An audiogram is not sufficient to indicate cochlear dead regions.To investigate cochlear dead regions in sensorineural hearing loss subjects using the TEN test. Site: CEDALVI/ HRAC-USP-Bauru/Sao Paulo/Brazil, August 2003 to February 2004.A contemporary cross-sectional cohort study. Material and Methods: The TEN test was applied in three groups: G1; G2; G3.In the G1 group the TEN value required to eliminate the test tone was, on average, close to the absolute threshold for all frequencies. No dead regions were found in the ears tested in group G2. 76 ears were tested in group G3, and six showed no evidence of dead regions in the cochlea.The TEN test was an effective test to indicate a dead region in the cochlea of subjects with sloping sensorineural hearing loss. There is evidence that pure-tone detection is different for subjects with high frequency sensorineural hearing loss and flat hearing loss; we observed a significant difference between the masked threshold and the absolute threshold only in sloping hearing loss and not for flat hearing loss. The cochlear region where internal ciliated cells (ICC) are injured, inactive or absent, and the neurons that innervate this region are inactive or even degenerate, has been named dead region of the cochlea, or dead region.3Although the dead region concept was elaborated many years ago by Gravendeel and Plomp,6Audiogram configurations have also been considered as evidence or dead regions, such as sloping audiograms, where thresholds worsen abruptly with increasing pitch (over 50 dB/octave) at high frequencies. Hearing losses of 40-50 dB at low frequencies and hearing close to normal in medium and high frequencies may suggest dead regions at low frequencies. This may also be the case when there is hearing loss over 50 dB at low frequencies with improved thresholds at higher frequencies. A U-shaped audiogram, with better hearing at low and high frequencies and a dead region at middle frequencies, is rare and generally does not interfere significantly with speech recognition. Residual hearing allows good recognition, meaning that some of these cases do not require hearing aids .Huss et al. investigated pure tone perception in subjects with and without dead regions diagnosed by psychophysical tuning curves and the threshold-equalizing-noise (TEN) test, in which participants were asked to score the sharpness of a pure tone on a scale from sharp (1) to noisy (7). The authors concluded that the pure tone subjective impression of sharpness is not a consistent indicator of dead regions in the cochlea, as higher scores were seen not only in ears with dead regions but also in normal ears at similar frequencies and at higher thresholds.It is clear that patient reports or the audiogram by themselves are not sufficient to establish or exclude the presence and extension of dead regions. Thus, masking has been used in some studies to investigate this condition in greater detail.The investigation of dead regions uses ipsilateral masking, in which the test signal and the masking noise are presented to the same ear. The idea is to raise the threshold of non-tested frequencies so that they do not respond to excitation diffusion of the signal that is being tested at a specific frequency.Moore et al. developed the Threshold-Equalizing-Noise test to investigate the presence of dead regions. Although masking is defined as a procedure used in audiological evaluation, and not a test in itself, TEN is considered a test by its authors. It is based on the detection of pure tones presented simultaneously with a wide band noise (TEN) which produces practically the same level of masking (measured in dBNPS) throughout audiogram frequencies  in normal hearing or hearing-impaired individuals with no dead regions.5In Brazil Eguti investigated the efficiency of the masking technique using white noise to identify dead regions in the cochlea of 32 adult individuals that presented acquired sensorineural or mixed hearing loss and a sloping audiometric configuration. Moore'sThe diagnosis of dead regions in the cochlea is important in medical practice, as studies have shown that its presence limits the use of hearing aids, since these regions respond minimally or not at all to sound amplification.9Our aim was to use the TEN test to investigate the presence of dead regions in the cochlea in subjects with sensorineural hearing loss seen at the Center for Audition, Vision and Language Disorders (CEDALVI) of the Sao Paulo University Craniofacial Anomalies Rehabilitation Hospital/Bauru Campus (HRAC-USP-Bauru).This study was undertaken at the CEDALVI/HRAC/USP-Bauru between August 2003 and February 2004. According to the routine at CEDALVI, patients are monitored by a multidisciplinary audiology team. Participants were assessed by an ENT specialist who did otoscopy and decided whether conditions were appropriate for the TEN test, namely intact tympanic membranes and no external and middle ear compromise.Cases for this study are shown on a)G1: pure tone air conduction thresholds within normal limits (≤15dBNA);b)G2: mild flat configuration sensorineural hearing loss (Davis and Silverman);c)G3: moderate to severe sloping sensorineural hearing loss.The selection of subjects was based on the following inclusion criteria:Participants signed a free informed consent form approved by the Research Ethics Committee of the HRAC-USP, number 263/2004-UEP-CEP, agreeing to take part in the study and to allow publication of collected data.a)TEN biological calibrationWe used the Veniarb)TEN testThe TEN test was applied using a two-channel audiometer, PAC 2000 model (Acustica Orlandi) with a TDH-39 P supra-aural earphone and a Sony Discman digital MEGA BASS model, in a soundproofed cabin. Moore et al. (2000) recorded the test on a CD with noise (TEN) on one channel and a digitally generated test signal (pure tone) recorded on another CD channel. This was done to simplify and increase the clinical availability of the TEN test.(1)the CD player output plug was connected to the audiometer right and left input socket . The first CD track is a calibrating tone, which was played to adjust the audiometer right and left channel VU meters at -5 dB, or 5 dB below zero; this was done to calibrate the signal-to-noise ratio, or the intensity difference between the pure tone and noise. For the remaining tracks the noise intensity level in the right channel was read as 10 dB better than the indicated level on the dial, and the left channel pure tone was read as 10 dB worse. We could then adjust the audiometer controls for the desired intensities for pure tones and noise. Thus, if the pure tone and noise intensities were 50 and 40dBNPS, the dial was adjusted to 40 and 50 dB, respectively.(2)Air conduction absolute thresholds (ATs) and masked thresholds (MTs) were obtained by manual audiometry, according to Carhart and Jerger,(3)MTs with noise (TEN) were assessed at the same frequencies, according to the audiometric configuration, with both channels operating. The TEN intensity depended on the AT by frequency in each patient, and was 10 dB better than the AT, that is, if the AT was 50 dB, the TEN was adjusted to 40 dB on the dial.(4)If the MT was only 5 dB worse than the AT, the test was repeated with higher masking noise intensity.The following test procedures were undertaken: a)Biological calibration of the TENThe minimal effective masking level considered was the amount of noise (TEN) that was needed to eliminate the test tone (30 dBNPS) for each frequency.b)TEN testAccording to Moore et al.'sc)Statistical methodThe following statistical tests were used: t test , and Spearman's rank correlation coefficient. The null hypothesis rejection value (Ho) was 5%.As seen on According to Moore et al.'sA frequency of occurrence analysis was made in group G3 , as show Statistical analysis using the t test (Ho= A=B and H1= B>A) was applied to group G3, in which the tests suggested dead regions in the cochlea according to Moore et al.'sThe t test (Ho= A=B e H1= B>A) was applied to group G3 to check whether there was any significant difference between the MT and the AT after applying the TEN test. No significant difference was seen between the MT and the AT at all frequencies.Spearman's rank correlation coefficient (rs) (Ho= rs = 0 and H1= rs ≠ 0) was calculated to check whether dead regions could be correlated with different frequencies, in cases of sloping sensorineural hearing loss. Differences between the MT and the AT were reported by frequency in group G3.The correlation is significant only for differences between the MT and the AT between frequencies of 1 kHz and 2 kHz, 6 kHz and 8 kHz, and 3 kHz and 4 kHz for group G3. The correlation is not significant between frequencies of 1 kHz and 8 kHz, and 1 kHz and 6 kHz.A 5dB variation over the absolute auditory threshold following presentation of TEN was considered as indicating the absence of dead regions in the cochlea. This variation was observed in the biological calibration procedure for ten normal ears (group G1; According to Moore et al.'sStatistical analysis showed that the difference between the MT and the AT was higher only for 2 kHz when compared to 1 kHz. Moore et al.Equality relations between 8 kHz and 6 kHz, and 4 kHz and 3 kHz could be associated with the closeness of theses frequencies; a larger difference has been observed from 2 kHz upwards, which suggests constancy in the variation of masked thresholds for high frequencies.Inconclusive results were seen more frequently at frequencies of 6 kHz and 8 kHz , mostly Spearman's rank correlation coefficient was calculated for group G3 to check whether dead regions could be associated with different frequencies in sloping sensorineural hearing loss. There was no significant correlation between distant frequencies, such as between 1 kHz and 8 kHz, and 1 kHz and 6 kHz. For close frequencies, such as 1 kHz and 2 kHz, and 3 kHz and 4 kHz for G3, and 6 kHz and 8 kHz, there was a significant correlation, which could be explained by the excitation diffusion phenomenon in the case of dead regions within the nervous fiber response area. According to Henderson et al.,White noise tests for dead regions, however, require further validation, such as comparisons with PTC results. PTC is a proven and internationally accepted method to study cochlear tuning curves (HalpinSummers et al.The TEN test described by Moore et al.During the TEN test dial readings can by somewhat confusing; according to Moore el al.'sTo address these difficulties in the TEN test, MooreAlthough we saw a significant difference between the MT and the AT at all frequencies, the number of inconclusive results  and the As Moore-Dead regions in the cochlea were present in 92% of cases of sloping sensorineural hearing loss and absent in flat hearing loss.-The TEN test is effective to suggest the presence of dead regions in the cochlea in individuals with sloping sensorineural hearing loss.A critical analysis of our results in this study allows us to conclude that:


This study seeks to investigate the relationship between Obstructive Sleep Apnea-Hypopnea Syndrome (OSAHS) and hearing impairment by meta-analysis.Cochrane Library, PubMed, Embase, Web of Science and other databases are searched from their establishment to July 1st, 2022. Literature on the relationship between OSAHS and hearing loss is collected, and two researchers independently perform screening, data extraction and quality evaluation on the included literature. Meta-analysis is performed using RevMan 5.4.1 software. According to the heterogeneity between studies, a random-effects model or fixed-effects model is used for meta-analysis.P < 0.001), and the average hearing threshold of OSAHS patients is higher than that of the control group . After stratifying the included studies according to hearing frequency, the meta-analysis shows that the OSAHS group has a higher threshold of 0.25, and the response amplitudes at frequencies 2, 4, 6, and 8 kHz are all higher than those of the control group.A total of 10 articles are included, with 7,867 subjects, 1,832 in the OSAHS group and 6,035 in the control group. The meta-analysis shows that the incidence of hearing impairment in the OSAHS group is higher than in the control group (OR = 1.38; 95% CI 1.18–1.62, Z = 4.09, Compared with the control group, the OSAHS group has a higher incidence of hearing loss, mainly high-frequency hearing loss. Thus, OSAHS is closely associated with and a risk factor for hearing loss. Obstructive Sleep Apnea-Hypopnea Syndrome (OSAHS) refers to either hypoventilation (reduced airflow during sleep) or apnea (complete cessation of airflow during sleep) due to the repeated collapse and obstruction of the upper airway , 2. Its Several studies have explored the relationship between OSAHS and hearing loss, and the understanding of the relationship between the two is relatively late. A search of the PubMed database found that most studies on this topic were published after 2003. The conduction mechanism of the inner ear and the transmission of nerve impulses are highly dependent on the supply of oxygen. The conduction mechanism of the inner ear and the transmission of nerve impulses along the hearing are highly dependent on the supply of oxygen, so any significant reduction in local oxygen may have a significant effect on hearing sensitivity. The possibility that OSAHS interferes with the generation and transmission of nerve impulses at the level of the auditory system is currently under consideration –14. KayaThe primary purpose of this systematic review was to explore the OSAHS-based alteration in the auditory system function by conducting a quantitative analysis of presently published data. We compared PTA thresholds in OSAHS patients and controls. Besides, the results of this review may provide an insight into the potential association between OSAHS and hearing loss, thereby providing evidence-based medical evidence for early prevention of hearing loss in patients with OSAHS.According to the Meta-analysis of Observational Studies in Epidemiology (MOOSE) statement  and StanInclusion criteria: ① the subjects were adults over 18 years old; ② the subjects were OSAHS patients clinically diagnosed in the case group, and those without OSAHS in the control group; ③ the study types were cohort study, case-control study and cross-sectional study, which were compared with the control group; ④ the exposure factors were the number of patients with hearing loss and/or pure-tone audiometry indicators.Exclusion criteria: ① repeated published and included studies; ② studies without a control group; ③ reviews, conference abstracts or literature for which the full text could not be obtained; ④ studies with incomplete data or data that could not be extracted wherein the original authors were contacted and did not respond or could not provide meta-analysis data; ⑤ studies with little research information, incomplete data or inconsistent outcome indicators; ⑥ studies in which the subjects had other diseases that could affect hearing function, such as outer or middle ear diseases, high environmental noise exposure or ototoxic drug exposure; ⑦ studies in languages other than English.The data was screened and extracted from all eligible studies by two researchers, then checked against each other. Differences in extracted data were discussed among authors, and studies with differences were provided to a third researcher for analysis to decide whether they should be included. The methodological quality of the included literature was assessed.The Newcastle-Ottawa scale (NOS) was used to evaluate the quality of the included cohort studies and cross-sectional studies . NOS assData extracted from the literature included the first author, study area, publication time, sample size, the total number of cases and mean and standard deviation of study effect indicators. After data extraction, the two datasets were checked and inconsistent data was extracted again. After checking, the data was analyzed.OSAHS was assessed by polysomnography (PSG) or sleep questionnaire, and hearing loss was assessed by pure-tone audiometry. In the included studies, the total number of cases of hearing loss and mean and standard deviation of pure-tone hearing thresholds in the OSAHS and control groups were counted.I2 statistical value test and Q test. If the heterogeneity was not statistically significant (I2 ≤ 50% and P ≥ 0.1), a fixed-effects model was used; if there was heterogeneity (I2 > 50% and P < 0.1), a random-effects model was used , so a fixed-effects model was used for analysis. The results showed that the incidence of hearing loss in the OSAHS group was higher than that in the control group . Thus, OSAHS is associated with and a risk factor for hearing loss.A total of four studies were included , 25, 28 P < 0.001, I2 = 91%), so a random-effects model was used for analysis. The results showed that compared with the control group, the OSAHS group had a significantly higher pure-tone hearing threshold  with statistical significance. To further reduce clinical heterogeneity and explore the effects of OSAHS on pure-tone hearing thresholds at different audiometric test frequencies, a subgroup analysis of pure-tone hearing thresholds with different hearing test frequencies was conducted. A total of four studies reported pure-tone hearing threshold data of different hearing test frequencies between the OSAHS group and the control group. The total heterogeneity was , so a random-effects model was used for analysis , and the pure-tone hearing threshold of the OSAHS group was higher than that of the control group .A total of seven studies were included , 26–30 wanalysis . There wanalysis , 28 on tP < 0.001, I2 = 87%). The results showed that the pure-tone hearing threshold of the OSAHS group was higher than that of the control group , but the results were not statistically significant.There were three studies on the 500 Hz hearing test , 29, 30 P < 0.001, I2 = 87%). The results showed that the pure-tone hearing threshold of the OSAHS group was higher than that of the control group , but the results were not statistically significant.There were four studies on the 1,000 Hz hearing test , 29, 30 P < 0.001, I2 = 91%). The results showed that the pure-tone hearing threshold of the OSAHS group was higher than that of the control group .There were four studies on the 2,000 Hz hearing test , 29, 30 P = 0.02, I2 = 71%). The results showed that the pure-tone hearing threshold of the OSAHS group was higher than that of the control group .There were four studies on the 4,000 Hz hearing test , 29, 30 P = 0.52, I2 = 0%). The results showed that the pure-tone hearing threshold of the OSAHS group was higher than that of the control group .There were three studies on the 8,000 Hz hearing test , 29, 30 In conclusion, the pure-tone hearing thresholds at 250, 2,000, 4,000, and 8,000 Hz in OSAHS patients were higher than those of the control group. After subgroup analysis, the heterogeneity of the pure-tone hearing threshold correlation between the OSAHS group and control group was reduced, especially at 4,000 and 8,000 Hz, suggesting that hearing tests of different frequencies may be the source of heterogeneity.In the sensitivity analysis of the incidence of hearing loss in the OASHS group and control group , the pooP = 0.488 > 0.05, indicating no significant publication bias. There were seven studies on the pure-tone hearing threshold between the OASHS group and the control group. The result of Egger's method : On the one hand, OSAHS episodes are characterized by upper airway pressure imbalance during inspiration. Specifically, this imbalance occurs in the oropharynx and nasopharynx, where the Eustachian tube sits and connects the nasopharynx to the middle ear . OpeningMetabolic disorders: In recent years, an increasing number of scholars have held that the possible mechanism of OSAHS-induced hearing loss is obesity and abnormal hemorheology associated with OSAHS. This leads to a series of pathophysiological changes, which in turn lead to severe metabolic disorders and multi-system damage in the body. Dhanda and Taheri  reportedThis meta-analysis study explored the correlation of OSAHS with the incidence of hearing loss and pure-tone hearing thresholds, and further analyzed the pure-tone hearing thresholds of different frequencies. In addition, Kayabasi et al.  found thLimitations of this study: ① this study only included English literature, which may have generated a certain selection bias; ② the sample size of the included studies on the correlation between OSAHS and hearing loss was relatively small, there was a lack of large-scale, multi-center randomized controlled trials, etc.; ③ part of the study population and control population were based on patients who visited a doctor; thus participants were not randomly selected, so there may have been a selection bias; ④ the included studies were case-control studies with weak causality demonstration ability and low overall level of evidence-based medicine; ⑤ Although high-quality literature was included and strict inclusion and exclusion criteria were established, large heterogeneity was still found, and heterogeneity remained after subgroup analysis according to different hearing test frequencies. After analysis, we did not further study the severity of OASHS in the study population, and it may have had a huge impact on the outcome indicators. Due to the limited number of studies, the relevant subgroup analysis could not be carried out, which affected the reliability of the results and may have caused some bias. Therefore, the conclusions herein need to be interpreted with caution, and a large-sample prospective cohort study should be used for further verification in the future.In conclusion, the prevalence of hearing loss in OSAHS patients is high, and OSAHS can cause hearing loss. Foreign studies have shown that even with timely symptomatic treatment after the onset of deafness, hearing damage is often irreversible . With thThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.CW, FX, MC, CL, XC, XS, and YZ are the guarantor of the manuscript and take responsibility for the content of this manuscript. ZZZ, YW, RC, and CW contributed to the design of the study. QW, HC, JZ, JL, YL, and XO were involved in the data analysis. ZHZ, SL, and RC contributed to the acquisition of primary data. ZZZ, CW, and RC wrote the initial draft of the manuscript. YW, RC, and FX contributed significantly to the revision of the manuscript. All authors read and approved the final manuscript.This study was funded by the Natural Science Foundation of Guangdong Province (2021A1515011373).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.


