Susceptibility of Oryctes rhinoceros adults to Metarrhizium anisopliae

The susceptibility of Oryctes rhinoceros adults to infections due to entomopathogenic fungi (Beauveria bassiana, Beauveria tenella, Metarrhizium anisopliae, Paecilomyces fumoso-roseus, and Spicaria rileyi) was studied by spraying titrated spores suspensions on the insect integument. The results show a definite susceptibility of the adults to Metarrhizium anisopliae strains of the major type only.     

First Report of a New Isolate of <Emphasis Type="Italic">Metarhizium rileyi</Emphasis> from Maize Fields of Quivicán,Cuba

Metarhizium rileyi (Farlow) Samson is an important entomopathogenic fungus of more than 30 species of Lepidoptera larvae. The aim of this research was to characterize isolate of M. rileyi from Quivicán, Cuba on the basis of morphological and molecular approaches. The fungus was isolated from samples of S. frugiperda larvae collected from maize fields of Quivicán municipality, Mayabeque province, Cuba, and it was cultured on PDA?+?Ampicillin solid media for morphological characterization. The DNA was isolated using CTAB method and internal transcribed spacer (ITS1, ITS4) were used as the primers for the amplification. The amplified products of 1335 bp were purified and sequenced at CINVESTAV-IPN in both the directions using the above primers. A consensus sequence was obtained by alignment of the forward and reverse sequences for this region and deposited in GenBank (MG637450). The fungus produced slightly cottony colony of pale green color and dispersed conidia and septal mycelium were observed under the optical microscope. A BLAST search of the sequence in GenBank revealed a 99% of identity with several strains of N. rileyi (e.g., AF368501.1, AB268359.1 and EU553337.1) and M. rileyi (e.g., KY436756.1). This is the first report of M. rileyi isolate from maize fields of Quivicán in Cuba and this is important for biodiversity studies and is another possibility for Integrated Pest Management.     

Mise au point d'un milieu nutritif pour la culture profonde du champignon entomopathogèneNomuraea rileyi

Résumé La blastogenèse deNomuraea rileyi (Farlow) Samson requiert des besoins très spécifiques. La présence de Tween et une forte concentration en extrait de levure sont indispensables. Contrairement aux conidies, les blastospores obtenues ne sont pas capables de traverser le tégument de l'h?te alors qu'il est démontré que les blastospores de quelques autres espèces sont effectivement entomopathogènes par la voie tégumentaire.

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Summary Nomuraea rileyi (Farlow) Samson blastogenesis requires very specific factors which may change according to the biotype. Limiting components of the medium can be the presence of Tween 80 and a high concentration of yeast extract. The blastospores obtained are not infective when applied topically.

     

Influence of fungal elicitation with Nomuraea rileyi (Farlow) Samson in the metabolism of acclimatized plants of Hypericum polyanthemum Klotzsech ex Reichardt (Guttiferae)

Changes in biomass and bioactive metabolites after treatment with the fungus Nomuraea rileyi (Farlow) Samson added as dried culture (DC) or as dried autoclaved cell powder (DACP) for short and long periods of time have been investigated in Hypericum polyanthemum plants grown under controlled conditions and after 18 weeks of field acclimatization. Plants grown under controlled conditions and treated with DACP showed increased concentrations of the benzopyrans HP1 (6-isobutyryl-5,7-dimethoxy-2,2-dimethylbenzopyran), HP2 (7-hydroxy-6-isobutyryl-5-methoxy-2,2-dimethylbenzopyran), and HP3 (5-hydroxy-6-isobutyryl-7-methoxy-2,2-dimethylbenzopyran) while DC affected negatively or did not alter the synthesis of these compounds. Lower levels of the phloroglucinol derivative uliginosin B were detected in all treatments. Long time treatment with DACP of acclimatized plants triggered doubling of biomass as compared to control and chemical analyses demonstrated significant increase on total phenolic compounds in vegetative parts of plants. Moreover, the higher performance liquid chromatography analysis showed different pattern of metabolites accumulation, with higher yields of HP1, HP2, HP3 and uliginosin B in the reproductive parts of the plants treated with DACP during all experiments. The elevation of bioactive metabolites levels in response to the elicitor suggests that these compounds are inducible as part of the defense response of H. polyanthemum.     

