Transcriptomic analysis of entomopathogenic fungus Beauveria bassiana infected by a hypervirulent polymycovirus BbPmV-4.

Polymycoviridae is a recently established family of mycoviruses. Beauveria bassiana polymycovirus 4 (BbPmV-4) was previously reported. However, the effect of the virus on host fungus B. bassiana was not clarified. Here, a comparison between virus-free and virus-infected isogenic lines of B. bassiana revealed that BbPmV-4 infection of B. bassiana changes morphology and could lead to decreases in conidiation and increases in virulence against Ostrinia furnacalis larvae. The differential expression of genes between virus-free and virus-infected strains was compared by RNA-Seq and was consistent with the phenotype of B. bassiana. The enhanced pathogenicity may be related to the significant up-regulation of genes encoding mitogen activated protein kinase, cytochrome P450, and polyketide synthase. The results enable studies of the mechanism of interaction between BbPmV-4 and B. bassiana.

Effect of ultraviolet radiation on Beauveria bassiana virulence and development of protective formulations.

Locusta migratoria is a serious agricultural pest in China. Beauveria bassiana is one of the most important pathogens of grasshoppers and locusts. The effects of ultraviolet light were evaluated on the B. bassiana strain BbZJ1. The results showed that 253.7 and 360 nm wavelength UV (Ultra Violet) did not affect the germination of B. bassiana after its recovery from UV treatments. Nevertheless, the virulence of B. bassiana BbZJ1 after its recovery from radiation of UV (253.7 nm) increased. The mortality rates were 85.00% for the BbZJ1 control, was 96.67% for BbZJ1 recovered from radiation of UV (253.7 nm) for 60 min. After treatment with 253.7 nm UV radiation for 60 min, the expression levels of stress-resistant genes BbAlg9 and Bbadh2 in BbZJ1 strain were 2.68 and 2.29 times higher than those in the control group, respectively. Meanwhile, the B. bassiana prepared in 5% groundnut oil showed highest tolerance levels to the ultraviolet radiation. The 5% groundnut oil was the most suitable potential UV-protectant for B. bassiana in terms of cost and availability.

[Effect of VvLaeA on the growth and development of Beauveria bassiana].

It is unclear how VvLaeA functions in regulating the growth and development of Volvariella volvacea (Bull. ex. Fr.) Sing.. Firstly, bioinformatics analysis of VvLaeA was carried out in this study. Subsequently, the Vvgpd promoter and the open reading frame (ORF) fragment of VvlaeA were amplified and fused by polymerase chain reaction (PCR). The fusion fragment was cloned into the pK2 (bar) plasmid. The recombinant construct pK2(bar)-OEVvlaeA was transfected into Beauveria bassiana by Agrobacterium tumefaciens-mediated transformation. Finally, the growth and development of the transformants were examined. The results showed that VvLaeA shared a low homology with similar proteins in other fungi. Compared with the wild type, the colony diameter of the transformant was significantly increased. However, the pigment deposition, conidial yields and germination rates were significantly decreased. The overexpression strains were more sensitive to stresses than that of the wild type. Further studies showed the conidial cell wall properties of the transformants were altered, and the expressions of genes related to the conidial development were significantly down-regulated. Collectively, VvLaeA increased the growth rate of B. bassiana strains and negatively regulated the pigmentation and conidial development, which shed a light for the functional identification of straw mushroom genes.

Pathogenesis and defense mechanism while Beauveria bassiana JEF-410 infects poultry red mite, Dermanyssus gallinae.

