Isolation of Paecilomyces lilacinus (Thom) Samson (Ascomycota: Hypocreales) from the Chagas disease vector, Triatoma infestans Klug (Hemiptera: Reduviidae) in an endemic area in Argentina

A survey for entomopathogenic fungi of the Chagas disease vector Triatoma infestans was conducted in two provinces of Argentina from March–December 2003. Field-collected insects that died in the laboratory were individually maintained in moist chamber and incubated at 22 °C. Triatominae adults infected with the fungus Paecilomyces lilacinus were found at El Quebracho (27°34′S–64°31′W), Santiago del Estero province, Argentina, in December 2003. Paecilomyces lilacinus was cultured and isolated from infected insects in SDAY, PYG and MEA media. Pathogenicity tests were conducted and positive results were recorded. The median survival time (MST) of T. infestans exposed to a P. lilacinus conidial suspension was 12.8 days, and 100% mortality occurred at 30 days post-treatment. This is the first record of natural infection caused by P. lilacinus in T. infestans in the world.     

Toluene gas phase biofiltration by Paecilomyces lilacinus and isolation and identification of a hydrophobin protein produced thereof

Paecilomyces lilacinus consumed toluene as the sole carbon source in a gas-phase biofilter packed with perlite obtaining an average elimination capacity of 50 g m(-3) h(-1), a removal efficiency of 53%, and a final biomass of 31.6 mg biomass g dry support(-1). Hydrophobin proteins from the mycelium produced in the biofilter were purified by formic acid extraction and precipitated by electrobubbling, and the molecular weight was found to be 10.6 +/- 0.3 kDa. The peptide mass fingerprinting analysis of the purified hydrophobin by matrix-assisted laser desorption/ionization time-of-flight resulted in the identification of two peptides that presented high homology with sequences of class I hydrophobin proteins from other ascomycetous fungi when compared against the National Center for Biotechnology Information database. The yield of hydrophobin (PLHYD) from P. lilacinus was 1.1 mg PLHYD g biomass(-1). These proteins modified the hydrophobicity of Teflon by lowering the contact angle from 130.1 (+/-2) degrees to 57.0 (+/-5) degrees supporting hot sodium dodecyl sulfate washing. This work is the first report about biodegradation of toluene by the nematophagous fungus P. lilacinus in a gas-phase biofilter and the identification of its hydrophobin protein.     

Interaction of Glomus mosseae and Paecilomyces lilacinus on Meloidogyne javanica of tomato

The effects of Glomus mosseae and Paecilomyces lilacinus on Meloidogyne javanica of tomato were tested in a greenhouse experiment. Chicken layer manure was used as a carrier substrate for the inoculum of P. lilacinus. The following parameters were used: gall index, average number of galls per root system, plant height, shoot and root weights. Inoculation of tomato plants with G. mosseae did not markedly increase the growth of infected plants with M. javanica. Inoculation of plants with G. mosseae and P. lilacinus together or separately resulted in similar shoots and plant heights. The highest root development was achieved when mycorrhizal plants were inoculated with P. lilacinus to control root-knot nematode. Inoculation of tomato plants with G. mosseae suppressed gall index and the average number of galls per root system by 52% and 66%, respectively, compared with seedlings inoculated with M. javanica alone. Biological control with both G. mosseae and P. lilacinus together or separately in the presence of layer manure completely inhibited root infection with M. javanica. Mycorrhizal colonization was not affected by the layer manure treatment or by root inoculation with P. lilacinus. Addition of layer manure had a beneficial effect on plant growth and reduced M. javanica infection.     

Neglected and emerging fungal infections: review of hyalohyphomycosis by Paecilomyces lilacinus focusing in disease burden, in vitro antifungal susceptibility and management

Paecilomyces lilacinus is an emerging pathogenic fungus that can cause different clinical manifestations ranging from cutaneous and sub-cutaneous infections to severe oculomycosis. This review discusses infections caused by P. lilacinus, as well as their symptoms and correlates of immune responses, morphological characteristics of the fungus, therapies, in vitro susceptibility tests, laboratory diagnosis and the experimental models available.     

