Proposal of Lactobacillus amylovorus subsp. animalis subsp. nov. and an emended description of Lactobacillus amylovorus.

Seven novel lactic acid bacterial strains (BF125T, BF186, TKL145, YK3, YK6, YK10 and NSK) were isolated from the fresh faeces of Japanese black beef cattle and weanling piglets, spent mushroom substrates, or steeping water of a corn starch production plant. These strains are rod-shaped, Gram-stain-positive, non-motile, non-spore-forming, catalase-negative, cytochrome oxidase-negative, facultatively anaerobic, and homofermentative. Strain BF125T did not produce any gas from glucose; both d- and l-lactate were produced as end-products of glucose (D/L, 40 : 60). Growth occurred at 30-45 °C (optimum, 37 °C), pH 5.0-8.0 (optimum, pH 6.0), and with NaCl concentration of 1.0-3.0% (w/v). The G+C content of genomic DNA of strain BF125T was 37.8 mol% (whole-genome analysis). The major fatty acids were C16 : 0, C18 : 1 ω9c, C19 cyclopropane 9, 10, and summed feature 10. The 16S rRNA gene in strain BF125T showed high similarity to that of the type strain of Lactobacillus amylovorus (99.93%), and the other isolates were also identified as L. amylovorus based on these similarities. A phylogenetic tree based on the core genomes of L. amylovorus strains (n=54), including the seven isolates, showed that they could be divided into two clusters. Strains YK3, YK6, YK10, and NSK were in the first cluster, along with the type strain DSM 20531T, while the second cluster included isolates BF125T, BF186, TKL145, and other strains isolated from various animal origins. Phenotypic differences in fermentability were observed for lactose, salicin, and gentiobiose between these two groups. The intergroup digital DNA-DNA hybridization values (72.9-78.6%) and intergroup average nucleotide identity values (95.64-96.92%) were comparable to values calculated using datasets of other valid subspecies of the genus (ex-) Lactobacillus. In light of the physiological, genotypic, and phylogenetic evidence, we propose a novel subspecies of L. amylovorus, named Lactobacillus amylovorus subsp. animalis subsp. nov. (type strain BF125T=MAFF 212522T=DSM 115528T). Our findings also led to the automatic creation of Lactobacillus amylovorus subsp. amylovorus subsp. nov. and an emended description of the species L. amylovorus.

Lactobacillus corticis sp. nov., isolated from hardwood bark.

During a study on the biodiversity of bacteria that inhabit woody biomass, we isolated a strain coded B40T from hardwood bark used as a compost ingredient in Japan. The strain, characterized as B40T, is a Gram-stain-positive, rod-shaped, non-motile, non-spore-forming and catalase-negative bacterium. This novel isolate showed growth at 30-50 °C, at pH 3.5-7.5 and in the presence of up to 4 % (w/v) NaCl. Its major fatty acids include C16:0, C18:1 ω9c and summed feature 8. The genomic DNA G+C content of strain B40T is 42.2 mol%. Results of 16S rRNA gene sequence-based phylogenetic analysis indicated that strain B40T belongs to the genus Lactobacillus and the closest neighbours of strain B40T are Lactobacillus gigeriorum 202T (95.7 %), Lactobacillus pasteurii CRBIP 24.76T (95.6 %), Lactobacillus psittaci DSM 15354T (95.4 %), Lactobacillus fornicalis TV1018T (95.4 %) and Lactobacillus jensenii ATCC 25258T (95.2 %). The amino acid sequence-based phylogenetic analyses of 489 shared protein-encoding genes showed that the strain forms a phylogenetically independent lineage in the genus Lactobacillus but could not be assigned to any known species. Strain B40T has an average nucleotide identify of <70.2 % and a digital DNA-DNA hybridization value of 19.2 % compared with the strains of other closely related Lactobacillus species. Differential genomic, phenotypic and chemotaxonomic properties, in addition to phylogenetic analyses, indicated that strain B40T represents a novel species of the genus Lactobacillus, for which the name Lactobacillus corticis sp. nov. is proposed. The strain type is B40T (=JCM 32597T=DSM 107967T).

Lentilactobacillus fungorum sp. nov., isolated from spent mushroom substrates.