Pinteraction < 0.0396 and 0.0017, respectively). Positive associations between Ln-transformed 3-PBA and both low-frequency and high-frequency hearing thresholds were observed in participants aged 20–39 years after adjusting confounders  with the highest tertile (≥ 0.884 μg/g creatinine) of 3-PBA compared with the lowest tertile (< 0.407 μg/g creatinine). The possibility of interaction between 3-PBA and age on the hearing threshold shifts indicated that pyrethroid insecticides were prone to be more toxic to auditory system in younger adults than in older ones. Further studies will be required to confirm these findings.Hearing loss (HL) is a global health problem with a high prevalence and profound socioeconomic impact. Pyrethroids are one of the most commonly used insecticides. Although previous studies have reported the relationship between pyrethroids and neurotoxicity, little is known about the effect of pyrethroid exposure on the auditory system among the general population. This study is aimed to investigate the association of pyrethroid exposure with hearing threshold shifts of adults in the United States. A total of 726 adults, aged from 20 to 69 years from the 2011–2012 National Health and Nutrition Examination Survey (NHANES) data were included in the study. Urinary 3-phenoxybenzoic acid (3-PBA), a general pyrethroid metabolite, was used as a biomarker for pyrethroid exposure. HL was defined as a pure-tone average (PTA) at 0.5, 1, 2, 4 kHz ≥ 20 dB in the better ear. Analyses by using multivariate linear regressions were conducted to explore the associations of urinary 3-PBA with PTA hearing threshold shifts. There were no statistically significant correlations between Ln-transformed 3-PBA and either low-frequency or high-frequency hearing thresholds after adjusting for age, gender, race/ethnicity, education level, firearm noise exposure, occupational noise exposure, recreational noise exposure, serum cotinine, BMI, hypertension, and diabetes. However, associations of 3-PBA with both low-frequency and high-frequency hearing thresholds depended on age ( Hearing loss (HL) is the most common human sensory deficit. It is estimated that over five percent of the world’s population (~360 million people) are currently living with a disabling HL . HL is aPyrethroid insecticides are synthetic chemicals derived from natural pyrethrins extracted from the flowers of Chrysanthemum, and they are more stable in the sunlight than pyrethrin. They are the second generation of insecticides which were developed mostly during World War II . Due to More people, including children, are exposed to pyrethroids as the consumption of these synthetic chemicals increases. Previous studies have suggested associations between exposure to pyrethroids and various neurotoxic impairment in humans such as numbness, dizziness, motor neuron disorder, and memory impairment –13. Thouhttps://wwwn.cdc.gov/nchs/nhanes/continuousnhanes/default.aspx?BeginYear=2011). The protocols for NHANES 2011–2012 were approved by the National Center for Health Statistics (NCHS) of the Centers for Disease Control and Prevention (CDC) Institutional Review Board in accordance with the provisions of the Declaration of Helsinki. Written informed consents were obtained from all subjects.All NHANES data for analysis in this study were publicly available on the NHANES website .Urinary concentrations of pyrethroid metabolites, including 3-phenoxybenzoic acid (3-PBA), 4-fluoro-3-phenoxybenzoic acid (4F-PBA) and trans-3--2,2-dimethylcylopropane carboxylic acid (trans-DCCA) were measured using high-performance liquid chromatography with a gradient elution program. Sensitive detection of the analytes is performed by a triple quadrupole mass spectrometer with a heated electrospray ionization source. The approach was based on a modification of the method of Beeson, et al. and Olsson et al. ,21. Morehttps://wwwn.cdc.gov/Nchs/Nhanes/2011-2012/ALB_CR_G.htm).The detection rate of 3-PBA was about 92.1%, whereas only a small percentage of the other two metabolites was observed above the limit of detection (LOD) . Therefohttps://wwwn.cdc.gov/Nchs/Nhanes/2011-2012/AUX_G.htm). Pure-tone air condition average (PTA) hearing thresholds were calculated at low-frequency  and high-frequency  in the better ear. HL was defined as PTA across 0.5, 1, 2 and 4 kHz ≥ 20 dB in the better ear [All audiometric measurements were conducted in a dedicated sound-isolating booth by a trained technician. Hearing thresholds were obtained for both ears of the subjects at frequencies between 500 and 8000 Hz. The detailed audiometry procedure and analytical methods can be found on the NHANES website , serum cotinine, body mass index (BMI), hypertension, and diabetes. Information on age, gender, race/ethnicity, education level, noise exposures, hypertension, and diabetes were obtained from self-reported questionnaires. BMI data were calculated from weight and height data collected during the physical examination.P value of less than 0.05 was considered to be statistically significant.The study used Environmental Subsample C Weights (WTSC2YR) of 2011–2012 NHANES cycle to estimate representative measures for the United States general population, following the analytic guidelines of NCHS ,24. UrinThe study sample comprised 726 adults . The means ± SD of low-frequency and high-frequency PTA hearing thresholds in participants were 7.62 ± 7.43 and 18.53 ± 15.26 dB, respectively. Overall, 11.15% adults had HL. P value for trend shows that 3-PBA levels were positively associated with low-frequency PTA hearing threshold shifts. In the fully adjusted model (model 2), significant P for trend was not observed among tertiles of 3-PBA levels and hearing threshold shifts .Pinteraction = 0.0396 and 0.0017, respectively). People aged 20–39 years in the highest tertile of 3-PBA levels had higher low-frequency and high-frequency PTAs compared to those in the lowest tertile of 3-PBA levels after adjusting for age, gender, race/ethnicity, education level, BMI, hypertension, diabetes, serum cotinine level, noise exposures  (ctively) . Gender ctively) .The current study examined the relationship between urinary pyrethroid metabolite 3-PBA and hearing threshold shifts of adults in the United States in a nationwide cross-sectional study. In this research, we have described a positive association between 3-PBA and both low-frequency and high-frequency PTAs in the 20–39 years old people. To the best of our knowledge, this is the first cross-sectional study that explores the relationship between individual urinary pyrethroid metabolites and hearing threshold shifts of adults in the United States. The findings indicated that pyrethroid exposure might have detrimental effects on human hearing status and that pyrethroid insecticides were prone to be more toxic to auditory system in younger adults than in older ones.The metabolites of pyrethroid insecticides are excreted in urine which makes urine the most suitable matrix for assessing the exposure of humans to pyrethroid insecticides . The detHuman studies to evaluate the effect of pyrethroids exposure on HL are few and contradictory ,17–19,29The mechanisms of increasing pyrethroid exposure to be risk for hearing is still unknown. One study showed ototoxicity of subchronic inhalation exposure to cypermethrin (type II pyrethroid pesticide) in Wistar rats by testing Distortion Product Otoacoustic Emissions (DPOAE) amplitudes . DPOAE iOur study has several strengths. First, the analyses were based on a large and nationally representative sample from the NHANES. Second, pure-tone hearing threshold was measured instead of self-reported HL. Third, confounding factors including age, gender, race, education level, BMI, diabetes, hypertension, serum cotinine level, and noise exposure were adjusted in our analyses for that could result in a misinterpretation .Despite these strengths, some limitations exist. First, the results of this study did not permit a cause-effect relation to be examined because the NHANES is of cross-sectional design. It will be important to continue examining these relationships in longitudinal follow-up studies. Second, we only analyzed data of people in the United States in this study. Statistical analysis of data from broad-ranging surveys in different countries, like China can provide more evidence to support the findings in our investigation. Third, though being a standard approach, pure tone audiometry was used to measure HL in this nationally representative survey, the data would be with more value if evaluation with more sophisticated methods such as DPOAE would have been added to the survey design. In addition, only main confounders reported in previous studies based on prior knowledge and the correlations between exposures and outcomes were included in calculation in the analytical models of this study ,35–41, wWe observed a positive relationship between 3-PBA and both low-frequency and high-frequency hearing threshold shifts in adults aged 20–39 years old in the United States, which indicated the vulnerability of young adults to the toxicity of pyrethroid insecticides. Further studies will be required to confirm these findings.S1 Table(DOCX)Click here for additional data file.


