Fungistatic Mechanism of Ammonia against Nematode-Trapping Fungus Arthrobotrys oligospora, and Strategy for This Fungus To Survive Ammonia.

Soil fungistasis is a phenomenon in which the germination and growth of fungal propagules is widely inhibited in soils. Although fungistatic compounds are known to play important roles in the formation of soil fungistasis, how such compounds act on soil fungi is little studied. In this study, it was found that ammonia (NH3) induced global protein misfolding marked by increased ubiquitination levels of proteins (ubiquitylome data and Western blot verification). The misfolded proteins should trigger the endoplasmic reticulum (ER) stress, which was indicated by electron microscope image and proteome data. Results from the mutants of BiP and proteasome subunit alpha 7 suggested that ER stress played a mechanistic role in inhibiting conidial germination. Results from proteome data indicated that, to survive ammonia fungistasis, conidia first activated the unfolded protein response (UPR) to decrease ER stress and restore ER protein homeostasis, and the function of UPR in surviving ammonia was confirmed by using mutant strains. Second, ammonia toxicity could be reduced by upregulating carbon metabolism-related proteins, which benefited ammonia fixation. The results that metabolites (especially glutamate) could relieve the ammonia fungistasis confirmed this indirectly. Finally, results from gene knockout mutants also suggested that the fungistatic mechanism of ammonia is common for soil fungistasis. This study increased our knowledge regarding the mechanism of soil fungistasis and provided potential new strategies for manipulating soil fungistasis. IMPORTANCE Soil fungistasis is a phenomenon in which the germination and growth of fungal propagules is widely inhibited in soil. Although fungistatic compounds are known to play important roles in the formation of soil fungistasis, how such compounds act on soil fungi remains little studied. This study revealed an endoplasmic reticulum stress-related fungistatic mechanism with which ammonia acts on Arthrobotrys oligospora and a survival strategy of conidia under ammonia inhibition. Our study provides the first mechanistic explanation of how ammonia impacts fungal spore germination, and the mechanism may be common for soil fungistasis. This study increases our knowledge regarding the mechanism of soil fungistasis in fungal spores and provides potential new strategies for manipulating soil fungistasis.

Methylglyoxal Has Different Impacts on the Fungistatic Roles of Ammonia and Benzaldehyde, and Lactoylglutathione Lyase Is Necessary for the Resistance of Arthrobotrys oligospora to Soil Fungistasis.

Biocontrol of root-knot nematode has attracted increasing attention over the past two decades. The inconsistent field performance of biocontrol agents, which is caused by soil fungistasis, often restricts their commercial application. There is still a lack of research on the genes involved in biocontrol fungi response to soil fungistasis, which is important for optimizing practical applications of biocontrol fungi. In this study, the lactoylglutathione lyase-encoding AOL_s00004g335 in the nematophagous fungi Arthrobotrys oligospora was knocked out, and three mutant strains were obtained. The hyphal growth of mutants on the three media was almost the same as that of the wild-type strain, but mutants had slightly higher resistance to NaCl, SDS, and H2O2. Methylglyoxal (MG) significantly increased the resistance of A. oligospora to ammonia, but decreased the resistance to benzaldehyde. Furthermore, the resistance of the mutants to soil fungistasis was largely weakened and MG could not increase the resistance of A. oligospora to soil fungistasis. Our results revealed that MG has different effects on the fungistatic roles of ammonia and benzaldehyde and that lactoylglutathione lyase is very important for A. oligospora to resist soil fungistasis.

[Responses of the germinability of Sphagnum spores in peat to drainage in Baijianghe Peatland, China]. / 白江河泥炭地泥炭藓孢子萌发力对排水的响应.

Drainage severely changes the environment and ecological process in peatlands, but how does it affect the germinability of Sphagnum spores in peat remains unclear. In this study, we took two peat cores from a near-pristine stand dominated by Sphagnum and a drained stand dominated by dwarf shrubs in Baijianghe Peatland in the Changbai Mountains as experimental materials. Those peat cores were cut into slices. Physicochemical characteristics were measured while Sphagnum spores from each slice were extracted to count spore density and test spore germinability. After dating and determining relationship between peat depth and age, we tried to figure out the mechanism underlying the responses of Sphagnum spore germinability to drainage. The average number of spores in the near-pristine stand was slightly higher than that in the drained stand. There was no difference in average spore germinability between the two stands. The drained stand showed higher peat bulk density, total carbon and total nitrogen relative to the near-pristine stand. Upper peat core showed no significant difference in spore accumulation rate between the two stands after drainage (in 1987), with lower average spore germinability (34%) in the near-pristine stand relative to the drained stand (72%). For the whole peat cores, C/N was positively correlated with spore ger-minability in the near-pristine stand while total carbon, pH and burial time were negatively correlated with spore germinability in the drained stand. The drainage 30 years ago had limited effect on spore accumulation, but improved germinability of spores in shallow peat by changing physicochemical properties of peat due to accelerating decomposition, and thus reduced the persistence of spore bank. This may reduce the persistent regeneration potential of Sphagnum after catastrophic distur-bances.

