Antimicrobial Activities and Molecular Signature of Endophytic Fungi of Opuntia ficus-Indica Cacti and the Cactus-Like Plant Aloe vera

Endophytes are fungi that colonize the internal tissues of plants without causing immediate adverse effects. Saudi Arabia (SA) is rich in Opuntia ficus-indica cacti and the cactus-like plant Aloe vera, which grow in the southern and western areas of SA. This study aimed to isolate and identify endophytic fungi from cacti and cactus-like plants in the Jeddah, Taif, and Al Baha regions KSA and then determine their effects on pathogenic fungal and bacterial growth. The isolates were groupedAloe Vera; Opuntia Ficus-Indica; Endophytic Fungi; Antimicrobial Activities; Pathogenic Bacteria. into 16 distinct operational taxonomic units based on the sequence of the internal transcribed spacer in the rDNA gene with the primers ITS1 and ITS4. Mucor circinelloides was the endophytic fungus found most frequently, with a relative frequency of 20.43%, followed by Talaromyces funiculosus, with a relative frequency of 16.12% when isolated from Opuntia ficus-indica and Aloe vera. Nine out of sixteen endophytic fungi exhibited strong antifungal activity against all the tested pathogens. P. funiculosum, Aspergillus versicolor, Penicillium janthinellum, and Fusarium oxysporum showed vigorous antimicrobial activities against the human pathogenic bacteria Escherichia coli, Shigella sp., and Salmonella typhimurium.

Sterigmatocystin Limits Plasmodium falciparum Proliferation and Transmission

As part of our drug discovery program against malaria, the Penicillium janthinellum extract was discovered to inhibit P. falciparum proliferation in blood and transmission to mosquitoes. Bioactivity-guided fractionation of P. janthinellum extraction was carried out using chromatographic techniques. We determined the activities of fractions against Plasmodium falciparum asexual stage parasite proliferation in culture and sexual stage parasite transmission to mosquitoes using standard membrane feeding assays (SMFA). One active compound was isolated. Based on mass spectrometry and nuclear magnetic resonance profiles, the compound was structurally determined to be sterigmatocystin. Sterigmatocystin inhibited P. falciparum proliferation in the blood with an IC50 of 34 µM and limited the sexual parasites to infect mosquitoes with an IC50 of 48 µM. Meanwhile, sterigmatocystin did not show any acute toxicity to human kidney cells at a concentration of 64 µM or lower. Sterigmatocystin can be used as a drug lead for malaria control and as a probe to understand molecular mechanisms of malaria transmission.

Ultrasound-Assisted Production of Xylo-Oligosaccharides From Alkali-Solubilized Corncob Bran Using Penicillium janthinellum XAF01 Acidic Xylanase

A novel treatment involving enzymatic hydrolysis using an acidic xylanase coupled with ultrasound was performed to improve the xylo-oligosaccharides (XOS) yield from corncob bran. The acidic xylanase (XynB) was purified to a most suitable pH, temperature, and operational parameters for ultrasound-assisted hydrolysis were determined. A preliminary mechanistic investigation was performed through circular dichroism (CD) spectroscopy, scanning electron microscope (SEM) and a laser particle size analyzer, and the effects of ultrasound on enzyme (XynB) and substrate (corncob bran) were assessed. The results show that the maximum XOS yield was 20.71% when the reaction pH and temperature were 4.3 and 50°C, the ultrasonic parameters were 50 kHz and 0.40 W/cm2, which was 2.55 fold higher than that obtained using a non-ultrasound-assisted enzymatic preparation. Mechanism studies indicated that ultrasonic pretreatment could reduce the β-fold content and increase the random coil content. Changes in structure and size of substrate were observed. The specific surface area of the XAC molecules is easy to carry out enzymatic reaction, which is beneficial to the production of XOS.

SELEÇÃO DE FUNGOS PRODUTORES DE CELULASE PELO TESTE DO VERMELHO CONGO

Introdução: A celulose é um dos principais constituintes da parede celular vegetal, sendo de grande interesse econômico por ser uma alternativa para a produção de bioetanol, um combustível de fontes renováveis de ampla utilização mundial. O processo industrial de produção de bioetanol utiliza enzimas celulases fúngicas para converter a celulose em glicose, que será o substrato para a fermentação alcoólica. Entretanto, o modelo de interação de enzima-substrato pode variar de acordo com a fonte de celulose utilizada. O presente estudo visou uma seleção de fungos produtores de celulase para uma posterior avaliação da fermentação com celulose obtida a partir de algodão. Objetivos: Obtenção de um parâmetro qualitativo para screening de fungos celulolíticos. Material e métodos: As culturas foram induzidas a produção de celulase pelo meio de triagem cuja a única fonte de carbono foi a carboximetilcelulose com nutrição de sais. o período de cultivo foi de 7 dias em 30º C. As mesmas condições foram mantidas nos cultivos para o teste do vermelho congo. A ausência de detergentes diminuiu a indução dos halos de hidrolise. No teste, as colônias foram submetidas a choque térmico em 50 ºC por 4 h e uma solução de vermelho congo 2,5 g/L foi adicionada. Após 30 minutos a solução foi descartada e lavada com NaCL para realce dos halos. O índice enzimático (i.e) foi obtido pela razão dos diâmetros do halo de hidrolise com o halo de colônia. Resultados: Das 26 amostras, 4 testaram negativo para a hidrolise de celulose, os maiores índices enzimáticos ocorreram no fungo B12 proveniente da amostra de Buriti e Penicillium Janthinellum com i.e 2, 37 e 2,18 respectivamente. Conclusão: Devido à ausência de detergentes, fungos com maior potencial podem ter sido mascarados uma vez que os detergentes auxiliam na formação do complexo enzima-substrato. Observou-se uma maior relação entre a falta de detergentes e o halo de hidrolise em fungos Trichoderma spp o que se comprova na bibliografia.