Different noise regimes can reduce the ability to detect sounds of conspecifics due to an upward shift of the hearing threshold, a phenomenon termed masking. We therefore investigated the masking effect of white noise on the auditory thresholds in female croaking gouramis . We hypothesized that noise would influence the detection of conspecific vocalizations and thus acoustic communication. The auditory evoked potentials (AEP) thresholds were measured at six different frequencies between 0.1 and 4 kHz using the AEP recording technique. Sound pressure level audiograms were determined under quiet laboratory conditions (no noise) and continuous white noise of 110 dB RMS. Thresholds increased in the presence of white noise at all tested frequencies by 12–18 dB, in particular at 1.5 kHz. Moreover, hearing curves were compared to spectra of conspecific sounds to assess sound detection in the presence of noise in various contexts. We showed that masking hinders the detection of conspecific sounds, which have main energies between 1.0 and 1.5 kHz. We predict that this will particularly affect hearing of female’s low-intensity purring sounds during mating. Accordingly, noise will negatively affect acoustic communication and most likely reproductive success.An increasing level of anthropogenic underwater noise (shipping, drilling, sonar use,  Malurus cyaneus, showed that background noise affected the response to alarm calls, probably due to acoustic masking rather than distraction or changes in vigilance  up to several kHz and lowers the auditory thresholds over the entire frequency range  we compared unmasked and masked hearing thresholds to the spectra of conspecific sounds. These comparisons will clarify the extent to which noise reduces the ability of T. vittata to detect conspecific sounds and correctly assess opponents and mates (The aim of the study is twofold: (1) we measured unmasked and masked hearing thresholds to determine the extent to which standardized white noise deteriorates the sound pressure sensitivity in a vocal hearing specialist, the anabantoid nd mates .Animals were handled as described previously in Ten adult female croaking gouramis were used for the experiments , obtained from a local aquarium store in Vienna. Sexing of fish was based on the presence of the whitish ovary in females . FemalesAll applicable national and institutional guidelines for the care and use of animals were followed .−1(1−2.5 µg g−1) and enabled the fish to breathe during the experiment but with only slight opercula movement that prevented an excessive myogenic noise level, which could interfere with the recordings. Individuals that started to move prior to the end of the measurements were not immobilized a second time. This explains the different numbers of females measured at different frequencies  recording technique , which rested on a vibration-isolated concrete plate. The entire setup was enclosed in a semi-soundproof room constructed as a Faraday cage  to a dual-cone speaker , which was placed 1 m above the tub. Sound stimuli were presented as tone bursts at a repetition rate of 21 per second. Hearing thresholds were determined at frequencies of 0.1, 0.5, 1, 1.5, 2 and 4 kHz, presented in random order. All bursts were gated using a Blackman window. The stimuli were presented at opposite polarities (180° phase shifted) for each test condition and the corresponding AEPs were averaged by the BioSig RP software in order to eliminate stimulus artefacts. The sound pressure level (SPL) of tone-burst stimuli was reduced in 4 dB steps until the AEP waveform was no longer apparent . The lowi.e., center) sinusoid of a particular tone-burst recording was determined. This RMS voltage was then used to calculate the absolute SPL re 1 µPa based on the sensitivity of the hydrophone and the amplification factor of the hydrophone amplifier and of the TDT system  was placed at the same position as the fish, after the AEP measurements (relative hearing thresholds at different frequencies with and without white noise). Using BioSig RP, the RMS voltage of the largest  to a 30-band equalizer (Alesis DEQ 230) to obtain a flat noise spectrum underwater and fed to the second channel of a signal mixer (SM5 of TDT System 3). The tone burst signals were fed to the first channel of the signal mixer. Both signals were then fed via the Alesis RA 300 amplifier to the dual-cone speaker (Tannoy System 600). The SPLs of the masking noise were measured at the position of the fish using the hydrophone, which was connected to a power supply (Brüel & Kjaer 2804) and a sound level meter (Brüel & Kjaer 2238 Mediator). We determined L-weighted (5 Hz to 20 kHz) equivalent continuous SPL (LLeq) averaged over 1 min measuring time. The Leq is a measure of the averaged energy in a varying sound level and commonly used to assess environmental noise. We also measured background noise levels in the experimental test tank . After SPL measurements, the background noise and the white masking noise were recorded via an external sound card  on a PC. Recording and analyzing were done using S_Tools-STX 3.7.8, an acoustics, speech, and signal processing application developed by the Acoustics Research Institute at the Austrian Academy of Sciences, Vienna. Sound spectra of 1 min recordings were calculated by an FFT analysis using a filter bandwidth of 1 Hz. Absolute spectral values were calculated from the relative spectral values  and in the presence of continuous white noise played back at a sound level of 110 dB re 1 µPa (RMS)  and Bonferroni Audiograms of the different experimental groups (no noise and white noise) were compared by two-factor analysis of variance (ANOVA) using a general linear model where one factor was noise-treatment and the other was frequency. The noise-treatment group factor alone should indicate overall differences between different treatments of animals, and in combination with the frequency factor if different tendencies exist at different frequencies of the audiograms. All calculations were done using SPSS 26 .T. vittata revealed best hearing ability at 1.5 kHz (average threshold: 86 dB re 1 µPa) and lowest sensitivity at 4 kHz (average threshold: 109 dB re 1 µPa) (The lab noise (No Noise) SPL-audiogram of e 1 µPa) , Fig. 2.e 1 µPa) .p < 0.001) and a significant interaction between noise condition and frequency . Thus, changes in thresholds showed different trends at different frequencies.Playback of white noise drastically lessened auditory sensitivity. Comparing audiograms by a two-factor ANOVA revealed significant overall differences , but conducted on fish from different orders possessing hearing specializations  significantly increased the auditory thresholds, and that this masking effect was maximal within the most sensitive hearing range of rmatulus  and 11 daculatus . This inaculatus . SimilarLagodon rhomboides and the African mouth-breeder Tilapia macrocephaIa revealed that hearing thresholds depend on background noise levels at particular frequencies  showed that traffic noise from streets may masks acoustic signaling in fish such as shiners (family Cyprinidae) inhabiting shallow freshwater streams.How do these observations and current results relate to hearing in croaking gouramis in their natural habitats? This species inhabits shallow, still, densely vegetated waters in South-East Asia. Labyrinth fishes overcome the lack of oxygen in such still tropical waters by airbreathing using the suprabranchial chamber dorsally of the gills . Still wT. vittata under laboratory noise conditions. The spectral sound energy was at least 20 dB above the baseline hearing thresholds at the most sensitive frequency of 1.5 kHz. Hearing provides fishes with important information in a broad range of environments, making sound a key cue for perhaps most aquatic animals , probably failed to detect male vocalizations during courtship. In croaking gouramis, both sexes emit croaking sounds during dyadic contests, which differ only in SPL because male sounds are 4-5 dB louder than those of females. In contrast, only females produce purring sounds prior to spawning, which are only half as long and half as loud as their croaking sounds (6 dB lower in SPL)  . We assu in SPL) . Note thC. chromis as well as the brown meagre, conservative calculations regarding the distance at which conspecific sounds are detectable (brown meagre) yielded brief decreases from 500 m under ambient noise conditions to about 1 m in the presence of noise . Together with the current study, these data indicate that noise does influence the hearing ability in fish and may therefore negatively impact communication distances, reproductive success, and aquatic ecosystems in general.Similar findings in the male Lusitanian toadfish demonstrate that the detectability of their boatwhistle calls decreased considerably in the presence of noise: sound frequencies were detectable only up to 300 Hz, indicating that noise decreases the ability of females to find nest sites of calling male nest sites and assess mates . In the of noise . MoreoveTurdus merula) and great tits (Parus major) apparently benefit from closing the distance between them, simply by moving upward to a higher perch  an increase of hearing thresholds by approximately 20 dB in the presence of boat noise, but no such effect under either quiet laboratory or ambient noise conditions. Similar observations were made in the Lusitanian toadfish: the main energies of ferry-boat noise were within the most sensitive hearing range and increased its auditory threshold by up to 36 dB at most frequencies tested. This indicates that acoustic communication is affected by masking of their hearing abilities (In general, an upcoming problem in natural environments is the masking effect due to the expansion of traffic noise, ship noise, and aquatic industrial activities. All have increased in recent years and have led to concern about the effects of man-made sounds on aquatic life . Birds eer perch . In sevebilities . In the bilities .amount of acoustic and visual signaling during agonistic behaviour. But T. vittata is unable to adapt its sound characteristics, and our conclusion is that noise can therefore negatively affects sound detection by masking the thresholds of female T. vittata. This would impact acoustic communication during agonistic and mating behaviour and affect the assessment of opponents and mates during territory defense and reproduction. A future study on the effect of noise on males would help clarify the extent to which noise impacts their hearing ability and potentially influences reproduction success.This is the first study to investigate the effect of noise on the AEP thresholds in the croaking gourami, a highly vocal labyrinth fish species. We know from a former study  that noi10.7717/peerj.14230/supp-1Supplemental Information 1All SPLs are given in dB re 1 µPa.Click here for additional data file.10.7717/peerj.14230/supp-2Supplemental Information 2Click here for additional data file.


