Discovery of non-squalene triterpenes.

All known triterpenes are generated by triterpene synthases (TrTSs) from squalene or oxidosqualene1. This approach is fundamentally different from the biosynthesis of short-chain (C10-C25) terpenes that are formed from polyisoprenyl diphosphates2-4. In this study, two fungal chimeric class I TrTSs, Talaromyces verruculosus talaropentaene synthase (TvTS) and Macrophomina phaseolina macrophomene synthase (MpMS), were characterized. Both enzymes use dimethylallyl diphosphate and isopentenyl diphosphate or hexaprenyl diphosphate as substrates, representing the first examples, to our knowledge, of non-squalene-dependent triterpene biosynthesis. The cyclization mechanisms of TvTS and MpMS and the absolute configurations of their products were investigated in isotopic labelling experiments. Structural analyses of the terpene cyclase domain of TvTS and full-length MpMS provide detailed insights into their catalytic mechanisms. An AlphaFold2-based screening platform was developed to mine a third TrTS, Colletotrichum gloeosporioides colleterpenol synthase (CgCS). Our findings identify a new enzymatic mechanism for the biosynthesis of triterpenes and enhance understanding of terpene biosynthesis in nature.

Crystal structure of activating sulfotransferase SgdX2 involved in biosynthesis of secondary metabolite sungeidine.

Microorganisms synthesize a plethora of complex secondary metabolites, many of which are beneficial to human health, such as anticancer agents and antibiotics. Among these, the Sungeidines are a distinct class of secondary metabolites known for their bulky and intricate structures. They are produced by a specific biosynthetic gene cluster within the genome of the soil-dwelling actinomycete Micromonospora sp. MD118. A notable enzyme in the Sungeidine biosynthetic pathway is the activating sulfotransferase SgdX2. In this pathway, SgdX2 mediates a key sulfation step, after which the product undergoes spontaneous dehydration to yield a Sungeidine compound. To delineate the structural basis for SgdX2's substrate recognition and catalytic action, we have determined the crystal structure of SgdX2 in complex with its sulfate donor product, 3'-phosphoadenosine 5'-phosphate (PAP), at a resolution of 1.6 Å. Although SgdX2 presents a compact overall structure, its core elements are conserved among other activating sulfotransferases. Our structural analysis reveals a unique substrate-binding pocket that accommodates bulky, complex substrates, suggesting a specialized adaptation for Sungeidine synthesis. Moreover, we have constructed a substrate docking model that provides insights into the molecular interactions between SgdX2 and Sungeidine F, enhancing our understanding of the enzyme's specificity and catalytic mechanism. The model supports a general acid-base catalysis mechanism, akin to other sulfotransferases, and underscores the minor role of disordered regions in substrate recognition. This integrative study of crystallography and computational modeling advances our knowledge of microbial secondary metabolite biosynthesis and may facilitate the development of novel biotechnological applications.

Lincosamide Antibiotics: Structure, Activity, and Biosynthesis.

Lincosamides are naturally occurring antibiotics isolated from Streptomyces sp. Currently, lincomycin A and its semisynthetic analogue clindamycin are used as clinical drugs. Due to their unique structures and remarkable biological activities, derivatizations of lincosamides via semi-synthesis and biosynthetic studies have been reported. This review summarizes the structures and biological activities of lincosamides, and the recent studies of lincosamide biosynthetic enzymes.

Duration of PCR positivity by type of respiratory virus among children using a multiplex PCR test.

Prolonged positive polymerase chain reaction (PCR) results, irrespective of the transmission risk, can lead to prolonged restrictions on daily activities and infection precaution interventions. Studies evaluating the duration of PCR positivity for multiple pathogens in a single patient cohort are scarce. This study aimed to evaluate and compare the durations of PCR positivity for multiple respiratory viruses among children and adolescents. This retrospective study was conducted between April 2018 and March 2024 using a multiplex PCR respiratory panel for symptomatic children and adolescents who had at least two tests within 90 days of study period, with the first PCR test positive. The rate and likelihood of persistent PCR positivity were evaluated for multiple respiratory viruses. For 1325 positive results, repeat tests were conducted within 90 days. The persistent PCR positivity rate at repeat testing decreased over time (60.6%, Days 1-15 and 21.7%, Days 76-90, after the first test). In multivariate logistic regression analysis, an increased likelihood of persistent PCR positivity was observed for rhinovirus/enterovirus and adenovirus, whereas decreased likelihood of persistent positivity was seen in influenza and seasonal coronaviruses, compared with parainfluenza viruses. Persistent PCR positivity is common for multiple respiratory viruses in symptomatic children.