The poultry red mite, Dermanyssus gallinae (Mesostigmata: Dermanyssidae), is a major pest that causes great damage to chicken egg production. In one of our previous studies, the management of red mites using entomopathogenic fungi was evaluated, and the acaricidal fungus Beauveria bassiana JEF-410 was selected for further research. In this study, we tried to elucidate the pathogenesis of B. bassiana JEF-410 and the defense mechanisms of red mites at a transcriptome level. Red mites collected from a chicken farm were treated with B. bassiana JEF-410. When the mortality of infected red mites reached 50%, transcriptome analyses were performed to determine the interaction between B. bassiana JEF-410 and red mites. Uninfected red mites and non-infecting fungus served as controls. In B. bassiana JEF-410, up-regulated gene expression was observed in tryptophan metabolism and secondary metabolite biosynthesis pathways. Genes related to acetyl-CoA synthesis were up-regulated in tryptophan metabolism, suggesting that energy metabolism and stress management were strongly activated. Secondary metabolites associated with fungal up-regulated DEGs were related to the production of substances toxic to insects such as beauvericin and beauveriolide, efflux pump of metabolites, energy production, and resistance to stress. In red mites, physical and immune responses that strengthen the cuticle against fungal infection were highly up-regulated. From these gene expression analyses, we identified essential factors for fungal infection and subsequent defenses of red mites. These results will serve as a strong platform for explaining the interaction between B. bassiana JEF-410 and red mites in the stage of active infection.

Gut bacterial microbiota of Lymantria dispar asiatica and its involvement in Beauveria bassiana infection.

The gut bacterial microbiota of insects has been shown to play essential roles in processes related to physiology, metabolism, and innate immunity. In this study, we firstly performed a broad analysis of the gut bacteria in Lymantria dispar asiatica, one of the most devastating forestry defoliators. We analyzed the bacterial composition among different individuals from lab-reared or wild-collected using 16 s rRNA-sequencing, revealing that the gut bacteria of wild-collected larvae were highly diverse, while lab-reared larvae were only associated with a few genera. We found Lactobacillus sp. present in all the gut samples, which indicates that it is part of the core microbiome in the caterpillar. Further Beauveria bassiana infection-based assays showed that the mortality of non-axenic L. dispar asiatica larvae was significantly higher than that of axenic larvae at 72 h. Moreover, we isolated several bacteria from the hemolymph of the non-axenic larvae infected by B. bassiana, which may be caused by the translocation of gut bacteria from the gut to the hemocoel. Reintroduction of Enterococcus sp., Pseudomonas sp., Enterobacter sp., and Microbacterium sp. into axenic larvae recurred the larval mortality in their non-axenic counterpart. Taken together, our study demonstrates that the gut bacteria of L. dispar asiatica are highly volatile, and different bacteria taxa can promote host infection by entomopathogenic fungus, providing a new strategy for the pest management.

Symbiotic Bacteria System of Locusta migratoria Showed Antifungal Capabilities against Beauveria bassiana.

The stability of symbiotic flora is an important indicator of the health of an organism. Symbiotic bacteria have been proven to be closely involved in the immune process of organisms. The pathogenicity of Beauveria bassiana was studied in relation to symbiotic bacteria on the surface and inside of the migratory locust (Locusta migratoria). The results showed that the surface disinfection of test locusts contributed to the pathogenicity of B. bassiana to locusts. Most of the surface bacteria of L. migratoria caused some inhibition of B. bassiana growth, and LM5-4 (Raoultella ornithinolytica), LM5-2 (Enterobacter aerogenes), and LM5-13 (Citrobacter freundii) showed the highest inhibitory effect on the growth of B. bassiana. The inoculation of locusts with additional surface symbiotic bacteria reduced the virulence of B. bassiana to L. migratoria. Infection by different strains of B. bassiana caused similar changes in the symbiotic flora of migratory locusts. The inoculation of locusts with additional intestinal symbiotic bacteria (Enterobacter sp.) reduced the virulence of B. bassiana to L. migratoria. These findings illustrate the effect of bacterial communities on fungal infections in L. migratoria when seen from the perspective of ecology in a microenvironment. The active antifungal substances of such bacteria and their mechanisms of action need further study.

Diversity of entomopathogenic fungi from soils of eucalyptus and soybean crops and natural forest areas.

Soils present high fungal diversity, including entomopathogenic species. These fungi are used in pest control, providing easy production, multiplication, application, and dispersion in the field. The objective of the present study was to evaluate entomopathogenic fungal diversity in soils from eucalyptus and soybean crops and natural forest areas. These fungi were isolated using the "Bait Method" with Tenebrio molitor (Linnaeus, 1758) (Coleoptera: Tenebrionidae) larvae from 10 soil samples per area, collected at 10 cm deep in a zig-zag pattern. The isolated entomopathogenic fungi were cultivated in Petri dishes using PDA medium and their mycelia separated after seven days of incubation in a BOD-type chamber. Species of Aspergillus, Beauveria, Cordyceps, Fusarium, Metarhizium, Penicillium and Purpureocillium were identified. The "Bait Method" with T. molitor larvae is efficient to isolate entomopathogenic fungi with higher diversity from soils of the natural forest than the cultivated area.

