Long non-coding RNAs (lncRNAs) play a significant role in stress responses. To date, only a few studies have reported the role of lncRNAs in insect-pathogenic fungi. Here, we report a genome-wide transcriptional analysis of lncRNAs produced in response to heat stress in Metarhizium robertsii, a model insect-pathogenic fungus, using strand-specific RNA sequencing. A total of 1655 lncRNAs with 1742 isoforms were identified, of which 1081 differentially expressed (DE) lncRNAs were characterized as being heat responsive. By characterizing their genomic structures and expression patterns, we found that the lncRNAs possessed shorter transcripts, fewer exons, and lower expression levels than the protein-coding genes in M. robertsii. Furthermore, target prediction analysis of the lncRNAs revealed thousands of potential DE lncRNA–messenger RNA (mRNA) pairs, among which 5381 pairs function in the cis-regulatory mode. Further pathway enrichment analysis of the corresponding cis-regulated target genes showed that the targets were significantly enriched in the following biological pathways: the Hippo signaling pathway and cell cycle. This finding suggested that these DE lncRNAs control the expression of their corresponding neighboring genes primarily through environmental information processing and cellular processes. Moreover, only 26 trans-regulated lncRNA–mRNA pairs were determined. In addition, among the targets of heat-responsive lncRNAs, two classic genes that may be involved in the response to heat stress were also identified, including hsp70 (XM_007821830 and XM_007825705). These findings expand our knowledge of lncRNAs as important regulators of the response to heat stress in filamentous fungi, including M. robertsii.

For the high-value utilization of tuna liver, the effects of acid-aided (Acid-pH) and alkali-aided pH-shifting (Alkali-pH) on the physicochemical and functional properties of the protein powder prepared by pH-shifting and freeze-drying were studied. As expected, the protein powder with high purity could be obtained through Acid-pH or Alkali-pH followed by freeze drying, while the Alkali-pH led to a higher protein yield, higher protein ratio, lower lipid ratio and lower heavy metal content than Acid-pH. The amino acid profile of the protein powder prepared by Alkali-pH (Alkali-PP) was similar with that prepared by Acid-pH (Acid-PP). In addition, compared with Acid-PP, the Alkali-PP possessed the greater capacities in emulsion activity, foaming capacity and fat absorption capacity. Furthermore, the foaming capacity, foam stability and fat absorption capacity of Alkali-PP was better than soy protein powder. Therefore, Alkali-pH followed by freeze-drying would be a better alternative to prepare high-quality protein powder from tuna liver in the food industry.

Metarhizium spp. are well-known biocontrol agents used worldwide to control different insect pests. Keto-acid reductoisomerase (ILVC) is a key enzyme for branched-chain amino acid (BCAA) biosynthesis and regulates many physiological activities. However, its functions in insect pathogenic fungi are poorly understood. In this work, we identified MrilvC in M. robertsii and dissected their roles in fungal growth, conidiation, germination, destruxins biosynthesis, environmental stress response and insecticidal virulence. BCAA metabolism affects conidial yields and germination. However, BCAAs cannot recover the conidia germination of MrilvC deficient strain. Further feeding assays with intermediates showed that some conidia of ΔMrilvC start to germinate. Therefore, it is the germination defect that causes the complete failures of conidial penetration and pathogenicity in ΔMrilvC mutant. In conclusion, we found intermediates in BCAA biosynthesis are indispensable for Metarhizium robertsii conidial germination. This study will advance our understanding of the fungal germination mechanism. IMPORTANCE Branched-chain amino acid (BCAA) metabolism plays a significant role in many biological activities beyond protein synthesis. Spore germination initiates the first stage of vegetative growth, which is critical for the virulence of pathogenic fungi. In this study, we demonstrated that the keto-acid reductoisomerase MrILVC, a key enzyme for BCAA biosynthesis, from the insect pathogenic fungus Metarhizium robertsii is associated with conidial germination and fungal pathogenicity. Surprisingly, the germination of ΔMrilvC mutant was restored when supplemented with the intermediates of BCAA metabolism rather than three BCAAs. The result was significantly different from that of plant pathogenic fungi. Therefore, this report highlights that the intermediates in BCAA biosynthesis are indispensable for conidial germination of M. robertsii.

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Silymarin, a known extract, is used in the treatment of liver diseases with various origins, but its current administration form cannot target the liver because of its poor oral bioavailability. A new type of oral silymarin proliposome aimed at improving silymarin’s poor bioavailability and hepatoprotective effects, is introduced in this work. Silymarin-loaded liquid proliposome were prepared using a simple dissolving process. The morphology, particle size, zeta potential, and entrapment efficiency of the silymarin liposomes were analysed. The everted gut sac transport model was used to measure the intestinal transport of liposomes. The liposomal hepatoprotective activity was evaluated in three types of experimental hepatitis animal models. After staining with haematoxylin and eosin, the livers were microscopically examined to analyse any pathological changes. The prepared silymarin proliposome formed silymarin liposomes with a multilayer liposome structure and improved intestinal transport. In an injured liver, the silymarin liposomes produced a stronger hepatoprotective effect through a significant decrease in both the aminotransferase and MDA levels and a significant increase in the SOD and GSH-PX levels compared to orally administered silymarin tablets. This effect was also confirmed histopathologically. In a word, incorporation of silymarin into a liposomal carrier system increased intestinal absorption and showed better hepatoprotective effects compared to silymarin tablets.