Structural and Mechanistic Insights into the C-C Bond-Forming Rearrangement Reaction Catalyzed by Heterodimeric Hinokiresinol Synthase.

Hinokiresinol synthase (HRS) from Asparagus officinalis consists of two subunits, α and ß, and catalyzes an unusual decarboxylative rearrangement reaction of 4-coumaryl 4-coumarate to generate (Z)-hinokiresinol with complete stereoselectivity. Herein, we describe the mechanism of rearrangement catalysis and the role played by the heterodimeric HRS, through structural and computational analyses. Our results suggest that the HRS reaction is unlikely to proceed via the previously hypothesized Claisen rearrangement mechanism. Instead, we propose that the 4-coumaryl 4-coumarate substrate is first cleaved into coumarate and an extended p-quinone methide, which then recombine to generate a new C-C bond. These processes are facilitated by proton transfers mediated by the basic residues (α-Lys164, α-Arg169, ß-Lys168, and ß-Arg173) in the cavity at the heterodimer interface. The active site residues, α-Asp165, ß-Asp169, ß-Trp17, ß-Met136, and ß-Ala171, play crucial roles in controlling the regioselectivity of the coupling between the fragmented intermediates as well as the stereoselectivity of the decarboxylation step, leading to the formation of the (Z)-hinokiresinol product.

Mechanistic Analysis of Stereodivergent Nitroalkane Cyclopropanation Catalyzed by Nonheme Iron Enzymes.

BelL and HrmJ are α-ketoglutarate-dependent nonheme iron enzymes that catalyze the oxidative cyclization of 6-nitronorleucine, resulting in the formation of two diastereomeric 3-(2-nitrocyclopropyl)alanine (Ncpa) products containing trans-cyclopropane rings with (1'R,2'R) and (1'S,2'S) configurations, respectively. Herein, we investigate the catalytic mechanism and stereodivergency of the cyclopropanases. The results suggest that the nitroalkane moiety of the substrate is first deprotonated to produce the nitronate form. Spectroscopic analyses and biochemical assays with substrates and analogues indicate that an iron(IV)-oxo species abstracts proS-H from C4 to initiate intramolecular C-C bond formation. A hydroxylation intermediate is unlikely to be involved in the cyclopropanation reaction. Additionally, a genome mining approach is employed to discover new homologues that perform the cyclopropanation of 6-nitronorleucine to generate cis-configured Ncpa products with (1'R,2'S) or (1'S,2'R) stereochemistries. Sequence and structure comparisons of these cyclopropanases enable us to determine the amino acid residues critical for controlling the stereoselectivity of cyclopropanation.

Structure-based engineering of α-ketoglutarate dependent oxygenases in fungal meroterpenoid biosynthesis.

Non-heme iron- and α-ketoglutarate-dependent oxygenases (αKG OXs) are key enzymes that play a major role in diversifying the structure of fungal meroterpenoids. They activate a specific C-H bond of the substrate to first generate radical species, which is usually followed by oxygen rebound to produce cannonical hydroxylated products. However, in some cases remarkable chemistry induces dramatic structural changes in the molecular scaffolds, depending on the stereoelectronic characters of the substrate/intermediates and the resulting conformational changes/movements of the active site of the enzyme. Their molecular bases have been extensively investigated by crystallographic structural analyses and structure-based mutagenesis, which revealed intimate structural details of the enzyme reactions. This information facilitates the manipulation of the enzyme reactions to create unnatural, novel molecules for drug discovery. This review summarizes recent progress in the structure-based engineering of αKG OX enzymes, involved in the biosynthesis of polyketide-derived fungal meroterpenoids. The literature published from 2016 through February 2022 is reviewed.

Prognostic awareness in Japanese patients with advanced cancer: a follow-up cohort study.