Acquired middle ear cholesteatoma can be classified as primary or secondary. Although both can result in hearing loss, it is still controversial whether there is an association between the type of cholesteatoma and the degree of hearing loss.To analyze the association between hearing loss and the type of acquired cholesteatoma, and the status of the ossicular chain.This was a cross-sectional historical cohort study involving patients diagnosed with acquired cholesteatoma who were surgically treated. Air and bone conduction thresholds, air–bone gaps and the status of the ossicular chain were analyzed for both types of cholesteatoma.Eighty patients aged 5–57 were included in the study. Fifty-one patients had primary cholesteatoma and 29 had secondary cholesteatoma. Both types of cholesteatoma determined greater air–bone gaps at 0.5 kHz. Secondary cholesteatoma determined greater hearing loss in all analyzed frequencies and higher air conduction and air–bone gap means.There was association between hearing loss and the type of cholesteatoma. Secondary cholesteatoma resulted in greater hearing impairment. Cholesteatoma is defined as the presence of skin in any air-filled area of the temporal bone, and can be congenital or acquired.8Studies have shown that the incidence and prevalence of cholesteatoma are much higher in developing countries, highlighting its association with low socioeconomic status, poor hygiene, and delay in seeking health care or poor health services.10Cholesteatomas determine mechanical compression of adjacent structures and have cells with hyperproliferative characteristics , which may cause temporal bone and ossicular chain erosion, resulting in hearing loss.There is no consensus in the literature regarding the influence of the type of cholesteatoma on hearing loss, or concerning the frequencies most affected.17Most studies have shown that there is an association between ossicular chain erosion and magnitude of hearing loss, and that the most affected ossicle is the incus.17The aim of this study was to analyze the association of primary and secondary acquired cholesteatoma, as well as ossicular chain erosion with hearing loss.The study was approved by the Ethics Committee of the institution under No. 03856512.4.0000.5479.This was a historical cohort study with cross-sectional design, based on data obtained from medical records of patients treated at the Otorhinolaryngology Outpatient Clinic of a medical teaching institution, between January of 2010 and October of 2013.Inclusion criteria: patients who were diagnosed with acquired cholesteatoma and submitted to otological surgery.Exclusion criteria: patients with other concomitant ear diseases or those who had been previously submitted to otological surgery.Prior to surgery, all patients underwent pure tone audiometry with determination of air thresholds at 0.25, 0.5, 1–4, 6, and 8 kHz, and bone thresholds at 0.5 and 1–4 kHz. Air and bone thresholds and air–bone gaps were evaluated separately at the frequencies 0.5, 1, 2, and 4 kHz. The means were also evaluated for each patient at those frequencies.The variables studied were: (a) age and gender; (b) type of cholesteatoma: defined according to clinical history, physical examination data, and intraoperative findings described in the standardized surgical description form, classified as primary cholesteatoma when originating from pars flaccida retraction and secondary when originating from the involvement of pars tensa of the tympanic membraneStatistical analysis was performed using Stata™ software, version 11. The association between hearing loss and the assessed variables was performed using Pearson's chi-squared test for categorical variables, and the nonparametric Wilcoxon–Mann–Whitney test for continuous variables, considering a 95% confidence interval and 5% level of significance.The present study included 80 patients, 31 females and 49 males, aged between 5 and 57 years (mean age of 26.5 ± 14.75). The left ear was affected in 52.5% of the patients and the right ear, 47.5%. Primary cholesteatoma was found in 51 (63.8%) patients and secondary cholesteatoma in 29 (36.2%) patients. There was no statistically significant difference between the two groups in relation to gender and age .Table 1DAs for the hearing impairment at the different frequencies, it was observed that the bone threshold was higher at frequencies 2 and 4 kHz, both in primary and secondary cholesteatoma ; the airAs for the degree of hearing loss, it was observed that secondary cholesteatoma caused greater loss than primary cholesteatoma at all analyzed frequencies, as well as a larger gap at the frequencies of 1 and 2 kHz , Table 4Only five (6.25%) patients had an intact ossicular chain during surgery, two of them with primary cholesteatoma and three with secondary cholesteatoma. Patients with intact ossicular chain had a mean gap of 26.25 dB (SD ± 7.55 dB) and patients with ossicular chain erosion, 31.77 dB (SD ± 13.54 dB).The aim of this study was to primarily investigate the association of the types of cholesteatoma with hearing loss and, secondarily, the association between the ossicular chain status and hearing impairment.The two types of cholesteatoma caused a greater gap at the low frequencies, and 0.5 kHz was affected to a greater degree. Thus, in common with that reported in the literature,The difference in hearing loss between both types of cholesteatoma is still controversial in the literature, as the different methodologies make it difficult to compare results.19The literature indicates that the degree of hearing loss is not a good predictor of ossicular chain status.The present study was performed in a public health service, where consultations with specialists, complementary examinations, and surgical treatment still require considerable time.This study has some limitations. The study includes patients treated at an outpatient clinic rather than a random patient sample, which reduces its ability to be generalized. Additionally, the lack of information on the socioeconomic characteristics of patients and disease duration makes it impossible to investigate possible associations of these variables with hearing loss and the development of explanations for the high prevalence of ossicular chain erosion.The type of acquired cholesteatoma resulted in statistically significant differences regarding hearing loss. Secondary cholesteatoma results in greater air threshold impairment and air–bone gap than the primary cholesteatoma. Additionally, both types of cholesteatoma resulted in a greater air–bone gap at the lower frequencies.10.13039/501100003593CNPq.This study was funded by the PIBIC Scientific Initiation Grant – The authors declare no conflicts of interest.


Taeniopygia guttata) and a precocial  bird species. We found that pHisH3-immunoreactive apical and non-apical mitoses were abundant in the T. guttata retina at the hatching stage. In contrast, pHisH3 immunoreactivity almost disappeared from the quail retina at the embryonic day 10 (E10). Furthermore, we also found that the onset of the appearance of non-apical mitoses occurred at later stages in the altricial bird species than in the precocial one. The disappearance of apical mitoses and the spatiotemporal distribution of non-apical mitoses followed central to peripheral and dorsal to ventral gradients, similar to gradients of cell differentiation described in the retina of birds. Therefore, these results suggest that retinal neurogenesis is active at the hatching stage in T. guttata, and that horizontal cell differentiation is delayed in the altricial bird species compared to the precocial one. Together, this study reveals important insights into the timing differences that regulate bird retinal maturation and provides a better understanding of the evolution of avian altriciality and precociality.During development of the vertebrate retina, mitotic activity is defined as apical when is located at the external surface of the neuroepithelium or as non-apical when is found in more internal regions. Apical mitoses give rise to all retinal cell types. Non-apical mitoses are linked to committed horizontal cell precursors that subsequently migrate vitreo-sclerally, reaching their final position in the outer surface of the inner nuclear layer, where they differentiate. Previous studies have suggested differences in the timing of retinal maturation between altricial and precocial bird species. In the present study we analyze qualitatively and quantitatively the mitotic activity in the developing retina of an altricial (zebra finch,  Developmental events in the retina, such as cell proliferation, cell migration, and cell differentiation occur in a precise and predetermined order . The retIn nature, there is a wide spectrum of avian hatchlings from altricial to precocial extreme . At hatcTaeniopygia guttata, Vieillot 1817)  is a notable avian model species that belongs to the order of Galliformes. It has been widely used by researchers to study several aspects of visual development, such as the entry of microglia in the developing retina (The chicken  and the ot 1817) ,2021b cog retina  and the g retina . HoweverStudies on detailed direct comparison of different aspects of retinal development between precocial and altricial bird species are sparse. The aim of the present study is to compare patterns of retinal development, specifically mitotic activity, in precocial and altricial bird species. The arised results could provide new insights into the timing of cell proliferation and cell differentiation in the bird retina and a greater understanding of the evolutionary mechanisms involved in the process of speciation within the precocial-altricial spectrum.T. guttata (n = 63) and C. coturnix (n = 45) eggs were placed within a rotating egg incubator designed for chicken eggs  and were maintained at 37.5 ± 1°C and 80–90% humidity. Eggs were placed in a small padded dish on the rotating bars of the incubator. They roll within the dish preventing embryo adhesion to the shell. A total of 34 T. guttata embryos and 34 embryos of C. coturnix and 2 newly hatched animals were used in the present study . Experimental protocols were approved by the Bioethics Committee for Animal Experimentation of the University of Extremadura (Ref 264/2019). Fertilized nt study . The oldSections were subjected to an antigen retrieval process with citrate buffer (pH 6.0) at 90°C during 30 min. Sections were chilled at room temperature for 20 min. Slides were washed one time in 0.1% Triton-X-100 in PBS (PBS-T) and two times in 0.2% gelatin, 0.25% Triton-X-100 in PBS (PBS-G-T) and then pre-blocked in 0.2% gelatin, 0.25% Triton-X-100, Lys 0.1 M in PBS (PBS-G-T-L) for 1 h. Sections were incubated with the primary antibody over night at RT in a humidified chamber:1.rabbit anti-phisH3  polyclonal antibody, that identifies cells in late G2/M-phase in the chicken  and zebr2.rabbit anti-Prox1 polyclonal antibody , that identifies a transcription factor that is present in all horizontal cells in the chicken  and in t3.mouse anti-Islet1 (Isl1) monoclonal antibody , that labels a subpopulation of horizontal cells in the retina of the chicken  and zebrAfter incubation with the primary antibody, slides were washed several times in PBS-T and PBS-G-T and incubated with Alexa Fluor 488 goat anti-mouse IgG antibody , Alexa Fluor 488 goat anti-rabbit IgG antibody  and Alexa Fluor 594 goat anti-rabbit IgG antibody  during 2 h at RT in a humidified chamber in darkness. Sections were washed two times in PBS-T and one in PBS-G-T in darkness. The tissue was incubated for 10 min with DAPI at RT and then washed three times in PBS. Slides were mounted with Mowiol .Immunofluorescence sections were observed with a bright field and epifluorescence Nikon Eclipse 80i microscope and photographed using an ultra-high-definition Nikon DXM1200F digital camera. Images were processed with Adobe Photoshop CS4.1 accessed on 28 January 2021). The density profiles were expressed as the mean ± sem of the number of pHisH3-positive mitoses per square millimeter (mitosis/mm2). Statistical analyses and the graphs were performed with the R programming language. To analyze if there were significative differences, the Mann–Whitney U test was used. Differences between groups were considered as significant when P < 0.05 and highly significant when P < 0.001.The eyes of the embryos were sectioned parallel to the sagittal plane , allowinToluidine blue-stained semi-thin sections in the undifferentiated quail retina showed abundant mitoses mainly localized in the scleral surface of the neuroblastic layer (NbL), but sparse non-apical mitoses were also observed. We also evaluated the specificity of primary antibodies in the quail retina by comparison with published examples of labeling results and immunoassays in other bird species, and we found similar expression patterns for pHisH3, Prox1, and Isl1 antibodies see . None ofC. coturnix retina did not show immunoreactivity against pHisH3 neither in central  the laminated structure of the retina could be distinguishable in the Dc sector . At thisNon-apical mitoses were mainly detected in the Vc and Vcp sectors by E10-E13 , 6A, coiAt E13, non-apical mitoses were absent from the dorsal retina , 6A and C. coturnix, showed that non-apical mitoses were mainly concentrated in the dorsal sectors of the NbL, but also in the Vc sector (The St27 (E5) retina, the first stage analyzed in c sector , 11A. Prc sector . Betweenc sector , 11A, buc sector , 11A, coc sector , 9D and c sector , 9E immuc sector .At E8-9 non-apical mitoses almost disappeared from the Dc, Dcp, Vc, and Vcp sectors, but they were still detected in the Dp and Vp sectors , 11A. AcTherefore, non-apical divisions in the developing retina of birds spread following central to peripheral and dorsal to ventral gradients, and their appearance coincides chronotopographically with the onset of Prox1 immunoreactivity. When the non-apical divisions disappear, Prox1-immunoreactivity is located mainly in the nuclei of differentiated horizontal cells. Additionally, the disappearance of apical mitoses followed the same gradients described above. Finally, while abundant proliferative activity is detected in the retina of altricial birds at hatching, residual mitotic figures are only observed in the peripheral region of the precocial retina from E10 until the hatching stage (E16.5).G. gallus . However, mitotic activity almost disappears from the apical surface in the entire quail retina at E10. Similar results were found when monitoring the disappearance of non-apical cell divisions. By these embryonic stages, the quail retina is fully differentiated and the main retinal cell types could be characterized immunohistochemically . Then, our findings are in line with previous studies showing that neurogenesis ceases at embryonic stages in the precocial bird retina, and hatchlings present a mature and functional visual system .G. gallus than in T. guttata (T. guttata take place at E6 (St30), while in C. coturnix non-apical mitoses were numerous at E5 (St27). Furthermore, the density of non-apical mitosis in the central region of the T. guttata retina reached a peak at E8-9, whereas the highest density values were found by E7.5-8 in C. coturnix (present study) and in G. gallus . Although we never detected co-labeling of these horizontal cell markers with non-apical pHisH3 mitoses, there was a close spatiotemporal relationship between the onset of the appearance of non-apical pHisH3-immunoreactive mitoses and the onset of Prox1-expression in the different retinal sectors, in concordance with the results obtained in the chicken (Non-apical mitoses have been linked to horizontal cell differentiation in fish  and bird guttata  and in t chicken . These fIn conclusion, the present study provides comprehensive data on distinct patterns of retinal development between the altricial zebra finch and the precocial Japanese quail, which may serve as empirical reference in future studies. While the order of different events involved in retinal maturation is the same during development, their specific timing differs between the altricial and precocial bird species. The retina of the precocial birds exhibits higher degree of maturation at hatching, thus providing evidence for the notions that precocial species might have acquired the morphological machinery required to attain their higher functional state at hatching. Together, our results expand our current understanding of the timing and cellular differences that regulate patterns of avian retinal growth and maturation, and provides a better understanding of the evolution of avian altriciality and precociality.The original contributions presented in the study are included in the article/The animal study was reviewed and approved by Comité de Ética en Experimentación Animal.JF-M, GM-P, and JR-L contributed to conception and design of the study. JM-R, AM, and YG organized the database. GM-P performed the statistical analysis. JF-M wrote the first draft of the manuscript. GÁ-H wrote sections of the manuscript and performed histological and immunohistochemical analysis. AA performed the immunohistochemical analysis. All authors contributed to manuscript revision, read, 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.


