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We aimed to evaluate the relationship between hearing ability and cognitive domains and determine how the relationship changes after 6 months of introducing a hearing aid.We conducted a 6-month hearing aid lending study between September 2014 and March 2019, including 59 older participants who visited the Memory Clinic at the National Center for Geriatrics and Gerontology. The hearing level was assessed using pure tone audiometry. Speech intelligibility was measured using the monosyllabic word discrimination score. We assessed the relationship between hearing ability and cognitive domains using the Mini-Mental State Examination (MMSE) total score and four subscale scores (orientation, memory, attention, and language). Differences in the cognitive function between baseline (pre-) and 6 months later (post-) after introducing a hearing aid were also assessed.The pre-orientation score was significantly associated with the pure-tone average (p = 0.013), and the pre-language score was significantly associated with speech intelligibility (p = 0.006) after adjusting for confounders. None of the MMSE subscale scores were significantly different between pre- and post-scores, however, an expectation of improvement with continuous hearing aid use was implied in the attention domain.We found a significant association between hearing ability and cognitive domains in individuals whose cognitive functions were not considered healthy. The presence of a potential relationship between cognitive domains, hearing ability, and auditory compensation is suggested.

Fucoidans are hetero-sulfated polysaccharides that are widely distributed in brown algae and have been extensively studied for their various biological activities. The structure-function relationship of fucoidans remains unclear but can be studied using fucoidan-degrading enzymes (fucoidanases). Here, we isolated and identified Flavobacterium sp. SW as a microbial strain that can grow on fucoidan from Cladosiphon okamuranus as the sole carbon source. Genomic analysis of this strain revealed the presence of two genes, swfct and swfcn2, that are homologous to fct114 from Luteolibacter algae H18 and fcnA from Psychromonas sp. SW5A, respectively. The gene products were produced in Escherichia coli and showed significantly different specificities for fucoidan. Swfct catalyzed the degradation of deacetylated fucoidan from C. okamuranus, and Swfcn2 degraded fucoidans from Saccharina sculpera and Macrocystis pyrifera. The general properties of Swfct were examined by measuring the amounts of reducing ends produced by the enzymatic reaction, and the enzyme properties of Swfcn2 were evaluated by carbohydrate-polyacrylamide gel electrophoresis. Our findings indicate that one microbial strain can harbor genes encoding two different types of fucoidanases.

IntroductionSensory dysfunctions and cognitive impairments are related to each other. Although a relationship between tinnitus and subjective olfactory dysfunction has been reported, there have been no reports investigating the relationship between tinnitus and olfactory test results.MethodsTo investigate the relationship between tinnitus and olfactory test results, we conducted sensory tests, including hearing and visual examinations. The subjects included 510 community-dwelling individuals (295 women and 215 men) who attended a health checkup in Yakumo, Japan. The age of the subjects ranged from 40 to 91 years (mean ± standard deviation, 63.8 ± 9.9 years). The participants completed a self-reported questionnaire on subjective tinnitus, olfactory function, and hearing function, as well as their lifestyle. The health checkup included smell, hearing, vision, and blood examinations.ResultsAfter adjusting for age and sex, the presence of tinnitus was significantly associated with subjective olfactory dysfunction, poor olfactory test results, hearing deterioration, vertigo, and headache. Additionally, high serum calcium levels and a low albumin/globulin ratio were significantly associated with low physical activity and nutrition. Women scored higher than men in olfactory and hearing examinations, but there was no gender difference in vision examinations.ConclusionSubjective smell dysfunction and poor smell test results were significantly associated with tinnitus complaints. Hearing and vision were associated even after adjusting for age and sex. These findings suggest that evaluating the mutual relationships among sensory organs is important when evaluating the influence of sensory dysfunctions on cognitive function.

Geobacillus thermodenitrificans K1041 is an unusual thermophile that is highly transformable via electroporation, making it a promising host for screening genetic libraries at elevated temperatures. In this study, we determined its biological properties, draft genome sequence, and effective vectors and also optimized the electroporation procedures in an effort to enhance its utilization. The organism exhibited swarming motility but not detectable endospore formation, and growth was rapid at 60°C under neutral and relatively low-salt conditions. Although the cells showed negligible acceptance of shuttle plasmids from general strains of Escherichia coli, methylation-controlled plasmids from dam mutant strains were efficiently accepted, suggesting circumvention of a restriction-modification system in G. thermodenitrificans K1041. We optimized the electroporation procedure to achieve efficiencies of 10(3) to 10(5) CFU/μg for five types of plasmids, which exhibited the different copy numbers and segregational stabilities in G. thermodenitrificans K1041. Some sets of plasmids were compatible. Moreover, we observed substantial plasmid-directed production of heterologous proteins in the intracellular or extracellular environments. Our successful construction of a library of promoter mutants using K1041 cells as hosts and subsequent screening at elevated temperatures to identify improved promoters revealed that G. thermodenitrificans K1041 was practical as a library host. The draft genomic sequence of the organism contained 3,384 coding genes, including resA and mcrB genes, which are involved in restriction-modification systems. Further examination revealed that in-frame deletions of resA increased transformation efficiencies, but mcrB deletion had no effect. The ΔresA mutant exhibited transformation efficiencies of >10(5) CFU/μg for some plasmids. IMPORTANCE Geobacillus thermodenitrificans K1041 has yet to be fully characterized. Although it is transformable via electroporation, it rarely accepts Escherichia coli-derived plasmids. This study clarified the biological and genomic properties of G. thermodenitrificans K1041. Additionally, we developed an electroporation procedure resulting in efficient acceptance of E. coli-derived plasmids. This procedure produced transformants using small amounts of plasmids immediately after the ligation reaction. Thus, G. thermodenitrificans K1041 was identified as a host for screening promoter mutants at elevated temperatures. Furthermore, because this strain efficiently produced heterologous proteins, it could serve as a host for screening thermostable proteins encoded in random mutant libraries or metagenomes. We also generated a ΔresA mutant that exhibited transformation efficiencies of >10(5) CFU/μg, which were highest in cases of electroporation-based transformation of Geobacillus spp. with E. coli-derived plasmids. Our findings provide a new platform for screening diverse genetic libraries at elevated temperatures.

