Discovery of microRNA-like Small RNAs in Pathogenic Plant Fungus Verticillium nonalfalfae Using High-Throughput Sequencing and qPCR and RLM-RACE Validation

Verticillium nonalfalfae (V. nonalfalfae) is one of the most problematic hop (Humulus lupulus L.) pathogens, as the highly virulent fungal pathotypes cause severe annual yield losses due to infections of entire hop fields. In recent years, the RNA interference (RNAi) mechanism has become one of the main areas of focus in plant—fungal pathogen interaction studies and has been implicated as one of the major contributors to fungal pathogenicity. MicroRNA-like RNAs (milRNAs) have been identified in several important plant pathogenic fungi; however, to date, no milRNA has been reported in the V. nonalfalfae species. In the present study, using a high-throughput sequencing approach and extensive bioinformatics analysis, a total of 156 milRNA precursors were identified in the annotated V. nonalfalfae genome, and 27 of these milRNA precursors were selected as true milRNA candidates, with appropriate microRNA hairpin secondary structures. The stem-loop RT-qPCR assay was used for milRNA validation; a total of nine V. nonalfalfae milRNAs were detected, and their expression was confirmed. The milRNA expression patterns, determined by the absolute quantification approach, imply that milRNAs play an important role in the pathogenicity of highly virulent V. nonalfalfae pathotypes. Computational analysis predicted milRNA targets in the V. nonalfalfae genome and in the host hop transcriptome, and the activity of milRNA-mediated RNAi target cleavage was subsequently confirmed for two selected endogenous fungal target gene models using the 5′ RLM-RACE approach.

Plantas medicinais no tratamento de ansiedade e depressão: Uma revisão

Há uma grande diversidade de plantas no Brasil, e muitas delas, ao serem estudadas, apresentam elevado teor nutricional, presença de antioxidante, entre outras propriedades, podendo ter um potencial farmacológico significativo e consequentemente gerando aplicações em diversas áreas como saúde, cosmética e culinária. As plantas medicinais são usadas por grande parte da população mundial, como um recurso medicinal alternativo para o tratamento de diversas doenças, visto que em muitas comunidades, representam um recurso mais acessível em diversos sentidos em relação aos medicamentos alopáticos. Destaca-se a utilização das plantas medicinais para fins terapêuticos, em que muitas atividades biológicas conferidas às plantas foram demonstradas, dentre elas a ação no sistema nervoso central, sendo utilizadas para o tratamento e/ou prevenção de transtornos psicossociais como ansiedade e depressão. Sendo assim, o presente estudo tem como objetivo realizar uma revisão bibliográfica sobre as plantas medicinais mais utilizadas no tratamento da ansiedade e depressão. Verificou-se que nos tratamentos, as plantas medicinais e os fitoterápicos são vistos como uma alternativa viável para o tratamento dos quadros de ansiedade e depressão, visto que possuem menos efeitos colaterais. Sendo assim, a presente pesquisa revisou as espécies: Hypericum perforatum L.; Passiflora incarnata L.; Melissa Officinalis L.; Chamomilla recutita L.; Piper methysticum G.; Erythrina verna; Humulus lupulus L. e Crataegus oxyacantha L. Foi possível apresentar a efetividade de todas as espécies citadas, com as descrições encontradas em estudos da literatura.

Evaluation of resistance to Pseudoperonospora humuli and of the content of alpha acids and hop oils in hops of selected genetic resources of hop Humulus lupulus L.

Twenty hop genotypes were selected for the evaluation of resistance to primary and secondary Pseudoperonospora humuli infection and of alpha acid and hop oil content in the hops. From the wild hop genotypes, two from Canada and one from Belgium showed resistance. Among the registered hop varieties, the Czech varieties Kazbek and Boomerang were the most resistant. Both wild hop genotypes from Canada showed the highest content of alpha acids among the wild hop entries, namely 4% w/w. The lowest variability of the alpha acid content in the wild hop category was found in two wild hop varieties from the Caucasus, one from Austria and one from Lithuania. The highest content of hop oils was determined in two hop genotypes from Canada and two from Belgium. Wild hop genotypes from the Caucasus have the lowest variability of hop oils among the wild hop entries. Two hop genotypes from Canada and one from Belgium were selected for breeding aimed at drought resistance.

Establishment of an Efficient Micropropagation System for Humulus lupulus L. cv. Cascade and Confirmation of Genetic Uniformity of the Regenerated Plants through DNA Markers

The demand for virus-free hop planting material has increased in the last few years due to its multipurpose uses. The present study aimed to establish an effective protocol for clonal propagation of cv. Cascade using only the cytokinins as PGRs in all stages of micropropagation: (i) in vitro culture initiation using single-node micro-cuttings inoculated on modified Murashige and Skoog (MSm) medium solidified with Plant agar and supplemented with 0.5 mg L−1 6-benziyladenine (BA) with 76% recorded viability of nodal explants; (ii) in vitro multiplication of multinodal shoots on MSm medium gelled with Plant agar and supplemented with different types and concentrations of cytokinins: 2 mg L−1 kinetin (KIN), 0.7 mg L−1 1-(2-Chloro-4-pyridyl)-3-phenylurea) (1 CPPU), 2 mg L−1 meta-topoline (mT) and 0.5 mg L−1 BA, which was the best variant for shoot proliferation (9.48 ± 0.78 shoots/explant); (iii) rooting and acclimatization with the best results obtained by ex vitro rooting and acclimatization of plants in the same stage in perlite (96.00 ± 0.60% acclimatized rooted plants with 100% survival under greenhouse conditions). The true-to-type nature of in vitro raised plants with the mother plant was assessed by Random Amplified Polymorphic DNA (RAPD) and Start Codon Target Polymorphism (SCoT) molecular markers, and then their genetic uniformity were confirmed.

Microwave-Assisted Hydrodistillation of Hop (Humulus lupulus L.) Terpenes: A Pilot-Scale Study

Interest in essential oils has consistently increased in recent years. Essential oils have a large variety of applications in multiple fields, including in the food, cosmetics and pharmaceutical industries. The volatile fraction (VF) in hops (Humulus lupulus L.) fits within this domain as it is primarily used in the brewery industry for the aromatization of beer, and is responsible for the floral and fruity tones. This work aims to design an optimized extraction protocol of the VF from hops, using microwaves. Microwave-assisted hydrodistillation (MAHD) has been developed to reduce energy and time consumption in lab-scale reactors up to industrial-scale systems. Hops are principally available in three forms, according to a brewery’s applications: (i) fresh (FH); (ii) dried (DH) and (iii) pelletized (PH). In this work, all three forms have therefore been studied and the recovered volatiles characterized by means of GC-MS. The optimized lab-scale MAHD protocol gave the best extraction yield of 20.5 mLVF/kgdry matrix for FH. This value underwent a slight contraction when working at the highest matrix amount (3 kg), with 17.3 mLVF/kgdry matrix being achieved. Further tests were then performed in a pilot reactor that is able to process 30 kg of material. In this case, high yield increases were observed for PH and DH; quadruple and double the lab-scale yields, respectively. In addition, this industrial-scale system also provided marked energy savings, practically halving the absorbed kJ/mLVF.