Identification of the pigment and its role in UV resistance in Paecilomyces variotii, a Chernobyl isolate, using genetic manipulation strategies.

Fungi produce secondary metabolites that are not directly involved in their growth, but often contribute to their adaptation to extreme environmental stimuli and enable their survival. Conidial pigment or melanin is one of the secondary metabolites produced naturally by a polyketide synthesis (PKS) gene cluster in several filamentous fungi and is known to protect these fungi from extreme radiation conditions. Several pigmented or melanized fungi have been shown to grow under extreme radiation conditions at the Chernobyl nuclear accident site. Some of these fungi, including Paecilomyces variotii, were observed to grow towards the source of radiation. Therefore, in this study, we wanted to identify if the pigment produced by P. variotii, contributes to providing protection against radiation condition. We first identified the PKS gene responsible for synthesis of pigment in P. variotii and confirmed its role in providing protection against UV irradiation through CRISPR-Cas9 mediated gene deletion. This is the first report that describes the use of CRISPR methodology to create gene deletions in P. variotii. Further, we showed that the pigment produced by this fungus, was not inhibited by DHN-melanin pathway inhibitors, indicating that the fungus does not produce melanin. We then identified the pigment synthesized by the PKS gene of P. variotii, as a naptho-pyrone Ywa1, by heterologously expressing the gene in Aspergillus nidulans. The results obtained will further aid in understanding the mechanistic basis of radiation resistance.

Evaluation of antileishmanial activity of harzialactone a isolated from the marine-derived fungus Paecilomyces sp.

Fractionation of extracts from the culture broth of the marine-derived fungus, Paecilomyces sp. 7A22, resulted in the isolation of the harzialactone A (HA), a known compound previously isolated from fungi of marine environments. The chemical structure of HA was determined by spectroscopic analyses. Upon evaluation of HA on antileishmanial assays against Leishmania amazonensis, HA exhibited significant activity against promastigotes forms with IC50 of 5.25 µg mL-1 and moderate activity against intracellular amastigotes with IC50 of 18.18 µg mL-1. This is the first report on the antileishmanial activity of HA, and the effects of HA presented in this work suggest that this class of compounds are suitable for future biological in vitro and in vivo studies for the search of natural products with activity against Leishmania spp. Furthermore, the present results corroborate marine-derived fungi as a promising source of natural products with antiparasitic activity.

Posaconazole in the treatment of refractory Purpureocillium lilacinum (former Paecilomyces lilacinus) keratitis: the salvation when nothing works.

We report a case of a 41-year-old woman, wearer of contact lenses who was presented to the emergency room with a 2-month history of pain and red eye. She presented with a severe keratitis refractory to quinolones, fortified antibiotics and clotrimazole. Due to the risk of perforation, a tectonic penetrating keratoplasty (PK) was performed. Clinical signs of keratitis recurrence were observed and cultures were positive for Purpureocillium lilacinum (former Paecilomyces lilacinus) The patient did not improve on topical amphotericin B and intracameral voriconazole. Worsening of clinical condition required a new PK. Oral posaconazole was initiated postoperatively and suspended at the fourth postoperative month. The cornea remains clear until the last follow-up visit, 12 months after the second graft. To our knowledge, this is the second case report that documents the effectiveness of oral posaconazole in a refractory P. lilacinus keratitis, resistant to other second-generation triazoles and conventional antifungals.

Recalcitrant Paecilomyces keratitis.

A 54-year-old Caucasian woman presented with corneal ulcer of the right eye of 4 weeks duration after scratching her cornea while removing her contact lens and artificial eye lashes. Her visual acuity was 20/32 (left eye) and finger counting (right eye). She had a 3x3 mm epithelial defect with underlying corneal oedema and hypopyon. Right eye cultures grew Paecilomyces species. Topical and systemic antifungal agents were initiated. Due to the sight-threatening disease, the patient underwent surgical intervention with intrastromal injection of amphotericin B and a large conjunctival flap covering 75% of the right eye corneal ulcer. After 3 months of therapy, she had near-complete resolution of the corneal ulcer. Unfortunately, recurrence of the corneal ulcer occurred within 3 weeks of cessation of therapy, prompting reinitiation of ophthalmic and systemic antifungal agents. The patient was advised to continue therapy for 6 months with regular follow-up.