Olfactory impairment is associated with cognitive impairment. However, differences between subjective and objective olfactory impairment measurements in older adults and their relationship with cognitive impairment are unclear.We assessed the relationship between cognitive and olfactory impairments via a single-center observational study in 2020. Forty-seven individuals who visited our memory clinic were enrolled and asked to complete objective and subjective olfactory tests. Participants had dementia, mild cognitive impairment, or normal cognition. We administered the Open Essence (a simple card-type odor identification test) and self-administered odor questionnaire to assess objective and subjective olfaction, respectively, and the Mini-Mental State Examination to assess cognitive function.Older age was related to decreased Open Essence scores (P 0.001). Compared with the mild cognitive impairment and normal cognition groups, the dementia group had significantly lower Open Essence scores (P 0.001). Cognitive impairment was also associated with decreased Open Essence scores after adjusting for age, sex, and education (P 0.001). Participants with dementia did not obtain lower self-administered odor questionnaire scores than those with normal cognition, and they tended to be unaware of their olfactory impairment.Open Essence scores significantly decreased according to the degree of cognitive impairment. Participants with dementia were unaware of their olfactory impairment, as indicated by odor questionnaire scores. Objective and subjective scores of olfaction differed in participants with dementia. Concomitant assessment of objective and subjective olfaction is useful to screen older adults with both olfactory and cognitive impairments. Geriatr Gerontol Int 2022; 22: 924-929.

The bio-logging technique is extensively used in the fields of ecology and ethology, wherein a data logger, such as a sensor or camera, is attached to the target animal’s body to collect the required data. In this method, the efficiency of recovery of the data logger is not ideal. In this study, we proposed a new recovery method, with the aim of addressing the aforementioned problem in bio-logging. The authors previously fabricated a data-logger separator, which weighed approximately 10 g, and was targeted at small seabirds. Because there were some problems associated with the circuit board and the separation performance of this device, we modified the device to overcome the previous drawbacks. We fabricated a flexible printed circuit to improve the operation of the mounted actuator and wireless microcomputer, and improve the efficiency of the fabrication process. We conducted an experiment to determine the proper length and position at which the actuator is attached, in order to achieve a stable motion. We thus fabricated a new prototype with these improvements and performed an operational test at low temperatures from a particular distance, simulating actual usage in a natural environment. The results demonstrated that separation occurred without failure, thus indicating that the separator can be efficiently used in practical environment.

Obesity and aging are known to affect the skeletal muscles. Obesity in old age may result in a poor basement membrane (BM) construction response, which serves to protect the skeletal muscle, thus making the skeletal muscle more vulnerable. In this study, older and young male C57BL/6J mice were divided into two groups, each fed a high-fat or regular diet for eight weeks. A high-fat diet decreased the relative gastrocnemius muscle weight in both age groups, and obesity and aging individually result in a decline in muscle function. Immunoreactivity of collagen IV, the main component of BM, BM width, and BM-synthetic factor expression in young mice on a high-fat diet were higher than that in young mice on a regular diet, whereas such changes were minimal in obese older mice. Furthermore, the number of central nuclei fibers in obese older mice was higher than in old mice fed a regular diet and young mice fed a high-fat diet. These results suggest that obesity at a young age promotes skeletal muscle BM formation in response to weight gain. In contrast, this response is less pronounced in old age, suggesting that obesity in old age may lead to muscle fragility.

Aim Olfactory impairment as a prodromal symptom, as well as sarcopenia, frailty and dependence as geriatric syndromes, is often associated with cognitive decline in older adults with progression of Alzheimer's disease. The present study aimed to evaluate the associations of olfactory and cognitive decline with these geriatric syndromes, and with structural changes of the brain in older adults.

In ecology and ethology, a technique called ‘bio-logging’ has been used to record the behaviour of wild animals by attaching a small logger, which includes a global positioning system (GPS) or acce...

An examination of taste and olfactometry in Yakumo-Cho inhabitants’ examination carried out in 2005, and the result reported in Academia Journal of Medicinal Plants 2018. This study examined olfactory function. A personal function test is calculated from the Yakumo study database, and the odor stick identification tests administered to healthy older adults. The participants were community dwellers who voluntarily participated in the Yakumo Study and had managed everyday life by themselves. The participants were engaged in a variety of jobs, not only white-collar but also agriculture, fishery,and forestry. Therefore, the city regarded as a representative of today's Japanese society. From the database, 298 participants (169 females and 129 males) were selected form data in August, 2019. The Odor Stick Identification Test (OSIT-J) was used to assess odor perception. The aromas used in the OSIT-J include curry, perfume, Japanese cypress, India ink, menthol, rose, wood, Smelly socks/ sweat, fried garlic, condensed milk, gas for cooking, and Japanese mandarin aromas.