BACKGROUND: Patients with advanced cancer have been reported to be more likely to receive goal-concordant care if they have accurate prognostic awareness. However, many patients do not have this awareness. This study aimed to examine the prognostic awareness among Japanese patients with advanced cancer. METHODS: This single-center, follow-up cohort study included Japanese patients with advanced cancer who received chemotherapy at Tohoku University Hospital between January 2015 and January 2016. Patients were surveyed at enrollment and followed up for clinical events for 5 years thereafter. We compared (i) the patients' prognostic awareness with both actual survival time and physician's prediction of survival and (ii) physician's prediction of survival time with actual survival. Factors associated with accurate prognostic awareness were identified by univariate analysis. RESULTS: Of the 133 patients eligible for the study, 57 patients were analyzed. Only 10 (17.5%) patients had accurate prognostic awareness. Forty-three patients (75.4%) were optimistic about their prognosis; >80% of patients were more optimistic than their physicians about their prognosis. The physicians' predictions were accurate in for patients (37.5%). Accurate prognostic awareness was associated with physician's explanation of the prognosis and patients' perception of a good death. CONCLUSIONS: A majority of the patients with advanced cancer in this study had prognostic awareness that was more optimistic in comparison with their actual survival, and most were more optimistic than their physicians about their prognosis. Further research is needed to develop programs to facilitate the discussion of life expectancy with patients in a manner that is consistent with their preferences.

K2P2.1 (TREK-1)-activator complexes reveal a cryptic selectivity filter binding site.

Polymodal thermo- and mechanosensitive two-pore domain potassium (K2P) channels of the TREK subfamily generate 'leak' currents that regulate neuronal excitability, respond to lipids, temperature and mechanical stretch, and influence pain, temperature perception and anaesthetic responses. These dimeric voltage-gated ion channel (VGIC) superfamily members have a unique topology comprising two pore-forming regions per subunit. In contrast to other potassium channels, K2P channels use a selectivity filter 'C-type' gate as the principal gating site. Despite recent advances, poor pharmacological profiles of K2P channels limit mechanistic and biological studies. Here we describe a class of small-molecule TREK activators that directly stimulate the C-type gate by acting as molecular wedges that restrict interdomain interface movement behind the selectivity filter. Structures of K2P2.1 (also known as TREK-1) alone and with two selective K2P2.1 (TREK-1) and K2P10.1 (TREK-2) activators-an N-aryl-sulfonamide, ML335, and a thiophene-carboxamide, ML402-define a cryptic binding pocket unlike other ion channel small-molecule binding sites and, together with functional studies, identify a cation-π interaction that controls selectivity. Together, our data reveal a druggable K2P site that stabilizes the C-type gate 'leak mode' and provide direct evidence for K2P selectivity filter gating.

Functions, Structures, and Engineering of the Teleocidin Biosynthetic Enzymes.

Teleocidins are natural products belonging to the indole alkaloid family and show potent protein kinase C activation activity. The structural feature of teleocidins is an indole-fused nine-membered lactam ring structure. Due to their unique structures and strong biological activities, many total synthesis and biosynthetic studies of teleocidins have been performed. Teleocidin biosynthesis involves interesting enzymatic reactions that are challenging in organic synthesis, including oxidative intramolecular C-N bond-forming reactions, regio- and stereo-selective reverse prenylation reactions, and methylation-triggered terpene cyclization. This review summarizes the recent research on functional and structural analyses, as well as enzyme engineering, of teleocidin biosynthetic enzymes.

Clinical analysis of gender and pre-existing diabetes mellitus in patients with polymyalgia rheumatica: A retrospective study in a Japanese population.

OBJECTIVE: This study clarifies the involvement of gender and pre-existing diabetes mellitus (DM) in the clinical characteristics of polymyalgia rheumatica (PMR). METHODS: The clinical records of patients diagnosed with PMR in our department between January 2011 and June 2021, especially in terms of gender and DM were retrospectively analysed. RESULTS: We identified 89 patients with the median age of 75.37 cases were men and 52 cases were women. Pre-existing DM was found in 21 patients (23.6%). Male PMR patients exhibited a higher complication rate of pre-existing DM and C-reactive protein (CRP) levels at diagnosis (p = .04 and p < .01, respectively) than female patients, and men were more common in the patient group with pre-existing DM (p = .04). The CRP levels of male PMR patients without pre-existing DM were higher than female PMR patients without pre-existing DM. CONCLUSION: Male PMR patients might have a varying pathophysiology from female patients in terms of high inflammation levels accompanied by a high prevalence rate of pre-existing DM and need a gender-specific approach.

Structure-Function Analysis of the S-Glycosylation Reaction in the Biosynthesis of Lincosamide Antibiotics.