Infections by entomopathogenic fungi in common green iguanas (Iguana iguana) in captivity in Brazil.

Entomopathogenic fungi, widely available biological agents used to control agricultural pests, are sporadically reported to cause focal or disseminated infection in reptiles and mammals, including humans. This study summarizes the clinical presentation, histopathological and molecular findings by panfungal polymerase chain reaction and sequencing of four cases of hypocrealean fungal infections in captive common green iguanas (Iguana, iguana). One case of granulomatous pneumonia, hepatitis and serositis was related to Metarhizium flavoviride complex infection. Two disseminated fungal infection cases, with scarce inflammatory cell infiltration, were caused by Beauveria bassiana while there was one case of multifocal granulomatous and necrotizing pneumonia by Purpureocillium spp. To the best of our knowledge, this is the first report of fatal mycosis infection due to entomopathogenic fungi in captive common green iguanas.

Beauveria bassiana interacts with gut and hemocytes to manipulate Aedes aegypti immunity.

BACKGROUND: Mosquito-borne diseases affect millions of people. Chemical insecticides are currently employed against mosquitoes. However, many cases of insecticide resistance have been reported. Entomopathogenic fungi (EPF) have demonstrated potential as a bioinsecticide. Here, we assessed the invasion of the EPF Beauveria bassiana into Aedes aegypti larvae and changes in the activity of phenoloxidase (PO) as a proxy for the general activation of the insect innate immune system. In addition, other cellular and humoral responses were evaluated. METHODS: Larvae were exposed to blastospores or conidia of B. bassiana CG 206. After 24 and 48 h, scanning electron microscopy (SEM) was conducted on the larvae. The hemolymph was collected to determine changes in total hemocyte concentration (THC), the dynamics of hemocytes, and to observe hemocyte-fungus interactions. In addition, the larvae were macerated to assess the activity of PO using L-DOPA conversion, and the expression of antimicrobial peptides (AMPs) was measured using quantitative Real-Time PCR. RESULTS: Propagules invaded mosquitoes through the midgut, and blastopores were detected inside the hemocoel. Both propagules decreased the THC regardless of the time. By 24 h after exposure to conidia the percentage of granulocytes and oenocytoids increased while the prohemocytes decreased. By 48 h, the oenocytoid percentage increased significantly (P < 0.05) in larvae exposed to blastospores; however, the other hemocyte types did not change significantly. Regardless of the time, SEM revealed hemocytes adhering to, and nodulating, blastospores. For the larvae exposed to conidia, these interactions were observed only at 48 h. Irrespective of the propagule, the PO activity increased only at 48 h. At 24 h, cathepsin B was upregulated by infection with conidia, whereas both propagules resulted in a downregulation of cecropin and defensin A. At 48 h, blastospores and conidia increased the expression of defensin A suggesting this may be an essential AMP against EPF. CONCLUSION: By 24 h, B. bassiana CG 206 occluded the midgut, reduced THC, did not stimulate PO activity, and downregulated AMP expression in larvae, all of which allowed the fungus to impair the larvae to facilitate infection. Our data reports a complex interplay between Ae. aegypti larvae and B. bassiana CG 206 demonstrating how this fungus can infect, affect, and kill Ae. aegypti larvae.

The Potential of a New Beauveria bassiana Isolate for Mosquito Larval Control.

