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  • A total of 74 subjects born before 1911 participated in this study. Hearing tests were part of the physical examinations (including electrocardiogram and laboratory tests of blood and liver function) and mental health evaluation for the centenarians. Their ages varied between 100 and 106 years, with a mean age of 102 years. A questionnaire about ear- and hearing-related medical history, noise exposure (during leisure and work), and self-perceived hearing function was administered as an interview. All participants were farmers with no history of leisure- and/or work-related high intensity noise exposure. Questionnaire data on socioeconomic status, medical history, lifestyle factors, and medication use were obtained as part of the examination. Subjects with a family history of hearing loss and/or a history of ototoxic drug usage were excluded from the study. After excluding subjects with apparent middle ear diseases after otoscopic examination, 68 subjects (21 male and 47 female) were included in the report. The mean age of this group was 102 years. Figure 1 summarizes their lifestyle factors and medical conditions that are known risk factors for aggravating age-related hearing loss. For comparison, we also recruited normal subjects aged between 20 and 25 years and 60 and 65 years. Twenty subjects (equal number of males and females) for each of these two age groups were examined in the same condition as centenarian listeners. The same questionnaire that was used for centenarian subjects was used to obtain medical history and noise exposure history from the younger subjects before hearing tests. Subjects with a family history of hearing loss, history of otologic diseases, and treatment with ototoxic drugs were excluded. Written informed consent was obtained from all participants or their guardians. The procedures described in the present study have been approved by the Institutional Review Board of the Shaoxing Second Hospital. Figure data removed from full text. Figure identifier and caption: 10.1371/journal.pone.0065565.g001 Lifestyle factors and health condition of the centenarian participants.A: Percentage of centenarian subjects who had some of the risk factors for age-related hearing loss. Smoking was defined as consumption of at least half a pack of cigarettes a day for more than a year within the past 10 years. Drinking was defined as consumption of more than 50 ml wine or alcohol on the daily basis for more than a year within the past 10 years. COPD: Chronic obstructive airways disease (diagnosed by a physician). B: Distribution of centenarian subjects (presented as percentage) with different blood pressure and glucose levels. Glucose level presented was based on blood collected 2 to 3 hours after meal. The numbers indicated inside the plots are the percentage. Pure-tone air-conduction thresholds were obtained in both ears of participants at the frequencies of 250, 500, 1,000, 2,000, 4,000, and 8,000 Hz using a diagnostic audiometer (MADSEN Midimate 622). Bone-conduction thresholds were also obtained at the frequencies of 500, 1,000, 2,000, and 4,000 Hz. No masking was used during test. The audiometer was calibrated in accordance with international (ISO) standards. Testing was completed in a room that met standard requirements. To measure evoked DPOAEs, a MADSEN Capella Cochlear Emissions Analyzer was used. Test frequencies were 500, 1,000, 2,000, 4,000, 6,000, and 8,000 Hz with an f2/f1 ratio of 1.2. The level of f1 was set at 70 dB (maximum output), while the level of f2 was set at 65 dB. DPOAE response was regarded as detectable if the “cubic” distortion tone (DP1) and the “quadratic” distortion tone (DP2) were 6 dB above noise floor. For the assessment of middle ear function, tympanometry was used with a 226 Hz probe tone. An otoscopic examination was performed before all the tests to ensure that the ear canal was clear and that there were no obvious signs of middle ear infection or perforation in the tympanic membrane. Middle ear impedance, pure tone thresholds, and DPOAEs were obtained from both ears of each participant. When a participant was unable to hear a tone, 5 dB above the highest audiometer output level was recorded as the threshold. Data were presented as mean and standard deviation (SD) and evaluated with student's t-tests. Statistical significance was assigned to P values of less than 0.01. A p value larger than 0.5 was considered as statistically insignificant. Data presented in this study reflected a sample size of 136, 40, and 40 ears for the centenarian group and the two groups aged 60–65 and 20–25, respectively.
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