The S-glycosyltransferase LmbT, involved in the biosynthesis of lincomycin A, is the only known enzyme that catalyzes the enzymatic incorporation of rare amino acid L-ergothioneine (EGT) into secondary metabolites. Here, we show the structure and function analyses of LmbT. Our in vitro analysis of LmbT revealed that the enzyme shows promiscuous substrate specificity toward nitrogenous base moieties in the generation of unnatural nucleotide diphosphate (NDP)-D-α-D-lincosamides. Furthermore, the X-ray crystal structures of LmbT in its apo form and in complex with substrates indicated that the large conformational changes of the active site occur upon binding of the substrates, and that EGT is strictly recognized by salt-bridge and cation-π interactions with Arg260 and Trp101, respectively. The structure of LmbT in complex with its substrates, the docking model with the EGT-S-conjugated lincosamide, and the structure-based site-directed mutagenesis analysis revealed the structural details of the LmbT-catalyzed SN 2-like S-glycosylation reaction with EGT.

Stereoselectivity and Substrate Specificity of the Fe(II)/α-Ketoglutarate-Dependent Oxygenase TqaL.

Non-heme iron enzymes are versatile catalysts in the biosynthesis of medicinal natural products and have attracted increasing attention as practical catalytic tools in chemical synthesis due to their ability to perform chemically challenging transformations. The Fe(II)/α-ketoglutarate-dependent oxygenase TqaL catalyzes unusual aziridine formation from l-Val via cleavage of the unactivated Cß-H bond. However, the mechanistic details as well as the synthetic potential of TqaL-catalyzed ring closure remain unclear. Herein, we show that the TqaL-catalyzed aziridination of l-Val proceeds with an atypical, mixed stereochemical course involving both the retention and inversion of the C3(Cß) stereocenter. It is also demonstrated that TqaL accepts l-Ile and l-allo-Ile to generate the same diastereomeric pairs of aziridine products via an enzyme-controlled, stereoconvergent process. Our mutagenesis studies reveal that the reaction type (aziridination versus hydroxylation) and the stereochemical outcome are regulated by Ile343 and Phe345. Proper substitutions of Ile343 or Phe345 also make TqaL highly active toward the oxidation of α-amino acid substrates. This work provides mechanistic insights into the stereoselectivity and substrate specificity of the TqaL reactions.

Regional variation in long-term care spending in Japan.

BACKGROUND: Health inequalities are widening in Japan, and thus, it is important to understand whether (and to what extent) there is a regional variation in long-term care (LTC) spending across municipalities. This study assesses regional variation in LTC spending and identifies the drivers of such variation. METHODS: We conducted a cross-sectional study using publicly available municipality-level data across Japan in 2019, in which the unit of analysis was municipality. The outcome of interest was per-capita LTC spending, which was estimated by dividing total LTC spending in a municipality by the number of older adults (people aged ≥ 65). To further identify drivers of regional variation in LTC spending, we conducted linear regression of per-capita spending against a series of demand, supply, and structural factors. Shapley decomposition approach was used to highlight the contribution of each independent variable to the goodness of fit of the regression model. RESULTS: In Fiscal 2019, per-capita LTC spending varied from 133.1 to 549.9 thousand yen (max/min ratio 4.1) across the 1460 municipalities analyzed, showing considerable regional variation. The included covariates explained 84.0% of the total variance in LTC spending, and demand-determined variance was remarkably high, which contributed more than 85.7% of the overall R2. Specifically, the highest contributing factor was the proportion of severe care-need level and care level certification rate. CONCLUSIONS: Our results demonstrate that, even after adjusting for different municipalities' age and sex distribution, there is a large variation in LTC spending. Furthermore, our findings highlight that, to reduce the spending gap between municipalities, the issues underlying large variations in LTC spending across municipalities must be identified and addressed.

Effect of COVID-19 infection related experiences on social behaviors when a state of emergency is declared: a cohort study.

BACKGROUND: Restricting the movement of the public to gathering places and limiting close physical contact are effective measures against COVID-19 infection. In Japan, states of emergency have been declared in specific prefectures to reduce public movement and control COVID-19 transmission. We investigated how COVID-19 infection related experiences including people with a history of infection, people with a history of close contact, and people whose acquaintances have been infected, affected self-restraint from social behaviors during the second state of emergency in Japan. METHODS: A prospective cohort study was conducted among workers aged 20-65 years using data from an internet survey. The baseline survey was conducted on December 22-25, 2020, and a follow-up survey was on February 18-19, 2021. There were 19,051 participants who completed both surveys and were included in the final analysis. We identified eight social behaviors: (1) eating out (4 people or fewer); (2) eating out (5 people or more); (3) gathering with friends and colleagues; (4) day trip; (5) overnight trip (excluding visiting home); (6) visiting home; (7) shopping for daily necessities; and (8) shopping for other than daily necessities. We set self-restraint regarding each social behavior after the second state of emergency was declared in January 2021 as the dependent variable, and COVID-19 infection related experiences as independent variables. Odds ratios were estimated using multilevel logistic regression analyses nested in the prefecture of residence. RESULTS: Significant differences by COVID-19 infection related experiences were identified: compared to people without COVID-19 related experiences, people with a history of COVID-19 were less likely self-restraint from most social behaviors. People whose acquaintance had been diagnosed with COVID-19 were significantly more likely to refrain from most social behaviors. There was no significant difference in any social behaviors for people with a history of close contact only. CONCLUSION: To maximize the effect of a state of emergency, health authorities should disseminate information for each person in the target population, taking into account potential differences related to the infection related experiences.