The African malaria mosquito, Anopheles gambiae Giles (Diptera: Culicidae), and the Asian tiger mosquito, Aedes albopictus Skuse (Diptera: Culicidae) are of public health concern due to their ability to transmit disease-causing parasites and pathogens. Current mosquito control strategies to prevent vector-borne diseases rely mainly on the use of chemicals. However, insecticide resistance in mosquito populations necessitates alternative control measures, including biologicals such as entomopathogenic fungi. Here we report the impact of a new Beauveria bassiana (Balsamo) Vuillemin (Hyprocreales: Cordycipitaeceae) isolate, isolated from field-collected Ae. albopictus larvae on mosquito survival and development. Larval infection bioassays using three B. bassiana conidial concentrations were performed on the second and third larval instars of An. gambiae and Ae. albopictus mosquitoes. Larvae were monitored daily for survival and development to pupae and adults. Our results show that B. bassiana MHK was more effective in killing An. gambiae than Ae. albopictus larvae. We further observed delays in development to pupae and adults in both mosquito species exposed the varying concentrations of B. bassiana as compared to the water control. In addition, larval exposure to B. bassiana reduced adult male and female survival in both mosquito species, further contributing to mosquito population control. Thus, this study identifies a new B. bassiana isolate as a possible biological control agent of two mosquito species of public health concern, increasing the arsenal for integrated mosquito control.

Beauveria australis finds a new host in French dung beetles introduced to Australia.

Exotic dung beetles have been introduced to Australia for over 50 years to mitigate issues caused by dung produced by livestock. This study aims at identifying fungi affecting a beetle colony and investigating their source. Fungal hyphae emerging from the cuticle of dead beetles were cultured and a multigene phylogeny showed that Beauveria bassiana and B. australis both infected these introduced beetles, likely acquired through local soil or dung. This is the first record of B. australis infecting family Scarabaeidae. This study highlights the importance of fungal outbreak management in insect rearing before release, and challenges associated with new pathogens.

Genetic variability of Metarhizium isolates from the Ticino Valley Natural Park (Northern Italy) as a possible microbiological resource for the management of Popillia japonica.

The natural occurrence of entomopathogenic fungi (EPF) was investigated along the Ticino River (Ticino River Natural Park, Novara Province, Piedmont, Italy), at the center of the area of the first settlement of the invasive alien pest Popillia japonica. Using Zimmermann's "Galleria bait method", EPF were successfully isolated from 83 out of 155 soil samples from different habitats (perennial, cultivated, or uncultivated meadows, woodlands, and riverbanks). Sequencing of the 5' end of the Translation Elongation Factor 1 alfa (5'-TEF) region allowed the assignment of 94% of the isolates to Metarhizium spp., while 8% and 7% were assigned to Beauveria spp. and Paecilomyces spp., respectively. Four Metarhizium species were identified: Metarhizium robertsii was the most common one (61.5% of the isolates), followed by M. brunneum (24.4%), M. lepidiotae (9%), and M. guizhouense (5.1%). Microsatellite marker analysis of the Metarhizium isolates revealed the presence of 27 different genotypes, i.e., 10 genotypes among M. robertsii, 8 among M. brunneum, 5 among M. lepidiotae, and 4 among M. guizhouense. Metarhizium brunneum appeared to be associated with woodlands and more acid soils, while the other species showed no clear association with a particular habitat. Laboratory virulence tests against P. japonica 3rd instar larvae allowed the identification of one M. robertsii isolate that showed efficacy as high as 80.3%. The importance of this kind of study in the frame of eco-friendly microbiological control is discussed.

The AreA Nitrogen Catabolite Repression Activator Balances Fungal Nutrient Utilization and Virulence in the Insect Fungal Pathogen Beauveria bassiana.

In many fungi, the AreA GATA-type transcription factor mediates nitrogen catabolite repression affecting fungal development and, where applicable, virulence. Here, we investigated the functions of AreA in the fungal entomopathogen and plant endophyte Beauveria bassiana using knockdown of gene expression. The antiAreA mutants were impaired in nitrogen utilization and showed increased sensitivities to osmotic stressors but increased tolerances to oxidative/hypoxia stresses. Repression of BbAreA caused overall minimal effects on fungal virulence. The minor effects on virulence appeared to be due in part to competing secondary effects where host defense phenoloxidase activity was significantly decreased, but production of the fungal metabolite oosporein was increased and hyphal body development was impaired. Knockdown of BbAreA expression also resulted in impairment in ability of the fungus to associate with host plants. These data implicate that BbAreA likely acts as a regulator to balance fungal nutrient utilization, pathogenicity, and mutualism, facilitating the fungal occupation of host niches.