Gustatory function is closely related to chewing and swallowing; however, there are currently no reports regarding gustatory function in persons with cerebral palsy (CP). The aim of this study was to compare the gustatory function between persons with CP and healthy controls. We investigated sweet, salty, sour and bitter tastes using the whole-mouth method and measured the electrogustometric thresholds in the chorda tympani nerve area. Twelve participants with CP (6 women and 6 men) for whom gustatory testing was possible at our facilities, and 17 age-matched controls (9 women and 8 men) were included. The mean age ± standard deviation was 58.6 ± 8.1 years and 58.5 ± 8.7 years in subjects with CP and controls, respectively. Taste detection and identification were significantly worse in persons with CP compared with the controls. Taste identification was more impaired than taste detection. At the highest concentration, taste identification was impossible 11 times out of 48 (12 persons × 4 kinds of tastes) in persons with CP but such a deficit was not observed in 68 attempts (17 persons × 4 kinds of tastes) involving controls. The electrogustometric thresholds were not significantly different between the groups. Gustatory function associated with chewing and swallowing is worse in persons with CP compared to the controls.

We investigated the associations between hearing aids (HA) and the maintenance of cognitive function among community-dwelling older adults with moderate hearing loss. A total of 407 participants aged 60 years or older with moderate hearing loss were recruited from the National Institute for Longevity Sciences, Longitudinal Study for Aging (NILS-LSA). Moderate hearing loss was defined as a pure-tone average of 40–69 dB at 500, 1000, 2000, and 4000 Hz of the better ear, according to the definition proposed by the Japan Audiological Society. Cognitive function was evaluated using the four subtests of the Japanese version of the Wechsler Adult Intelligence Scale-Revised Short Forms (WAIS-R-SF): Information, Similarities, Picture completion, and Digit Symbol Substitution (DSST). A longitudinal analysis of 1192 observations with a mean follow-up period of 4.5 ± 3.9 years was performed. The HA use rate at any time during the follow-up period was 31.4%, and HA users were significantly younger (t-test, p = 0.001), had worse hearing (p < .0001) and higher education (p = 0.001), participated more frequently in the survey (p < .0001), and were less depressed (χ2 test, p = 0.003) than the older adults not using HA. General linear mixed models consisted of the fixed effects of HA use, follow-up time, and an HA use × time interaction term adjusted for age and pure-tone average thresholds at baseline, sex, education, and other possible confounders. HA use showed significant main effects on the scores for Picture completion and DSST after adjustment; scores were better in the HA use group than in the no HA use group. The HA use × time interaction was significant for the Information score (p = 0.040). The model-predicted 12-year slope with centralizing age indicated that the no HA use group showed greater decline over time on Information scores than did HA use group. The slopes did not differ between HA users and non-users for the Similarities, Picture completion and DSST. In conclusion, HA use may have a protective effect on the decline in general knowledge in older adults with moderate hearing loss.

Geobacillus kaustophilus is a thermophilic bacterium that grows at temperatures ranging between 42 and 74 °C. Here, we modified this organism to produce the thermolabile protein (PyrFA) or its thermostable variant (PyrFV) and analyzed the transcriptome and growth efficiency profiles of the resultant strains. In the producer of PyrFA, the transcriptome profile was changed to facilitate ATP synthesis from NADH without pooling reduced quinones. This change implies that PyrFA production at elevated temperatures places an energy burden on cells potentially to maintain protein homeostasis. This was consistent with the observation that the PyrFA producer grew slower than the PyrFV producer at > 45 °C and had a lower cellular fitness. Similar growth profiles were also observed in the PyrFA and PyrFV producers derived from another thermophile (Geobacillus thermodenitrificans) but not in those from Escherichia coli at 30 °C. Thus, we suggest that the production of thermolabile proteins impairs host survival at higher temperatures; therefore, thermophiles are under evolutionary selection for thermostable proteins regardless of whether their functions are associated with survival advantages. This hypothesis provides new insights into evolutionary protein selection in thermophiles and suggests an engineering approach to select thermostable protein variants generated via random gene mutagenesis.

Geobacillus kaustophilus HTA426 is a thermophilic bacterium whose genome harbors numerous insertion sequences (IS). This study was initially conducted to generate mutant genes for thermostable T7 RNA polymerase in G. kaustophilus; however, relevant experiments unexpectedly identified that the organism transposed multiple IS elements and produced derivative cells that expressed a silent gene via transposition. The transposed elements were diverse and included members of the IS4, IS701, IS1634, and ISLre2 families. The transposition was relatively active at elevated temperatures and generated 4–9 bp of direct repeats at insertion sites. Transposition was more frequent in proliferative cells than in stationary cells but was comparable between both cells when sigX, which encodes an extra-cytoplasmic function sigma factor, was forcibly expressed. Southern blot analysis indicated that IS transposition occurred under growth inhibitory conditions by diverse stressors; however, IS transposition was not detected in cells that were cultured under growth non-inhibitory conditions. These observations suggest that G. kaustophilus enhances IS transposition via sigX-dependent stress responses when proliferative cells were prevented from active propagation. Considering Geobacillus spp. are highly adaptive bacteria that are remarkably distributed in diverse niches, it is possible that these organisms employ IS transposition for environmental adaptation via genetic diversification. Thus, this study provides new insights into adaptation strategies of Geobacillus spp. along with implications for strong codependence between mobile genetic elements and highly adaptive bacteria for stable persistence and evolutionary diversification, respectively. This is also the first report to reveal active IS elements at elevated temperatures in thermophiles and to suggest a sigma factor that governs IS transposition.

