This study aimed to analyze the hearing improvement and prognosis factors of idiopathic sudden sensorineural hearing loss (ISSNHL) in different ages with initial total hearing loss.We reviewed the medical records of 5,711 hospitalized patients with ISSNHL from 2016 to 2021 in our center. All of the patients had been treated with uniform combination drug therapy. After excluding the patients with initial partial hearing loss and those diagnosed with clear etiology, 188 patients were enrolled in this study and divided into six age groups . In all groups, decreases in pure-tone average (PTA) 1 month posttreatment, effective rate, and clinical characteristics  were analyzed.Among the 188 enrolled patients, 86% had vertigo. Complete recovery was seen in 0.5% of patients, and marked recovery was seen in 43% of patients. The mean 1 month posttreatment PTAs were as follows: 18–30 years: 80 ± 7.5 dB; 31–40 years: 100 ± 9.0 dB; 41–50 years: 99 ± 8.3 dB; 51–60 years: 101 ± 8.6 dB; 61–70 years: 96 ± 9.6 dB; and ≥ 71 years: 88 ± 13.0 dB. Compared with the other groups, the 18–30- years group showed better recovery of hearing threshold in five frequencies , and the recovery of hearing threshold at 0.25 and 0.5 Hz was better than the recovery at 1, 2, and 4 kHz. According to the results of the chi-square test statistical analysis, vertigo and comorbidities were associated with a poor prognosis of ISSNHL.In summary, the treatment outcomes of patients with ISSNHL with initial total hearing loss were poor. There was a significant age-related difference with respect to marked recovery 1 month posttreatment, and the 18–30- years group showed better recovery than the other age groups. Idiopathic sudden sensorineural hearing loss (ISSNHL) is an otologic emergency. Hearing loss and vertigo are the main complaints associated with it. ISSNHL has been defined as a sensorineural hearing loss > 30 dB over at least 3 contiguous audiometric frequencies within 3 days or less with no identifiable causes . DespiteIdiopathic sudden sensorineural hearing loss is a heterogeneous disease in terms of clinical symptoms , causes , the severity of hearing loss , and prognosis. Profound ISSNHL has a particularly poor prognosis, regardless of the treatment type and hospital stay. Although numerous studies concerning severe to profound ISSNHL have been published , most diTherefore, we evaluated hearing improvement after treatment in different-age patients who presented with total hearing loss. We paid special attention to the improvement in the pure-tone average (PTA) 1 month posttreatment.This retrospective study included patients with initial total hearing loss due to ISSNHL hospitalized at the Department of Otologic Center, Shandong Provincial ENT Hospital between January 2016 and August 2021. Some patients were referred from other hospitals for further treatment in our department because of poor recovery. This study was approved by the hospital ethics review board and performed in accordance with the Declaration of Helsinki.The patient inclusion criteria were as follows: (1) age ≥ 18 years, (2) diagnosis of unilateral ISSNHL, where the initial pure-tone audiometry produced no responses , (3) follow-up ≥ 1 month, and (4) treatment for ≥ 14 days.The exclusion criteria were as follows: (1) bilateral sudden hearing loss, (2) age < 18 years, (3) history of hearing loss, (4) middle ear disease, and (5) other distinct diagnoses related to known causes of rapid hearing loss, such as injury, labyrinthine fistula, cytomegalovirus infection, Hunt syndrome, psychogenic deafness, Meniere’s disease, congenital and genetic hearing loss (large vestibular aqueduct), systemic immunological disease, vestibular Schwannoma, and noise-induced hearing loss.All patients were hospitalized for ≥ 14 days . The combination therapy administered was as follows: (1) intravenous drip dexamethasone: 10 mg/day consecutive for 3 days and 5 mg/day consecutive for 4 days; (2) postauricular injection of methylprednisolone (40 mg) once every 2 days until discharge; (3) Ginkgo biloba extract: 105 mg/day intravenous drip for 14 days; and (4) monosialoganglioside 80 mg/day intravenous drip for 7 days. After discharge, the patients received oral mecobalamin and Ginkgo tablets.The patients gave a full medical history and underwent a comprehensive audiological evaluation that includes otoscopic examination, pure-tone audiometry , tympanometry (Grason Stadler Tympstar), distortion products otoacoustic emissions , auditory brainstem responses , and other systematic examinations for hearing impairment during hospitalization. Inner ear magnetic resonance imaging (MRI) was performed on a 3T clinical scanner . A 3D-FLAIR sequence and a 3D-T2WI sequence were employed. The following variables that might influence improvement were recorded: age, sex, affected side, onset season, time between ISSSHL onset and the start of treatment, hospital stay, vertigo, tinnitus, MRI of the inner ear, arterial hypertension, diabetes, dyslipidemia, and thyroid disorders.Pure-tone averages  at octave or semioctave frequencies under air conduction and 0.25–4 kHz under bone conduction were measured two times per week during treatment and once every 2 weeks after the completion of treatment. The average air-conduction PTAs were obtained through the thresholds at frequencies of 0.25, 0.5, 1, 2, and 4 kHz, respectively, following the “Kanzaki criteria” ; Table 1p < 0.05.Statistical analyses were performed using SPSS statistical software (version 26). The Fisher’s exact test and the chi-square test were applied to categorical data, variance analysis to quantitative data, and the Mann–Whitney and Kruskal–Wallis tests to quantitative and ordinal data. Chi-square test was conducted to assess the effects of different variables on hearing prognosis. Statistical significance was determined at a confidence level of n = 5,365), (2) bilateral ISSNL (n = 8), (3) age < 18 years (n = 35), (4) previous hearing loss (n = 3), (5) middle ear disease (n = 3), (6) injury (n = 2), (7) cytomegalovirus infection (n = 1), (8) Hunt syndrome (n = 1), (9) psychogenic deafness (n = 3), (11) large vestibular aqueduct (n = 3), (12) vestibular Schwannoma (n = 2), (13) lack of 30-day follow-up after treatment (n = 94), and (14) inner ear malformation (n = 3). Finally, 188 patients were enrolled and divided into 6 age groups   the initial hearing loss was not total (: n = 8) . All patn = 94). There were no significant differences in side of involvement, sex, time from onset to treatment (≤ 7 days is not listed in Patient demographics are summarized in p = 0.009). When comparing the marked recovery (II) status in all groups, there were significant differences between the following groups (p = 0.001), 18–30 and 41–50-year groups , and 18–30 and 51–60-years groups . The chi-square test was used to compare the recovery  and non-recovery groups  among the six age groups. There was a significant difference in the 18–30-years  and 31–40-years  groups and the 18–30 and 51–60-years groups . However, there was no significant difference in the other groups. There were no statistical differences between the 18–59 and ≥ 60 y in I, II, III, and IV or in I + II, III + IV . The mean 1 month posttreatment PTAs in the six groups were as follows: 18–30 years: 80 ± 7.5 dB; 31–40 years: 100 ± 9.0 dB; 41–50 years: 99 ± 8.3 dB; 51–60 years: 101 ± 8.6 dB; 61–70 years: 96 ± 9.6 dB; and ≥ 71 years: 88 ± 13.0 dB. The Kruskal–Wallis test showed a significant difference among all the six age groups (g groups : 18–30- p = 0.001); the hearing gain in the 18–30-years group was higher than in the other groups . The recovery of hearing at 0.25 and 0.5 Hz was better than the recovery at 1, 2, and 4 kHz. The mean PTA in all groups 1 month posttreatment is shown in Comparisons of hearing gain at five frequencies  revealedp = 0.014) and comorbidity (p = 0.000) were scored as prognostic factors , including those aged less than18 years, presented with total initial hearing loss. We excluded 35 patients aged less than 18 years because the recovery rate of ISSNHL in pediatric patients is higher than in adults , and twoThere was no statistical difference in sex, affected side, and tinnitus; a result matching those of previous studies . A signin = 8). Considering the hearing improvement and the cost and adverse effects of the drugs used, these average PTAs indicate that more medication and longer treatment duration should not be applied to patients with an initial no-response audiogram after more than 2 weeks of treatment in the hospital.In this study, the subjects’ mean 1 month posttreatment 5-pure-tone hearing thresholds showed poor improvement in general . One (0.The mean hearing thresholds showed better improvement in the 18–30-years group than in the other five groups , 5. ThisIn all patients, 0.25 and 0.5 kHz responses improved more than the 1, 2, and 4 kHz responses . AccordiOur study had several limitations. First, it was a single-center retrospective study, which could have introduced bias; this bias could be reduced by consistent hearing tests and treatments being performed by the same clinicians. Second, the age group that is 71 or more than 71 y included eight patients only; as the main inclusion criterion was a complete lack of response at the initial pure-tone threshold, we had few patients aged 71 or more than 71 years, and we will continue collecting the data of these patients in our ward. We divided the patients into six groups per 10 years because we wanted to know how many dBHL lead to hearing improvement in different ages. Third, subtotal hearing loss was not set as a control group. We will conduct further research regarding this topic. Fourth, 94 patients were lost to the 1-month follow-up. The patients lost trust and were not monitored continuously because of their poor recovery. Self-testing using smartphones, technology similar to that used for detecting middle ear fluid using smartphones, would be more convenient for patients . This maWe reviewed the treatment outcomes and hearing status of large patient groups that suffered from ISSNHL with initial total hearing loss. Our findings showed that the mean PTAs 1 month posttreatment in different age groups were still severe to profound. Although there was a significant age-related difference with respect to marked recovery 1 month posttreatment, and the 18–30-years group showed better recovery than the other age groups; considering the hearing improvement and the cost and adverse effects of the drugs used, these average PTAs suggest that more medication and longer treatment duration should not be used for patients with an initial no-response audiogram.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.Ethical review and approval was not required for the study on human participants in accordance with the local legislation and institutional requirements. Written informed consent from the patients/participants or patients/participants legal guardian/next of kin was not required to participate in this study in accordance with the national legislation and the institutional requirements.WX: conceptualization and writing original draft preparation. QD: statistical analysis. YW, ZH, KL, XS, and FD: data collection. HW: conceptualization and funding acquisition. DZ: conceptualization, supervision, writing and reviewing. MW: conceptualization, supervision, writing, reviewing, and editing. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