The medial inclination of the proximal tibia is associated with the external knee adduction moment in advanced varus knee osteoarthritis.

PURPOSE: Whether the inclined articular surface on the medial proximal tibia and the external knee adduction moment (KAM) correlate remains unclear. The hypothesis was that a steeper inclined articular surface correlated with a larger KAM in advanced knee osteoarthritis (OA). METHODS: A total of 44 females (non-OA, 9 knees; early OA, 14 knees; advanced OA, 21 knees; mean age, 58 ± 16 years) were examined. Three-dimensional (3D) assessment was used on biplanar long-leg radiographs and 3D bone models using a 3D to 2D image registration technique. The approximation plane in the proximal tibia was determined using the least-square method. The joint moments were mathematically calculated in a gait analysis, applying a motion capture system and force plates. The main evaluation parameters were the femorotibial angle (FTA), the coronal inclination of the approximation plane in the medial proximal tibia (coronal inclination), and internal knee joint moments. The KAM means the external moments balanced with the internal knee abduction moments. RESULTS: The advanced OA showed a larger internal abduction moment (p = 0.017) at the loading response than the other groups. The larger FTA and steeper coronal inclination correlated with the larger internal abduction moment (FTA, p < 0.001; coronal inclination, p = 0.003) at the loading response. CONCLUSIONS: As the clinical relevance, the association among the coronal inclination of the medial proximal tibia, lower extremity alignment, and KAM is one of the key factors to help better understand the etiology of knee OA. LEVEL OF EVIDENCE: III.

The coronal inclination of the medial tibial plateau affects coronal gait kinematics for varus osteoarthritic knees.

PURPOSE: This study aims to (1) measure the kinematics of lower extremity alignment and the bony position relative to the ground during walking, focusing on the coronal plane, and (2) determine the correlation between the kinematics and coronal inclination of the medial tibial plateau (coronal inclination) for healthy and varus knee osteoarthritis (OA). METHODS: In this study, 43 women (non-OA, 9 knees; early OA, 13 knees; advanced OA, 21 knees; mean age 58 ± 17 years) were examined. The knee phenotypes in varus knee OA were varied. Three-dimensional (3D) knee kinematics were calculated in gait analysis by combining the motion capture system and the 3D lower extremity alignment assessment system via biplanar long-leg X-rays, applying the 3D-2D registration technique. The main parameters were the kinematics of the bony axes relative to the ground in the coronal plane during the stance phase of the gait. The differences in overall kinematics were assessed using repeated measures ANOVA with Tukey's post hoc test. The association between kinematic parameters and coronal inclination was evaluated by multiple linear regression after univariate analysis. RESULTS: The tibia tilted laterally during the loading response, and a plateau area subsequently appeared until the terminal stance phase, whereas the femur slowly tilted laterally until the terminal stance phase. The dynamic alignment showed a relatively large varus angular change during the loading response in all groups. The trend of motion was similar among all groups (p = n.s.), although to varying degrees. The coronal inclination was the more dominant factor than the Kellgren-Lawrence (K-L) grades (ß = - 0.423, p = 0.005) when the change in dynamic alignment was determined. CONCLUSIONS: The TAA plateau area after the loading response implies that the tibial articular surface may become horizontal. The femur slowly tilted laterally until the terminal stance phase in response to the tibial motion. Consequently, the dynamic alignment showed a varus angular change, in which coronal MCT was more involved than K-L grades.

Oral hypofunction and its relation to frailty and sarcopenia in community-dwelling older people.