A case of recurrent keratitis caused by Paecilomyces lilacinus and treated by voriconazole / Uso do voriconazol na ceratite recorrente causada por fungo Paecilomyces lilacinus

ABSTRACT - We describe here a case of a 21-year-old woman who presented with low visual acuity, pain, and hyperemia in the left eye for 45 days. Her eye had extensive corneal infiltrate, with melting and a central perforation that was glued with cyanoacrylate, but with Seidel (+). She underwent tectonic corneal transplantation, and anterior chamber lavage with subconjunctival infiltration with voriconazole, as well as intracameral injections of amphotericin B. Laboratory tests revealed Paecilomyces lilacinus as the infectious agent. The patient was then maintained with oral voriconazole and eye drops for three months, after which the infection was considered cured. However, in the sixth postoperative month she presented with endothelial rejection, and two weeks later signs of recurrence of the fungal infection. She was treated with two further washes of the anterior chamber and subconjunctival injection of voriconazole, followed by intravenous voriconazole that was replaced with drops after ten days. The infection initially worsened, but then regressed, and at last follow-up, the patient was still infection-free.

RESUMO - Descrevemos aqui um caso de uma mulher de 21 anos que apresentou baixa acuidade visual, dor e hiperemia no olho esquerdo por 45 dias. O olho apresentava infiltrado corneano extenso, com fusão e perfuração central colada com cianoacrilato, mas com Seidel (+). Ela foi submetida a transplante de córnea tectônica e lavagem de câmara anterior com infiltração subconjuntival com voriconazol, além de injeções intracamerais de anfoterecina B. Testes laboratoriais revelaram Paecilomyces lilacinus como agente infeccioso. A paciente foi então mantida com voriconazol oral e colírio por período de três meses, após o qual a infecção foi considerada curada. No entanto, no sexto mês de pós-operatório, ela apresentou rejeição endotelial e, duas semanas após, sinais de recidiva de infecção fúngica. Ela foi tratada com mais duas lavagens de câmara anterior e injeção subconjuntival de voriconazol, seguida por voriconazol intravenoso que foi substituído por gotas após 10 dias. A infecção piorou inicialmente, mas depois regrediu e, no último seguimento, o paciente ainda estava livre de infecção.

A case of recurrent keratitis caused by Paecilomyces lilacinus and treated by voriconazole.

We describe here a case of a 21-year-old woman who presented with low visual acuity, pain, and hyperemia in the left eye for 45 days. Her eye had extensive corneal infiltrate, with melting and a central perforation that was glued with cyanoacrylate, but with Seidel (+). She underwent tectonic corneal transplantation, and anterior chamber lavage with subconjunctival infiltration with voriconazole, as well as intracameral injections of amphotericin B. Laboratory tests revealed Paecilomyces lilacinus as the infectious agent. The patient was then maintained with oral voriconazole and eye drops for three months, after which the infection was considered cured. However, in the sixth postoperative month she presented with endothelial rejection, and two weeks later signs of recurrence of the fungal infection. She was treated with two further washes of the anterior chamber and subconjunctival injection of voriconazole, followed by intravenous voriconazole that was replaced with drops after ten days. The infection initially worsened, but then regressed, and at last follow-up, the patient was still infection-free.

Identification of hypocrealean reptile pathogenic isolates with MALDI-TOF MS.