Hearing loss is a risk factor for cognitive impairment, and the use of a hearing aid (HA) may prevent cognitive decline alongside hearing loss. We aimed to elucidate the prevalence of self-reported HA usage in Japanese community-dwelling elders with hearing loss, and the effect of hearing and HA on cognitive impairment.A total of 1193 participants, who had audiometric defined hearing loss and were aged 60 years or over, had their cumulative 3260 observations followed up for 10 years from a large cohort of a Japanese study. Association between hearing (pure-tone average threshold level at 500, 1000, 2000, and 4000 Hz from the better hearing ear: PTABHE) and HA usage with cognitive impairment (total score of Mini-Mental State Estimation was under 27 or diagnosed as dementia) was analyzed using generalized estimating equations.The HA usage rate of the 1193 community-dwelling elders with hearing loss was 6% during the first involvement. The majority (59.2%) of HA users always used an HA. HA usage rate was 0.7% for the mild hearing loss group and 32.4% for the moderate or greater hearing loss group in the latest participating wave. PTABHE was significantly associated with cognitive impairment (odds ratio for every 10 dB 1.36; 95% CI 1.21-1.53, p0.0001) after adjusting for age, sex, education, depressed mood, smoking status, alcohol intake, income, activity, obesity, histories of hypertension, dyslipidemia, ischemic heart disease, diabetes, stroke, ear disease, and occupational noise exposure. PTABHE was also significantly associated with cognitive impairment in the mild hearing loss group (odds ratio for every 10 dB 1.34; 95% CI 1.05-1.72, p = 0.020) and moderate hearing loss group (odds ratio for every 10 dB 1.82; 95% CI 1.27-2.61, p = 0.001). HA use showed a significant suppressive effect on cognitive impairment in those with moderate hearing loss who always use an HA (odds ratio 0.54; 95% CI 0.30-1.00, p = 0.049).The prevalence of HA usage among Japanese community-dwelling elders with hearing loss is consistent, at around 10%. The hearing level remained a primary risk factor for cognitive impairment among elders with hearing loss after adjusting for several confounding factors. Regular HA use may have a protective effect on cognitive impairment in those with moderate hearing loss.

Gaussian Process Regression (GPR) is one of popular supervised learning techniques that map input data sequence into the continuous outputs with Gaussian probability distribution. Since GPR can quantify uncertainty in the predicted values unlike other machine learning techniques, it is becoming used to predict behaviors of mission critical applications such as satellites or gas turbines. However, its computation requirement is very high due to the extensive use of matrix operations. In this paper, we focus on the mean value prediction part as a major time-consuming one of Gaussian process regression and attempt to implement a custom hardware accelerator on FPGAs using high-level synthesis technique. Assuming the target dataset is onedimensional time series data, we design the accelerator specific to the single feature. Following the reference design using Python’s scikit-learn, we prepare a straightforward implementation in C++, and optimize it for an FPGA with high-level synthesis technique. From the results of implementation on the Ultra96 MPSoC board, we find that our accelerator can achieve a 250x speedup and 177 times less energy compared with the reference Python code for the CPU on the target board.

Unravelling the secrets of wild animals is one of the biggest challenges in ecology, with bio-logging (i.e., the use of animal-borne loggers or bio-loggers) playing a pivotal role in tackling this challenge. Bio-logging allows us to observe many aspects of animals’ lives, including their behaviours, physiology, social interactions, and external environment. However, bio-loggers have short runtimes when collecting data from resource-intensive (high-cost) sensors. This study proposes using AI on board video-loggers in order to use low-cost sensors (e.g., accelerometers) to automatically detect and record complex target behaviours that are of interest, reserving their devices’ limited resources for just those moments. We demonstrate our method on bio-loggers attached to seabirds including gulls and shearwaters, where it captured target videos with 15 times the precision of a baseline periodic-sampling method. Our work will provide motivation for more widespread adoption of AI in bio-loggers, helping us to shed light onto until now hidden aspects of animals’ lives., Joseph Korpela et al. demonstrate the use of machine-learning assisted bio-loggers on black-tailed gulls and streaked shearwaters. As video recording is only activated through variations in movement detected by low-cost accelerometers, this method represents improvements to runtime and precision over existing bio-logging technology.

Pelagic seabirds have evolved physiological and morphological characteristics that enable them to fly in an energy efficient manner over the oceans. Although there are some exceptions (e.g., Delord et al. 2020), most species come ashore only to breed or rest. However, recent tracking technologies have revealed some unexpected overland journeys. For example, brown pelicans Pelecanus occidentalis (Lamb et al. 2017), Arctic terns Sterna paradisaea (Redfern & Bevan 2020), and the focal species of this study, fledglings of streaked shearwaters Calonectris leucomelas (Yoda et al. 2017a; Fig. 1a), are known to migrate overland.