In the United States, up to two-thirds of older adults have hearing loss. Untreated hearing loss can have significant health outcomes, yet less than 20% of adults with hearing loss use hearing aids. In this study, we examined potential factors associated with hearing aid use, including detailed measures of health status, access to care, patient engagement, and technology use, in a nationally representative sample of Medicare beneficiaries.Cross-sectional study using the 2017 Medicare Current Beneficiary Survey. Participants with self-reported hearing loss were included. The primary outcome was hearing aid use. Factors potentially associated with hearing aid use included: sociodemographics, health determinants, access to care, patient activation, and technology access/use.Overall, 5,146 participants were included. Of them, 27% reported using hearing aids. In a multivariable logistic regression model, predisposing factors associated with greater odds of hearing aid use included older age, identifying as a man, identifying as White, having completed college, having 3 or more chronic conditions, having dementia, not having trouble seeing, not having limitations in activities of daily living, having moderate relative to low information-seeking scores, and having a personal computer at home (range of odds ratios [ORs]: 1.22–4.46). Enabling factors associated with greater odds of hearing aid use included higher income, living alone relative to living with family members other than a spouse, and having a usual source of care (range of ORs: 1.43–1.54).In addition to addressing previously identified factors associated with hearing aid use, improving access to health care, technology, and information about hearing aids may improve the uptake of hearing aids. These findings help further inform our understanding on how to address low treatment levels of hearing loss in the community by identifying new populations to target and potentially modifiable risk factors for hearing aid use. Translational Significance: Health care behaviors such as having a routine place for health care and seeking health information may translate into hearing aid uptake among those with hearing loss. Medical providers who are more forthcoming with information on hearing loss and treatments may help bridge this access gap.Hearing loss affects up to two-thirds of older adults in the United States . It is aDespite this, less than 20% of adults with hearing loss in the United States use hearing aids . A key bPatient engagement and health-seeking behaviors may be key factors that fuel health behaviors . HoweverThe MCBS is an ongoing survey of the Medicare program’s impact on beneficiaries. Analyses were limited to participants who reported hearing difficulty, defined through their use of a hearing aid or not using a hearing aid but having a little or a lot of trouble hearing. Participants were asked: “Do you use a hearing aid? Yes, no,” followed by: “Which statement best describes your hearing [with a hearing aid]? No trouble, a little trouble, a lot of trouble.”The primary outcome was self-reported hearing aid use . Covariates were identified using the Andersen–Aday Behavioral Model of Health Services Use framework, which describes drivers of health care utilization fitting into one of the following categories: predisposing characteristics, enabling factors, and perceived need . PredispA multivariable logistic regression model examined the association of hearing aid use with the predisposing characteristics and enabling factors. Coefficients were exponentiated to odds ratios (ORs) for ease of interpretation. To consistently report on factors associated with greater (rather than lower) odds of hearing aid use, ORs below 1.00 were converted to their inverse and categories were reversed when describing the results in the text. Survey weights were used to account for the MCBS sampling design and survey nonresponse. Analyses were conducted using Stata/SE (Version 14). The Institutional Review Board of NORC at the University of Chicago approved the MCBS study.In total, 5,146 participants were included, representing a weighted sample of 21,341,218 Medicare beneficiaries with self-reported hearing loss. Overall, 42.2% were 75 years and older, 51.9% were women, and 86.9% identified as White. Less than a third (27.0%) reported using hearing aids. A greater proportion of participants who used hearing aids were 75 years and older, men, and identified as White .Greater odds of hearing aid use were associated with being between the ages 65 and 74 years relative to younger than 65 years , and being 75 years or older relative to being between the ages 65 and 74 years , identifying as a man relative to a woman , identifying as White relative to Black , having completed college relative to not having completed high school , having three or more chronic conditions , having dementia relative to not , not having trouble with vision relative to having trouble , not having limitations in ADLs relative to having one limitation , having moderate information-seeking scores relative to low scores , and having a personal computer at home relative to not , living alone relative to living primarily with children/other family members , and having a usual source of care relative to not having one  were associated with higher odds of hearing aid use. Urban area of residence and having hearing insurance coverage were not associated with hearing aid use .In a nationally representative sample of Medicare beneficiaries with hearing loss, only 27% reported using hearing aids. In addition to previously identified factors associated with hearing aid use, we found that having access to technology, a usual source of care and moderate relative to low health information-seeking scores were associated with greater odds of use.Having a usual source of care has been shown to have a positive effect on health care utilization, including preventive care  were associated with greater odds of hearing aid use, but functional impairments  were associated with lower odds of use. This is consistent with general patterns of health care use, whereby disability is generally associated with access barriers , and mulIn our study, access to technology  was associated with greater odds of hearing aid use, but technology use (using the Internet to get information) was not. Prior studies have demonstrated that people with higher technology commitment scores, defined as better technology competence, acceptance, and control, were more likely to use hearing aids . TechnolConsistent with previous studies, we found that older age, identifying as White, having higher educational attainment, and higher income were associated with hearing aid use, and living in an urban versus rural area was not associated with hearing aid use . We alsoOur study has limitations that should be taken into account when interpreting the results. First, our outcome measure does not distinguish between regular and nonregular hearing aid use, or hearing aid ownership. Second, only those with self-reported hearing trouble or hearing aid use were included in the study; thus, results may not be generalizable to those who do not know that they could benefit from hearing aids. Finally, temporality between the variables assessed and hearing aid use cannot be established based on these cross-sectional data.In conclusion, in addition to the previously reported factors associated with hearing aid use, we found that technology use and health care behaviors such as information seeking and having a usual source of care may influence hearing aid uptake. These findings could help better target those who may benefit from interventions to improve hearing aid use.


Positionally, fullbacks and centre forwards in a 4–3–3 formation exhibited the greatest frequency of high-intensity accelerations and decelerations. Very large differences were observed between the frequency of high-intensity decelerations compared to accelerations in games won (g = 2.37), drawn (g = 2.99) and lost (g = 3.59). The highest team frequencies of high-intensity accelerations (n = 3330) and decelerations (n = 6482) were completed in games won. Conclusions: The frequency of high-intensity accelerations and decelerations has a significant impact on match performance outcomes in an elite English League Two football team. Consideration needs to be given to specific conditioning and recovery strategies to optimise high-intensity acceleration and deceleration performance in games. Caution should be taken as these findings are representative of one team within the EFL.Objectives: Previous research has highlighted the frequency of high-intensity accelerations and decelerations in elite football. The influence of these actions on match performance outcomes has not been established. The aim of the present study was to identify the influence of high-intensity accelerations and decelerations on match performance outcomes . Comparisons were also made between team and positional high-intensity accelerations and decelerations recorded within the games. Methods: 26 elite outfield footballers from an elite English Football League (EFL) Two team completed the present study. Global Positioning System (GPS) technology was utilised to quantify high-intensity accelerations and decelerations during 45 games in a competitive season. Magnitude analysis and the effects of results, positions and fixture periods were observed. Results: Significant effects of results, periods and positions were observed ( The high-performance sport of football and its demand for optimum levels of physical, technical, tactical and mental ability from players is well established ,2. FootbOf recent interest is the effect of high-intensity accelerations and decelerations on the physiological and mechanical demands and performance outcomes of players ,12. ThisThe influence of high-intensity decelerations on player load demands during match play is of particular note as high eccentric braking actions seem to account for the highest magnitude of mechanical load per metre than any other match play locomotor activity . AccordiDespite advancements in periodisation and injury prevention practices in elite sports, incidence rates of non-contact injuries in elite football have not improved . To our n = 12) to minimise the effect of formation on physical demands. Players were presented with information of the project protocol and provided informed consent for the use of match data in accordance with the Helsinki Declaration. All data was anonymised prior to data analysis to ensure player confidentiality. Ethical approval was provided by the host university (STEMH 0094).Twenty-six elite outfield football players from an English Football League team  participated in the study during the 2019/2020 season. The sample included five centre forwards, four centre halves, seven central midfielders, five fullbacks and five wingers. The formation of the team was consistent throughout all games analysed and represented a 4–3–3. In total, 57 competitive games were completed in this season. Games where formation was changed from the 4–3–3 were excluded (n = 45) using global positioning analysis (GPS) units  to provide data at 10 Hz. Mean high-intensity accelerations and decelerations were derived from PlayerLoad™ instantaneously. Reliability of this system was previously reported, indicating excellent intra-device reliability. ICC values ranged from 0.77 (95% CI: 0.62–0.89) (very large) to 1.0 (95% CI: 0.99–1.0) (nearly perfect) [External workload data was collected from competitive matches (perfect) . The uni−2 for accelerations and <−3 m/s−2 for decelerations were respectively utilised [Physical parameters of interest which were analysed in this study included frequency of accelerations and decelerations for each player during matches. Pre-determined thresholds criteria of >3 m/sutilised .2 and decelerations <3 m/s2. Following each game, data were downloaded into Catapult, Australian specialised analysis software . Occurrence of accelerations and decelerations above or below the relative thresholds set were extracted for mid-week games, Saturday games following a mid-week game and Saturday games in isolation that were not preceded with a mid-week game across the season and utilised for analysis.Data were collected for analysis from each GPS unit, including accelerations >3 m/sg corrected effect size with 95% confidence intervals (95% CI), calculated using an online statistical spreadsheet [Differences in total team frequency of high-intensity accelerations and decelerations across matches with win, draw and lost performance outcomes were examined using Hedges’ eadsheet  and inteeadsheet  as: trivη2) were calculated to estimate effect sizes for all significant main effects and interactions. As recommended by Cohen [Further statistical analyses were conducted using IBM Statistical Package for the Social Sciences  with the statistical significance accepted at the 0.05 level. A univariate analysis of variance (ANOVA) was conducted to quantify main effects for result, period and position. Interaction effects were also quantified and any significant main effects associated with result, period and position were explored using post hoc pairwise comparisons with a Bonferroni correction factor. The assumptions associated with the statistical model were assessed to ensure model adequacy. To assess residual normality for each dependant variable, Q–Q plots were generated using stacked standardised residuals. Scatterplots of the stacked unstandardised and standardised residuals were also utilised to assess the error of variance associated with the residuals. Mauchly’s test of sphericity was also completed for all dependent variables, with a Greenhouse–Geisser correction applied if the test was significant. Partial eta squared , Saturday following a midweek game (n = 15) and a Saturday game where no midweek game preceded it (n = 15)). The mean and standard deviation (SD) are reported for each game type. The total number of accelerations and decelerations performed (>3 m/s2 and <−3 m/s2) are reported for all games along with mean and SD for all games (n = 45). Further to this, 2 and <−3 m/s2, respectively, in relation to game type-highlighting outputs for wins, draws or losses. Finally, 2 and <−3 m/s2) over 45 competitive League Two fixtures.g = 2.37–3.59) between the frequencies of high-intensity decelerations compared to accelerations, with the greatest difference observed in matches lost (g = 3.59). The highest total team frequency of both high-intensity accelerations and decelerations were completed during matches won (185 ± 48 and 360 ± 90) in comparison to games drawn (146 ± 37 and 291 ± 55) and lost (152 ± 31 and 326 ± 59), respectively. Trivial-to-moderate differences (g = 0.17–0.86) were observed between high-intensity accelerations during matches with win, draw and lost performance outcomes. Similar effect size differences were also observed when comparing the frequency of high-intensity decelerations between matches won, drawn and lost (g = 0.43–0.86).Total team frequency and effect size differences between high-intensity accelerations and decelerations completed during matches with win, draw and lost performance outcomes are presented in F = 2.286, p = 0.01, η2 = 0.14; Decelerations: F = 1.474, p = 0.04, η2 = 0.08) and game type . There was a result x game type interaction for accelerations , and no interaction was identified for decelerations (p > 0.05). The data set was collapsed to consider the result and game type in isolation. Significant differences were identified between wins and draws  and wins and losses  for both accelerations and decelerations. No significant differences were identified between draws and losses (p > 0.05). Significant differences were identified for accelerations from mid-week games to the Saturday following a mid-week game (p = 0.01) and mid-week games compared to Saturday games which were not preceded by a mid-week game (p = 0.01). Decelerations only displayed significant differences between mid-week games and the subsequent Saturday games (p = 0.002).Analysis of accelerations and decelerations over 45 games in the season identified significant effects for result . The collapsing of the data set to consider positions in isolation detailed that fullbacks and centre forwards outputted significantly higher accelerations and decelerations than any of the other positions (p ≤ 0.05) .Positional analysis of accelerations and decelerations identified a significant effect of position . Additionally, significantly more high-intensity accelerations and decelerations were displayed when games were won, supporting the proposed hypothesis. Interestingly, the present study also highlighted that the frequency of high-intensity accelerations and decelerations performed by the team was reduced in periods of fixture congestion, suggesting potential fatigue effects. Finally, analysis of positional effects in a 4–3–3 formation highlighted that fullbacks and centre forwards outputted significantly more high-intensity accelerations and decelerations than any other position. Although these present findings support previous work detailing that match outcome strongly influences physical performance metrics, differences do exist in relation to positional outputs . This diOur findings show that high-intensity decelerations were performed much more frequently than high-intensity accelerations in all matches regardless of the match performance outcome . These findings agree with previous studies that have also reported high-intensity decelerations (of varying thresholds) to be significantly higher than equivalent intense accelerations during elite football match play ,15,19. FImportantly, significantly more accelerations and decelerations were exhibited by the team when games were won compared to drawn or lost. Interestingly, however, no significant differences were found between games drawn and lost. Accordingly, the significant moderate-to-large effect size for ‘result’ seems to support a greater frequency of accelerations and decelerations being performed in winning games. These findings may be best explained by the chaotic nature of the game and the increasing demand to perform high-intensity actions over a 90-minute period when both in and out of possession in order to successfully impact the match performance outcome ,33. The Findings within the current body of work further emphasised a moderate effect for game type. Saturday games following a mid-week fixture displayed significantly lower high-intensity accelerations and decelerations than the preceding mid-week game. Contextualising this information against the results presents some interesting findings. Notably, for Saturday games that followed a mid-week fixture, the team only won 1 game out of the 15 games that appeared in this scenario. This is a notable effect with average team high-intensity accelerations and decelerations both being shown to be significantly lower in these Saturday games. This is also observed when analysing the Saturday games that were completed in isolation and did not follow a mid-week fixture. Fixture-congested periods have been shown to have no effect on performance outcome measures such as total distance ; howeverThese considerations have important ramifications for players that lack the capacity to perform these important high-intensity actions, particularly decelerations, during fixture congested periods. For example, a reduced deceleration capacity would result in players not being able to perform sharp, suddenly imposed change of direction/speed manoeuvres, which could lead to potential detriments in team tactical goals  and overall performance outcomes, whilst increasing injury risk ,33,36. Tn = 10) and mean scores highlighted the highest frequency of high-intensity accelerations and decelerations occurred in this game type. Effect sizes for results were reported as small-to-moderate. That being said, all players exhibited significantly higher acceleration and deceleration outputs in games when the team won, again aligning with the overall team findings described. Collectively, these findings further highlight the importance of high-intensity accelerations and decelerations to match performance outcomes and the importance of closely monitoring positional differences in these metrics to ensure optimal preparation of players for forthcoming fixtures.Positional analysis over the 45 games investigated in the current study identified that fullbacks and centre forwards exhibit significantly more high-intensity accelerations and decelerations than any other positions. Effect sizes for positions were highlighted as being moderate to large, indicating a strong association between high-intensity acceleration and deceleration output and position. These findings agree with Tierney et al.  who repo2) and decelerations (<−3m/s2) is of particular importance [To our knowledge, this is the first study to specifically examine the effect of high-intensity accelerations and decelerations on match outcome. Although a variety of acceleration and deceleration thresholds have been examined previously, the significance of high-intensity accelerations (>3m/sportance ,15. Thesportance . They alportance ,35. Thatportance  whilst dHigh-intensity accelerations and decelerations are important match actions that allow players to adapt to tactical demands, win duals and create/defend goal attempts, all critical for match outcome. This is the first study to investigate the influence of high-intensity accelerations and decelerations on team performance over a competitive season in an English League Two football team. Importantly, our findings suggest there is an impact of these high-intensity actions on team performance, in that outputs were highest when the team won the game and lowest during draws or losses. Across the competitive season, players performed more high-intensity declarations than accelerations. Thus, in contemporary soccer match play particular consideration may be needed to prepare players for the demands of high-intensity decelerations. In addition, the positional demands of high-intensity accelerations and decelerations are important to consider when preparing and conditioning players for competition throughout the season. Our findings have important implications for practitioners and provide evidence for the utility of these important metrics during the competitive season to optimise team performance.