In Japan, during a screening of lactic acid bacteria in spent mushroom substrates, an unknown bacterium was isolated and could not be assigned to any known species. Strain YK48GT is Gram-stain-positive, rod-shaped, non-motile, non-spore-forming and catalase-negative. The isolate grew in 0-4 % (w/v) NaCl, at 15-37 °C (optimum, 30 °C) and at pH 4.0-8.0 (optimum, pH 6.0). The genomic DNA G+C content of strain YK48GT was 42.5 mol%. Based on its 16S rRNA gene sequence, strain YK48GT represented a member of the genus Lentilactobacillus and showed the highest pairwise similarity to Lentilactobacillus rapi DSM 19907T (97.86 %). Phylogenetic analyses based on amino acid sequences of 466 shared protein-encoding genes also revealed that the strain was phylogenetically positioned in the genus Lentilactobacillus but did not suggest an affiliation with previously described species. The average nucleotide identity and digital DNA-DNA hybridization values between strain YK48GT and the type strains of phylogenetically related species were 72.2-76.6% and 19.0-21.2 %, respectively, indicating that strain YK48GT represents a novel species within the genus Lentilactobacillus. Phenotypic data further confirmed the differentiation of strain YK48GT from other members of the genus Lentilactobacillus. According to the results of the polyphasic characterization presented in this study, strain YK48GT represents a novel species of the genus Lentilactobacillus, for which the name Lentilactobacillus fungorum sp. nov. is proposed. The type strain is YK48GT (=JCM 32598T=DSM 107968T).

Lactobacillus salitolerans sp. nov., a novel lactic acid bacterium isolated from spent mushroom substrates.

A taxonomic study of a Gram-stain-positive, rod-shaped, non-motile, non-spore-forming, catalase-negative bacterium, strain YK43T, isolated from spent mushroom substrates stored in Nagano, Japan was performed. Growth was detected at 15-45 °C, pH 5.0-8.5, and 0-10 % (w/v) NaCl. The genomic DNA G+C content of strain YK43T was 43.6 mol%. The predominant fatty acids were C16 : 0, C18 : 1 ω9c and summed feature 8. Based on 16S rRNA gene sequence analysis, the type strains of Lactobacillus acidipiscis (sequence similarity, 97.6 %) and Lactobacillus pobuzihii (97.4 %) were most closely related to YK43T. The average nucleotide identities were 74.1 % between strain YK43T and L. acidipiscis DSM 15836T and 74.0 % between YK43T and L. pobuzihii E100301T. Based on a multilocus sequence analysis, comparative genomic analysis and a range of phenotypic and chemotaxonomic characteristics, strain YK43T represents a novel species of the genus Lactobacillus, for which the name Lactobacillussalitolerans sp. nov. is proposed. The type strain is YK43T (=JCM 31331T = DSM 103433T).

Genetic dissection of black grain rice by the development of a near isogenic line.

Rice (Oryza sativa L.) can produce black grains as well as white. In black rice, the pericarp of the grain accumulates anthocyanin, which has antioxidant activity and is beneficial to human health. We developed a black rice introgression line in the genetic background of Oryza sativa L. 'Koshihikari', which is a leading variety in Japan. We used Oryza sativa L. 'Hong Xie Nuo' as the donor parent and backcrossed with 'Koshihikari' four times, resulting in a near isogenic line (NIL) for black grains. A whole genome survey of the introgression line using DNA markers suggested that three regions, on chromosomes 1, 3 and 4 are associated with black pigmentation. The locus on chromosome 3 has not been identified previously. A mapping analysis with 546 F2 plants derived from a cross between the black rice NIL and 'Koshihikari' was evaluated. The results indicated that all three loci are essential for black pigmentation. We named these loci Kala1, Kala3 and Kala4. The black rice NIL was evaluated for eating quality and general agronomic traits. The eating quality was greatly superior to that of 'Okunomurasaki', an existing black rice variety. The isogenicity of the black rice NIL to 'Koshihikari' was very high.

Deep learning-based flocculation sensor for automatic control of flocculant dose in sludge dewatering processes during wastewater treatment.

In sludge dewatering of most wastewater treatment plants (WWTPs), the dose of polymer flocculant is manually adjusted through direct visual inspection of the flocs without the aid of any instruments. Although there is a demand for the development of automatic control of flocculant dosing, this has been challenging owing to the lack of a reliable flocculation sensor. To address this issue, this study developed a novel image sensor for measuring the degree of flocculation (DF) based on deep learning. Two types of sludge samples were used in the laboratory-scale flocculation tests: excess sludge and mixtures of excess sludge and raw wastewater. To search for a deep learning regression model suitable for DF inference, ten models, including convolutional neural networks, vision transformers, and a multilayer perceptron MLP mixer, were comparatively analysed. The ConvNeXt and MLP mixer models showed the highest accuracies with coefficients of determination (R2) greater than 0.9. The region contributing to the DF inference in the flocculation images was visualised using a modulus-weighted, gradient-weighted regression activation map. A prototype of the flocculation sensor was constructed using an inexpensive EdgeAI device. This device comprises a single-board computer and an integrated graphical processing unit (GPU) and is equipped with a quantised ConvNeXt model. The maximum inference speed of the sensor was 12.8 frames per second (FPS). The flocculation control tests showed that the sensor could control the DF to the target value by adjusting the polymer flocculant dose. Therefore, the flocculation sensor can provide a data-driven approach to the management of the flocculation process, thereby facilitating the automation of WWTPs.