Meloidogyne incognita Survival in Soil Infested with Paecilomyces lilacinus and Verticillium chlamydosporium

Meloidogyne incognita-infected tomato seedlings were transplanted into sterilized soil or unsterilized soil collected from 20 California tomato fields to measure suppression caused by Paecilomyces lilacinus, Verticillium chlamydosporium, and other naturally occurring antagonists. Unsterilized soils Q, A, and H contained 35, 39, and 55% fewer M. incognita second-stage juveniles (J2) than did sterilized soil 1 month after infected tomato seedlings were transplanted to these soils and placed in a greenhouse. Three months after infected seedlings were transplanted to unsterilized or sterilized soil, unsterilized soils K, L, and Q had 97, 62, and 86% fewer J2 than the corresponding sterilized soils. Unsterilized soils of M. incognita-infected seedlings that were maintained 1 month in a greenhouse followed by 1 or 2 months of post-harvest incubation contained J2 numbers equal to, or greater than, numbers in the corresponding sterilized soil. The most suppressive of the unsterilized soils, K and Q, were not infested with V. chlamydosporium. Paecilomyces lilacinus and V. chlamydosporium increased in colony forming units in unsterilized soil of all bioassays, but they were not associated with lower numbers of J2.     

Acremonium-like submerged conidiation inPaecilomyces nostocoides andP. lilacinus

Paecilomyces nostocoides, in which conidia of smaller or larger sizes appear in chains, was newly isolated from Japan. In addition to the typicalPaecilomyces-type conidiation, the Japanese isolate showed additionalAcremonium-like submerged conidiation in and/or on some agar media. The submerged conidiation was also observed in the ex-type strains, but not in the type specimens ofP. nostocoides. The same submerged conidiation was observed inP. lilacinus, the species most similar toP. nostocoides. The species definitions ofP. nostocoides andP. lilacinus were emended to include the submerged conidiation.Paecilomyces-type conidia were uninucleate in bothP. nostocoides andP. lilacinus. Paecilomyces nostocoides andP. lilacinus had the Q-10(H₂) ubiquinone system.     

Growth of Isolates of Paecilomyces lilacinus and Their Efficacy in Biocontrol of Meloidogyne incognita on Tomato

The potential of 13 Paecilomyces lilacinus isolates from various geographic regions as biocontrol agents against Meloidogyne incognita, the effects of temperature on their growth, and the characterization of the impact of soil temperature on their efficacy for controlling this nematode were investigated. Maximum fungal growth, as determined by dry weight of the mycelium, occurred from 24 to 30 C; least growth was at 12 and 36 C. The best control of M. incognita was provided by an isolate from Peru or a mixture of isolates of P. lilacinus. As soil temperatures increased from 16 to 28 C, both root-knot damage caused by M. incognita and percentage of egg masses infected by P. lilacinus increased. The greatest residual P. lilacinus activity on M. incognita was attained with a mixture of fungal isolates. These isolates effected lower root-galling and necrosis, egg development, and enhanced shoot growth compared with plants inoculated with M. incognita alone.     

Association of Verticillium chlamydosporium and Paecilomyces lilacinus with Root-knot Nematode Infested Soil

Population densities of Meloidogyne incognita and the nematophagous fungi, Paecilomyces lilacinus and Verticillium chlamydosporium, were determined in 20 northern California tomato fields over two growing seasons. Paecilomyces lilacinus was isolated from three fields, V. chlamydosporium was isolated from one field, and both fungi were isolated from 12 fields. Verticillium chlamydosporium numbers were positively correlated with numbers of M. incognita and P. lilacinus. Paecilomyces lilacinus numbers were positively correlated with V. chlamydosporium numbers, but they did not correlate with M. incognita numbers. The correlation coefficients were low (R < 0.5) but significant (P < 0.05). All P. lilacinus and V. chlamydosporium field isolates parasitized M. incognita eggs in vitro. In a greenhouse study, numbers of V. chlamydosporium and P. lilacinus increased more in soils with M. incognita-infected tomato plants than in soil with uninfected tomato plants. After 10 weeks, the Pf/ Pi of second-stage juveniles in soils infested with P. lilacinus, V. chlamydosporium, and M. incognita was 47.1 to 295.6. The results suggest V. chlamydosporium and P. lilacinus are not effectively suppressing populations of M. incognita in California tomato fields.     

Biological Control of Meloidogyne incognita by Paecilomyces lilacinus and Pasteuria penetrans

The root-knot nematode Meloidogyne incognita was controlled more effectively and yields of host plants were greater when Paecilomyces lilacinus and Pasteuria penetrans were applied together in field microplots than when either was applied alone. Yields of winter vetch from microplots inoculated with the nematode and with both organisms were not statistically different from yields from uninoculated control plots.     