Caenorhabditis elegans olfactory neuron amphid wing “C” cell (AWC), while two opposite cGMP signaling pathways are responsible for odor-sensing in olfactory neuron amphid wing “B” cell (AWB): (1) a depolarizing Gα (GPA-3)- phosphodiesterase (PDE) – cGMP pathway which can be activated by low concentrations of isoamyl alcohol (IAA), and (2) a hyperpolarizing Gα (ODR-3)- GC- cGMP pathway sensing high concentrations of IAA. Besides, IAA induces Gα (ODR-3)-TRPV(OSM-9)-dependent currents in amphid wing “A” cell (AWA) and amphid neuron “H” cell with single ciliated sensory ending (ASH) neurons with different thresholds. Our results demonstrate that an elaborate combination of multiple signaling machineries encode the intensity of olfactory input, shedding light on understanding the molecular strategies on sensory transduction.Various odorants trigger complex animal behaviors across species in both quality- and quantity-dependent manners. However, how the intensity of olfactory input is encoded remains largely unknown. Here we report that isoamyl alcohol (IAA) induces bi-directional currents through a Gα- guanylate cyclase (GC)- cGMP signaling pathway in  Animals sense numerous volatile compounds from environmental cues to locate food sources, avoid predators and pathogens, and communicate with each other . For exaCaenorhabditis elegans offers a powerful model to mechanistically study how the nervous system responds to various odorants at the molecular, cellular and circuit levels. Equipped with a compact nervous system with only 302 neurons and 56 glial cells, C. elegans can sense a vast number of odors and execute a wide range of olfactory-related behaviors such as chemotaxis to food sources, pathogen avoidance, social feeding, olfactory associativelearning and imprinting (C. elegans- amphid wing “A” cell (AWA), amphid wing “C” cell (AWC), amphid wing “B” cell (AWB), amphid neuron “H” cell with single ciliated sensory ending (ASH), and amphid neuron “L” cell with dual ciliated sensory endings (ADL) (The nematode printing . Five pags (ADL) . AWC andgs (ADL) .2+-activated Cl– channels  are employed as olfactory receptors (ORs) . Variouschannels . In Dros binding . Previou elegans . Notably elegans .C. elegans exhibits distinct behaviors to certain odorants depending on their concentrations  triggered similar responses in AWCOFF but not AWCON , and that of IAA removal-induced inward currents was 1.25 ± 0.35 s (n = 12)  , which a elegans , supportunc-13(e51), unc-13(e1091), unc-31(e928), and unc-18(e81) mutants, in which the release of neurotransmitter and/or neuropeptide is blocked (A previous study has reported that AWC is suppressed by other neurons under high concentrations of IAA . We then blocked . Remarka blocked . These runc-13(e51) mutants (unc-13(e51) or unc-31(e928) mutants were equal to that in wild-type worms (We next checked IAA-induced currents in AWB neurons. High concentration of IAA (1%) induced outward currents in AWB and inward currents upon removal, which were similar to those in AWC . Strikin mutants . Additiope worms . Taken todr-3(n2150) or osm-9(ky10) mutants  mutants, suggesting that OSM-9 is required for the IAA-induced olfactory responses in AWA neurons  mutant worms, the cGMP-induced currents were abolished in both AWC and AWB neurons (tax-2(p691) or tax-4(ks11) mutant background, IAA failed to induce any detectable current in either AWC or AWB neurons (tax-2(p691) mutants as compared to wild-type worms and the input resistances were higher (A cyclic nucleotide-gated ion channel (CNG) encoded by two subunits of tax-2 (α-subunit) and tax-4 (β-subunit) is required for olfaction, thermosensation, phototransduction, gustation and O elegans . By dial elegans . In tax- neurons . Further neurons . We alsoe higher , indicatcng-1 (jh111);cng-3 (jh113) double mutants were similar to that of wild-type animals, suggesting that CNG-1 and CNG-3 are not required for olfactory transduction in AWC (Data not shown).Additional CNG channel homologs CNG-1 and CNG-3 are expressed by several amphid neurons including AWC . HoweverC. elegans genome encodes 21 Gα, 2 Gβ, and 2 Gγ subunits. The Gi/o-like Gα protein ODR-3, GPA-2, GPA-3, GPA-5, and GPA-13 have been implicated in mediating olfactory-related behaviors in C. elegans  mutants, and the residual currents were eliminated in odr-3(n1605);gpa-3(pk35) double mutants. In gpa-3(pk35) mutants, the outward currents were slightly reduced, but the inward currents were unaffected ;gpa-3(pk35) double mutants, suggesting that ODR-3 and GPA-3 are also required for the IAA-mediated olfactory responses in AWB (gpa-3(pk35) mutants but kept normal in odr-3(n2150) mutants (gpa-3(pk35) mutants, and the deficit was fully rescued by expressing of ODR-3 specifically in AWB neurons  mutants instead of the outward currents in the wild-type worms (odr-3(n2150) mutants were similar to those induced by 0.01% IAA in wild-type worms  or daf-11(ks67) mutants, suggesting that cGMP synthesis by ODR-1 and DAF-11 is essential for the olfactory transduction in AWC  mutants ;pde-2(nj58);pde-3(nj59);pde-5(nj49) quadruple mutants were not significantly different from those in wild-type worms ;pde-6(ok3410) double mutants, suggesting that cAMP is not required for IAA-sensing in AWC neurons (Data not shown).Decrease of cGMP can be achieved by more degradation, less synthesis, or both. There are six PDEs in ave cGMP . PDEs ha elegans . Howeverpe worms . Howeverpe worms . Loss of animals . These olar cGMP . In addipde-1(nj57);pde-2(nj58); pde-3(nj59); pde-5(nj49) quadruple mutants, the amplitude of 1% IAA addition-induced outward currents was increased and that of 1% IAA removal-induced inward currents was reduced  mutant animals, but not in either pde-1(nj57) or pde-2(nj58) or pde-3(nj59) mutants, suggesting that PDE-5 plays a role in 0.01% IAA-induced responses in AWB neurons (pde-5(nj49) mutants ;pde-2(nj58);pde-3(nj59);pde-5(nj49) quadruple mutants , but did not change the amplitude of ThRCs .Requests for information and for resources and reagents should be directed to and will be fulfilled by the Lead Contact, Lijun Kang (Escherichia coli using standard methods previously described (tax-2(p691), tax-4(ks11), cng-1(jh111);cng-3(jh113), unc-13(e51), unc-13(e1091), unc-31(e928), unc-18(e81), odr-3(n2150), gpa-3(pk35), odr-3(n1605);gpa-3(pk35), odr-1(n1936), daf-11(ks67), osm-9(ky10), pde-1(nj57), pde-2(nj58), pde-3(nj59), pde-5(nj49), pde-1(nj57);pde-2(nj58);pde-3(nj59);pde-5(nj49) and pde-4(nj60);pde-6(ok3410) were provided by the Caenorhabditis Genetic Center (CGC).All nematode strains were cultivated at 20°C on nematode growth medium (NGM) plates seeded with the OP50 strain of escribed . Well-fePodr-1:RFP (AWC and AWB), Podr-1:DsRed2b (AWC and AWB), Pstr-2:DsRED (AWCon), Pstr-1:DsRED (AWB), Psra-6:DsRED (ASH and ASI), Pstr-3:YFP (ASI), Podr-10:mCherry (AWA) and Psrh-220:mCherry (ADL) transgenic strains were used to identify AWC, AWB, ASH, AWA and ADL neurons, respectively. Pstr-1:GCaMP5.0 was constructed for calcium imaging in AWB neurons.Promoters were PCR-amplified from N2 genomic DNA and then recombined with specific donor vector fragments using the In-Fusion PCR Cloning Kit (TaKaRa Inc.). Chemotaxis assays were performed essentially as described . Briefly2, 1 MgCl2, 20 D-glucose and 10 HEPES . The pipette solution constitutes (mM) of 145 potassium gluconate, 2.5 KCl, 5 MgCl2, 10 HEPES, 0.25 CaCl2, 10 glucose, 5 EGTA, 5 Na2ATP and 0.5 NaGTP .Patch-clamp recording was carried out on an Olympus microscope (BX51WI) with an Axon 700A patch-clamp amplifier and Digidata 1320A interface (Axon Instruments). Signals were filtered at 2 kHz using amplifier circuitry, sampled at 10 kHz, and analyzed using Clampex 10.4 (Axon Instruments). Recording pipettes were pulled from borosilicate glass capillaries  to a resistance of 15–20 MΩ on a P-97 micropipette puller (Sutter Instruments). Worms were glued on the surface of a sylgard-coated cover glass. A small piece of cuticle in the head of the worm was cut open and glued to the coverslip to expose the neurons of interest for recording as previously described . OdorantAnimals were immersed in bath solution  and subsequently glued on a glass coverslip with a medical grade cyanoacrylate-based glue  . All odot-test or two-way ANOVA was used to compare data sets. If the data were not normally distributed, the Wilcoxon test was used. P < 0.05 was considered to be statistically significant. Sample sizes were determined by the reproducibility of the experiments and are similar to the sample sizes generally used in the field.Data analysis was performed using GraphPad Prism 6. All data were presented as mean ± SEM. Unpaired two-tailed Student’s The original contributions presented in the study are included in the article/LK and SD designed the experiments. HC conducted the calcium imaging experiments. YL conducted the electrophysiological experiments. YX, JS, ZT, and SL conducted the other experiments. HC and YL contributed equally to this study. YL, HC, and LK analyzed and interpreted the results. YL and LK wrote the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Background: There is no guideline for hearing compensation after temporal bone resection. This study aimed to retrospectively analyze surgical cases with reconstruction for hearing preservation after temporal bone malignancy resection and propose a new alternative to compensate for hearing loss. Methods: We retrospectively reviewed the medical records of 30 patients who underwent lateral temporal bone surgery for temporal bone malignancy at our institution and examined their hearing abilities after surgery. Result: The hearing outcomes of patients with an external auditory meatus reconstruction varied widely. The mean postoperative air–bone gap at 0.5, 1, 2, and 4 kHz ranged from 22.5 dB to 71.25 dB. On the other hand, the average difference between the aided sound field thresholds with cartilage conduction hearing aid and bone conduction thresholds at 0.5, 1, 2, and 4 kHz ranged from −3.75 to 41.25. More closely located auricular cartilage and temporal bone resulted in smaller differences between the aided sound field and bone conduction thresholds. Conclusions: There is still room for improvement of surgical techniques for reconstruction of the auditory meatus to preserve hearing after