Humulane-Type Macrocyclic Sesquiterpenoids From the Endophytic Fungus Penicillium sp. of Carica papaya

Three new humulane-type sesquiterpenoids, penirolide A (1), penirolide B (2), and 10-acetyl-phomanoxide (3), together with three known compounds aurasperone A (4), pughiinin A (5), and cyclo(l-Leu-l-Phe) (6) were isolated from the endophytic fungus Penicillium sp. derived from the leaves of Carica papaya L. Their structures including their absolute configurations were determined based on the analysis of NMR and HRESIMS spectra, NMR chemical shifts, and ECD calculations. Compounds 2, 3, 5, and 6 significantly inhibited glucagon-induced hepatic glucose production, with EC50 values of 33.3, 36.1, 18.8, and 32.1 μM, respectively. Further study revealed that compounds 2, 3, 5, and 6 inhibited hepatic glucose production by suppression of glucagon-induced cAMP accumulation.

Клонирование и экспрессия гена целлюлазы Penicillium sp. ‘occitanis’ в Komagataella phaffii T07, выделение и анализ свойств

Проведены клонирование и анализ свойств целлюлазы Penicillium sp. ‘occitanis’. Ген был обнаружен в геномных данных базы NCBI. Получена плазмида pPZL-4xBGLU-Pocc, несущая четыре копии конструкции, экспрессирующей ген целлюлазы P. sp. ‘occitanis’. Плазмида была клонирована в штамм дрожжей Komagataella phaffii T07 под контроль промотора и терминатора гена AOX1. Было клонировано четыре копии экспрессирующей конструкции. В результате получен штамм K. phaffii T07 BGLU-4xPocc. Максимальный уровень экспрессии этого штамма составил ~4 000 000 Ед/л культуральной жидкости при культивировании в 5-литровом биореакторе. Полученный белок, согласно данным гельэлектрофореза, дает полосу ~39.5 кДа, что соответствует расчетной массе, и обширную область примерно от 42 до 70 кДa. Формирование этой области говорит о возможном гликозилировании белка при экспрессии в K. phaffii. Анализ свойств фермента показал, что температурный оптимум активности составляет 55–65 °С, рН оптимум – примерно 1.5–2. Фермент сохраняет 50 % активности при pH от 1.5 до 6 и от 30 до 80 °С. Он стабилен при температуре ниже 75 °С в течение 4 ч, но при 85 °С за 4 ч теряет 50 % активности. Фермент устойчив при рН от 4 до 7.5.

New compounds from Sarcophyton glaucom-derived Penicillium sp.

Further investigation of the residual bioactive compounds produced by the soft coral Sarcophyton glaucom-derived Penicillium sp. MMA afforded five new compounds assigned as 9-methoxy-penicyrone A (1), 9-methoxy-penicyrone B (2), 3-hydroxy-2,2,4-trimethyl-pentyl ester (3), 3-hydroxy-1-isopropyl-2,2-dimethyl-propyl ester (4), and 3-isobutyryloxy-2,2,4-trimethyl-pentyl linoleate (5). Additional six known compounds were isolated: penicyrones A–B (6, 7), 4-(2-hydroxy-3-butynoxy)benzoic acid (8), cyclopenol (9), aspermytin A (10), and aurantiomide A (11). Structures of the new compounds (1–5) were identified by 1D (1H & 13C) and 2 D (1H–1H COSY, HMBC and NOESY) NMR and HRESI-MS spectroscopic data. Biologically, the antimicrobial activities of the obtained compounds were studied as well.

Fungos anemófilos associados ao ambiente das enfermarias em unidade hospitalar do Cabo de Santo Agostinho-PE, Brasil

Os fungos anemófilos contaminantes de ambientes hospitalares, aparentemente inofensivos, têm se tornado importantes agentes oportunistas a serem considerados como possíveis precursores de manifestações alérgicas e de infecções graves em indivíduos com o sistema imunológico comprometido, por isso, conhecer a microbiota anemófila das unidades de saúde tem sido tão importante para o desenvolvimento de ações que reduzam a proliferação dos biocontaminantes, bem como de programas de controle de infeções. O objetivo deste estudo foi isolar e identificar os fungos anemófilos associados ao ambiente das enfermarias de uma unidade de saúde situada no município do Cabo de Santo Agostinho – PE. A coleta foi realizada pelo método de sedimentação passiva de esporos, onde placas de Petri, contendo meio de cultura específico, foram expostas abertas em pontos estratégicos durante 20 min. Após este período as placas foram fechadas, etiquetadas e incubadas por 7 dias a 28 ºC, com leituras diárias para o acompanhamento do crescimento fúngico. Foram encontrados os fungos filamentosos: Penicillium sp., Aspergillus sp., Fusarium sp., Rhizopus sp., Cladosporium sp., Acremonium sp., Tritirachium sp., Curvularia sp., Nigrospora sp., Cunninghamella sp., além de leveduras e fungos não-esporulantes. Os gêneros mais frequentes foram Penicillium e Aspergillus, os quais são possíveis agentes de doenças disseminadas em pacientes imunodeprimidos. Os resultados encontrados neste estudo subsidiaram o desenvolvimento e a implantação de programas e estratégias de controle de patógenos fúngicos e de prevenção das infecções nosocomiais pela Comissão de Controle de Infecção Hospitalar (CCCIH) do hospital analisado.