Survival of Paecilomyces lilacinus in Selected Carriers and Related Effects on Meloidogyne incognita on Tomato

Laboratory and microplot experiments were conducted to determine the influence of carrier and storage of Paecilomyces lilacinus on its survival and related protection of tomato against Meloidogyne incognita. Spores of P. lilacinus were prepared in five formulations: alginate pellets (pellets), diatomaceous earth granules (granules), wheat grain, soil, and soil plus chitin. Fungal viability was high in wheat and granules, intermediate in pellets, and low in soil and chitin-amended soil stored at 25 ± 2 C. In 1985 P. lilacinus in field microplots resulted in about a 25% increase in tomato yield and 25% gall suppression, compared with nematodes alone. Greatest suppression of egg development occurred in plots treated with P. lilacinus in pellets, wheat grain, and granules. In 1986 carryover protection of tomato against M. incognita resulted in about a threefold increase in tomato fruit yield and 25% suppression of gall development, compared with plants treated with nematodes alone. Higher numbers of fungus-infected egg masses occurred in plots treated with pellets (32%) than in those treated with chitin-amended soil (24%), wheat (16%), granules (12%), or soil (7%). Numbers of fungal colony-forming units per gram of soil in plots treated with pellets were 10-fold greater than initial levels estimated at planting time in 1986.     

Purification and characterization of a serine protease and chitinases from Paecilomyces lilacinus and detection of chitinase activity on 2D gels

The filamentous fungus Paecilomyces lilacinus is currently developed as a biocontrol agent against plant parasitic nematodes. Nematode eggs and cuticles are the infection sites for biocontrol agents that penetrate by the production of lytic enzymes. P. lilacinus was cultured in liquid media and proteases and chitinases were induced by the introduction of egg yolk and chitin, respectively. A serine protease was purified from a culture medium using Sepharose-bacitracin affinity column. The protease occurred in three forms, two of which were C-terminally truncated. Chitinase activity was also observed in the culture supernatant, and after separation by isoelectric focusing six proteins were detected that showed activity. Chitinase activity was further confirmed on non-denaturing one-dimensional (1D) and two-dimensional (2D) gels using a sandwich assay with glycol chitin as a substrate. Two of the proteins had similarities with endochitinases as shown by their N-terminal amino acid sequences.     

Histology of the Interactions of Paecilomyces lilacinus with Meloidogyne incognita on Tomato

Excised tomato roots were examined histologically for interactions of the fungus Paecilomyces lilacinus and Meloidogyne incognita race 1. Root galling and giant-cell formation were absent in tomato roots inoculated with nematode eggs infected with P. lilacinus. Few to no galls and no giant-cell formation were found in roots dipped in a spore suspension of P. lilacinus and inoculated with M. incognita. Numerous large galls and giant cells were present in roots inoculated only with M. incognita. P. lilacinus colonized the surface of epidermal cells as well as the internal cells of epidermis and cortex. The possibility of biological protection of plant surfaces with P. lilacinus against root-knot nematodes is discussed.     

Oxidation and ring cleavage of dibenzofuran by the filamentous fungus <Emphasis Type="Italic">Paecilomyces lilacinus</Emphasis>

The ability of the imperfect soil fungus Paecilomyces lilacinus to transform the environmental pollutant dibenzofuran was investigated. Transformation of dibenzofuran and related derivatives lead to 14 products, which were identified by UV spectroscopy, mass spectrometry, and proton nuclear magnetic resonance spectroscopy. Biotransformation was initiated by two separate hydroxylation steps, leading to the accumulation of 4-monohydroxylated and 4-dihydroxylateddibenzofurans. Hydroxylation at both aromatic rings produced 2,7-dihydroxydibenzofuran, 3,7-dihydroxydibenzofuran, and 2,8-dihydroxydibenzofuran. Further oxidation yields ring cleavage of dibenzofuran, which has not been described before for filamentous fungi. The ring fission products were identified as benzo[b]furo[3,2-d]-2-pyrone-6-carboxylic acid and [2-(1-carboxy-methylidene)-benzofuran-3-ylidene]-hydroxy-acetic acid and its derivatives hydroxylated at carbon 7 and 8 at the non-cleaved ring. Other metabolites were riboside-conjugates of 2-hydroxydibenzofuran and 3-hydroxydibenzofuran. The results showed that P. lilacinus transforms the hydrophobic compound dibenzofuran by phase I/phase II reactions to produce hydroxylated products and excretable sugar conjugates.     