Biotyper analysis of Nannizziopsis guarroi, a fatal fungal pathogen in lizards, was described recently. Hypocrealean fungal infections in captive reptiles appear with an increasing frequency during the last decade. Therefore, the aim of this study was to proof Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) as diagnostic tool for the identification of reptile pathogenic hypocrealean fungi. Ten fungal isolates obtained from nine reptiles with fungal glossitis, disseminated visceral mycosis, pneumomycosis, and fungal keratitis were analyzed. Phylogeny consisted of fragments of the large subunit of nuclear encoded ribosomal DNA (D1/D2, LSU) and the internal transcribed spacer region 1 of nuclear encoded ribosomal DNA (ITS1) as well as the protein coding gene translation elongation factor 1 alpha (TEF). Results revealed unanimously two Metarhizium granulomatis genotypes in a total of three isolates, various M. viride genotypes (n = 3), two different Purpureocillium lilacinum isolates as well as one isolate of each P. lavendulum and Beauveria bassiana. Purpureocillium lilacinum and B. bassiana are likewise frequently employed as a mycoinsecticide and mycoacaricide in agriculture on a worldwide scale and have occasionally been reported in man, causing fungal keratitis, sclerokeratitis, nosocomial infections in immunosuppressed patients, as well as cavitary pulmonary disease and cutaneous hyalohyphomycosis in immunocompetent patients. According to the results establishment of Biotyper analysis for faster differentiation of reptile-associated fungal pathogens is entirely justified.

Potential of endophytic fungi as biocontrol agents of Duponchelia fovealis (Zeller) (Lepidoptera: Crambidae) / Fungos endofíticos para o biocontrole de Duponchelia fovealis (Zeller) (Lepidoptera: Crambidae)

Abstract This study reports the first assessment of endophytic fungi isolated from strawberry leaves and selection of isolates for the control of Duponchelia fovealis, a new pest of strawberries. A total of 400 strawberry leaves of the cultivar 'Albion' were collected in four commercial farms. Leaves were disinfected, cut in fragments, and placed on Petri dishes containing potato dextrose agar media with tetracycline and incubated for 30 days. Following this time, 517 fungal colonies were isolated, and thirteen genera were identified: Cladosporium, Aspergillus, Nigrospora, Fusarium, Trichoderma, Chaetomium, Alternaria, Paecilomyces, Penicillium, Ulocladium, Bipolaris, Diaporthe, and Phoma. Eight isolates belonging to the genera Aspergillus, Diaporthe, Paecilomyces, and Cladosporium were selected for pathogenicity bioassays against third instar larvae of D. fovealis. Isolates of Paecilomyces induced the highest mortality rates.

Resumo Este trabalho apresenta a primeira contribuição no isolamento de fungos endofíticos de folhas de morangueiro e na seleção de isolados para controle de Duponchelia fovealis, uma nova praga do morangueiro. Foram coletadas 400 folhas da cultivar 'Albion' em quatro lavouras comerciais de morangueiro. As folhas foram desinfetadas, cortadas em fragmentos e depositadas em placas de Petri contendo Ágar Batata Dextrose como meio, em conjunto com tetraciclina e incubados durante 30 dias. Um total de 517 colônias fúngicas e treze gêneros foram isolados: Cladosporium, Aspergillus, Nigrospora, Fusarium, Trichoderma, Chaetomium, Alternaria, Paecilomyces, Penicillium, Ulocladium, Bipolaris, Diaporthe e Phoma. Oito isolados pertencentes aos gêneros Aspergillus, Diaporthe, Paecilomyces e Cladosporium foram selecionados para os bioensaios de virulência contra larvas de 3º instar de D. fovealis. Isolados de Paecilomyces causaram as maiores taxas de mortalidade.

Keratomycosis due to Purpureocillium lilacinum: A case report from Sub-Himalayan region of Uttarakhand.

The fungus Purpureocillium lilacinum previously known as Paceliomyces lilacinus is an emerging pathogen that can cause severe human infections including devastating oculomycosis. Treatment with traditional antifungals often fails, and the organism shows variable susceptibility to novel triazoles. We hereby report a case of keratomycosis caused by Pur. lilacinum in an immunocompetent male patient following trauma. The patient was successfully treated with voriconazole. The drug shows good activity against Pur. lilacinum and could be a promising therapeutic alternative to treat infections caused by this fungus, which generally shows resistance to conventional antifungal agents including novel triazoles.