The activities of daily living (ADL) ability level of an elderly patient is an important indicator in determining the patient’s degree of degenerative brain disease and is mainly evaluated through face-to-face interviews with doctors and patients in hospitals. It is impossible to determine the exact ADL ability of a patient through such a temporary interview, and the pursuit of accurate ADL ability evaluation technology is a very important research task worldwide. In this paper, in order to overcome the limitations of the existing ADL evaluation method mentioned above, first of all, a self-organized IoT architecture in which IoT devices autonomously and non-invasively measure a patient’s ADL ability within the context of the patient’s daily living place was designed and implemented. Second, a remote rehabilitation treatment concept for enhancing the patient’s ADL ability we call an “e-coaching framework”, in which a doctor remotely gives an instruction in a specific ADL scenario, and the patient’s ability to understand and perform the instruction can be measured on-line and in real time, was additionally developed on top of the self-organized IoT architecture. In order to verify the possibility of remote rehabilitation treatment through the proposed architecture, various remotely directed ADL scenarios were performed and the accuracy of the measurements was verified. If it is possible to remotely rehabilitate patients with degenerative brain disease who are not able to move freely, it will prove to be a very important and effective solution in the current COVID-19 pandemic era. In order for this technology to become possible, a technology that accurately identifies a patient’s daily living ability at home must be developed.Activities of daily living (ADL) refers to the abilities essential for daily life, such as moving, cooking, and using tools . BecauseIn this paper, in order to overcome the limitations of the existing ADL measurement method mentioned above, first of all, a self-organized IoT platform in which IoT devices autonomously and non-invasively measure a patient’s ADL ability in their daily living place was designed and implemented. Second, a remote rehabilitation treatment concept for enhancing the patient’s ADL ability, we call an “e-coaching framework”, which is a framework in which a doctor remotely gives an instruction in a specific ADL scenario, and the patient’s ability to understand and perform the instruction can be measured on-line and in real time, was additionally developed on top of the self-organized IoT platform.The ADL measurement scenarios required for the proposed e-coaching framework were implemented as rule knowledge-based systems . For theThis paper is organized as follows. A “smart home” is a residential environment, such as an apartment or house, with wired and wireless communication networks consisting of various IoT devices and a central control unit called a hub . CurrentCurrently, various studies are being conducted on services that objectively evaluate users’ conditions through the ADL measurements of patients with cognitive impairment using various sensors and IoT platforms. For instance, one study measured the movement state of users in a living environment and warned them according to the ADL evaluation results . It usedAnother research study focused on an integrated system that detects user behavior from sensors placed in the homes of dementia patients and transmits and records the patient’s ADL through cloud services . This alThe basic concept for the remote e-coaching framework is shown in The user’s daily behavior consists of several unit-behaviors. For example, to do “flush the toilet and turn off the light,” small units of actions such as “enter the bathroom”, “flush the toilet”, “turn off the light”, and “get out of the bathroom” must be performed in order. In this paper, this kind of daily behavior  is referred to as an “ADL scenario”. Every ADL scenario in the system consists of small goals composed of several stages, and the user must perform each stage accurately, in order, and in time. Each stage that constituted the scenario was named a “step”, and the goal unit-behavior of each step was named the “target behavior”.First, in an “abnormal” situation, after the analysis of which part of the user’s behavior does not match the target behavior, coaching is immediately provided so that the correct behavior can be performed. Moreover, in a “timeout” situation, when the user does not do anything within the time limit of the step, the proposed system coaches the user on the target behavior of the step. As can be seen from First, “sensor” devices are attached to several home appliances and furniture in the home. When the user uses a home appliance or furniture, sensor data are collected from an “IoT device” that communicates with the sensors which is either wired or wireless. This IoT device is located in each unit-space, such as the living room, kitchen, or toilet, so it can combine the data with its own location information, which allows it to determine the user’s current location and behavior. The “e-coaching hub” is a device that implements the e-coaching framework (a system proposed by this paper) and that is placed in a suitable position for stable communication with all of the IoT devices.For all the inference processes that occurred during ADL evaluation, a relevant knowledge base was implemented as a rule-based system software tool called CLIPS (C Language Integrated Productions System) . CurrentNumerous situations can occur as a user performs each scenario for ADL evaluation. Therefore, implementing rules for all these situations is not only difficult but also inefficient. Thus, this paper modularized the rules for several inferences required in the proposed system in order to solve this problem and to further consider the scalability of users, sensors, and ADL scenarios.This paper provides ADL scenarios consisting of unit-behaviors that can be sufficiently known only by using the sensor values of IoT devices  and implements these as objects to be used efficiently in the modular rules described above.Every ADL scenario consists of several steps. If all steps are performed accurately in order, the ADL evaluation for a scenario ends. When a timeout situation occurs because a user does not act during an ongoing step, the proposed system provides coaching about the step at the appropriate time based on the time limit value of the user table and the CLOCK object representing the current time.In this paper, in order to verify the effectiveness of the proposed e-coaching framework and knowledge base system, a testbed in a real environment was constructed and tested, as shown in the The sensors in this testbed are mobile tags that can be attached to home appliances, and a current sensor to check whether a hairdryer is used has also been installed. The mobile tag has 6-axis acceleration, 3-axis gyro, and ambient light sensors, and has a built-in BLE communication function. An IoT device in a unit space performs BLE bidirectional communication with mobile tags, and an IoT device uses an e-coaching hub and MQTT protocol. In order to minimize the patient’s behavior time error due to the delay rate caused by MQTT messaging, the time measured based on the e-coaching hub time for communication between sensors and the IoT device is regarded as the patient’s execution time. A time stamp for each time value is embedded in the MQTT message and sent as data to the e-coaching hub. Therefore, the effect of the communication delay between the e-coaching hub and the IoT device is not included in the patient behavior measurement time.Does the system accurately recognize the numerous unit-behaviors performed within the time limit?How long does it take for the system to recognize a unit-behavior?In performing the ADL scenario, determining how quickly and accurately the proposed system recognizes the user’s behavior is necessary. Thus, the evaluation factors were as follows:T_act, the time it took for the system to recognize each unit-behavior, was obtained following the method for obtaining T in T_(act.avg)”, was also obtained.Eleven unit-behaviors were able to be performed while using the objects in the environment in T_(act.avg). Each unit-behavior was performed 30 times—for a total of 330 times—to obtain accurate results. The time limit for each behavior was arbitrarily set to 3 s.T_act was measured as 416ms.A total of 309 times, actions were accurately recognized, and the accuracy was 93.6%. The percentage of behavior recognition failure due to defective sensors or problems in the data transmission process was 3.9%. The ratio of behavior non-recognition within the time limit was 2.5%. Moreover, the time T_act values than those of the other behaviors. The sensors connected to the induction and the hair dryer used in the performance evaluation measure the power consumption, so they recognize that they are in a “TURN_ON” or “TURN OFF” state when the power value gradually changes and reaches a certain value. Therefore, the proposed system recognizes the ON/OFF state of home appliances relatively slowly. Thus, it is confirmed that the accuracy levels for these behaviors were less than those for the other sensors simply because they exceeded the time limit.The proposed system should not only recognize the user ADL, but also be able to provide quick and appropriate coaching services depending on the situation. Thus, the coaching service performance was evaluated by appropriately placing abnormal situations that may occur during ADL evaluation in each step of the scenario in an IoT test environment such as that in the previous performance evaluation.The first line in When the user performs an abnormal behavior within the time limit, is coaching provided immediately?The evaluation factors are as follows:Is coaching provided as soon as the time limit for the ongoing step is reached?The remote e-coaching framework proposed in this paper enabled real-time ADL evaluation with the help of an external doctor so that a patient with cognitive impairment could perform rehabilitation treatment in a residential environment equipped with various sensors. It can thus be applied to patient rehabilitation by being used to provide real-time coaching services for abnormal situations, furthering several existing studies that recognized or evaluated user behavior using sensor data. It is a more convenient and objective approach than evaluating ADL within existing medical facilities, as the use indirect methods, such as taking personal videos and doing evaluations with a close caregiver. Additionally, this system can be applied not only to a user’s home but also to care facilities which have many patients with similar diseases. The proposed system showed high accuracy in the area of behavioral cognition or coaching services using a rule-based system. However, its determination of ADLs is restricted by the limited use of sensors. Moreover, even if numerous different types of sensors were used in IoT platform environments, there would still be difficulties in accurately recognizing all the users’ behaviors using only sensors. These problems can be sufficiently solved by deep learning based on various sound data or real-time change values in the environmental data that occur during daily activities. In these cases, more diverse ADLs will be recognized, and practical ADL evaluation scenarios can be constructed and implemented.We hope that the proposed e-coaching framework can aid patients with cognitive impairment in living stable lives in their homes with the help of IoT devices.