Impact of Paecilomyces lilacinus Inoculum Level and Application Time on Control of Meloidogyne incognita on Tomato

Microplot experiments were conducted to evaluate the effects of inoculum level and time of application of Paecilomyces lilacinus on the protection of tomato against MeIoidogyne incognita. The best protection against M. incognita was attained with 10 and 20 g of fungus-infested wheat kernels per microplot which resulted in a threefold and fourfold increase in tomato yield, respectively, compared with tomato plants treated with this nematode alone. Greatest protection against this pathogen was attained when P. lilacinus was delivered into soil 10 days before planting and again at planting. Yield was increased twofold compared with yield in nematode-alone plots and plots with M. incognita plus the fungus. Percentages of P. lilacinus-infected egg masses were greatest in plots treated at midseason or at midseason plus an early application, compared with plots treated with the fungus 10 days before planting and (or) at planting time.     

Infection of plant-parasitic nematodes by Paecilomyces lilacinus and Monacrosporium lysipagum

Studying the mode of infection of a biocontrol agent is important in order to assess its efficiency. The mode and severity of infection of nematodes by a soil saprophyte Paecilomyces lilacinus (Thom) Samson and a knob-producing nematode trapping fungus Monacrosporium lysipagum (Drechsler) Subram were studied under laboratory conditions using microscopy. Infection of stationary stages of nematodes by P. lilacinus was studied with three plant-parasitic nematodes Meloidogyne javanica (Treub) Chitwood, Heterodera avenae Wollenweber and Radopholus similis (Cobb) Thorne. Paecilomyces lilacinus infected eggs, juveniles and females of M. javanica by direct hyphal penetration. The early developed eggs were more susceptible than the eggs containing fully developed juveniles. As observed by transmission electron microscopy, fungal hypha penetrated the M. javanica female cuticle directly. Paecilomyces lilacinus also infected immature cysts of H. avenae including eggs in the cysts and the eggs of R. similis. Trapping and subsequent killing of mobile stages of nematodes by M. lysipagum were studied with the above three nematodes. In addition, plant-parasitic nematodes Pratylenchus neglectus (Rensch) Chitwood and Oteifa and Ditylenchus dipsaci (Kuhn) Filipjev were tested with M. lysipagum. This fungus was shown to infect mobile stages of all the plant-parasitic nematodes. In general, juveniles except those of P. neglectus, were more susceptible to the attack than adults.     

Nematophagous fungus <Emphasis Type="Italic">Paecilomyces lilacinus</Emphasis> (Thom) Samson is also a biological agent for control of greenhouse insects and mite pests

Pathogenicity of nematophagous fungus Paecilomyces lilacinus (Thom) Samson in control of the most destructive greenhouse pests such as: greenhouse whitefly, Trialeurodes vaporariorum, glasshouse red spider mite, Tetranychus urticae, the cotton aphid, Aphis gossypii and western flower thrips, Frankliniella occidentalis was examined in laboratory and pot experiments. The fungus showed the greatest efficacy in controlling winged and wingless forms of the cotton aphid. The cotton aphid’s population was almost totally eliminated. In controlling the greenhouse whitefly, P. lilacinus was most successful when applied against nymphal growth stages (L₃-L₄). Control of the western flower thrips was most efficient against prepupal and pupal stages when the fungus was applied as a water spore suspension to the soil. When the fungus was applied at temperatures below 10 °C, it was able to reduce a glasshouse red spider mite population by 60%.     

Thiol-dependent serine proteinase from <Emphasis Type="Italic">Paecilomyces lilacinus</Emphasis>: Purification and catalytic properties