Endophytic fungus Paecilomyces formosus LHL10 produces sester-terpenoid YW3548 and cyclic peptide that inhibit urease and α-glucosidase enzyme activities.

Endophytic fungi have been used to obtain novel bioactive secondary metabolites with potential applications in medical and agricultural sectors, which can also act as lead targets for pharmaceutical and medicinal potential. In the present study, the endophytic fungus Paecilomyces formosus LHL10 isolated from the root of cucumber plant was tested for its enzyme inhibitory potential. The ethyl acetate (EtOAc) extract of LHL10 was screened for its inhibitory effect on acetylcholinesterase (AChE), α-glucosidase, urease, and anti-lipid peroxidation. The findings suggest that the EtOAc extract from LHL10 possesses significant inhibitory potential against urease and α-glucosidase. The EtOAc extract was thus, subjected to advanced column chromatographic techniques for the isolation of pure compounds. The structure elucidation was carried out through spectroscopic analysis and comparison with literature data, and these compounds were confirmed as known a sester-terpenoid (1) and a known cyclic peptide (2). The enzyme inhibition bioassay indicated that Compounds 1 and 2 exhibited remarkable inhibitory rate against α-glucosidase and urease, with an IC50 value of 61.80 ± 5.7, 75.68 ± 6.2 and 74.25 ± 4.3, 190.5 ± 10.31 µg/g, respectively. Thus, the current study concludes the enzyme inhibitory potential of endophyte LHL10 and provides the basis for further investigations of bioactive compounds, which could be used as potent drugs for enzyme inhibition.

[Calcitriol induced hypercalcemia in a hunting dog with a disseminated Paecilomyces variotii infection]. / Calcitriol-bedingte Hyperkalzämie bei einem Jagdhund mit disseminierter Paecilomyces variotii-Infektion.

INTRODUCTION: A 5-year old hunting dog was presented with reduced appetite, weight loss and polyuria/polydipsia. Hematology and clinical chemistry revealed anemia, leukocytosis, increased liver enzymes, hypoalbuminemia and hypercalcemia. The cytological, pathohistological and microbiological examination identified a disseminated infection with the saprophytic mould fungus Paecilomyces variotii in the biopsies of the spleen and a lymph node. Determination of vitamin D metabolites confirmed a calcitriol induced hypercalcemia.

Cutaneous ulcerations caused by Paecilomyces variotii in a renal transplant recipient.

Skin infections caused by Paecilomyces species have been rarely described in patients with solid organ transplantation. Cutaneous manifestations are highly variable and include erythematous macules, nodules, pustules, and vesicular and necrotic lesions. The diagnosis of these infections is generally made by examination of a skin biopsy. Management of these fungal infections is difficult due to the immunocompromised state of the patients. Moreover, antifungal therapy and immunosuppressive drug interactions should be considered during treatment management. Herein, we reported a case of cellulitis caused by Paecilomyces variotii in a 56-year-old man who had undergone a kidney transplantation. Erythematous macular and nodular lesions on the left hand and left foot appeared first; within 2 months the skin lesions became ulcerated, hemorrhagic, and progressively painful and the patient was admitted to our hospital. The diagnosis was made by skin biopsy and tissue culture. The skin lesions resolved by the sixth week of the treatment with voriconazole.

Potential of endophytic fungi as biocontrol agents of Duponchelia fovealis (Zeller) (Lepidoptera:Crambidae).