temporal bone resection. The cartilage conduction hearing aid may provide non-invasive postoperative hearing compensation after lateral temporal bone resection. Malignant tumors of the temporal bone are rare with an extremely low incidence rate ,2. The mAdditionally, each facility may have various treatment strategies to compensate for hearing loss after temporal bone resection. Patients with temporal bone malignancies are often provided with treatment options that result in hearing loss. Hearing loss leads to the deterioration of patients’ quality of life. To compensate, hearing improvement after surgery is desirable, but there is no standard protocol or guideline on this issue.It has been shown that negative margin resection for temporal bone malignancies provides excellent long-term tumor-free survival. While the use of hearing preservation surgery with auditory canal reconstruction and tympanoplasty after temporal bone resection has recently been reported, surgical results remain under discussion and in need of improvement. Morita et al. reported favorable results in eight cases of auditory canal reconstruction using split-thickness skin grafts for surgically treated early temporal bone malignancies . HoweverNishimura’s group first introduced cartilage conduction hearing in clinical practice. A cartilage conduction hearing aid (CCHA) includes both the cartilage–bone sound pathway and the cartilage–air and direct air pathway. This small and non-invasive device was considered as an option for hearing compensation after lateral temporal bone surgery ,7,8,9,10In this report, to discuss options for hearing compensation after lateral temporal bone resection (LTBR), we report the postoperative hearing progression of cases with external auditory canal reconstruction after lateral temporal bone resection (LTBR) and the results of our examination of the effectiveness of the CCHA after LTBR.A retrospective review of the patients treated at the Department of Otorhinolaryngology, Head and Neck Surgery at the Kyushu University Hospital from January 1993 to July 2020 was performed. A total of 181 patients were treated for temporal bone-related malignancies. A total of 161 cases of malignancies originated from the temporal bone. LTBR cases with postoperative hearing compensation were selected for this review. The final dataset included nine patients who underwent LTBR with the reconstruction of the external auditory meatus and tympanoplasty. Furthermore, we obtained audiometric data from 16 cases aided with CCHAs. Approval from the ethics review committee of Kyushu University Hospital (permit no. 29–43) was obtained.All patients with temporal bone squamous cell carcinoma were treated with LTBR. When a postoperative pathological examination revealed a positive resection margin or if it was highly suspected intraoperatively, postoperative chemoradiotherapy was added. When the tumor was considered resectable with free negative margins on preoperative computed tomography (CT) and magnetic resonance imaging (MRI) scans, reconstruction of the external auditory canal with a free flap was planned for the patient undergoing hearing-preserving surgery.Audiometry with a pure-tone audiometer  was conducted in a soundproof booth by experienced audiologists. Pure-tone thresholds were measured at 0.125, 0.25, 0.5, 1, 2, 4, and 8 kHz frequencies for air conduction and at 0.25, 0.5, 1, 2, 4 kHz for bone conduction with masking as appropriate. The results of both preoperative and postoperative hearing thresholds are included in our dataset. The hearing level was evaluated based on pure-tone audiograms as a follow-up to postoperative hearing levels in patients with auditory canal reconstruction. Pure-tone air and bone conduction thresholds averages were obtained. For pure-tone averages, the thresholds measured were 0.5, 1, 2, and 4 kHz. Air-bone gaps (ABGs) were calculated using air and bone conduction averages from the same test. To test the hearing level in patients that underwent surgery with the bone–cartilage anchoring technique, ipsilateral pure-tone hearing thresholds were tested while the patients wore commercial CCHAs  with appropriate masking for the contralateral side. We calculated and averaged the difference between the aided sound field thresholds and bone conduction thresholds at 0.5, 1, 2, and 4 kHz, which is referred to as “aided ABG.”An axial image of CT was used to measure the closest distance between the auricular cartilage and temporal bone after surgery.Our study included 30 patients that underwent LTBR, among which nine cases underwent the reconstruction of the external auditory meatus and tympanoplasty and 12 cases underwent the closure of the external auditory meatus. Five out of 12 cases underwent LTBR with the bone–cartilage anchoring technique to establish firm contact between the cartilage and the temporal bone. We obtained audiometric data from 16 cases aided with CCHAs after surgery. Pathology, clinical T stage (based on the modified Pittsburgh classification), sex, age, affected side, type of surgical approach, type of free flap for reconstruction, operation time, surgeon, resection margin examination, adjuvant radiotherapy, and aided ABG are summarized in The surgical steps for the reconstruction of the external auditory meatus with a free flap are shown in The postoperative follow-up for hearing levels was reviewed in all nine patients with external auditory canal reconstruction. Mean postoperative air–bone gap varied from 22.5 dB to 71.25 dB A. At 2 kPostoperative air conduction level also varied from 25 dB to 90 dB at 0.5 Hz, from 30 dB to 95 dB at 1 kHz, from 45 dB to 110 dB at 2 kHz, and from 65 dB to 115 dB at 4 kHz C. The aup = 0.0021 R2 = 0.503; We examined the audiometric data of 16 patients wearing the CCHAs. In four cases with and 12 cases without external auditory reconstruction, we obtained audiometric data postoperatively using the CCHAs. The results showed that the average difference between the aided sound field thresholds and bone conduction thresholds at 0.5, 1, 2, and 4 kHz ranged from −3.75 to 41.25. There was a moderate correlation between the distance between the auricular cartilage and the temporal bone around the triangular fossa and the postoperative difference between the aided sound field thresholds and bone conduction thresholds. Here, the closer the distance, the smaller the difference  reconstruction with a local flap, (2) reconstruction with a free pedicled flap, (3) middle ear implant or BAHA, and (4) bone conduction hearing aid. As of 1 April 2021, neither the middle ear implant or the BAHA for unilateral hearing deficit were covered by health insurance in Japan. Therefore, patients must choose from the other three options. Each option has advantages and disadvantages, as shown in In 2013, Iida et al. reported the reconstruction of the external auditory canal with a free flap after LTBR . To recoIt is well known that, in general, the volume of the free flap decreases gradually after surgery. However, not all cases involving free flap reconstruction follow the same clinical course. We encountered a case in which the volume of the free flap was conserved, resulting in stenosis of the reconstructed external auditory meatus (Case 6). Thus, the surgeon should preoperatively explain the possibility of a staged surgery to reduce the flap volume if necessary. Additionally, there is a possibility of deviation of the columella, resulting in worsening conductive hearing loss. Furthermore, hearing results depend on the patient. Only two patients achieved <30 dB of a mean postoperative ABG. Considering that hearing results largely rely on both patient factors and surgeon skills, there is a significant amount of room for improving this surgical procedure.Adjuvant radiotherapy can cause osteoradionecrosis, the elevation of the sensorineural hearing thresholds, and radiation-induced otitis media and externa (dermatitis). Thus, LTBR with external auditory canal reconstruction and tympanoplasty is recommended only in cases with a high possibility of a margin-free resection based on the preoperative radiological evaluation. However, we cannot predict the result of a postoperative histopathological examination of all surgical cases. In our series, four out of nine patients had a positive margin resection, although the preoperative radiological assessment seemed to predict negative surgical margins. To prevent postoperative osteoradionecrosis of the temporal bone, the best option is to fill the surgical defect with well-vascularized tissue. Considering these aspects, an alternative method for postoperative hearing compensation is needed for cases with a high possibility of adjuvant radiotherapy.Because a reconstructed auditory canal does not have a natural self-cleaning mechanism, cleaning the reconstructed auditory canal after surgery should also be considered. A preoperative explanation should be given to patients on how the reconstructed ear canal should be cleaned regularly for the rest of their life. On the other hand, strong contact with the skin and the pressure exerted against the cranial bone from the bone conduction hearing aids can cause skin erosion and pain. To overcome these disadvantages, we used a CCHA in a case with LTBR to maintain hearing postoperatively. In Japan, CCHAs have become commercially available ,7,8,9,10The hearing results of the patient with a CCHA after LTBR implied that the distance separating the auricular cartilage and the temporal bone is a potential factor for improving the effectiveness of sound transfer using the CCHAs after LTBR. With the cartilage anchored to the temporal bone, as shown in Nishimura’s group reported several advantages of CCHAs ,7,8,9,10A limitation of this study was its small sample size. Further studies are warranted to validate our preliminary data. However, based on the present data, we predict that CCHAs will be introduced to more patients treated using the bone–cartilage anchoring technique.This paper presented the hearing outcomes and options for hearing compensation after LTBR. The information obtained from our review can be extrapolated to offer guidance on reconstruction for these patients. Surgeons should consider hearing compensation for surgical cases of temporal bone malignancies as well as curative surgical resection.


