructicola</Emphasis> and <Emphasis Type="Italic">Monilinia laxa</Emphasis> isolates

Monilinia fructicola, the most destructive pathogen of the genus Monilinia, has recently been introduced into Serbia and many other European countries. Since then, many studies have been conducted to evaluate the characteristics of Monilinia species that have a role in the establishment and survival of the pathogen in new areas. The present study assessed the capacity of M. fructicola to repress and replace Monilinia laxa in Serbia based on: fungicide sensitivity, growth rate and aggressiveness at different temperatures, as well as frost hardiness of the isolates of both species. The results showed that the isolates of M. fructicola, compared to M. laxa, were significantly less sensitive to the following fungicides: iprodione, tebucanozole, chlorothalonil, azoxystrobin, fluopyram, and boscalid. In addition, M. laxa isolates exhibited little variation in sensitivity to all of the tested fungicides, whereas M. fructicola isolates displayed a wide range of sensitivity. The temperature of 5°C favored M. laxa growth and aggressiveness, while at 30°C M. fructicola grew faster and had higher lesion expansion rate. These results support an assumption that M. fructicola will continue to spread in Serbian orchards in coming years, particularly on stone fruits harvested during hot summer weather.     

Possibilities of biological and chemical control of Verticillium wilt in pepper

In pathogen populations in Serbia, the incidence, pathogenic and morphological characters ofVerticillium spp. were studied. Biological and chemical control ofVerticillium was investigated in pepper plants (Capsicum annuum L. cv. ‘Soroksari’) with the biofungicide Polyversum^® (Pythium oligandrum) and the conventional fungicides benomyl and propamocarb-hydrochloride. On the basis of macroscopic and microscopic characters of the isolates originating from eight localities in Serbia, it was established that they apparently belong to the speciesVerticillum dahliae. The isolates differed in their pathogenic characters. However, all of them caused marked wilting symptoms on pepper plants 40 days after inoculation, conducted when there were more than nine fully developed leaves on the primary stem. The fungicides were applied either before or after inoculation. Benomyl was the most efficient fungicide in wilt control (88.2% when applied after inoculation and 94.6% when applied before inoculation). Polyversum proved more efficient (66.6%) when applied before rather than after inoculation. Propamocarb-hydrochloride provided sufficient Verticillium wilt control; its efficacy and that of Polyversum were similar, and less efficient than benomyl, but still significantly different from the disease control.     

Assessing the Belgian potato <Emphasis Type="Italic">Alternaria</Emphasis> population for sensitivity to fungicides with diverse modes of action

Early blight and brown spot, caused by respectively Alternaria solani and Alternaria alternata, can lead to severe yield losses in potato-growing areas. To date, fungicide application is the most effective measure to control the disease. However, in recent years, a reduced sensitivity towards several active ingredients has been reported. To shed light on this issue, Alternaria isolates were collected from different potato fields in Belgium during two growing seasons. Subsequently, the sensitivity of these isolates was assessed using four widely used fungicides with different modes of action. Demethylation inhibitors, quinone outside inhibitors, a dithiocarbamate and a carboxylic acid amide were included in this study. Although all fungicides reduced spore germination and vegetative growth of Alternaria species to some extent, the interspecies sensitivity was very variable. In general, A. solani was more suppressed by the fungicides compared to A. alternata. The effectiveness of the dithiocarbamate mancozeb was high, whereas the quinone outside inhibitor azoxystrobin showed a limited activity, especially towards A. alternata. Therefore, a subset of the A. alternata and A. solani isolates was tested for the presence of, respectively, the G143A substitution and the F129L substitution in the cytochrome b. The frequency of A. alternata isolates bearing the resistant G143A allele (approximately 65%) was comparable in both sampling years, although sensitivity of isolates decreased during the growing season. This finding points to a shift of the population towards resistant isolates. Both the European genotype I and American genotype II were present in the A. solani population, with genotype I being the most prevalent. None of the genotype I isolates carried the F129L substitution, whereas in 83% of the genotype II isolates this substitution was present. Our results demonstrate for the first time that the Belgian Alternaria population on potato comprises a considerable broad spectrum of isolates with different sensitivity to fungicides.     