Accumulation of pyrethroids induces changes in metabolism of the entomopathogenic fungus Beauveria bassiana-Proteomic and lipidomic background.

Advances in the agrochemical industry, such as using plant protection products e.g. pyrethroid insecticides, lead to environmental pollution via the accumulation of toxic compounds in soil. An interesting approach to overcoming this threat is using biopreparations based on entomopathogenic fungi that come into contact with the residues of the insecticides in the environment. The aim of this study was to determine whether the soil-dwelling entomopathogenic fungus Beauveria bassiana ARSEF 2860 is capable of accumulating pyrethroids (λ-cyhalothrin, α-cypermethrin and deltamethrin) and to identify the metabolomics and proteomic implications of this process. In this work, we demonstrated for the first time that the tested fungus accumulated pyrethroids as early as on day 2 of incubation with an average efficiency of 90%. Pyrethroids accumulated in large quantities in the mycelium of B. bassiana induced oxidative stress and interacted differently with the enzymes of the basic metabolic pathways, enzymes associated with the organization of the actin cytoskeleton and cell walls, as well as extracellular enzymes responsible for the infectious abilities (α-cypermethrin caused a 61% decrease in PR1, λ-cyhalothrin - a 31% decrease in PR2, which are proteolytic enzymes with a confirmed role in the infectious process). This study also revealed that the accumulated pyrethroids decreased the activity of phospholipase C, which increased the triacylglycerols/diacylglycerols (TAG/DAG) ratio, especially in mycelium in which α-cypermethrin was accumulated. It should be emphasized that the accumulation of pyrethroids in the environment is not fully understood, and current research suggests that entomopathogenic fungi may be part of the process.

OTU7B Modulates the Mosquito Immune Response to Beauveria bassiana Infection via Deubiquitination of the Toll Adaptor TRAF4.

The Aedes aegypti mosquito transmits devastating flaviviruses, such as Zika, dengue, and yellow fever viruses. For more effective control of the vector, the pathogenicity of Beauveria bassiana, a fungus commonly used for biological control of pest insects, may be enhanced based on in-depth knowledge of molecular interactions between the pathogen and its host. Here, we identified a mechanism employed by B. bassiana, which efficiently blocks the Ae. aegypti antifungal immune response by a protease that contains an ovarian tumor (OTU) domain. RNA-sequencing analysis showed that the depletion of OTU7B significantly upregulates the mRNA level of immunity-related genes after a challenge of the fungus. CRISPR-Cas9 knockout of OTU7B conferred a higher resistance of mosquitoes to the fungus B. bassiana. OTU7B suppressed activation of the immune response by preventing nuclear translocation of the NF-κB transcription factor Rel1, a mosquito orthologue of Drosophila Dorsal. Further studies identified tumor necrosis factor receptor-associated factor 4 (TRAF4) as an interacting protein of OTU7B. TRAF4-deficient mosquitoes were more sensitive to fungal infection, indicating TRAF4 to be the adaptor protein that activates the Toll pathway. TRAF4 is K63-link polyubiquitinated at K338 residue upon immune challenge. However, OTU7B inhibited the immune signaling by enzymatically removing the polyubiquitin chains of mosquito TRAF4. Thus, this study has uncovered a novel mechanism of fungal action against the host innate immunity, providing a platform for further improvement of fungal pathogen effectiveness. IMPORTANCE Insects use innate immunity to defend against microbial infection. The Toll pathway is a major immune signaling pathway that is associated with the antifungal immune response in mosquitoes. Our study identified a fungal-induced deubiquitinase, OTU7B, which, when knocked out, promotes the translocation of the NF-κB factor Rel1 into the nucleus and confers enhanced resistance to fungal infection. We further found the counterpart of OTU7B, TRAF4, which is a component of the Toll pathway and acts as an adaptor protein. OTU7B enzymatically removes K63-linked polyubiquitin chains from TRAF4. The immune response is suppressed, and mosquitoes become much more sensitive to the Beauveria bassiana infection. Our findings reveal a novel mechanism of fungal action against the host innate immunity.