Mycoplasma (M.) hyopneumoniae interacts with the respiratory microbiota and facilitates colonization of other pathogens. The present study investigated the pulmonary and nasal microbiota of M. hyopneumoniae-infected and M. hyopneumoniae-free pigs. Sixty-six pigs from three commercial herds were selected at the end of the finishing phase: 44 originated from two M. hyopneumoniae-positive herds and 22 from a M. hyopneumoniae-negative farm. At the slaughterhouse, samples of nasal turbinate (NT) and bronchus-alveolar lavage fluid (BALF) were collected. DNA was extracted with a commercial kit and the infection status was confirmed by qPCR. All samples from the same herd were pooled, and next-generation sequencing based on the hypervariable region V3–V4 of the 16 s bacterial rDNA was performed. Data analysis included the taxonomic analysis, Alpha diversity indexes, and Principal coordinates analysis (Pcoa) using Jaccard, Bray–Curtis, Weighted Unifrac, and Unweighted Unifrac distances. All pigs from the infected herds tested PCR positive for M. hyopneumoniae, whereas all pigs from the negative farm were negative. There was a greater diversity of microorganisms in BALF when compared to NT samples in all the farms. BALF samples from infected animals showed higher abundance of M. hyopneumoniae than NT samples and a predominance of Pasteurella multocida among the main species identified, which was also abundant in the M. hyopneumoniae-free herd. PCoa diagrams indicated that for most of the samples, dissimilarity on bacterial composition was observed, regardless of infection status and sample type. Therefore, the lung microbiota was modulated by M. hyopneumoniae infection, which could play a role in the pathogenesis of M. hyopneumoniae-disease.The online version contains supplementary material available at 10.1186/s13567-022-01038-9. Mycoplasma (M.) hyopneumoniae, Mycoplasma (M.) flocculare, and Mycoplasma (M.) hyorhinis. Mycoplasma hyopneumoniae is considered the primary agent of Swine Enzootic Pneumonia (SEP), a disease that results in chronic pneumonia, non-productive dry cough, growth retardation, and a negative impact on productive indicators [M. hyopneumoniae can also play an important role in the Porcine Respiratory Disease Complex (PRDC), a multifactorial disorder where several infectious agents can be involved, such as porcine reproductive and respiratory syndrome virus (PRRSv), porcine circovirus type 2 (PCV2), and Pasteurella (P.) multocida [Mycoplasmas are small bacteria without a cell wall that affect several animals, including pigs. Different species are known to infect swine, among them dicators . Besidesultocida , 2.Mycoplasma hyopneumoniae infections are commonly associated with a decrease in performance and an increase in costs related to control and treatment [Actinobacillus (A.) pleuropneumoniae, Streptococcus (S.) suis, P. multocida, and Glaesserela (G.) parasuis [reatment , 4. The parasuis , 3.M. hyorhinis has shown to be associated with M. hyopneumoniae infection and PRDC [M. flocculare, another commensal bacterium in swine, has shown a negative correlation with the extension of macroscopic lung consolidation lesions (MLCL) when associated with M. hyopneumoniae infection [M. hyopneumoniae infection changes the environment in the respiratory tract, thereby creating unfavorable conditions for M. flocculare [Although considered commensal bacteria in the respiratory tract of swine, and PRDC , 5. In aand PRDC . Recentlocculare .M. hyopneumoniae, along with other bacterial genera  in the microbiome of macroscopically affected lungs, when compared to healthy lungs, where the main genera reported were Methylotenera, Prevotella, Sphingobium, and Lactobacillus [M. hyopneumoniae was positively associated with the extent of the lung lesions and reported as a taxonomic driver of functional shifts in the lungs with severe lung lesions [M. hyopneumoniae, M. flocculare, and M. hyorhinis in the lungs of pigs affected by enzootic pneumonia [Studies have shown higher abundance of bacillus . Additio lesions . Other aneumonia , 9. Despneumonia . Thus, kneumonia .M. hyopneumoniae was confirmed with a high prevalence when referring to community composition in lungs. Moreover, the authors suggest that the interactions among respiratory pathogens are numerous and complex, and therefore, can be additive or synergistic. In general, populations of microorganisms in organs of healthy individuals can help their functions and/or prevent colonization by pathogens. As an example, Lactobacillus spp. contribute to the formation of alveoli and mucus, and consequently they improve the functioning and protection of the respiratory tract [As reported by Siqueira et al. , M. hyopry tract .M. hyopneumoniae and other bacterial species in the respiratory tract of pigs, the present study investigated possible differences between the pulmonary and nasal microbiota of finishing pigs naturally infected with M. hyopneumoniae and M. hyopneumoniae-free pigs in order to better understand the interactions and pathogenesis.Considering the abundance of This study was submitted to the Ethics Committee in Animal Use of the Faculty of Agricultural and Veterinary Sciences of the São Paulo State University—Jaboticabal, and approved under the Protocol #2032/21.M. hyopneumoniae (H1) and the other two were infected with M. hyopneumoniae (H2 and H3), as reported by the herd veterinarian and the presence of MLCL, suggestive of enzootic pneumonia. Moreover, the M. hyopneumoniae-status of the three farms was confirmed by testing bronchus-alveolar lavage fluid (BALF) and nasal turbinate (NT) samples with qPCR [A. pleuropneumoniae and with vaccination against Influenza virus, Porcine Circovirus, Parvovirus, Leptospirosis, Erysipelas, and G. parasuis. The infected animals originated from two different commercial farms of 550 and 600 sows, farms H2 and H3, respectively. These two farms also used farrow-to-finish systems, with vaccination against M. hyopneumoniae, P. multocida, Influenza virus, Porcine Circovirus, Parvovirus, Leptospirosis, Erysipelas, and G. parasuis. The vaccination protocol is described in Additional file Three commercial swine herds (H), located in the state of São Paulo—Brazil, were selected for the study: one herd was free of ith qPCR , 6. The umoniae H and the M. hyopneumoniae-free pigs (H1). However, the rest pens were located in the same space, side by side, and separated by 1.5 m height concrete walls. Animals with M. hyopneumoniae-free status were the first to be slaughtered (H1), followed by the pigs from the M. hyopneumoniae-infected farms (H2 and H3).From each farm, 22 pigs (140-day-old average) were randomly selected and identified by an ear tag in the morning. The pigs were sent to the slaughterhouse later on the same day. Animals from the same farm were kept in the same rest pen for at least 3 h before slaughter, and animals from H2 and H3 were not mixed with the At the slaughterhouse, the corresponding head and lungs from the selected carcasses were separated for evaluation and sampling. The nose of the pigs was sawn off in the slaughter-line using a saw (disinfected by water at 90 °C) to access the NT. Fragments of the NT were collected directly from the carcass using sterile forceps and scalp. Then, the fragments were inserted into cryotubes free of DNAse and RNAse  and stored in a freezer at −80 °C.BALF was collected using a plastic pipette and automatic pipettor. For insertion of the pipette, an incision was made in the trachea, about 5 cm before the bifurcation of the bronchi. Directly into the bronchi, 20 mL of PBS  was dispensed, followed by a gentle massage through the lung parenchyma and aspiration with a pipette, recovering between 10 and 15 mL of lavage. The recovered content was transferred to 50 mL falcon tubes  and later aliquoted into 2 mL cryotubes free of DNAse and RNAse , which were then stored in a freezer at −80 °C until further processing.At the time of slaughter, the lungs of the study pigs were removed from the carcass and the macroscopic lung lesions compatible with SEP were scored according to the methodology described by Straw et al. . The apig for 20 min. After centrifugation, the supernatant was discarded and the pellet was used for DNA extraction. A total of 0.05 g of NT and the pellet from BALF samples were individually submitted to commercial DNA extraction using the DNeasy blood and Tissue kit , following the manufacturer’s instructions. The extraction product was evaluated by spectrophotometry in Nanodrop® 2000c  to measure concentration and purity.BALF samples were thawed and centrifuged at 4 °C  at 12 000 M. hyopneumoniae (p102), M. hyorhinis (p37), and M. flocculare (fruA) from DNA samples were quantified by qPCR as described elsewhere [7–101) was based on a synthetic DNA  containing fragments of 150 base pairs (bp) (p102), 101 bp (p37) and 119 bp (fruA), used as positive control [Gene fragments of lsewhere , with molsewhere . The sta control . Samples® .The extracted DNA samples from animals of the same herd  were grouped in equimolar amounts according to the concentration (ng∕µL) obtained by spectrophotometry. The pools from the control group were named H1NT and H1BALF, while the pools of samples from infected farms were named H2NT, H2BALF, and H3NT, H3BALF. After pooling and before submission to Illumina sequencing, the integrity and quality of the samples were evaluated using Bioanalizer®, USA).Pooled DNA samples were submitted to a PCR targeting the hypervariable region V3-V4 of the 16 s bacterial rDNA, using a previously described protocol , which fLibrary validation was performed in a Bioanalyzer 2100  and quantified using the Kappa Library Quant Kit for Illumina , following the manufacturer’s instructions. The library was then adjusted for equimolar concentration according to the manufacturer’s instructions . Then, 5Sequencing was performed using the MiSeq Reagent Kit v3 (600 cycles)  on an Illumina MiSeq platform , according to Caporaso et al. . The genMycoplasma spp. were evaluated according to the normality of the data, where Spearman’s Rank correlation test was used for non-parametric data (p < 0.05). To measure the strength of dependence between two variables, the Kendall’ rank correlation test was used (p < 0.05). The R software version 3.5.1  was used for data analysis.Correlations between MLCL and quantification of each p < 0.05.The contigs were aligned and assembled in silico using the software Quantitative Insights Into Microbial Ecology—QIMEE version 2019.10 [2 ranged from 0.992 to 0.999; the slope varied from −3.387 to −3.522. Detailed parameters are shown in Additional file M. hyopneumoniae on the qPCR while 21 samples (95.4%) from H2 and all samples (100%) from H3 were positive. For NT, all samples from H1 were M. hyopneumoniae-negative while 8 samples (36.4%) from H2 and 4 samples (18.2%) from H3 were M. hyopneumoniae-positive. One animal in H2 tested negative for M. hyopneumoniae in the BALF sample and tested positive in the NT sample, and therefore, is considered M. hyopneumoniae-positive. Among all NT samples, 81.8% (54/66) tested positive for M. flocculare and 59.1% (39/66) were positive for M. hyorhinis. Similarly, 80.3% (53/66) and 53% (35/66) of the BALF samples tested positive for M. flocculare and M. hyorhinis, respectively.The efficiency of qPCR reactions varied from 92.3% to 101.4%, RM. hyopneumoniae in BALF and NT were 8.73 × 105 copies/μL (2.98 × 102 to 4.51 × 107) and 4.20 × 102 copies/μL (9.51 × 100–4.20 × 103), respectively. Detailed information regarding quantification data are summarized in Additional file Median quantification values for M. hyopneumoniae-positive (H2 and H3) and M. hyopneumoniae-negative (H1) farms (p < 0.05). MLCL occurrence for H1, H2, and H3 was 68.18%, 90.9%, and 90.9%, respectively.The mean macroscopic lung lesion scores obtained from the different herds were: H1 − 0.5225 (SD = 2.58); H2 − 6.6025 (SD = 14.34); and H3 − 2.7075 (SD = 6.25). Significant differences in the mean macroscopic lesion score were detected between p < 0.05) between the quantification of M. hyopneumoniae in the BALF by qPCR and the macroscopic lung lesions. No significant correlations were observed between macroscopic lung lesions and M. hyorhinis or M. flocculare quantification results in BALF samples (p > 0.05). There were no significant correlations between macroscopic lung lesions and M. hyopneumoniae, M. hyorhinis, or M. flocculare quantification results in NT samples (p > 0.05).Spearman’s correlation showed a significant positive correlation , and no significant correlation between the quantification of M. hyopneumoniae and M. hyorhinis (p = 0.89). There was a significant positive correlation between the quantification of M. hyopneumoniae in BALF and NT samples .Kendall’s rank correlation test showed a negative correlation between the quantification of The NGS identified a total of 4364 features or operational taxonomic unit (OTUs) with sequence sizes ranging from 255 to 498 bp, and an average sequence size of 377.7 bp (± 68.8). The OTUs and readings obtained for each pool before rarefaction are shown in Table To estimate the species diversity in the samples, the sequences were rarefied to a depth of 69 100 reads. The samples showed varying numbers of reads, with H2NT and H2BALF showing the highest values, followed by H3 samples, both from infected herds , indicating, in general, a greater diversity of genders when compared to BALF samples (average: 572.6). In addition, Shannon values for BALF samples from infected animals were numerically lower (H2: 2.77 and H3: 1.68), indicating low diversity. Considering the uniformity aspect by the Pielou and Simpson metrics, the samples H2BALF and H3BALF