Low- and high-frequency hearing impairment were assessed using data from pure-tone audiometric testing. A Cox proportional hazards model was applied to estimate hazard ratio (HR) values for hearing impairment. Low eGFR did not increase the risk of low- or high-frequency hearing impairment. For males, multivariable-adjusted HR of high-frequency hearing impairment was 1.16  for the upper half of the ≥90 mL/min/1.73 m2; however, this positive association between high eGFR and high-frequency hearing impairment did not appear to be robust in a number of sensitivity analyses. We conclude that, among the Japanese working-age population, eGFR was not generally associated with hearing impairment in people of either sex.Several longitudinal studies have examined associations between renal dysfunction and hearing impairment. Here, we explored the longitudinal association between estimated glomerular filtration rate (eGFR) and hearing impairment among the working-age population in Japan. Participants were 88,425 males and 38,722 females aged 20–59 years, without hearing impairment at baseline (2013), who attended Japanese occupational annual health check-ups from 2013 to 2020 fiscal year. eGFR was categorized into four groups , 60–89, and <60 mL/min/1.73 m Similarly, the four categories < 60 mL/min/1.73 m2 were summarized as one group because few participants had eGFR < 60 mL/min/1.73 m2. For the final analyses, participants were classified into four groups: eGFR upper half of ≥90, lower half of ≥90 (reference), 60–89, and <60 mL/min/1.73 m2.Participants were classified into six categories , exercise was categorized as exercises causing light sweating exercises ≥30 min/time at frequency of ≥2 days/week, for ≥1 year, and job type was categorized as professional job, management, office job, sales, service, telecommunications, manufacturing, and other.Information on potential confounding factors that have been associated with hearing impairment ,24,25,262 group as the reference group. This analysis was performed according to participant sex because there are sex differences in renal and hearing impairments [Person-years were calculated from the day of the annual health check-up in the 2013 fiscal year (baseline) to the day of hearing impairment onset, or the day of the last annual health check-up, until March 2021 (whichever came first). A Cox proportional hazards model was applied to estimate hazard ratio (HR) and 95% confidence interval (CI) values for associations between eGFR and low- and high-frequency hearing impairment, relative to the lower half of the ≥90 mL/min/1.73 mairments ,32,33,342. Further, to avoid overestimation of eGFR, based on low serum creatinine level related to low skeletal muscle mass, we excluded participants with serum creatinine levels below the standard value .Several restricted analyses were performed as sensitivity analyses, to minimize the residual confounding, as follows: participants without diabetes, nonsmokers, those < 40 years old, those engaged in office jobs, and those with BMI 18.5–24.9 kg/mAll statistical analyses were performed using Stata version 14.2 .2, respectively. Mean follow-up period was approximately 5.3 years.Participant baseline characteristics, stratified according to eGFR group, are shown in 2 was positively associated with low-frequency hearing impairment in the crude model; however, this association disappeared after adjustment for age. Compared with those in the lower half of the eGFR ≥ 90 mL/min/1.73 m2 group, the HR values for low-frequency hearing impairment were 0.84  for eGFR 60–89 mL/min/1.73 m2 and 0.93  for eGFR < 60 mL/min/1.73 m2 in the multivariable model. Similarly, there was no association between eGFR < 60 mL/min/1.73 m2 and high-frequency hearing impairment . Meanwhile, in the upper half of the ≥90 mL/min/1.73 m2 group, an elevated risk of high-frequency hearing impairment was observed ; however, the association between high eGFR and high-frequency hearing impairment appeared not to be robust in several sensitivity analyses. Further, after exclusion of males with low creatinine level, the HR value was 1.14 . In females, eGFR appeared to increase the risk of low- and high-frequency hearing impairment in the crude model; however, after adjustment for several confounding factors, no association between eGFR and hearing impairment was detected.HR values for hearing impairment according to eGFR group are shown in 2), and anemia in males, and diabetes and underweight (BMI < 18.5 kg/m2) in females, were associated with increased risk of hearing impairment.HR values for hearing impairment related to covariates in the multivariable model are summarized in Among the working-age population in Japan, there was no association of eGFR with low-frequency hearing impairment in either sex or with high-frequency hearing impairment in females. For males, although high eGFR appeared to be associated with an elevated risk of high-frequency hearing impairment, this positive association was not consistent in sensitivity analyses.2, sex-combined multivariable-adjusted HR values for pure-tone average (PTA) > 25 dB hearing impairment at low frequencies  were 0.88  for eGFR < 60 mL/min/1.73 m2 and 1.19  for eGFR ≥ 90 mL/min/1.73 m2. Further, HR values for PTA > 25 dB hearing impairment at high frequencies  were 0.90  for eGFR < 60 mL/min/1.73 m2 and 1.02  for eGFR ≥ 90 mL/min/1.73 m2. These results were broadly consistent with the findings of our study. Similar to our hypothesis, several, but not all [To our best knowledge, only one cohort study  has expl not all ,12,13,14In this study, high eGFR appeared to likely be associated with high-frequency hearing impairment in males; however, the majority of sensitivity analyses , except 2 was low at approximately 3% for both males and females. Although most of signs and symptoms of CKD appears only when eGFR drops below 60 mL/min/1.73 m2, and several previous cross-sectional studies have reported that eGFR below 60 mL/min/1.73 m2 was associated with hearing impairment [2). Third, information on occupational risk factors for hearing impairment, such as exposure to noise and injurious work, were not available in our study; however, to minimize the confounding effect of noise, we performed sensitivity analysis restricted to participants with office jobs. Finally, the healthy worker effect should be considered due to the fact that our population comprised exclusively individuals of working age [The strengths of this study include its longitudinal design, spanning a maximum of 8 years; use of large-scale occupational health check-up data from approximately 127,000 Japanese workers aged 20–59 years old. Nevertheless, this study has some limitations. First, because the evaluation of hearing impairment was only a pure-tone test of limited frequencies, outcome misclassification unrelated to exposure was likely to occur. Such misclassification is known to result in “bias towards the null”. However, established or suspected risk factors, including age, manufacturing work with possible noise exposure, diabetes, BMI, smoking, and anemia, were associated with hearing impairment in this study ,23,24,40pairment ,13,14, wking age .eGFR is not generally associated with hearing impairment in either sex among the Japanese working-age population. To assess the tendency toward elevated risk of high-frequency hearing impairment among males in the highest eGFR group, further epidemiological studies of other populations will be required.