Activity of oil-formulated conidia of the fungal entomopathogens Nomuraea rileyi and Isaria tenuipes against lepidopterous larvae

The fungi Nomuraea rileyi and Isaria tenuipes (=Paecilomyces tenuipes) are ecologically obligate, widespread pathogens of lepidopterans. Bioassays were carried out to evaluate the activity of oil-suspended conidia of N. rileyi and I. tenuipes against larvae of Spodoptera frugiperda, Spodoptera exigua, Helicoverpa zea, and Heliothis virescens. The tests consisted of two bioassay sets. In the first set, conidia of N. rileyi and I. tenuipes were suspended in water + Tween 80, and in vegetable (canola, soybean) and mineral (proprietary mixture of alkanes and cyclic paraffins) oils, and tested against S. frugiperda. Both fungi were highly compatible with oils and caused mortalities near 100% in all oil treatments; the lowest LT₅₀ values were 4.7 days for N. rileyi in mineral oil and 6.0 days for I. tenuipes in soybean oil. The second set included additional fungal strains and oil formulations (mineral, canola, sunflower, olive and peanut oils) tested against larvae of S. exigua, S. frugiperda, H. zea and H. virescens. The highest activity was that of N. rileyi in mineral oil against Spodoptera spp., with LT₅₀ values of 2.5 days (strain ARSEF 135) and 3 days (strain ARSEF 762) respectively. For two different isolates of I. tenuipes the lowest LT₅₀ values (5.1-5.6 days respectively) were obtained with mineral oil formulations against Spodoptera spp. and H. zea respectively. Additionally, we tested both fungi against prepupae of all four lepidopteran species. Mortalities with I. tenuipes against S. exigua ranged from 90% to 100% (strains ARSEF 2488 and 4096); N. rileyi caused 95% mortality on S. frugiperda. The activity of formulations depended on host species and oil used; Spodoptera spp. was more susceptible to these fungi than Heliothis and Helicoverpa. The results indicate that a comprehensive evaluation of these entomopathogens in agriculture using oil application technologies is advisable, particularly, in organic and sustainable settings.     

Interactions between a braconid,Microplitis croceipes,and a fungus,Nomuraea rileyi,in laboratory-reared bollworm larvae

The parasite Microplitis croceipes required 1.1 days longer at 26°C to complete development in Heliothis zea larvae than was required for the fungus Nomuraea rileyi to kill the host larvae and sporulate. Host larvae parasitized by M. croceipes or infected with N. rileyi failed to complete a fifth larval molt or pupate. Of the remaining healthy larvae, one-half completed six larval stadia before popation. Larvae parasitized by M. croceipes were predisposed to infection by N. rileyi, but the fungus inhibited development of M. croceipes if host larvae were infected with N. rileyi within 1 day after parasitization.     

Susceptibility of the cabbage looper, Trichoplusia ni, and the velvetbean caterpillar, Anticarsia gemmatalis, to several isolates of the entomopathogenic fungus Nomuraea rileyi

Cabbage looper larvae, Trichoplusia ni, were equally susceptible to isolates of the entomogenous fungus Nomuraea rileyi from Missouri, Mississippi, and Brazil; an isolate from Florida was 7–17 times less active than the other isolates. Velvetbean caterpillars, Anticarsia gemmatalis, were either only slightly susceptible or not susceptible to the isolates of N. rileyi from Missouri, Florida, and Mississippi. In contrast, larvae of A. gemmatalis cultured from three locations (Missouri, Florida, and Brazil) all were equally susceptible to a Brazilian isolate of N. rileyi.     

Characterization of Chitinase Genes from an Alkaliphilic Actinomycete, Nocardiopsis prasina OPC-131

An alkaliphilic actinomycete, Nocardiopsis prasina OPC-131, secretes chitinases, ChiA, ChiB, and ChiBΔ, in the presence of chitin. The genes encoding ChiA and ChiB were cloned and sequenced. The open reading frame (ORF) of chiA encoded a protein of 336 amino acids with a calculated molecular mass of 35,257 Da. ChiA consisted of only a catalytic domain and showed a significant homology with family 18 chitinases. The chiB ORF encoded a protein of 296 amino acids with a calculated molecular mass of 31,500 Da. ChiB is a modular enzyme consisting of a chitin-binding domain type 3 (ChtBD type 3) and a catalytic domain. The catalytic domain of ChiB showed significant similarity to Streptomyces family 19 chitinases. ChiBΔ was the truncated form of ChiB lacking ChtBD type 3. Expression plasmids coding for ChiA, ChiB, and ChiBΔ were constructed to investigate the biochemical properties of these recombinant proteins. These enzymes showed pHs and temperature optima similar to those of native enzymes. ChiB showed more efficient hydrolysis of chitin and stronger antifungal activity than ChiBΔ, indicating that the ChtBD type 3 of ChiB plays an important role in the efficient hydrolysis of chitin and in antifungal activity. Furthermore, the finding of family 19 chitinase in N. prasina OPC-131 suggests that family 19 chitinases are distributed widely in actinomycetes other than the genus Streptomyces.     