You are what you eat: fungal metabolites and host plant affect the susceptibility of diamondback moth to entomopathogenic fungi.

Background: Beauveria are entomopathogenic fungi of a broad range of arthropod pests. Many strains of Beauveria have been developed and marketed as biopesticides. Beauveria species are well-suited as the active ingredient within biopesticides because of their ease of mass production, ability to kill a wide range of pest species, consistency in different conditions, and safety with respect to human health. However, the efficacy of these biopesticides can be variable under field conditions. Two under-researched areas, which may limit the deployment of Beauveria-based biopesticides, are the type and amount of insecticidal compounds produced by these fungi and the influence of diet on the susceptibility of specific insect pests to these entomopathogens. Methods: To understand and remedy this weakness, we investigated the effect of insect diet and Beauveria-derived toxins on the susceptibility of diamondback moth larvae to Beauveria infection. Two New Zealand-derived fungal isolates, B. pseudobassiana I12 Damo and B. bassiana CTL20, previously identified with high virulence towards diamondback moth larvae, were selected for this study. Larvae of diamondback moth were fed on four different plant diets, based on different types of Brassicaceae, namely broccoli, cabbage, cauliflower, and radish, before their susceptibility to the two isolates of Beauveria was assessed. A second experiment assessed secondary metabolites produced from three genetically diverse isolates of Beauveria for their virulence towards diamondback moth larvae. Results: Diamondback moth larvae fed on broccoli were more susceptible to infection by B. pseudobassiana while larvae fed on radish were more susceptible to infection by B. bassiana. Furthermore, the supernatant from an isolate of B. pseudobassiana resulted in 55% and 65% mortality for half and full-strength culture filtrates, respectively, while the filtrates from two other Beauveria isolates, including a B. bassiana isolate, killed less than 50% of larvae. This study demonstrated different levels of susceptibility of the insects raised on different plant diets and the potential use of metabolites produced by Beauveria isolates in addition to their conidia.

"Metabolomic diversity of local strains of Beauveria bassiana (Balsamo) Vuillemin and their efficacy against the cassava mite, Tetranychus truncatus Ehara (Acari: Tetranychidae)".

A desirable substitute for chemical pesticides is mycopesticides. In the current investigation, rDNA-ITS (Internal transcribed spacer) and TEF (Transcriptional Elongation Factor) sequencing were used for molecular identification of six Beauveria bassiana strains. Both, leaf discs and potted plant bioassaye were carried out to study their pathogenicity against the cassava mite, Tetranychus truncatus. LC50 and LC90 values of potential B. bassiana strains were estimated. We also discovered a correlation between intraspecific B. bassiana strains pathogenicity and comprehensive metabolome profiles. Bb5, Bb6, Bb8, Bb12, Bb15, and Bb21 strains were identified as B. bassiana by sequencing of rDNA-ITS and TEF segments and sequence comparison to NCBI (National Center for Biotechnology Information) GenBank. Out of the six strains tested for pathogenicity, Bb6, Bb12, and Bb15 strains outperformed against T. truncatus with LC50 values 1.4×106, 1.7×106, and 1.4×106 and with a LC90 values 7.3×107, 1.4×108, and 4.2×108 conidia/ml, respectively, at 3 days after inoculation and were considered as potential strains for effective mite control. Later, Gas Chromatography-Mass Spectrometry (GC-MS) analysis of the above six B. bassiana strains was done on secondary metabolites extracted with ethyl acetate revealed that the potential B. bassiana strains (Bb6, Bb12, and Bb15) have higher levels of acaricidal such as Bis(dimethylethyl)-phenol: Bb6 (5.79%), Bb12 (6.15%), and Bb15 (4.69%). Besides, insecticidal (n-Hexadecanoic acid), and insect innate immunity overcoming compound (Nonadecene) were also identified; therefore, the synergistic effect of these compounds might lead toa higher pathogenicity of B. bassiana against T. truncatus. Further, these compounds also exhibited two clusters, which separate the potential and non-potential strains in the dendrogram of Thin Layer Chromatography. These results clearly demonstrated the potentiality of the B. bassiana strains against T. truncatus due to the occurrence of their bioactive volatile metabolome.