Understanding the relationships among aging, cognitive function, and olfaction may be useful for diagnosing olfactory decline in older adults. Olfactory function declines in the early stage of neurodegenerative diseases, including Alzheimer's and Parkinson's diseases. Aging and cognitive impairment are associated with olfactory decline. Moreover, the assessment of hyposmia and anosmia is paramount to the diagnosis of neurodegenerative diseases. We aimed to assess the relationships among aging, cognitive function, and olfaction in patients with olfactory impairment.This observational study included 141 patients with olfactory deterioration who presented with dementia, mild cognitive impairment (MCI), age-related hyposmia, or postviral olfactory dysfunction (PVOD). The patients underwent TT olfactometry, a self-administered odor questionnaire (SAOQ), a visual analog scale (VAS), and a Mini-Mental State Examination.TT odor recognition thresholds decreased with aging (p0.01) and cognitive impairment (p 0.08). The average TT recognition thresholds were 5.1,4.6,4.2, and 3.7 in dementia, MCI, age-related hyposmia, and PVOD, respectively. Moreover, the average differences between the detection and recognition thresholds were 3.7, 2.8, 2.3, and 2.0 in dementia, MCI, age-related hyposmia, and PVOD, respectively. Hyposmia with dementia presented the highest recognition thresholds (p 0.05) and the largest differences between the TT detection and recognition thresholds, compared with age-related hyposmia and PVOD (p 0.05). Hyposmia with dementia had the highest SAOQ and VAS scores compared with the other groups (p 0.05).The possibility of dementia should be investigated in patients with hyposmia, including those with high TT recognition thresholds, a large difference between the TT detection and recognition thresholds, and high SAOQ and VAS scores.

The genus Geobacillus comprises gram-positive, moderate thermophilic, and aerobic or facultative anaerobic strains that can form endospores. The members of the genus are widely distributed in nature and have diverse habitats including cool environments. This is potentially due to not only their capacity for endospore formation but also their genetic plasticity that facilitates environmental adaptation, because Geobacillus spp. show remarkable genome diversification and have acquired beneficial properties that facilitate their exploitation of respective habitats. Such properties are often attractive for whole-cell applications performed at elevated temperatures. Consequently, extensive studies have been conducted to develop genetic tools for Geobacillus spp. and to generate their derivatives that are more practical for whole-cell applications. With the aim of highlighting the biotechnological platforms of Geobacillus spp., this chapter reviews genetic tools, notable properties, and biotechnological studies on Geobacillus spp. along with their history, habitats, and general characteristics. We also explore different perspectives from relevant studies.

Fucoidan is a hetero-sulfated polysaccharide found in brown algae and has received much attention as an ingredient in functional and health foods. The marine bacterial strain Luteolibacter algae H18 degrades fucoidan from Cladosiphon okamuranus. We purified the fucoidanase from a cell-free extract of L. algae H18, used it to decrease the molecular weight of deacetylated-fucoidan, determined the N-terminal amino acid sequence of the enzyme, and identified the gene involved in the degradation of fucoidan, fct114, in a draft genome sequence of strain H18. The gene product was heterologously produced in Escherichia coli and demonstrated to catalyze the degradation of deacetylated-fucoidan into lower molecular weight fragments. The mass of the gene product Fct114 is 112 kDa (1026 amino acid residues). The general properties of the enzyme were investigated by measuring the amount of reducing ends produced from deacetylated-fucoidan during the reaction. The enzyme was inactive toward fucoidans from other brown seaweed species or toward polysaccharides such as alginic acid, carrageenan, hyaluronic acid, and chondroitin sulfate. The amino acid sequence of Fct114 shared less than 25% identity and had no conserved motifs when compared with previously identified fucoidanases from other marine bacterial strains. These data suggest that Fct114 is a novel polysaccharide-degrading enzyme.

The genus Geobacillus comprises thermophilic bacilli capable of endospore formation. The members of this genus provide thermostable proteins and can be used in whole cell applications at elevated temperatures; therefore, these organisms are of biotechnological importance. While these applications have been described in previous reviews, the present paper highlights the environmental adaptations and genome diversifications of Geobacillus spp. and their applications in evolutionary-protein engineering. Despite their obligate thermophilic properties, Geobacillus spp. are widely distributed in nature. Because several isolates demonstrate remarkable properties for cell reproduction in their respective niches, they seem to exist not only as endospores but also as vegetative cells in diverse environments. This suggests their excellence in environmental adaptation via genome diversification; in fact, evidence suggests that Geobacillus spp. were derived from Bacillus spp. while diversifying their genomes via horizontal gene transfer. Moreover, when subjected to an environmental stressor, Geobacillus spp. diversify their genomes using inductive mutations and transposable elements to produce derivative cells that are adaptive to the stressor. Notably, inductive mutations in Geobacillus spp. occur more rapidly and frequently than the stress-induced mutagenesis observed in other microorganisms. Owing to this, Geobacillus spp. can efficiently generate mutant genes coding for thermostable enzyme variants from the thermolabile enzyme genes under appropriate selection pressures. This phenomenon provides a new approach to generate thermostable enzymes, termed as thermoadaptation-directed enzyme evolution, thereby expanding the biotechnological potentials of Geobacillus spp. In this review, we have discussed this approach using successful examples and major challenges yet to be addressed.