Orthologous peramine and pyrrolopyrazine-producing biosynthetic gene clusters in Metarhizium rileyi,Metarhizium majus and Cladonia grayi

Peramine is a non-ribosomal peptide-derived pyrrolopyrazine (PPZ)-containing molecule with anti-insect properties. Peramine is known to be produced by fungi from genus Epichloë, which form mutualistic endophytic associations with cool-season grass hosts. Peramine biosynthesis has been proposed to require only the two-module non-ribosomal peptide synthetase (NRPS) peramine synthetase (PerA), which is encoded by the 8.3 kb gene perA, though this has not been conclusively proven. Until recently, both peramine and perA were thought to be exclusive to fungi of genus Epichloë; however, a putative perA homologue was recently identified in the genome of the insect-pathogenic fungus Metarhizium rileyi. We use a heterologous expression system and a hydrophilic interaction chromatography-based analysis method to confirm that PerA is the only pathway-specific protein required for peramine biosynthesis. The perA homologue from M. rileyi (MR_perA) is shown to encode a functional peramine synthetase, establishing a precedent for distribution of perA orthologs beyond genus Epichloë. Furthermore, perA is part of a larger seven-gene PPZ cluster in M. rileyi, Metarhizium majus and the stalked-cup lichen fungus Cladonia grayi. These PPZ genes encode proteins predicted to derivatize peramine into more complex PPZ metabolites, with the orphaned perA gene of Epichloë spp. representing an example of reductive evolution.     

Steroids,fatty acids and ceramide from the mushroom Stropharia rugosoannulata Farlow apud Murrill

Stropharia rugosoannulata Farlow apud Murrill is an edible mushroom. Phytochemical investigation of the sporophore of Stropharia rugosoannulata resulted in isolation and identification of 16 compounds, including steroids (1-6), steroidal saponins (7), fatty acids (8–10), alkanes (11), ceramides (12), esters (13), pyrimidines (14), vitamins (15) and flavonoids (16). The structure elucidation of isolated compounds was achieved on the basis of NMR and mass spectral data. Compounds 4–16 were isolated for the first time from this genus and their chemotaxonomic significance was discussed. The isolation of steroids (4–5) and ceramides (12) might be used as chemotaxonomic markers for the genus of Stropharia.     

Disjunct performance and distribution in the sedge Carex prasina

Efforts to understand species distributions and predict responses to environmental changes depend on specifying how the abiotic environment determines distributions. At landscape scales, it is critical to distinguish effects of environmental factors from other mechanisms such as competition and dispersal limitation. We examined how environmental factors affect the distribution and performance of the sedge Carex prasina across a 10-km² old-growth forest in southern Québec. We isolated the effects of soil characteristics by conducting a greenhouse experiment that assessed the performance of C. prasina on soils from a range of wetland habitats where it could potentially occur. This allowed us to compare how the species’ performance and its distribution across the landscape relate to the same soil characteristics. In the experiment, the biomass and leaf chlorophyll content of C. prasina increased with increasing soil organic matter (OM). Across the landscape, however, the species’ probability of occurrence and abundance decreased with increasing soil OM. C. prasina had similar biomass on soils from sites where it did and did not occur, but it had higher leaf chlorophyll content on soils from sites where it did not occur. We found no evidence that differential performance across environments determines the distribution of this species, as C. prasina tended to occur on soils where it showed reduced performance. Rather, other processes such as competition or dispersal limitation likely override any direct effects of the soil environment on distribution. Our results caution against the common assumption that the environments where a species tends to occur or be most abundant are the environments where it performs best. C. prasina presents a clear example of a species whose performance, at least along edaphic gradients, cannot explain its distribution. This example highlights the importance of distinguishing the relative roles of biotic and abiotic factors that shape species distributions across landscapes.     

Soybean leaf consumption by Nomuraea rileyi (Fungi: Deuteromycotina)-infected Plathypena scabra (Lepidoptera: Noctuidae) larvae

To determine the impact of Nomuraea rileyi on consumption by the green cloverworm, Plathypena scabra, larvae were reared from eggs obtained from field-collected moths, inoculated with conidia, and placed individually in separate plastic vials with a piece of surface-sterilized soybean leaflet. No significant differences in consumption rates were found between N. rileyi-inoculated and control larvae until after 144 hr post-treatment. After this period, consumption by larvae inoculated as first, second, third, or fourth instars was significantly less than that of control larvae because of mortality. First or second instars inoculated with N. rileyi conidia had significantly longer subsequent stadia than did control larvae. The average LT₅₀ of all inoculated instar groups was 6.5 days, and there was little evidence of significantly different N. rileyi susceptibility among instar groups. In general during the N. rileyi infection period leading to their deaths, inoculated P. scabra larvae had consumption patterns that were very similar to those of healthy larvae.     