Whole Genome Sequence and CAZyme distribution of the cellulase hyper producing filamentous fungus Penicillium janthinellum NCIM 1366

Penicillium janthinellum NCIM 1366, capable of secreting cellulases that are highly efficient in the hydrolysis of lignocellulosic biomass, was sequenced to understand its cellulolytic machinery. De novo sequencing and assembly revealed a 37.6 Mb genome encoding 11,848 putative proteins, 93% of which had significant BLAST-P hits. The majority of the top hits (those with over 60% UniProt identity) belonged to P. brasilianum. Carbohydrate active enzymes (CAZymes) and other enzymes involved in lignocellulose degradation were also predicted from this strain and compared with those of the industrial workhorse of cellulase production- Trichoderma reesei RUT-C30. The comparison showed that the fungus encodes a far higher number of CAZYmes (422) as compared to T. reesei RUT-C30 (244), which gives a plausible explanation for its overall effectiveness in biomass hydrolysis. An analysis of the secreted CAZymes and annotated ligninases identified 216 predicted proteins which may be directly involved in the breakdown of lignocellulose

Sequencing and Comparative Genomic Analysis of a Highly Metal-Tolerant Penicillium janthinellum P1 Provide Insights Into Its Metal Tolerance

Heavy metal pollution is a global knotty problem and fungi hold promising potential for the remediation of wastewater containing heavy metals. Here, a new highly chromium-tolerance species, Penicillium janthinellum P1, is investigated. The genome of P1 was sequenced and assembled into 30 Mb genome size containing 10,955 predicted protein-coding genes with a GC content of 46.16% through an integrated method of Illumina short-read sequencing and single-molecule real-time Pacific Biosciences sequencing platforms. Through a phylogenetic analysis with model species of fungi, the evolutionary divergence time of Penicillium janthinellum P1 and Penicillium oxalicum 114-2 was estimated to be 74 MYA. 33 secondary metabolism gene clusters were identified via antiSMASH software, mainly including non-ribosomal peptide synthase genes and T1 polyketide synthase genes. 525 genes were annotated to encode enzymes that act on carbohydrates, involving 101 glucose-degrading enzymes and 24 polysaccharide synthase. By whole-genome sequence analysis, large numbers of metal resistance genes were found in strain P1. Especially ABC transporter and Superoxide dismutase ensure that the P1 fungus can survive in a chromium-polluted environment. ChrA and ChrR were also identified as key genes for chromium resistance. Analysis of their genetic loci revealed that the specific coding-gene arrangement may account for the fungus’s chromium resistance. Genetic information and comparative analysis of Penicillium janthinellum are valuable for further understanding the mechanism of high resistance to heavy metal chromium, and gene loci analysis provides a new perspective for identifying chromium-resistant strains.

Janthinoid A, an unprecedented tri-nor-meroterpenoid with highly modified bridged 4a,1-(epoxymethano)phenanthrene scaffold, produced by the endophyte of Penicillium janthinellum TE-43

Janthinoid A (1), an unprecedented C22 meroterpenoid featuring a highly modified bridged 4a,1-(epoxymethano)phenanthrene scaffold incorporating eight continuous quaternary carbons, along with its biosynthetic-related C25 analogue andrastone I (2), were isolated...

Penicillium janthinellum NCIM1366 shows improved biomass hydrolysis and a larger number of CAZymes with higher induction levels over Trichoderma reesei RUT-C30

Background Major cost of bioethanol is attributed to enzymes employed in biomass hydrolysis. Biomass hydrolyzing enzymes are predominantly produced from the hyper-cellulolytic mutant filamentous fungus Trichoderma reesei RUT-C30. Several decades of research have failed to provide an industrial grade organism other than T. reesei, capable of producing higher titers of an effective synergistic biomass hydrolyzing enzyme cocktail. Penicillium janthinellum NCIM1366 was reported as a cellulase hyper producer and a potential alternative to T. reesei, but a comparison of their hydrolytic performance was seldom attempted. Results Hydrolysis of acid or alkali-pretreated rice straw using cellulase enzyme preparations from P. janthinellum and T. reesei indicated 37 and 43% higher glucose release, respectively, with P. janthinellum enzymes. A comparison of these fungi with respect to their secreted enzymes indicated that the crude enzyme preparation from P. janthinellum showed 28% higher overall cellulase activity. It also had an exceptional tenfold higher beta-glucosidase activity compared to that of T. reesei, leading to a lower cellobiose accumulation and thus alleviating the feedback inhibition. P. janthinellum secreted more number of proteins to the extracellular medium whose total concentration was 1.8-fold higher than T. reesei. Secretome analyses of the two fungi revealed higher number of CAZymes and a higher relative abundance of cellulases upon cellulose induction in the fungus. Conclusions The results revealed the ability of P. janthinellum for efficient biomass degradation through hyper cellulase production, and it outperformed the established industrial cellulase producer T. reesei in the hydrolysis experiments. A higher level of induction, larger number of secreted CAZymes and a high relative proportion of BGL to cellulases indicate the possible reasons for its performance advantage in biomass hydrolysis.