This study reports the first assessment of endophytic fungi isolated from strawberry leaves and selection of isolates for the control of Duponchelia fovealis, a new pest of strawberries. A total of 400 strawberry leaves of the cultivar 'Albion' were collected in four commercial farms. Leaves were disinfected, cut in fragments, and placed on Petri dishes containing potato dextrose agar media with tetracycline and incubated for 30 days. Following this time, 517 fungal colonies were isolated, and thirteen genera were identified: Cladosporium, Aspergillus, Nigrospora, Fusarium, Trichoderma, Chaetomium, Alternaria, Paecilomyces, Penicillium, Ulocladium, Bipolaris, Diaporthe, and Phoma. Eight isolates belonging to the genera Aspergillus, Diaporthe, Paecilomyces, and Cladosporium were selected for pathogenicity bioassays against third instar larvae of D. fovealis. Isolates of Paecilomyces induced the highest mortality rates.

Fatal Purpureocillium lilacinum pneumonia in a green tree python.

A 10-y-old female green tree python ( Morelia viridis) died of fungal pneumonia caused by Purpureocillium lilacinum, which was confirmed histologically and by PCR and subsequent DNA sequencing. The same fungal species was cultivated from a swab taken from the terrarium in which the snake was housed. Clinical and environmental P. lilacinum isolates were indistinguishable by the typing method applied, strongly suggesting clonal relatedness of both isolates. Because no other underlying predisposing respiratory infection could be detected by virus-specific PCR or histopathology, P. lilacinum was considered a primary pulmonary pathogen in this tree python.

Paecilomyces lilacinus Keratitis in a Soft Contact Lens Wearer.

We describe a case of keratitis caused by Paecilomyces lilacinus in a contact lens wearer with a history of diabetes.

[Discussion on related problems of model species of fungus： Paecilomyces hepialid].

Paecilomyces hepiali is a new species of fungus isolated from a field collection of Ophiocordyceps sinensis from Baima snow mountain, Diqing Tibetan Autonomous Prefecture, Yunnan Province by the Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences. The specimen was identified and named as Paecilomyces hepiali by Qing-Tao Chen, the professor of the Institute of Microbiology, Chinese Academy of Sciences (Paecilomyces hepiali) (2008), who identified a dried culture of living strain 82-2 as the holotype. Until now, the holotype (the voucher specimen) was deposited in the Herbarium of the Institute of Chinese Materia Medica (HICMM), China Academy of Chinese Medical Sciences, Beijing. The P. hepiali neotype designated by the paper "Neotypification of P. hepiali (Hypocreales)" published in TAXON 64 (1) by Yao Yi-Jian et al. in February 2015 is untenable.

Fungal strains isolation, identification and application for the recovery of Zn(II) ions.

Fungal biomass proves to be highly efficient for the treatment of wastewater as well as recovery of metal ions from wastewater. Present investigation was aimed to evaluate the efficiency of indigenous fungal isolates for the sequestration of Zn(II) ions aqueous solution. Among twenty five fungal isolates, Aspergillus oryzae SV/09 (AO SV/09), Aspergillus flavus NA9 (AF NA9) and Paecilomyces formosus DTO 63f4 (PF DTO-63f4) were identified by gene sequencing of ITS regions of the ribosomal DNA (rDNA). The AO SV/09, AF NA9 and PF DTO-63f4 showed promising efficiency for the biosorption of Zn(II) ions. Zn(II) ions adsorption was endothermic in nature and data fitted will to the Freundlich isotherm with correlation coefficients values of 0.99, 0.98 and 0.99 for AO SV/09, AF NA9 and PF DTO-63f4, respectively. Pseudo-second order kinetic model explained well the Zn(II) adsorption kinetic of Zn(II) ions onto biosorbents. The adsorbed Zn(II) ions were desorbed using HCl and 85.5, 75.3, 73.7 (%) Zn(II) ions were recovered from AO SV/09, AF NA9 and PF DTO-63f4 sorbents, respectively. The fungal biosorbents were successfully recycled up to five cycles. Based on sorption, recovery and regeneration, the application of fungal bio-sorbents for the sequestration and recovery of Zn(II) ions is suggested from wastewater and could possibly be extended for the recovery of other heavy metal ions from wastewater.