Indigo is an insoluble blue dye historically used for dyeing textiles. A traditional approach for indigo dyeing involves microbial reduction of polygonum indigo to solubilize it under alkaline conditions; however, the mechanism by which microorganisms reduce indigo remains poorly understood. Here, we aimed to identify an enzyme that catalyzes indigo reduction; for this purpose, from alkaline liquor that performed microbial reduction of polygonum indigo, we isolated indigo carmine-reducing microorganisms. All isolates were facultative anaerobic and alkali-tolerant Bacillus spp. An isolate termed AO1 was found to be an alkaliphile that preferentially grows at pH 9.0–11.0 and at 30–35 °C. We focused on flavin-dependent azoreductase as a possible enzyme for indigo carmine reduction and identified its gene (azoA) in Bacillus sp. AO1 using homology-based strategies. azoA was monocistronic but clustered with ABC transporter genes. Primary sequence identities were

Seaweeds are a nonlignocellulosic biomass, but they are often abundant in unique polysaccharides that common microbes can hardly utilize; therefore, polysaccharide degradation is key for the full utilization of seaweed biomass. Here, we isolated 13 thermophiles from seaweed homogenates that had been incubated at high temperature. All of the isolates were Gram-positive and preferentially grew at 60-70 °C. Most formed endospores and were tolerant to seawater salinity. Despite different sources, all isolates were identical regarding 16S rRNA gene sequences and were categorized as Geobacillus thermodenitrificans. Their growth occurred on seaweed polysaccharides with different profiles but required amino acids and/or vitamins, implying that they existed as proliferative cells by utilizing nutrients on seaweed viscous surfaces. Among 13 isolates, strain OS27 was further characterized to show that it can utilize a diverse range of seaweed polysaccharides and hemicelluloses. Notably, strain OS27 degraded raw seaweeds while releasing soluble saccharides. The degradation seemed to depend on enzymes that were extracellularly produced in an inducible manner. The strain could be genetically modified to produce heterologous endoglucanase, providing a transformant that degrades more diverse seaweeds with higher efficiency. The draft sequences of the OS27 genome contained 3766 coding sequences, which included intact genes for 28 glycoside hydrolases and many hypothetical proteins unusual among G. thermodenitrificans. These results suggest that G. thermodenitrificans OS27 serves as a genetic resource for thermostable enzymes to degrade seaweeds and potentially as a microbial platform for high temperature seaweed biorefinery via genetic modification.

Stress-induced mutagenesis can assist pathogens in generating drug-resistant cells during antibiotic therapy; however, if and how antibiotics induce mutagenesis in microbes remains poorly understood. A non-pathogenic thermophile, Geobacillus kaustophilus HTA426, efficiently produces derivative cells resistant to rifampicin and streptomycin via rpoB and rpsL mutations, respectively. Here, we examined this phenomenon to suggest a novel mutagenic mode induced by antibiotics. Fluctuation analysis indicated that mutations occurred via spontaneous mutations during culture. However, mutations were much more frequent in growing cells than stationary cells, and mutation sites were varied through cell growth. These observations suggested that growing cells induced mutagenesis in response to antibiotics. An in-frame deletion of mfd, which governs transcription-coupled repair to correct DNA lesions on the transcribed strand, caused mutations that were comparable between growing and stationary cells; therefore, the mutagenic mechanism was attributable to DNA repair defects where growing cells depressed mfd function. Mutations occurred more frequently at optimal growth temperatures for G. kaustophilus than at a higher growth temperature, suggesting that the mutagenesis relies on active cellular activities rather than high temperature-associated DNA damage. In addition, the mutagenesis may involve a mutagenic factor targeting these sites, in addition to mfd depression, because rpoB and rpsL mutations were dominant at thymine and guanine sites on the transcribed strand. A similar mutagenic profile was observed for other Geobacillus and thermophilic Bacillus species. This suggests that Bacillus-related thermophiles commonly induce mutagenesis in response to rifampicin and streptomycin to produce resistant cells.

Physical characteristics: Original contains illustrations, Accepted: 2018-05-15, 資料番号: PA1920002000

Objective The influence of living alone on multiple food and nutrition behaviors across a range of ages and genders has not been sufficiently investigated. Moreover, to our knowledge, no studies have described both dietary status and medical examination findings in persons living alone. Among individuals who attended a health checkup in a rural town in Japan, we investigated dietary habits and medical examination findings in persons living alone and those living with one or more other persons. Methods The participants in this investigation were 501 community-dwelling individuals aged 40 to 91 y (mean, 63.8 ± 9.9 y). Thirty-four (16.4%) of the 207 men and 45 (15.3%) of the 294 women lived alone. Dietary intake frequency of 28 types of foods and drinks, including various vegetables, fruits, meat, seafood, and dairy products, was investigated based on responses to a self-completed questionnaire. During the physical examination, body weight, body mass index, body fat percentage, and blood pressure (systolic and diastolic) were measured. Blood examination included red blood cell count, white blood cell count, platelet count, hemoglobin, hematocrit, total protein, albumin/globulin ratio, total cholesterol, triglyceride, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, creatinine, blood urea nitrogen, uric acid, and serum calcium. Results Multivariate analysis revealed that living alone independently was associated with a reduced intake frequency of various vegetables and seafood, especially in men. Diastolic blood pressure and triglyceride levels were significantly higher in men living alone than in men living with other persons. The albumin/globulin ratio in women living alone was significantly lower than in women living with other persons. Conclusions It was found that living alone was associated not only with a reduced quality of dietary intake but also with an unfavorable trend in some of the medical examination findings.