OBJECTIVE: The purpose of this study was to examine the frequency of oral hypofunction in community-dwelling older people and determine its relationship with frailty and sarcopenia. BACKGROUND: Previous studies have shown that frailty and sarcopenia are associated with decreased oral function. However, these studies have only evaluated frailty or sarcopenia alone and have not evaluated their relationship with each other. MATERIALS AND METHODS: The participants were community-dwelling independent older people in Kyoto. Their oral function evaluation included seven items (oral hygiene, oral dryness, occlusal force, tongue-lip motor function, tongue pressure, masticatory function and swallowing function). Oral hypofunction was defined as abnormalities in at least three of these items. The frailty status was classified into three categories (robust, pre-frail and frail) according to the frailty phenotype and deficit-accumulation models. Sarcopenia was defined according to the Asian Working Group for Sarcopenia (AWGS) Consensus. The relationships between oral function and frailty were analysed using logistic regression analyses, after adjusting for sarcopenia. RESULTS: Among the 340 participants that were analysed (69 men, 271 women; average age: 75.0 years), 182 (53.5%) had oral hypofunction (40 men, 142 women; average age: 76.8 years). There was a significant relationship between oral hypofunction and deficit-accumulation model-assessed frailty, after adjusting for sarcopenia. CONCLUSION: Almost half of the community-dwelling older people have oral hypofunction, which is significantly related to comprehensive frailty and sarcopenia.

COVID-19 vaccination coverage by company size and the effects of workplace vaccination program in Japan: a cohort study.

BACKGROUND: Vaccination is considered the most effective control measure against COVID-19. Vaccine hesitancy and equitable vaccine allocation are important challenges to disseminating developed vaccines. To promote COVID-19 vaccination coverage, the government of Japan established the workplace vaccination program. However, while it appears that the program was effective in overcoming vaccine hesitancy, the program may have hindered the equitable allocation of vaccines because it mainly focused on employees of large companies. We investigated the relationship between company size and COVID-19 vaccination completion status of employees and the impact of the workplace vaccination program on this relationship. METHODS: We conducted an internet-based prospective cohort study from December 2020 (baseline) to December 2021. The data were collected using a self-administered questionnaire survey. Briefly, 27,036 workers completed the questionnaire at baseline and 18,560 at follow-up. After excluding ineligible respondents, we finally analyzed the data from 15,829 participants. At baseline, the participants were asked about the size of the company they worked for, and at follow-up they were asked about the month in which they received their second COVID-19 vaccine dose and the availability of a company-arranged vaccination opportunity. RESULTS: In each month throughout the observation period, the odds of having received a second COVID-19 vaccine dose were significantly lower for small-company employees than for large-company employees in the sex- and age-adjusted model. This difference decreased after adjusting for socioeconomic factors, and there was no significant difference after adjusting for the availability of a company-arranged vaccination opportunity. CONCLUSIONS: The workplace vaccination program implemented in Japan to control the COVID-19 pandemic may have been effective in overcoming vaccine hesitancy in workers; however, it may have caused an inequitable allocation of vaccines between companies of different sizes. Because people who worked for small companies were less likely to be vaccinated, it will be necessary to enhance support of vaccination for this population in the event of future infectious disease outbreaks. TRIAL REGISTRATION: Not applicable.

Association between willingness to receive the COVID-19 vaccine and sources of health information among Japanese workers: a cohort study.

BACKGROUND: It is important to achieve herd immunity by vaccinating as many people as possible to end the COVID-19 pandemic. We investigated the relationship between willingness to receive vaccination and sources of health information among those who did not want to be vaccinated against COVID-19. METHODS: This prospective cohort study collected data using a self-administered questionnaire survey. The baseline survey was conducted during December 22-25, 2020, and the follow-up survey during February 18-19, 2021. Participants were aged 20-65 years and worked at the time of the baseline survey (N = 33,087). After excluding 6,051 invalid responses, we included responses from 27,036 participants at baseline. In total, 19,941 people responded to the follow-up survey (74% follow-up rate). We excluded 7,415 participants who answered "yes" to the question "If a COVID-19 vaccine becomes available, would you like to get it?" in the baseline survey. We finally analyzed 12,526 participants. RESULTS: The odds ratio for change in willingness to be vaccinated from "no" to "yes" differed by source of health information. Compared with workers that used TV as a source of information, significantly fewer people who reported getting information from the Internet and friends/colleagues were willing to get the vaccine. CONCLUSIONS: It is important to approach workers who do not watch TV when implementing workplace vaccination programs. It is likely that willingness to be vaccinated can be increased through an active company policy whereby the top management recommend vaccination, coupled with an individual approach by occupational health professionals. TRIAL REGISTRATION: Not applicable.