FIRST EVIDENCE OF THE TICKICIDE EFFECT OF ENTOMOPATHOGENIC FUNGI ISOLATED FROM MEXICAN CATTLE FARMS AGAINST AMBLYOMMA MIXTUM.

This is the first study to empirically evaluate the mortality of Amblyomma mixtum larvae caused by native entomopathogenic fungi (EPF) from cattle farm soils and the impact of the physicochemical conditions of those soils on the ability of EPF to kill ticks. The efficacy of 25 EPF strains isolated from cattle farms soils in Veracruz, Mexico, against A. mixtum ticks was evaluated using a larval immersion test. Physicochemical analyses of the soils where the EPF were isolated were carried out, and the results were correlated with the obtained mortality. The MaV25 strain showed a mortality of 36.55% (P < 0.05), followed by MaV57 with 27.30%, MaV08 with 26.21%, and MaV31 with 24.32% (P < 0.05). Nitrogen and potassium are nutritional components of soils that seem to be associated with mortality caused by the evaluated fungal strains. This is the first study in Mexico where the effect of EPF against A. mixtum is evaluated and also is the first study in the world that uses native strains from livestock soils against larvae of this tick. It is necessary to determine the virulence characteristics of EPF on A. mixtum to improve knowledge of fungus-tick interactions (Metarhizium anisopliae-A. mixtum).

PULMONARY AND COELOMIC MYCOSES DUE TO METARHIZIUM AND BEAUVERIA SPECIES IN REPTILES.

This report documents cases of fatal pulmonary mycosis caused by entomopathogenic fungi in the genera Metarhizium and Beauveria (Order Hypocreales) in a loggerhead sea turtle (Caretta caretta), a Chinese alligator (Alligator sinensis), two gopher tortoises (Gopherus polyphemus), a Cuvier's dwarf caiman (Paleosuchus palpebrosus), a false gharial (Tomistoma schlegelii), a green sea turtle (Chelonia mydas), and a Kemp's ridley sea turtle (Lepidochelys kempii), and a case of granulomatous coelomitis in a hawksbill sea turtle (Eretmochelys imbricata). Fungi identified in these cases included Beauveria bassiana, Beauveria brongniartii, Metarhizium anisopliae, Metarhizium robertsii, and one case of infection by a novel Metarhizium species. The animals were either housed at zoos or brought into rehabilitation from the wild. Although the majority of animals had comorbidities, the fungal infections were believed to be the primary cause of death. Fungal susceptibility testing was performed on two Beauveria spp. isolates, and revealed lower minimum inhibitory concentrations for itraconazole and voriconazole when compared to terbinafine and fluconazole. This case series demonstrates that a variety of reptile species from different orders are vulnerable to infection with Metarhizium, and multiple species of sea turtle are susceptible to infection with Beauveria.

Distribution and pathogenicity of Beauveria bassiana in soil with earthworm action and feeding.

Earthworm action and feeding have an important impact on a variety of microorganisms in the soil. However, the effects of the earthworm on Beauveria bassiana, a common entomopathogenic fungus in the biological control of pests, have been little studied. In this study, the epigeic earthworm species Eisenia fetida (Savigny) was selected to evaluate its impact on B. bassiana TST05 including its distribution in soil and its pathogenicity to target insects. By testing B. bassiana TST05 distribution, biomass in soil, viable spore germination rate, and pathogenicity to insect larvae after passing through the earthworm gut, the results showed that the activity and feeding of E. fetida promoted the B. bassiana TST05 diffusing downwards in the soil, while decreasing active fungal spores. After passing through the earthworm gut and excretion, the living B. bassiana individuals still had activity and pathogenicity to insects. The germination rate of the viable fungal spores was 15.09% and the infection rate to the insect larvae of Atrijuglans hetaohei Yang reached 62.35%, 80.95% and 100% after infection at 7 d, 10 d, and 14 d, respectively. The results showed that action and feeding of earthworms promoted the distribution of B. bassiana TST05 in soil, but decreased B. bassiana viable spores. This study is important for understanding the interaction between earthworms and B. bassiana in soil and for guiding the scientific application of B. bassiana in the biological control of pests.