Activity of <Emphasis Type="Italic">ß</Emphasis>-1,3-glucanase and <Emphasis Type="Italic">ß</Emphasis>-1,4-glucanase in two potato cultivars following challenge by the fungal pathogen <Emphasis Type="Italic">Alternaria solani</Emphasis>

Early blight of potato, caused by Alternaria solani, is a ubiquitous disease in many countries around the world. Our previous screening of several Iranian potato cultivars found that variation in resistance exists between two cultivars: ‘Diamond’ and ‘Granula’. Cultivar Diamond is more resistant to multiple isolates of A. solani when compared to cv. Granula. Furthermore, we have found that different pathogen isolates have varying degrees of infection. We monitored the activities of two pathogen-related (PR) glucanase proteins in Diamond and Granula in response to two isolates of A. solani with different degrees of virulence. ß-1,3-glucanase and ß-1,4-glucanase activities were recorded in healthy and diseased leaves of potatoes up to 10 days after inoculation. Their activities were found to be higher in diseased leaves when compared to those of uninfected leaves. Our data suggest that significantly reduced activities of theses enzymes in potato could be related to a lower degree of resistance or an increased ability of a more aggressive isolate to suppress PR protein expression.     

Improved assessment of mycelial growth stimulation by low doses of mefenoxam in plant pathogenic <Emphasis Type="Italic">Globisporangium</Emphasis> species

Globisporangium Uzuhashi, Tojo & Kakish. (syn. Pythium Pringsheim) species cause many plant diseases, including Pythium damping-off, leaf and fruit blights, and root rots. Fungicide resistant isolates are selected by repeated use of a single active ingredient on infected crops without rotation. Previous studies demonstrated increased pathogenicity and radial growth in a mefenoxam resistant isolate of Pythium aphanidermatum when exposed to sub-lethal doses of fungicides and ethanol. In those studies, reproducibility of in vitro assays was difficult to achieve due to large variations among trials. This study aimed to examine two protocols for improved reproducibility during the assessment of biphasic dose-responses in mefenoxam-resistant isolates of Globisporangium ultimum and G. irregulare. Two different growth related endpoints, total growth area and total dry mass weight, were assessed. Assays were conducted using ten concentrations of mefenoxam ranging from 0.01 to 1,000 μg/ml. Statistically-significant stimulatory effects were observed in the two Globisporangium species using the two growth related endpoints. Because of its better reproducibility, mycelial growth area is recommended as an endpoint for future studies of chemical hormesis on growth of Globisporangium spp.     

Associated bacterial diversity of insecticide-susceptible and -resistant brown planthopper, <Emphasis Type="Italic">Nilaparvata lugens</Emphasis> (Homoptera: Delphacidae) analyzed by culture-dependent and -independent methods

Nilaparvata lugens (Stål) is one of the major pests of rice throughout tropical and temperate Asia. Indiscriminate use of insecticides for suppressing N. lugens has resulted in the development of resistance to multiple insecticide classes, causing frequent control failures in the field. Analysis of gut bacterial diversity within an insect host is the initial step towards understanding the ecological roles of the symbionts. Present study aimed to survey the bacterial diversity associated with laboratory-reared (insecticide-susceptible) and field-collected (insecticide-resistant) populations of N. lugens by culture-dependent and PCR-Denaturing Gradient Gel Electrophoresis (DGGE) methods. Seventeen bacterial isolates were obtained by the culture-dependent method. Molecular characterization using the 16S rRNA gene and phylogenetic analysis revealed that the isolates belonged to Firmicutes and Proteobacteria. Taxonomic assignment placed these isolates into seven families representing 10 genera. Enterobacteriaceae was the most dominant family with its occurrence in four out of the five populations studied. The DGGE profiles indicated a low complex gut bacteria associated with N. lugens with limited number of bands. The Shannon-Wiener index ranged from 0.898 in insecticide-susceptible population to 0.946–1.035 in resistant populations. Sequencing and phylogenetic analysis revealed that the DGGE bands belonged to Firmicutes, Proteobacteria and Bacteriodetes. Results of this study illustrated that gut bacterial community associated with N. lugens is dominated by Proteobacteria and Firmicutes. Present findings could provide the basis for future work on the possible role of the bacterial symbionts in insecticide resistance and to formulate potential resistance management strategies.     

Identification mating-type locus structure and distribution of <Emphasis Type="Italic">Cochliobolus lunatus</Emphasis> in China

Cochliobolus lunatus (teleomorph: Curvularia lunata) is an important plant pathogenic fungus that causes the maize foliar spot, resulting in serious yield losses. In ascomycetes, a single mating-type (MAT) locus with two idiomorphs controls sexual development. The structure and arrangement of the MAT genes were examined to understand the MAT locus of C. lunatus. MAT loci were MAT1–1-1 or MAT1–2-1, flanked upstream and downstream by regions encoding GTPase activating protein, pyridoxamine phosphate oxidase domain, and β-glucosidase. A MAT1–1 or MAT1–2 idiomorph was identified in single isolate, and sexual reproduction in vitro indicated that the species was heterothallic. In vitro crossing between isolates with opposite MATs produced perithecia, asci, and ascospores. A multiplex MAT-specific PCR method was developed and used to test mating-type genes in 177 C.lunatus isolates collected from China. The ratio of isolates of each mating-type in China was consistent with a 1:1 ratio.