Complete Genome Sequence of Clostridium kluyveri JZZ Applied in Chinese Strong-Flavor Liquor Production.

Chinese strong-flavor liquor (CSFL), accounting for more than 70% of both Chinese liquor production and sales, was produced by complex fermentation with pit mud. Clostridium kluyveri, an important species coexisted with other microorganisms in fermentation pit mud (FPM), could produce caproic acid, which was subsequently converted to the key CSFL flavor substance ethyl caproate. In this study, we present the first complete genome sequence of C. kluyveri isolated from FPM. Clostridium kluyveri JZZ contains one circular chromosome and one circular plasmid with length of 4,454,353 and 58,581 bp, respectively. 4158 protein-coding genes were predicted and 2792 genes could be assigned with COG categories. It possesses the pathway predicted for biosynthesis of caproic acid with ethanol. Compared to other two C. kluyveri genomes, JZZ consists of longer chromosome with multiple gene rearrangements, and contains more genes involved in defense mechanisms, as well as DNA replication, recombination, and repair. Meanwhile, JZZ contains fewer genes involved in secondary metabolites biosynthesis, transport, and catabolism, including genes encoding Polyketide Synthases/Non-ribosomal Peptide Synthetases. Additionally, JZZ possesses 960 unique genes with relatively aggregating in defense mechanisms and transcription. Our study will be available for further research about C. kluyveri isolated from FPM, and will also facilitate the genetic engineering to increase biofuel production and improve fragrance flavor of CSFL.

From waste to protein: a new strategy of converting composted distilled grain wastes into animal feed.

Distilled grain waste (DGW) is rich in nutrients and can be a potential resource as animal feed. However, DGW contains as much as 14% lignin, dramatically reducing the feeding value. White-rot fungi such as Pleurotus ostreatus could preferentially degrade lignin with high efficiency. However, lignin derivatives generated during alcohol distillation inhibit P. ostreatus growth. Thus, finding a new strategy to adjust the DGW properties to facilitate P. ostreatus growth is critical for animal feed preparation and DGW recycling. In this study, three dominant indigenous bacteria, including Sphingobacterium thermophilum X1, Pseudoxanthomonas byssovorax X3, and Bacillus velezensis 15F were chosen to generate single and compound microbial inoculums for DGW composting to prepare substrates for P. ostreatus growth. Compared with non-inoculated control or single microbial inoculation, all composite inoculations, especially the three-microbial compound, led to faster organic metabolism, shorter composting process, and improved physicochemical properties of DGW. P. ostreatus growth assays showed the fastest mycelial colonization (20.43 µg·g-1 ergosterol) and extension (9 mm/d), the highest ligninolytic enzyme activities (Lac, 152.68 U·g-1; Lip, 15.56 U·g-1; MnP, 0.34 U·g-1; Xylanase, 10.98 U·g-1; FPase, 0.71 U·g-1), and the highest lignin degradation ratio (30.77%) in the DGW sample after 12 h of composting with the three-microbial compound inoculation when compared to other groups. This sample was relatively abundant in bacteria playing critical roles in amino acid, carbohydrate, energy metabolism, and xenobiotic biodegradation, as suggested by metagenomic analysis. The feed value analysis revealed that P. ostreatus mycelia full colonization in composted DGW led to high fiber content retention and decreased lignin content (final ratio of 5% lignin) but elevated protein concentrations (about 130 g·kg-1 DM). An additional daily weight gain of 0.4 kg/d was shown in cattle feeding experiments by replacing 60% of regular feed with it. These findings demonstrate that compound inoculant consisting of three indigenous microorganisms is efficient to compost DGW and facilitate P. ostreatus growth. P. ostreatus decreased the lignin content of composted DGW during its mycelial growth, improving the quality of DGW for feeding cattle.

Development of a consortium-based microbial agent beneficial to composting of distilled grain waste for Pleurotus ostreatus cultivation.

BACKGROUND: Pleurotus ostreatus is an edible mushroom popularly cultivated worldwide. Distilled grain waste (DGW) is a potential substrate for P. ostreatus cultivation. However, components in DGW restrict P. ostreatus mycelial growth. Therefore, a cost-effective approach to facilitate rapid P. ostreatus colonization on DGW substrate will benefit P. ostreatus cultivation and DGW recycling. RESULTS: Five dominant indigenous bacteria, Sphingobacterium sp. X1, Ureibacillus sp. X2, Pseudoxanthomonas sp. X3, Geobacillus sp. X4, and Aeribacillus sp. X5, were isolated from DGW and selected to develop a consortium-based microbial agent to compost DGW for P. ostreatus cultivation. Microbial agent inoculation led to faster carbohydrate metabolism, a higher temperature (73.2 vs. 71.2 °C), a longer thermophilic phase (5 vs. 3 days), and significant dynamic changes in microbial community composition and diversity in composts than those of the controls. Metagenomic analysis showed the enhanced microbial metabolisms, such as xenobiotic biodegradation and metabolism and terpenoid and polyketide metabolism, during the mesophilic phase after microbial agent inoculation, which may facilitate the fungal colonization on the substrate. In accordance with the bioinformatic analysis, a faster colonization of P. ostreatus was observed in the composts with microbial inoculation than in control after composting for 48 h, as indicated from substantially higher fungal ergosterol content, faster lignocellulose degradation, and higher lignocellulase activities in the former than in the latter. The final mushroom yield shared no significant difference between composts with microbial inoculation and control, with 0.67 ± 0.05 and 0.60 ± 0.04 kg fresh mushroom/kg DGW, respectively (p > 0.05). CONCLUSION: The consortium-based microbial agent comprised indigenous microorganisms showing application potential in composting DGW for providing substrate for P. ostreatus cultivation and will provide an alternative to facilitate DGW recycling.

Complete genome sequence of Clostridium butyricum JKY6D1 isolated from the pit mud of a Chinese flavor liquor-making factory.

Clostridium butyricum is an important fragrance-producing bacterium in the traditional Chinese flavor liquor-making industry. Here the complete genome sequence of C. butyricum JKY6D1 isolated from the pit mud of a Chinese flavor liquor-making factory is presented. The genome is 4,618,327bp with the GC content of 28.74% and a plasmid of 8060bp. This is the first complete genome sequence of C. butyricum strains available so far.