Beauveria bassiana serves as a main source of global fungal insecticides, which are based on the active ingredient of formulated conidia vulnerable to solar ultraviolet (UV) irradiation and restrained for all-weather application in green agriculture. The anti-UV proteins Rad1 and Rad10 are required for the nucleotide excision repair (NER) of UV-injured DNA in model yeast, but their anti-UV roles remain rarely exploredin filamentous fungi. Here, Rad1 and Rad10 orthologues that accumulated more in the nuclei than the cytoplasm of B. bassiana proved capable of reactivating UVB-impaired or UVB-inactivated conidia efficiently by 5h light exposure but incapable of doing so by 24 h dark incubation (NER) if the accumulated UVB irradiation was lethal. Each orthologue was found interacting with the other and two white collar proteins (WC1 and WC2), which proved to be regulators of two photolyases (Phr1 and Phr2) and individually more efficient in the photorepair of UVB-induced DNA lesions than either photolyase alone. The fungal photoreactivation activity was more or far more compromised when the protein–protein interactions were abolished in the absence of Rad1 or Rad10 than when either Phr1 or Phr2 lost function. The detected protein–protein interactions suggest direct links of either Rad1 or Rad10 to two photolyase regulators. In B. bassiana, therefore, Rad1 and Rad10 tied to the photolyase regulators have high activities in the photoprotection of formulated conidia from solar UV damage but insufficient NER activities in the field, where night (dark) time is too short, and no other roles in the fungal lifecycle in vitro and in vivo. Strong UVB 280–320 nm) and UVA 320–400 nm) irradiations in summer sunlight  are ubiq80–320 nm00 nm irrAs a vital macromolecule, eukaryotic DNA is readily damaged by UV irradiation, which induces the formation of two distinctive cytotoxic photoproducts known as cyclobutane pyrimidine dimer (CPD) and (6-4)-pyrimidine-pyrimidone (6-4PP). The UV-induced CPD and 6-4PP DNA lesions harmful to cells ,12,13 foSaccharomyces cerevisiae  for estimation of parameters a and r. Lethal doses to inactivate 50% (LD50), 75% (LD75), and 95% (LD95) of tested conidia were computed as indices of conidial UVB resistance by solving the fitted equation at the respective Is values of 0.5, 0.25, and 0.05.Conidial UVB resistances of Δeviously ,7. Briefeviously  in the s2 in the Bio-Sun chamber. The irradiated plates were immediately incubated at 25 °C for the optimal 5 h [Further, the conidia of each strain smeared onto GM plates as above were differentially impaired at the UVB doses of 0.15, 0.3,and 0.4 J/cmimal 5 h  under whwc1 and Δwc2 mutants and their control strains, three 50 mL aliquots of a 106 conidia/mL SDBY were shaken (150 rpm) at 25 °C for 3 days. Cells from each culture were rinsed with sterile water and resuspended in 50 mL sterile water. The aliquots of 500 μL suspension were spread on cellophane-overlaid agar plates containing 0.1% yeast extract and exposed to a UVB dose of 0.4 J/cm2 for generation of intracellular CPD and 6-4PP DNA lesions. The irradiated plates were covered with lids and incubated at 25 °C for 5 h under white light (photorepair) or in the dark (NER). A Biospin Fungus Genomic DNA Extraction Kit  was used to extract genomic DNAs from the cells incubated under light and in the dark or directly from the irradiated cells not incubated (control). Anti-CPD and anti-6-4PP antibodies  were used in enzyme-linked immunosorbent assays (ELISAs) to quantify CPD and 6-4PP lesions accumulated in each DNA sample, respectively. Every 20 or 200 ng DNA was added to each well of 50 μL diluent system in a 96-well plate for quantification of CPD or 6-4P PDNA lesions following the manufacturer’s guide. The optical density (OD492) of each DNA dilution at 492 nm was used as an index of CPD or 6-4PP DNA lesions accumulated in the irradiated cells before and after photorepair or NER. Three independent DNA samples per strain were included in each treatment.For each of the Δrad1, rad10, wc1, or wc2 in targeted gene mutants and WT for verification of targeted gene recombination events; and (2) transcript levels of 22 DNA damage repair-required genes in Δwc1, Δwc2, and their control strains. The qPCR analysis withpaired primers (ExTaq (TaKaRa). The fungal β-actin gene was used as a reference. Relative transcript levels of analyzed genes in the mutants were computed with respect to the WT standard using a threshold-cycle (2−ΔΔCt) method.Total RNAs were extracted with a RNAiso Plus Kit  from three 3-day-old cultures of each strain grown on cellophane-overlaid SDAY plates at the optimal regime and reversely transcribed into cDNAs with a PrimeScript RT reagent kit (TaKaRa). The cDNA samples were used as templates to assess: (1) transcript levels of  primers  was perft test. All experimental data were subjected to one-factor analysis of variance and Tukey’s honestly significant difference (HSD) tests for differences among tested strains.Mean N/C-GFI ratios between Rad1-GFP and Rad10-GFP were compared with Student’s B. bassiana Rad1 (EJP63355) and Rad10 (EJP67637) were revealed to share higher sequence identities with the counterparts of entomopathogenic fungi (86–96% and 65–82%) than of non-entomopathogenic fungi (34–84% and 33–59%). Either Rad1 or Rad10 is different in molecular size between B. bassiana and S. cerevisiae but similar in domain architecture, as shown by the same size of ERCC4 domain (81 aa) and the similar size of Rad10 domain (114 or 118 aa), respectively  estimates indicative of virulence  for all tested strains (2 for NCI or CBI (50 means via NCI and CBI were 5.50 (±0.24) and 4.56 (±0.15) days (n = 9) for the three control strains but prolonged to 8.02 (±0.37) and 6.60 (±0.53) days (n =3) for Δrad1 and 8.98 (±0.30) and 8.20 (±0.18) days (n = 3) for Δrad10 , 0.275 (±0.008), and 0.370 (±0.012) J/cm2 (n = 9) for the control strains’ conidia , and 87.1% (±2.5), respectively, via 5 h light plus 19 h dark incubation at 25 °C  when the dark incubation was prolonged to 40 h  and Δphr2 (19%) mutants [rad1 and Δrad10 mutants’ conidia inactivated at 0.4 J/cm2 were lower or much lower than those seen in the previous Δphr1 (7%) and Δphr2 (37%) mutants’ conidia inactivated at 0.5 J/cm2 [B. bassiana. The NER activity of either Rad1 or Rad10 was observed in the control strains’conidia at the end of 24h dark incubation after exposure to ≤0.35 J/cm2 but not detectable after exposure to 0.4 J/cm2 unless the dark incubation exceeded 24 h, which is far beyond the nighttime of a circadian day. Therefore, Rad1 and Rad10 protect B. bassiana from solar UVB damage by photoreactivation. Their NER activities were seemingly extant but hardly sufficient for reactivation of UVB-imapred B. bassiana conidia under field conditions due to too short night (dark) time.The UVB LD mutants , highlig.5 J/cm2 . These dB. bassiana [M. robertsii [phr1 and phr2 (Photoreactivation relies on the photorepair of UVB-induced CPD and 6-4PP DNA lesions by Phr1 and Phr2 in bassiana  or by WCobertsii . Thus, Yand phr2 B. The dewc1 and Δwc2 mutants. The UVB irradiation at 0.4 J/cm2 resulted in similar levels of CPD lesions (control in wc1, Δwc2, and their control strains. After 5h light exposure for photorepair, the amount of CPD and 6-4PP lesions were lowered by 34–44% and 30–43% in the control strains’ cells, respectively. However, such photorepair led to insignificant changes in CPD lesions and slight decreases in 6-4PP lesions in the Δwc1 and Δwc2 mutants’ cells. After 5h dark incubation for NER, CPD lesions decreased in the DNA samples of the control strains slightly more than of Δwc1 and Δwc2, while 6-4PP lesions were moderately reduced in all tested strains except Δwc2, whose 6-4PP level was not affected in comparison to the control. These data demonstrate that, in B. bassiana, the role of either WC1 or WC2 in photorepairing both CPD and 6-4PP DNA lesions is beyond that of Phr1 specific to CPD or Phr2 specific to 6-4PP in B. bassiana [M. robertsii [Next, ELISAs with anti-CPD and anti-6-4PP antibodies were conducted to assess the accumulated CPD and 6-4PP lesions in the DNA samples isolated from irradiated hyphal cells of Δobertsii .phr1 and ph2 was verified by the nearly abolished expression of both phr1 and phr2 in either Δwc1 or Δwc2 (wc1 or Δwc2, suggesting active roles of WC1 and WC2 in transcriptional mediation of multiple anti-UV genes in B. bassiana.The role of WC1 or WC2 in the transcriptional activation of  or Δwc2 E, providB. bassiana, Rad1 and Rad10 were proven to co-localize in both the nucleus and the cytoplasm for nucleocytoplasmic shuttling and interact with each other as elucidated in S. cerevisiae [phr1 and phr2 and enable the photorepair of both CPD and 6-4PP DNA lesions, although their interactions with both photolyases were not evidenced in B. bassiana as shown previously in M. robertsii [phr1 and phr2 were nearly abolished in the Δwc1 or Δwc2 mutant. This is different from the abolished expression of phr2 in the absence of wc1 and of phr1 in the absence of wc2 in M. robertsii [wc1 and Δwc2 mutants were severely compromised in photorepair activity for both CDP and 6-4PP lesions, unlike the previous M. robertstii Δwc1 and Δwc2 mutants that were unable to photorepair 6-4PP and CPD DNA lesions, respectively [phr1 and phr2 in B. bassiana or interacts with both Phr1 and Phr2 in M. robertsii. The protein–protein interactions detected in this study suggest direct links of either Rad1or Rad10 to both WC1 and WC2 as photolyase regulators. Therefore, either Rad1 or Rad10 tied to two photolyase regulators displayed much higher UVB resistance and photoreactivation activity than a single photolyase or photolyase mediator in B. bassiana, although the involved mechanism remains to be explored.In revisiae ,54. The revisiae ,34,35,36obertsii . At the obertsii . Thus, tectively . The preB. bassiana cells like those of their yeast orthologues for UVC-impaired cells [2 and the abolished germination of the Δrad1 and Δrad10 mutants’ conidia incubated in the same fashion after exposure to ≤0.15 J/cm2. However, the control strains’ conidia impaired at the UVB dose of 0.4 J/cm2 were unable to be reactivated within 24 h of dark incubation, in contrast to their high photoreactivation rates of ~87%. This highlights an insufficient NER activity for either Rad1 or Rad10 in the field, where a much shorter nighttime is available for the reactivation of formulated conidia exposed to solar UVB irradiation that accumulates to ~2.5 J/cm2 during the daytime [B. bassiana from solar UVB damage mainly by photoreactivation.In the present study, Rad1 and Rad10 did exhibit NER activities for UVB-impaired ed cells ,34,35,36B. bassiana’s response and adaptation to solar UV irradiation in sunlight. The essential roles of Rad1 and Rad10 in preventing insecticidal fungal cells from solar UVB damage rely upon their high photoreactivation activities, whichcould have arisen from interactions of either with both WC1 and WC2 to govern photorepair via thetranscriptional mediation of both phr1 and phr2, and hence are distinguished from a dependence of the yeast orthologues’ anti-UV roles on NER. The NER activity of Rad1 or Rad10 is extant in B. bassiana, as known in the budding yeast [B. bassiana.In conclusion, Rad1 and Rad10 play essential roles in ng yeast , but harng yeast , which w