Sulfur compounds in fossil fuels are a major source of environmental pollution, and microbial desulfurization has emerged as a promising technology for removing sulfur under mild conditions. The enzyme TdsC from the thermophile Paenibacillus sp. A11-2 is a two-component flavin-dependent monooxygenase that catalyzes the oxygenation of dibenzothiophene (DBT) to its sulfoxide (DBTO) and sulfone (DBTO2) during microbial desulfurization. The crystal structures of the apo and flavin mononucleotide (FMN)-bound forms of DszC, an ortholog of TdsC, were previously determined, although the structure of the ternary substrate–FMN–enzyme complex remains unknown. Herein, we report the crystal structures of the DBT–FMN–TdsC and DBTO–FMN–TdsC complexes. These ternary structures revealed many hydrophobic and hydrogen-bonding interactions with the substrate, and the position of the substrate could reasonably explain the two-step oxygenation of DBT by TdsC. We also determined the crystal structure of the indole-bound enzyme because TdsC, but not DszC, can also oxidize indole, and we observed that indole binding did not induce global conformational changes in TdsC with or without bound FMN. We also found that the two loop regions close to the FMN-binding site are disordered in apo-TdsC and become structured upon FMN binding. Alanine substitutions of Tyr-93 and His-388, which are located close to the substrate and FMN bound to TdsC, significantly decreased benzothiophene oxygenation activity, suggesting their involvement in supplying protons to the active site. Interestingly, these substitutions increased DBT oxygenation activity by TdsC, indicating that expanding the substrate-binding site can increase the oxygenation activity of TdsC on larger sulfur-containing substrates, a property that should prove useful for future microbial desulfurization applications.

Geobacillus spp. are moderate thermophiles that have great potential for use in diverse applications. For effective utilization of the species, genetic tools have been extensively studied; however, an overexpression vector remains to be developed. Here we constructed a plasmid vector that can shuttle between Escherichia coli and Geobacillus spp., and which contained a maltose-inducible promoter from Geobacillus kaustophilus HTA426. Although the vector (termed pGKE119) was originally designed for basal gene expression, it surprisingly directed robust protein production in G. kaustophilus. Protein production essentially occurred in an auto-inducible manner without maltose; however, some proteins were produced more efficiently in the presence of maltose. Although the productivity was affected by culture conditions, three proteins were successfully produced with abundance ratios of 12–27% (on a total protein basis) and yields of 77–170 mg (per L culture). pGKE119 directed substantial protein production even in Geobacillus subterraneus, Geobacillus thermoglucosidasius, and Geobacillus thermoleovorans. This suggests that pGKE119 can use a range of Geobacillus spp. as hosts and widely expand their genetic toolbox. Because Geobacillus spp. are highly proliferative bacteria that are distinct from organisms used as protein production hosts, pGKE119 may also provide a novel platform for hyperproduction of recombinant proteins.

The anhydrofructose pathway is an alternate pathway for glycogen degradation by α-1,4-glucan lyase. The sugar 1,5-anhydro-D-fructose (1,5-AF) acts as the central intermediate of this pathway, but its physiological role of in mammals is unclear. Glycation reactions forming advanced glycation end-products (AGEs) are important in the development of complications of diabetes mellitus. We hypothesized that 1,5-AF may contribute to cellular damage by forming 1,5-AF-derived AGEs (AF-AGEs) with intracellular proteins. To clarify the role of 1,5-AF in protein modification, we created a novel antibody targeting AF-AGEs. Serum albumin modified by AF-AGEs was prepared by incubating rabbit serum albumin (RSA) or bovine serum albumin (BSA) with 1,5-AF. After immunizing rabbits with AF-AGEs-RSA, affinity chromatography of anti-AF-AGE antiserum was performed on a Sepharose 4B column coupled with AF-AGEs-BSA or N-(carboxymethyl)/N-(carboxyethyl)lysine-BSA. A novel immunopurified anti-AF-AGE antibody was obtained and was characterized using a competitive enzyme-linked immunosorbent assay. Then an AF-AGEs assay was established using this immunopurified antibody. This assay was able to detect AF-AGEs in human and animal serum samples. Finally, intracellular accumulation of AF-AGEs was shown to be associated with damage to cultured hepatocytes (HepG2 cells). This is the first report about in vivo detection of AF-AGEs with a novel structural epitope.

Animal-borne data loggers, i.e., biologgers, allow researchers to record a variety of sensor data from animals in their natural environments (Hussey et al. 2015; Kays et al. 2015). This data allows biologists to observe many aspects of the animals’ lives, including their behavior, physiology, social interactions, and external environment. However, the need to limit the size of these devices to a small fraction of the animal’s size imposes strict limits on the devices’ hardware and battery capacities (Kays et al. 2015). Here we show how AI can be leveraged on board these devices to intelligently control their activation of costly sensors, e.g., video cameras, allowing them to make the most of their limited resources during long deployment periods. Our method goes beyond previous works that have proposed controlling such costly sensors using simple threshold-based triggers, e.g., depth-based (Watanuki et al. 2007; Volpov et al. 2015) and acceleration-based (Nishiumi et al. 2018; Brown et al. 2012) triggers. Using AI-assisted biologgers, biologists can focus their data collection on specific complex target behaviors such as foraging activities, allowing them to automatically record video that captures only the moments they want to see. By doing so, the biologger can reserve its battery power for recording only those target activities. We anticipate our work will provide motivation for more widespread adoption of AI techniques on biologgers, both for intelligent sensor control and intelligent onboard data processing. Such techniques can not only be used to control what is collected by such devices, but also what is transmitted off the devices, such as is done by satellite relay tags (Cox et al. 2018).