Mapping and validation of QTLs for rice sheath blight resistance.

Sheath blight, caused by Rhizoctonia solani, is one of the most serious diseases of rice. Among 33 rice accessions, mainly from National Institute of Agrobiological Sciences (NIAS) Core Collection, we found three landraces from the Himalayas-Jarjan, Nepal 555 and Nepal 8-with resistance to sheath blight in 3 years' field testing. Backcrossed inbred lines (BILs) derived from a cross between Jarjan and the leading Japanese cultivar Koshihikari were used in QTL analyses. Since later-heading lines show fewer lesions, we used only earlier-heading BILs to avoid association with heading date. We detected eight QTLs; the Jarjan allele of three of these increased resistance. Only one QTL, on chromosome 9 (between markers Nag08KK18184 and Nag08KK18871), was detected in all 3 years. Chromosome segment substitution lines (CSSLs) carrying it showed resistance in field tests. Thirty F2 lines derived from a cross between Koshihikari and one CSSL supported the QTL.

Recovery and utilization of compost heat from an enclosed vertical-type composting facility development of warmed water supply system using compost heat.

This paper describes a compost heat utilization system that recovers the exhaust heat generated by an enclosed vertical-type composting facility (EVCF) installed in a dairy farm with 150 heads of dairy cows. The recovered exhaust heat warms the water supplied to dairy cows. This study determines the temperature of the supplied water and the heat utilization efficiency of the system. During operation, the EVCF generated a compost temperature of 60-70°C and an exhaust temperature of 60°C. The system provided warm (30°C) water throughout the day by raising the water temperature at night when the drinking water requirements were reduced and heat demand used for warming the water was low. Thermal energy generated 356% of the power consumption, and 49.3% of the generated energy was used for warming the water. The compost heat used to warm water was 2017 MJ d-1 , which was equivalent to 68.7 L d-1 of kerosene when the same amount of water was warmed in a boiler.

Quantitation of tocotrienol and tocopherol in various rice brans.

Rice bran is abundant in bioactive compounds including tocotrienol (T3, unsaturated vitamin E). T3 has been reported about its potential functionalities (i.e., antiangiogenic effect), so much attention has been paid on usability of rice bran T3. Hence, we developed a rapid screening method for T3-rich rice bran by one-step equilibrium direct solvent extraction followed by normal phase high-performance liquid chromatography (HPLC). The method gave high-extraction rate of rice bran T3 and tocopherol (above 90%), and the determination of vitamin E by HPLC was completed within 15 min. Using the method, an average of total T3 content in 109 kinds of rice bran samples was 830 mug/g dry wt. Kouchi-Akamai, Joushuu, and Wataribune were found as the T3-rich rice bran varieties (1350-1430 microg T3/g dry wt). According to T3 ratio against total vitamin E (wt %), the average ratio was 61%. Hirayama, Moritawase, and Kaneko were found as the varieties having the highest T3 ratio (80-86%). Since T3 content in Koshihikari rice bran (the leading variety in Japan) was a little above the average, we cross-fertilized Koshihikari with T3-rich varieties and found that T3 content or ratio in F1 was improved compared with Koshihikari. The varieties found rich in T3 could be used for nutraceutical purpose.

Cross-fertilization for enhancing tocotrienol biosynthesis in rice plants and QTL analysis of their F2 progenies.

As rice bran tocotrienol (T3) has been known to have a wide range of physiological functions (e.g., antiangiogenesis), we aimed at developing a T3-rich rice variety for nutraceutical purposes. T3 content in more than 250 kinds of rice bran samples were investigated, and Milyang23 was found as the best variety rich in T3. The variety was therefore chosen for cross-fertilization with Koshihikari. Among obtained F(2) progenies, some of them became improved in T3 content (up to 2-fold of reference Koshihikari). QTL analysis of the F(2) progenies revealed five putative loci corresponding to T3 biosynthesis, in which the main loci were located near a marker RM3827 on chromosome 6. The results show that cross-breeding is effective in improving rice bran T3 and provides more genetic understanding on T3 biosynthesis in rice plants.