showed the lowest values, indicating less uniformity between genera in the sample .The PCoa based on the unweighted Unifrac distance showed separation between BALF and NT samples, with greater dissimilarity for NT samples on axis 1. The H2NT and H3NT samples were close to each other, indicating qualitative similarity between the bacterial types present in both samples. Regarding BALF samples, no proximity was observed between any of the points regardless of the herd, with the greatest distance observed between H2BALF and H1BALF  explains a high percentage of variation in relation to the other axes. The samples of H2BALF, H3BALF, and H1BALF were close to each other and relatively far from H1NT and H2NT samples, and even further away from H3NT, possibly due to the low abundance in the sample. NT and BALF samples from uninfected animals were closer to each other when compared to corresponding samples in herds 2 and 3 (H2 and H3), as observed in Figure M. hyopneumoniae in samples H2BALF and H3BALF (62.39% and 68.61%). Mycoplasma hyopneumoniae was also detected in H1NT, H2NT, and H3NT, with the respective frequency of 0.15%, 0.09%, and 0.08%. Considering that all BALF samples were negative on the qPCR, it is possible that the occurrence of M. hyopneumoniae in H1NT is related to contamination during slaughter. In samples H2BALF, H3BALF, and H1BALF, relative frequencies of P. multocida were 18.21%, 4.83%, and 22.5%, respectively. Anoxybacillus was recorded in BALF and NT samples only in H3 and H1, ranging from 8.36% (H3BALF) to 36.36% (H3NT). Following the order of relative frequency, Actinobacillus indolicus appears in all samples, but only in H2NT, a high relative frequency of 40.2% was recorded, and in the others, the values (H3: 0.58% and H1: 1.74%) were not higher than 2%. Cilia-associated respiratory bacterium 95-15405 was identified in samples from H1 and H2, with the highest relative frequencies observed in H2NT and H2BALF, 21.36% and 7.28%, while in samples from H1 the values were not higher than 4% (H1NT: 3.72% and H1BALF: 0.94%). Unexpectedly, a high relative frequency of 67.40% of Escherichia/Shigella was identified in the H1NT sample while samples H1BALF, H2BALF, and H3BALF showed relative frequencies close to 0.3%. Mycoplasma flocculare was identified in all analyzed samples, and among the BALF samples the highest value was for H1BALF (12.09%) and in lower prevalence in the samples H2BALF (0.5%) and H3BALF (0.18%). On the other hand, NT samples showed a higher prevalence of M. flocculare in H2NT (3.41%) and H3NT (12.68%), while in H1NT the percentage was the lowest, 1.48%. Main results are shown in Figure The taxonomic analysis showed a high relative frequency of Mycoplasma hyopneumoniae is described as the primary agent in respiratory diseases in pigs, present worldwide, and directly related to the occurrence of pulmonary consolidation lesions in pigs [M. hyopneumoniae and low species diversity in alpha and beta diversity metric analyses, in addition to significant correlations between M. hyopneumoniae DNA quantification and macroscopic lung consolidated lesions (MLCL). in pigs . The preM. hyopneumoniae in H2 and H3 and high correlation with MLCL, which was also recently reported by Ferreira et al. [M. hyopneumoniae and, interestingly, the lower the concentration of M. flocculare in the lungs. Similarly, our results indicated that M. hyopneumoniae concentration in BALF is negatively correlated to M. flocculare concentration in BALF. Therefore, since these two Mycoplasma species share most of the surface proteins [M. hyopneumoniae cross-react with M. flocculare antigens, which could reduce the number of microorganisms. Considering its lower capacity of evading the immune system, M. flocculare would be more susceptible to be eliminated in case of co-infections with M. hyopneumoniae [The qPCR results showed a high occurrence of a et al. . In addiproteins  and are eumoniae .M. hyopneumoniae-free herd. The lesions were more severe in H2 than in H3, although M. hyopneumoniae relative frequency was higher in H3. However, P. multocida relative frequency was also higher in H2, which could have worsened the lung lesions in H2, as reported previously [P. multocida is a secondary pathogen that in combined infections with M. hyopneumoniae results in more severe pneumonia and clinical disease [P. multocida was higher. Although our results suggest that M. hyopneumoniae-like lung lesions were worsened when P. multocida co-infection occurred, co-infections by other respiratory pathogens such as Influenza and PCV2 cannot be excluded as they were not assessed in this study.Not surprisingly, MLCL occurrence was higher in the infected herds than in the eviously –24. P. m disease , 25, 26.M. hyopneumoniae, a predominance of the species in high relative frequency in BALF samples was shown, as well as lower numerical values in the diversity and uniformity metric, measured by the Pielou and Simpson analyses. Numerically, the H1BALF sample showed better results considering uniformity, according to Pielou and Simpson metrics, although Faith and OTUs values were low, possibly due to the low sequencing throughput (181 176 reads). Accordingly, previous studies have shown that animals infected with M. hyopneumoniae have a predominance of the species over other species, reducing bacterial diversity in the lung [M. hyopneumoniae stimulates the immune system, generating the release of various pro-inflammatory cytokines [M. hyopneumoniae had a higher degree of pulmonary involvement. Therefore, it is possible that the indirect effects on lung injuries could be associated to the reduction in the diversity of local microorganisms, either pathogenic or commensal.Regarding the animals infected by the lung , 27. It ytokines –31, induytokines , nitric ytokines , which pM. hyopneumoniae-infected animals, which may be an indicator of the lung’s health since higher diversity was observed in the non-infected animals (H1BALF). Other studies have shown that bacterial diversity helps to maintain balance by preventing the spread of pathogens and susceptibility to chronic diseases [Microbiome studies have demonstrated the beneficial effect of the diversity and uniformity of bacterial species, as well as the protective benefits the presence of certain species may have for the host . In our diseases , 36. In diseases .M. hyopneumoniae-infected pigs showed similarities in relation to the high abundance of M. hyopneumoniae, and the predominance of P. multocida among the main species of bacteria identified. Moreover, it seems that other bacterial genera are present in both infected and M. hyopneumoniae-free BALF samples due to the similarity observed in the PCoa metrics by Bray Curtis. Mycoplasma hyopneumoniae evasion characteristics and its ability to induce local inflammation are strategies that allow the pathogen to be present at higher relative frequencies [BALF samples from quencies , 20, 38.M. hyopneumoniae were detected in NT samples, as well as better results from the point of view of alpha diversity. This corroborates literature data that state that M. hyopneumoniae presents tropism for the lower parts of the respiratory tract of pigs [M. hyopneumoniae was identified, in a very low relative frequency, in the H1NT sample, but not in H1BALF or the individual qPCR results. Even though M. hyopneumoniae was identified in NT sample from a negative farm, it is possible that cross contamination occurred during the slaughter process, as reported by Marois et al. [M. hyopneumoniae DNA was found in nasal and tracheal swabs of SPF pigs that had no contact with other pigs at slaughter. In that case, the scalding water could have been a source of contamination since M. hyopneumoniae DNA was detected even before the onset of slaughter [M. hyopneumoniae DNA in SPF pigs that were in the rest area for 4 h before slaughter. Therefore, considering the similar conditions in our study, it is possible that infection occurred just before slaughter in the rest pen, when M. hyopneumoniae-infected and non-infected pigs were allocated close together, or during the slaughter process.In contrast to the BALF results, low relative frequencies of  of pigs , 40. Surs et al. , where Mlaughter . The autM. hyopneumoniae and sample type, with the exception of H2NT and H3NT samples that were relatively close to each other, indicating similarity between the compositions of microbiota in these samples. Although most of the points are equidistant, it was possible to notice in the PCoa diagram based on the Unweighted unifrac distance, that the distance was greater between BALF and NT samples. The greatest dissimilarity was registered between H1BALF and H2BALF, indicating qualitative differences between bacteria that make up both samples. In addition, the composition and diversity of the microbiota also seem to be closely related to environmental conditions. Differences in the swine rearing environment have been shown to have an influence on the animals’ respiratory and intestinal microbiota [Qualitative measures of dissimilarity, Jaccard, and Unweighted unifrac, observed in the PCoa diagrams indicated that for most of the samples dissimilarity was observed, regardless of infection status by crobiota , 43. TheFurthermore, the microbial composition of pig’s microbiota is dynamic and is related to several factors as environment, production system, diet, antimicrobial use and others . The impM. hyopneumoniae infection in BALF pools, but not in NT samples. The beta diversity analyses indicated differences between the abundance of microorganisms between the BALF samples when comparing M. hyopneumoniae-infected and non-infected herds, which is confirmed by observing the taxonomic analysis. Therefore, it is possible to observe different bacteria with approximate relative frequencies, which makes it more difficult to determine a single predominant species. On the other hand, samples H2BALF and H3BALF, demonstrated the predominance of M. hyopneumoniae. Nonetheless, further studies comparing the impact of different M. hyopneumoniae strains on the lung microbiota of infected animals are needed, as little is known about the virulence of the strains circulating in Brazil [In general, the abundance of microorganisms in the lung microbiota was affected by n Brazil , 47.In our study, sequencing pooled samples was a determining factor in making it possible to have an overview of the respiratory microbiota from the three groups, especially when considering the costs of the analyses. As shown by Ray et al. , poolingM. hyopneumoniae-infected pigs, some experimental limitations should be addressed and acknowledged, and the results reported here should be interpreted with caution. For instance, although NT samples showed higher diversity, the detection of M. hyopneumoniae was lower for these samples. Therefore, considering our results, the use of BALF samples seems more valuable when the aim is to assess possible associations between M. hyopneumoniae infection and the respective influence on the microbiota. Additionally, since sampling collection took place in the slaughterhouse, some sources of contamination such as the scalding water and the sawing machine were controlled only by the federal inspection system, and not by the researchers. Thus, possible contamination in the upper respiratory tract samples needs to be taken into account when interpreting the respective results. Even though internal negative controls were included and tested negative by qPCR for Mycoplasma [Although our results are important and can be useful to help understanding the respiratory microbiota modulation in coplasma , these sM. hyopneumoniae in the lungs of infected pigs at slaughter, and further studies are needed to investigate the significance of M. hyopneumoniae infection in the microbiome of diseased pigs in comparison to healthy animals.Overall, it has been observed that the presence or absence of a particular pathogen can affect directly or indirectly, not only promoting the development of lesions but also modulating the diversity, uniformity, and abundance of other bacterial populations in the respiratory tract of pigs. However, since the animals were assessed at slaughter, the results are not necessarily applicable for the situation during fattening period and earlier stages. Therefore, our results partially demonstrate the impact of M. hyopneumoniae in BALF of slaughter pigs seems to modulate the microbiota, which could facilitate dysbiosis and proliferation of other pathogens. Mycoplasma hyopneumoniae quantification was strongly correlated with macroscopic lung consolidated lesions, meaning that the severity of the lesions increases when there is a higher number of microorganisms.The presence of Additional file 1. Detailed vaccination protocol for farms H1, H2, and H3.Additional file 2. Parameters for qPCR assays based on the gene fragments of M. hyopneumoniae (p102), M. hyorhinis (p37), and M. flocculare (fruA). qPCR assays’ information for all plate runs, including efficiency (E), r2, slope, and Y-intercept for the three pathogens tested: M. hyopneumoniae (FAM), M. hyorhinis (Texas Red), and M. flocculare (Cy5).Additional file 3. Detailed information on the individual samples tested by multiplex-qPCR. Individual qPCR results of BALF and NT samples tested against M. hyopneumoniae, M. hyorhinis, and M. flocculare.Additional file 4. Detailed information on the taxonomic analysis with relative frequencies of each bacterial genera from pooled nasal tubinate (NT) and bronchoalveolar fluid (BALF) samples. Taxonomic analyses of pooled BALF and NT samples of M. hyopneumoniae-infected and non-infected pigs. The following results consider a taxonomic level 7 (species).