How do wild animals behave as they get older? We investigated the foraging behavior of black-tailed gulls (Larus crassirostris) of known age using GPS and acceleration loggers. We found that the older gulls reduced their mean flight speed and tended to feed on prey on land rather than on prey at sea. However, breeding performance of gulls did not change with age. Therefore, older gulls might experience senescence, but optimize their foraging behavior in relation to their age to maintain breeding performance.

Background and Purpose: In the horizontal canal benign paroxysmal positional vertigo (BPPV), cupulolithiasis shows apogeotropic direction changing nystagmus lasting more than 1 min, while canalolithiasis leads to geotropic direction changing nystagmus lasting < 1 min. The difference between cupulolithiasis and canalolithiasis is widely accepted to be the attachment of the displaced otoconia to the cupula of a semicircular canal. Several studies have shown a relationship between BPPV and vitamin D deficiency, but no studies have compared serum levels of vitamin D between canalolithiasis and cupulolithiasis patients. The purpose of this study was to clarify the difference in vitamin D serum level between canalolithiasis and cupulolithiasis of the horizontal canal. Methods: This retrospective study included 20 and 15 patients with canalolithiasis and cupulolithiasis of the horizontal canal, respectively. Serum levels of 25-hydroxyvitamin D [25(OH)D] during the acute phase of BPPV were measured. Results: The mean 25(OH)D serum level in patients with canalolithiasis and cupulolithiasis was 13.2 ± 1.4 and 20.4 ± 1.6 ng/mL, respectively, and the difference was statistically significant (p = 0.0014), also after adjusting for age and sex (p = 0.0351). Eighteen out of 20 (90%) and 5 of 15 (33%) patients were diagnosed with vitamin D deficiency in the canalolithiasis and cupulolithiasis groups, respectively, and this difference was also statistically significant (p = 0.0005). Conclusion: We found that serum vitamin D level in patients with canalolithiasis was significantly lower than that in patients with cupulolithiasis of the horizontal canal.

Revealing the route selection of wild animals is of fundamental importance in understanding their movements and foraging strategy. In this study, we attached GPS loggers to black-tailed gulls Larus crassirostris and recorded their movement trajectories during their foraging trips. Using inverse reinforcement learning (IRL), we analyzed the factors that affected their route selection. During the training phase, using pre-defined feature maps, we estimated a reward map that may affect the decision making of black-tailed gulls. The reward map can be used for predicting the trajectories of the gulls during the test phase. In addition, the resultant weight vector enabled us to analyze to which degree the black-tailed gulls favor each area.

Azoreductases reductively cleave azo linkages by using NAD(P)H as an electron donor. The enzymes are widely found in bacteria and act on numerous azo dyes, which allow various unique applications. This review describes primary amino acid sequences, structures, substrates, physiological roles, and biotechnological applications of bacterial azoreductases to discuss their remarkable diversification. According to primary sequences, azoreductases were classified phylogenetically into four main clades. Most members of clades I-III are flavoproteins, whereas clade IV members include flavin-free azoreductases. Clades I and II prefer NADPH and NADH, respectively, as electron donors, whereas other members generally use both. Several enzymes formed no clades; moreover, some bacteria produce azoreductases with longer primary structures than those hitherto identified, which implies further diversification of bacterial azoreductases. The crystal structures commonly reveal the Rossmann folds; however, ternary structures are moderately varied with different quaternary conformation. Although physiological roles are obscure, several azoreductases have been shown to act on metabolites such as flavins, quinones, and metal ions more efficiently than on azo dyes. Considering that many homologs exclusively act on these metabolites, it is possible that azoreductases are actually side activities of versatile reductases that act on various substrates with different specificities. In parallel, this idea raises the possibility that homologous enzymes, even if these are already defined as other types of reductases, widely harbor azoreductase activities. Although azoreductases for which their genes have been identified are not abundant, it may be simple to identify azoreductases of biotechnological importance that have novel substrate specificities.

Effective utilization of microbes often requires complex genetic modification using multiple antibiotic resistance markers. Because a few markers have been used in Geobacillus spp., the present study was designed to identify a new marker for these thermophiles. We explored antibiotic resistance genes functional in Geobacillus kaustophilus HTA426 and identified a thiostrepton resistance gene (tsr) effective at 50 °C. The tsr gene was further used to generate the mutant tsrH258Y functional at 55 °C. Higher functional temperature of the mutant was attributable to the increase in thermostability of the gene product because recombinant protein produced from tsrH258Y was more thermostable than that from tsr. In fact, the tsrH258Y gene served as a selectable marker for plasmid transformation of G. kaustophilus. This new marker could facilitate complex genetic modification of G. kaustophilus and potentially other Geobacillus spp.

Release from a high-altitude balloon is a cost-effective means of achieving supersonic flight, but there have been only two prior applications of this method to aircraft flight testing. The Japanese Aerospace Exploration Agency’s D–SEND#2 flight experiment is the latest of this kind of test. Due to the one-shot nature of the experiment, it was not sufficient to guarantee the stability of the test vehicle, but it was also necessary to achieve the mission objectives on the first flight. Accordingly, we performed an inclusive pre-flight evaluation of D–SEND#2 including the flight controller and operating conditions. This approach was very effective at enhancing the probability of mission success under off-design flight conditions. The actual flight results presented in this paper demonstrate the effectiveness of this approach.