Role of the exoprotease InA in the pathogenicity of Bacillus thuringiensis in pupae of Hyalophora cecropia

Two geographical biotypes of Nomuraea rileyi (from Ecuador and the United States) were topically bioassayed against seven lepidopteran species, i.e., Anticarsia gemmatalis, Heliothis zea, Heliothis virescens, Heliothis subflexa, Pseudoplusia includens, Spodoptera exigua, and Trichoplusia ni. There was an average difference of 1.7-fold in mortality in how cultures of the same insect species from different sources responded to topical applications of either biotype of N. rileyi. Regression equations and LC₅₀ values were obtained for each insect species and fungal biotype combination. Larvae of S. exigua were equally susceptible to both biotypes of N. rileyi. Although larvae of A. gemmatalis were moderately susceptible to the Ecuadoran biotype, they were relatively nonsusceptible to the Mississippian biotype. Species of Heliothis (H. zea, H. virescens, and H. subflexa) were about equally susceptible to the Mississippian biotype. Larvae of H. subflexa and H. virescens, however, were significantly less susceptible than H. zea to the Ecuadoran biotype. When the integumental barrier was breached via intrahemocoelic injections, larvae of H. virescens were as susceptible as H. zea larvae to blastospores of either biotype of N. rileyi.     

Laboratory studies of the entomopathogenic fungus Nomuraea rileyi: Soil-borne contamination of soybean seedlings and dispersal of diseased larvae of Trichoplusia ni

Germinating soybean seedlings were contaminated by soil-borne conidia of the entomogenous fungus Nomuraea rileyi. Although N. rileyi was detected on the unifoliate and trifoliate leaflets, most of the inoculum was found on the cotyledons. Diseased larvae of Trichoplusia ni released on the first trifoliate leaves of soybean plants dispersed to all trifoliates and died. In this way, inoculum is produced that infects other larvae feeding on these leaves.     

An older name for Lomentaria baileyana (Harvey) Farlow (Rhodymeniales, Rhodophyta)

The red alga Chrysymenia divaricata Durant, described from New York Harbor, is shown to belong to Lomentaria. The binomial Lomentaria divaricata (Durant) M.J. Wynne comb. nov. is proposed. This name includes the taxonomic synonyms L. baileyana (Harvey) Farlow and Chondrothamnion divaricatum Bailey ex Kützing.     

Micarea viridileprosa sp. nov., an overlooked lichen species from Western Europe

Abstract:A new lichen species, Micarea viridileprosa, is described from western Europe, where it is locally very common. This species, which has previously been much overlooked, is corticolous, lignicolous and terricolous and is usually found sterile. Some specimens of the new species were formerly included in Micarea prasina. The new combination Micarea levicula (Nyl.) Coppins is made for Lecidea levicula Nyl. from Cuba.     

A new xanthone-containing Micarea from Northwest Europe and the Pacific Northwest of North America

Abstract:Micarea xanthonica sp. nov. is described from the western British Isles, westernmost Norway and the Pacific Northwest of North America. It belongs to the M. prasina group, but differs from related species in possessing xanthones in the thallus.     

Characterization of an extracellular lipase from the biocontrol fungus, Nomuraea rileyi MJ, and its toxicity toward Spodoptera litura

An extracellular lipase from Nomuraea rileyi MJ was purified 23.9-fold with 1.69% yield by ammonium sulfate precipitation followed by Sephacryl S-100 HR column chromatography. By mass spectrometry and SDS-polyacrylamide gel electrophoresis, the molecular weight of the homogenous lipase was 81 kDa. The N-terminal sequence was determined as LeuSerValGluGlnThrLysLeuSerLysLeuAlaTyrAsnAsp and it showed no homology to sequences of known lipases. The optimum pH and temperature for activity were 8.0 and 35 °C, respectively. The enzyme was stable in the pH range 7.0-9.0 and at 15-35 °C for 1 h. Higher activity was observed in the presence of surfactants, Na⁺, NH₄⁺ ions, NaN₃ and ethylenediaminetetraacetic acid (EDTA), while Co²⁺ and Cu²⁺ ions, cysteine and dithiothreitol (DTT) strongly inhibited activity. The purified lipase hydrolyzed both synthetic and natural triglycerides with maximum activity for trilaurin and coconut oil, respectively. It also hydrolyzed esters of p-nitrophenol (pNP) with highest activity for p-nitrophenyl caprate (pNPCA). The purified lipase was found to promote N. rileyi spore germination in vitro in that germination reached 98% in conidial suspensions containing purified lipase at 2.75 U. Moreover, it enhanced toxicity of N. rileyi toward Spodoptera litura larvae with mortality via topical application reaching 63.3% at 4-10 days post-treatment which calculated to be 2.7 times higher than the mortality obtained using conidial suspensions alone.