An extracellular thiol-dependent serine proteinase was isolated from culture medium filtrate of the microscopic fungus Paecilomyces lilacinus with a yield of 33%. The enzyme is inactivated by specific inhibitors of serine proteinases, phenylmethylsulfonyl fluoride, as well as by chloromercuribenzoate and mercury acetate, but is resistant to chelating agents. The proteinase has broad specificity, hydrolyzes proteins and p-nitroanilides of N-acylated tripeptides, exhibiting maximal activity in hydrolysis of substrates containing long hydrophobic and aromatic residues (norleucine, leucine, phenylalanine) as well as arginine at the P1 position. The enzyme has a molecular weight of 33 kD. The enzyme is most active at pH 10.0-11.5; it is thermostable and is characterized by broad optimum temperature range (30-60 degrees C), displaying about 25% of maximal activity at 0 degrees C. The N-terminal sequence of the enzyme (Gly-Ala-Thr-Thr-Gln-Gly-Ala-Thr-Gly/Ile-Xxx-Gly) has no distinct homology with known primary structures of serine proteinases from fungi and bacilli. Based on its physicochemical and enzymatic properties, the serine proteinase from P. lilacinus can be classified as a thiol-dependent subtilisin-like enzyme.     

Control of plant-parasitic nematodes by Paecilomyces lilacinus and Monacrosporium lysipagum in pot trials

The common soil inhabiting nematophagous fungus Paecilomyces lilacinus (Thom) Samson and the nematode trapping fungus Monacrosporium lysipagum (Drechsler) Subram were assayed for their ability to reduce the populations of three economically important plant-parasitic nematodes in pot trials. The fungi were tested individually and in combination against the root-knot nematode Meloidogyne javanica (Treub) Chitwood, cereal cyst nematode Heterodera avenae Wollenweber, or burrowing nematode Radopholus similis (Cobb) Thorne on tomato, barley and tissue cultured banana plants, respectively. In all cases, nematode populations were controlled substantially by both individual and combined applications of the fungi. Combined application of P. lilacinus and M. lysipagum reduced 62% of galls and 94% of M.␣javanica juveniles on tomato when compared to the experiment with no fungi added. Sixty five percent of H. avenae cysts were reduced on barley by combined application of fungi. Control of R. similis on banana, both in the roots and in the soil, was greatest when M. lysipagum was applied alone (86%) or in combination with P. lilacinus (96%), using a strategy where the fungi were inoculated twice in 18 weeks growth period. Overall, combined application of P. lilacinus and M. lysipagum was the most effective treatment in controlling nematode populations, although in some cases M. lysipagum alone was as effective as the combined application of fungi, particularly against M. javanica.     

Evaluation of Paecilomyces lilacinus as a Biocontrol Agent of Meloidogyne javanica on Tobacco

The efficacy of the nematode parasite Paecilomyces lilacinus, alone and in combination with phenamiphos and ethoprop, for controlling the root-knot nematode Meloidogyne javanica on tobacco and the ability of this fungus to colonize in soil under field conditions were evaluated for 2 years in microplots. Combinations and individual treatments of the fungus grown on autoclaved wheat seed, M. javanica eggs (76,000 per plot), and nematicides were applied to specified microplots at the time of transplanting tobacco the first year. Vetch was planted as a winter cover crop, and the fungus and nematicides were applied again the second year to specified plots at transplanting time. The fungus did not control the nematode in either year of these experiments. The average root-gall index (0 = no visible galls and 5 = > 100 galls per root system) ranged from 2.7 to 3.9 the first year and from 4.3 to 5.0 the second in nematode-infested plots treated with nematicides. Plants with M. javanica alone or in combination with P. lilacinus had galling indices of 5.0 both years; the latter produced lower yields than all other treatments during both years of the study. Nevertheless, the average soil population densities of P. lilacinus remained high, ranging from 1.2 to 1.3 × 106 propagules/g soil 1 week after the initial inoculation and from 1.6 to 2.3 × 104 propagules/g soil at harvest the second year. At harvest the second year the density of fungal propagules was greatest at the depth of inoculation, 15 cm, and rapidly decreased below this level.     

Pathogenicity of hyphomycetous fungi against <Emphasis Type="Italic">Cyclocephala signaticollis</Emphasis>

Susceptibility of the white grub Cyclocephala signaticollis Burmeister (Coleoptera: Scarabaeidae: Dynastinae) larvae to seven isolates of Beauveria bassiana (Balsamo) Vuillemin, five of Metarhizium anisopliae (Metschnikoff) Sorokin and two of Paecilomyces lilacinus (Thom) Samson (Deuteromycotina: Hyphomycetes) was investigated. Among 14 fungal isolates screened the most virulent was a B. bassiana isolate (Bb 53) that caused 70% mortality of third instar larvae in 40 days after inoculation at 1 × 10⁸ conida/ml. Strains of M. anisopliae and P. lilacinus showed low efficacy or no virulence to the target host.