Evaluation of the antimicrobial and anti-Varroa destructor L. activity of the essential oil of clove (Syzygium aromaticum L. Myrtaceae)

Antimicrobial and anti-Varroa destructor L. activity of the essential oil (EO) of clove (Syzygium aromaticum L. Myrtaceae) was evaluated in this study. Antimicrobial activity concerned 9 bacterial strains (Bacillus cereus, Escherichia coli ATCC 25911, Staphylococcus aureus 29213 ATCC, Proteus mirabilis, Serratia marcescens, Enterobacter sakazakii, Pseudomonas aeruginosa ATCC, Escherichia coli and Acinetobacter sp.), a fungal strain (Penicillium sp.) and a yeast species (Candida albicans). The extraction of the EO was carried out by the method of hydrodistillation. Results showed that EO has no toxicity on bees Apis mellifera. EO demonstrated effective and stable anti-V. destructor activity, indicating the absence of possible resistance, in contrast to what was observed for Amitraz. The inhibitory activity of EO revealed an inhibition zone of diameter varying between 20 mm and 42 mm for bacteria. However, no inhibition zones were observed for Pseudomonas aeruginosa ATCC. The inhibitory activity of EO on Penicillium sp and Candida albicans revealed an inhibition zone of 39 mm in diameter. This study shows that the EO of cloves constitutes a simple and natural treatment, without inconveniences, with a high activity antimicrobial and anti-Varroa destructor which merits it to be proposed as a means to fight against varroosis and the tested pathogens.

Monitoring of mycotoxins in feed of the Krasnodar Territory and their combined effect on the microbiocenosis of the gastrointestinal tract of cattle

Diets for cattle are affected by fungi - producers of mycotoxins: hay, straw, silage, grain, concentrates. Highly productive cattle breeds are more sensitive to mycotoxins. It has been studied that mycotoxins have antibiotic properties, disrupting the biocenosis and functionality of the proventriculus of cattle. This leads not only to a disruption in the nutritional process of animals, but also contributes to a decrease in the enzymatic activity of the rumen microflora and thus facilitates the penetration of other mycotoxins into the animal body. When conducting sanitary and mycological studies of coarse and succulent feed, compound feed and feed additives for cattle, it was found that feed was affected by micromycetes in descending order Mucor sp., Fusarium sp., Aspergillus sp., Candida sp., Penicillium sp. In the study of feeds, it was found that the most frequently recorded mycotoxins in descending order, T-2 toxin, aflatoxin, zearalenonone, ochratoxin A, fumonisin. Monitoring studies found that feed for cattle not containing mycotoxins was 33.4%, containing one mycotoxin 34.8%, two mycotoxins 28.8%, three mycotoxins 3.03%. When conducting scientific and economic experiments, the content of two mycotoxins in feed was diagnosed, the concentration of which for zearalenone was 1.0 mg / kg, for aflatoxin - 0.04 mg / kg. To study the effect of mycotoxins on the microflora of the gastrointestinal tract (proventriculus), an experiment was carried out on 40 bull calves of 7 months of age. The experimental group of animals (20 animals) was given feed contaminated with mycotoxins, the control group received a diet without mycotoxins. the contents of the rumen were collected on days 10, 20, and 30 of the experiment. The clinical picture in experimental animals with combined mycotoxicosis during the experimental period was characterized by a decrease in feed consumption by 10-20%, weight gain by an average of 18-25%, exhaustion, dullness of the coat, and alopecia. The difference between the experimental and control group of lactobacilli was 28.0%; 157.1% and 215.0% log10 CFU, respectively, for study days. Bifidobacteria 13.0%; 86.2% and 192.3% log10 CFU, respectively. The decrease in ciliates in the experimental group in comparison with the control on the 10th day of the experiment was 7.1 thousand units / g, on the 20 day 27.9 thousand units / g, on the 30th day 77.2 thousand units / g. Thus, the content of the main joints of the biocenosis in the rumen of cattle on days 10, 20 and 30 of the experiment was significantly reduced in the experimental group, the diet of which contained the association of mycotoxins. The monitoring carried out indicates that mycotoxins (in descending order) are most often recorded in feed for farm animals in the Krasnodar Territory: T-2 toxin, aflatoxin B1, zearalenone, ochratoxin A, fumonisin. A high contamination of feed and raw materials with molds was established - Mucor sp., Fusarium sp., Aspergillus sp., Candida sp., Penicillium sp. A decrease in the main representatives of the biocenosis actively participating in the digestion of cattle: lactobacilli, bifidobacteria, ciliates, under the influence of the combined effect of mycotoxins, has been established.