All age and sex subgroups demonstrated a positive association between PPI use and hearing impairment. Severe hearing impairment showed consistently higher odds of a relation with PPI use. PPI use was associated with an increased rate of hearing impairment.This study investigated the association of previous use of proton pump inhibitors (PPIs) with the rate of hearing impairment. The ≥40-year-old population in the Korean National Health Insurance Service-Health Screening Cohort was enrolled. The 6626 registered hearing-impaired patients were matched with 508,240 control participants for age, sex, income, region of residence, and index date (date of hearing impairment diagnosis). The prescription histories of PPIs were collected for 2 years before the index date. The odds ratios of the duration of PPI use for hearing impairment were analyzed using conditional logistic regression. Subgroups of age/sex and severity of hearing impairments were additionally analyzed for the relation of PPI use with hearing impairment. PPI use for 30–365 days was associated with a 1.65-times higher odds of hearing impairment  = 1.47–1.86 for 30–365 days of PPI medication). PPI use for ≥365 days was also related to 1.52-times higher odds of hearing impairment (95% CI = 1.35–1.72,  Proton pump inhibitors (PPIs) have been widely used for the treatment of several acid-related disorders, such as gastroesophageal reflux disease (GERD), gastric ulcer, duodenal ulcer, erosive esophagitis, and laryngeal reflux diseases ,2. PPIs Hearing impairment is one of the most common sensorineural disorders worldwide, affecting approximately 6.1% of the world’s population . The etiBecause PPIs have adverse impacts on neurologic disorders via ischemia and electric imbalance, adverse effects of PPIs on cochlear function could be predicted ,14. TherThe present study was approved by the ethics committee of Hallym University . The requirement for written informed consent was exempted by the ethics committee of Hallym University. All studies were conducted according to the guidelines and regulations of the ethics committee of Hallym University.n = 6626). Participants were included in the control group if they were not defined as having hearing impairment from 2002 through 2015 . Participants who were diagnosed with other disabilities were excluded . To measure PPI history in the previous 2 years, we excluded participants with hearing impairment who were diagnosed with hearing impairment before 2003 (n = 2160). Hearing impairment participants were 1:4 matched with control participants for age, sex, income, and region of residence. The control participants were randomly selected. The date of hearing impairment diagnosis was defined as the index date, and the same index date was used for the matched control participants. The 447,019 control participants whose index date did not match that of the hearing impairment participants were excluded. A total of 4387 participants with hearing impairment and 17,548 control participants were enrolled  were categorized as previously described [n = 2 (0.009%)) and total cholesterol (n = 3 (0.013%)) values were substituted by the average values of the study participants.Age groups were classified into 5-year intervals. Income groups were divided into 5 classes (class 1 (lowest income) to 5 (highest income)). The region of residence was classified as urban or rural . Tobaccoescribed . The recThe Charlson Comorbidity Index (CCI) was calculated for 17 comorbidities as a continuous variable (0 (no comorbidities) through 29 (multiple comorbidities)) . DementiRegarding PPIs, the number of patients diagnosed with GERD  and the dates of H2 blocker prescription were additionally assessed. The number of patients diagnosed with GERD and the prescription dates of H2 blockers were assessed for the 2 years prior to the index date.t test for continuous variables.The hearing impairment and control groups were compared using the chi-square test for categorical variables and the independent Conditional logistic regression analysis was conducted, and the odds ratios (ORs) and 95% confidence intervals (Cis) of the prescription days of PPIs for hearing impairment were calculated. A crude model (simple), model 1 , model 2 , and model 3  were used. The matched variables were stratified.p-values less than 0.05 were defined as statistically significant. Statistical analyses were performed using SAS version 9.4 .Age and sex  subgroups were analyzed. We further analyzed the ORs of proton pump inhibitor prescription days for hearing impairment according to severity of hearing impairment. Two-tailed analyses were performed. p < 0.001, p < 0.05).The prescription days of PPI were different between the hearing impairment and control groups . The ≥365 days of PPI prescription was associated with 1.52 higher odds for hearing impairment .The participants with prior histories of PPI prescription demonstrated higher odds of hearing impairment . The ≥30The previous histories of PPI prescription were associated with higher odds for hearing impairment in all age and sex subgroups. The <70-year-old group who had a PPI prescription for ≥30 to <365 days showed 1.70 (95% CI = 1.49–1.95) higher odds of hearing impairment than those with a PPI prescription for ≥0 to <30 days. The men’s group who had a PPI prescription of ≥30 to <365 days showed 1.80 (95%CI = 1.57–2.07) higher odds of hearing impairment than those with a PPI prescription for ≥0 to <30 days.According to the degree of hearing impairment, the severe hearing impairment group, but not the profound hearing impairment group, exhibited a relationship between a longer duration of PPI prescription and a higher rate of hearing impairment .The long-term use of PPIs was linked with an increased rate of hearing impairment in the adult population. This relation of PPI with hearing impairment was maintained in all age and sex subgroups. This is a pioneering study on the potential impacts of PPI use on hearing impairment in a large population. We comprehensively considered possible confounders, such as lifestyle factors of smoking and alcohol consumption and comorbidities.Two previous studies suggested the association of PPIs with hearing impairment. The National Health and Nutrition Examination Survey study on the adverse effects of PPI revealed significantly increased risks of hearing impairment, dementia, migraine, and other peripheral neuropathies associated with PPI use . HoweverInsufficient blood supply could mediate ischemic injury in the inner ear . PPIs inMetabolic disturbances and the malabsorption of micronutrients may induce neuronal degeneration. A number of prior studies suggested that the metabolic disturbances associated with PPI use were linked with an increased risk of dementia ,28. The The dysfunction of H, K-ATPase in the cochlear lateral wall could induce electrical imbalance and disturb homeostasis of the endolymphatic fluid of the cochlea. It was reported that a proton pump identical to that in the stomach is expressed in the lateral wall of the cochlea . The cocThis study used data from a large nationwide representative cohort. Many control participants could be selected and matched for demographic and socioeconomic factors. The degree of hearing loss was based on three PTAs tests and auditory brainstem response test results. The use of multiple objective hearing measures prevented the misdiagnosis of hearing impairment in this study. Because registered hearing-impaired persons receive support for the cost of health care, including hearing aids, most hearing-impaired persons were included in our hearing-impaired group. However, some limitations should be considered when interpreting the current results. The degree and etiologies of hearing loss could not be detailed in this study. For histories of PPI prescriptions, the types and doses of PPIs could not be differentiated in the current data. Although past medical histories, including history of GERD, were adjusted, the potential confounding effects of reflux symptoms remained. Further studies on the associations of dose and types of PPI with the specific types of hearing loss will unravel the current questions. The analyses using machine learning approach could facilitate the manipulations of huge data.Longer durations of PPI use were related to hearing impairment in adult Koreans. This relation of PPIs with hearing impairment was valid in all age and sex groups. Therefore, patients who need long-term PPI medication should be consulted for their hearing preservation. Future study with randomized controlled trial study design could delineate the causal relation between PPI use and hearing impairment.