[Simulated study on the effects of smoke on Sphagnum spore germination]. / çƒŸæ°”å¯¹æ³¥ç‚­è—“å­¢å­èŒå‘å½±å“çš„æ¨¡æ‹Ÿç ”ç©¶.

Moderate smoke could facilitate seed germination, but its effects on bryophyte spore germination is still unknown. Here, we analyzed the effects of smoke, capsule size and storage time on the spore germination of Sphagnum squarrosum and S. magellanicum, with the capsules of which being collected from two peatlands of the Changbai Mountains. The smoke solution prepared by burning peatland plants was combined with the capsules with different sizes (large, 2.10-2.50 mm in diameter; small, 1.50-1.90 mm in diameter) and storage time (old, being stored for 4.3 or 6.3 a; new, being stored for 0.3 a) to conduct a factorial experiment. The spores were soaked with smoke solution for different durations and then cultured for germination. The results showed that smoke solution affected spore germination. After 10 d cultivation, germination rate of spores soaking with smoke solution for all duration was increased by more than 5-fold, with the small spores having higher germination rate. After 21 d cultivation, the facilitative effect was only observed in moderate soaking (3 d), and spore size showed no effect on germination. Smoke solution could not increase the germination of spores from the capsules with long storage time (4.3 and 6.3 a). Our results indicated that moderate smoke solution soaking might accelerate germination of Sphagnum spores including small pores. In the ecosystems with casual fire disturbance such as peatlands, similar with its effects on the seed plants, smoke might play a key role in the regeneration and persistence of bryophyte population.

Comparative Transcriptomics Reveals Features and Possible Mechanisms of Glucose-Mediated Soil Fungistasis Relief in Arthrobotrys oligospora.

Soil-borne pest diseases result in large annual agricultural losses globally. Fungal bio-control agents are an alternative means of controlling pest diseases; however, soil fungistasis limits the effect of fungal agents. Nutrients can relieve soil fungistasis, but the mechanisms behind this process remain poorly understood. In this study, we determined and quantified the transcriptomes of Arthrobotrys oligospora, a nematode-trapping fungus, derived from samples of fresh conidia, germinated conidia, soil fungistatic conidia, and glucose-relieved conidia. The transcriptomes of fungistatic and glucose-relieved conidia were significantly different from those of the other two conidia samples. KEGG pathway analyses showed that those genes upregulated in fungistatic and glucose-relieved conidia were mainly involved in translation and substance metabolism, and the downregulated genes were mainly involved in MAPK pathway, autophagy, mitophagy, and endocytosis. As being different from the transcriptome of fungistatic conidia, upregulated genes in the transcriptome of glucose-relieved conidia are also related to replication and repair, spliceosome, oxidative phosphorylation, autophagy, and degradation pathway (lysosome, proteasome, and RNA degradation). And the upregulated genes resulted from comparison of glucose-relieved conidia and fungistatic conidia were enriched in metabolic pathways, cycle, DNA replication, and repair. The differentially splicing events in the transcriptome of glucose-relieved conidia are far more than that of other two transcriptomes, and genes regulated by differentially splicing were analyzed through KEGG pathway analysis. Furthermore, autophagy genes were proved to play important role in resisting soil fungistasis and glucose-mediated soil fungistasis relief. These data indicate that, in addition to being a carbon and energy source for conidia germination, glucose may also help to relieve soil fungistasis by activating many cellular processes, including autophagy, DNA replication and repair, RNA alternative splicing, and degradation pathways.

The lysine acetylome of the nematocidal bacterium Bacillus nematocida and impact of nematode on the acetylome.

Bacillus nematocida B16 (B16) is a pathogenic bacterium that is nematotoxic to plant-parasitic nematodes. In this study, we performed a quantitative lysine acetylome analysis on B16 to understand the potential roles of protein lysine acetylation on this host-pathogen interaction. Altogether, we identified 529 acetylation sites in 349 proteins, quantified 411 sites in 288 proteins, determined that the acetylation levels of 18 sites were up-regulated and those of 19 sites were down-regulated during pathogenesis. The acetylated proteins mainly participated in metabolic processes, protein synthesis, and cell wall/membrane biogenesis. Moreover, these proteins are involved in more than twenty KEGG pathways. Eight peptide motifs of acetylated proteins were identified, five of which have been thus far found only in the B16 acetylome. Twenty-two acetylated proteins were found to be involved in the synthesis of nematode attractants, and two were found to be involved in the secretion of virulence factors. In addition, the acetylation levels of ten lysine sites were regulated significantly differently in the presence of nematodes. Our results reveal that lysine acetylation may play roles in regulating B16-nematode interaction. SIGNIFICANCE: B. nematocida B16 is a bio-control bacterium against nematodes. It lures nematodes to their death by a Trojan horse mechanism. But there is little understanding about the regulation of this "Trojan horse" like pathogenesis. Lysine acetylation was reported to regulate diverse cellular processes. Our results revealed that lysine acetylation played indeed roles in regulating the B16-nematodes interaction. Our data laid a foundation for studying the molecule mechanism of lysine acetylation in regulating this host-pathogen interaction.