Many youth and young adults experience high noise exposure compounded by lack of access to hearing health education. Although the need for hearing health education programs is evident, the efficacy of these programs for youth is unclear. We evaluated the literature for efficacy of various hearing conservation programs aimed at youth and young adults, and analyzed their strengths and limitations.Studies reporting results of hearing conservation or hearing loss prevention programs with youth or young adults, using randomized controlled trials, quasi-experimental designs, experimental design, or qualitative research, and published in peer-reviewed journals in English between 2001 and 2018 were included. Studies were found through searches of selected literature databases . Identified publications were assessed for relevance, and data were extracted from the studies deemed relevant.A total of 10 studies were included. Very little evidence of efficacy of hearing conservation educational programs was found in these studies. Several methodological limitations including lack of rigorous study designs, inadequate power, and application of inappropriate statistical analysis were noted. Some use of technology in programs  was observed, but conclusions as to the effectiveness of these tools were limited by the small number of studies and small sample sizes.The number of studies of educational hearing conservation programs for youth and young adults was low. The efficacy of the program was not reported in most studies, and it is difficult to draw public health conclusions from these studies due to their multiple methodological limitations. While use of technology in hearing conservation educational programs offers promise, its effectiveness has not been studied. Noise from recreational activities threatens the hearing ability and health of youth and young adults around the world. In the United States, approximately 5.2 million children and adolescents, and several million young adults, experience noise-induced hearing loss , 2. Non-Although the effects of high noise on hearing  are well-known , 10, higThe problem of hearing loss has been identified as high priority by leading national organizations. Healthy People 2020 includes a focus on reducing the prevalence and severity of disorders of hearing, and increasing “the proportion of adolescents aged 12 to 19 years who have ever used hearing protection devices  when exposed to loud sounds or noise” . The corUse of hearing protectors, maximizing the distance from the source of noise, and providing enclosure to minimize noise transmission are a few methods of mitigating the effects of high noise exposure in occupational and recreational settings . HoweverMethods for the systematic literature review were adapted from PRISMA guidelines  and inclStudies were found through searches of selected literature databases  using combinations of the following keywords: “hearing protection intervention” OR “hearing conservation intervention” OR “hearing protection” OR “hearing protection conservation” OR “hearing conservation” OR “hearing” OR “intervention” OR “noise induced hearing loss” AND “agricul*” OR “rural” OR “noise” AND “technology” AND “smartphone” OR “computer” AND “internet” AND “text messaging” AND “school based.” Multiple combinations of keywords were used to ensure that all relevant studies were discovered. Searches were conducted from October through December 2017. This resulted in a total of 23,306 reports (duplicates included), which were distributed between 6982 hits in PubMed, 16,293 hits in Google Scholar, 7 hits in NIOSH Toxline, and 24 hits in Scopus. A total of 7004 unique reports were screened by title and abstract, and out of those, 20 underwent full article screening. Ten studies met all inclusion criteria as listed above.The searches were conducted independently by two authors (KMK and SLB) in consultation with a librarian at Indiana University Bloomington who had expertise in scientific literature search in public health. Titles and abstracts were read independently, and relevance was assessed based upon the inclusion criteria. All authors participated in the full text review where the full article was read and discussed. All relevant studies were included, irrespective of their scientific quality. Information regarding study design, interventions conducted, study objective, randomization of subjects, data collection, analyses performed, and relevant results were extracted from reports.n = 3), quasi-experimental studies with crossover design with or without a control group (n = 4), and experimental studies with multiple groups (n = 3).As indicated in Fig. Studies included in this literature review demonstrated wide variations in study settings and subject characteristics . There were six studies of high-school children , 33–37, Interventions were exclusively educationally-based, directed at increasing users’ knowledge of noise and hearing, with an implicit goal of influencing hearing conservation behavior. Environmental modification and policy development approaches were not represented. Educational content on sources of loud noise, enhancing positive attitudes towards hearing conservation, noise-induced hearing loss, and use of hearing protection were the main focus areas for content of education in the selected studies. In two US studies in agricultural communities, hearing conservation was nested in health and safety education programs or curricula that also provided education to high school students on other farm safety hazards , 37. OthIn a limited number of studies, researchers employed various types of teaching and learning methods to accomplish the goals of the intervention projects. Most of these methods were interactive and conducted in field settings. Notable teaching/learning methods included hands-on demonstration of fitting of hearing protection devices , interacFindings regarding efficacy of programs are summarized in Table The use of technologies in educating youth about noise exposure and noise-induced hearing loss prevention was addressed in a few studies , 35, 38.n ≤ 100) that lacked adequate statistical power to detect group differences [Several limitations and weaknesses were observed in study design and data analysis Table . There wferences , 38–40. ferences  or even ferences , most stferences  or even ferences . Follow-ferences , or the ferences , 33, 36.ferences . The ranferences . Absenceferences , 35, 40,ferences . In seveferences , 39, nonferences , and lacThis review used a systematic and sensitive search strategy, together with multiple literature databases, to ensure that all studies meeting inclusion criteria were reviewed. Overall, we found a small number of hearing health intervention studies for youth with multiple methodological limitations. In addition to problems with study designs, the quality of reporting was low. Many studies failed to include essential elements needed to assess trial quality, e.g., robust design, presence of sufficient statistical information, specification of eligibility criteria, and random allocation of subjects . As a reIn the last 15 years, there have been only two previously published reviews of hearing conservation programs for youth; the most recent 5 years ago. Our review included some of the same studies included in previous reviews, but differed in quality. For example, we included a critical analysis of study methods and efficacy, which were not included in the previous review papers. Furthermore, we expanded the inclusion criteria to include a greater breadth of study populations and study settings compared to previous reviews , 43. In Despite the recognition of noise exposure as a priority occupational health problem, our extensive search of literature from 2001 onward produced only 10 reports that met study inclusion criteria. Weaknesses in study design and methods were common among reviewed studies, e.g., small sample sizes, lack of rigorous study designs , inadequate power, and application of inappropriate statistical analysis. These limitations seriously impaired the validity of study conclusions and generalizability of results. While several intervention studies included in the review demonstrated improvement in hearing conservation knowledge, attitude, and/or behavior from pre to post intervention, most post-tests were short-term ; long-term and sustained effects of such interventions were rare. There is a need for high-quality studies using robust trial designs to test longer-term effectiveness of hearing conservation programs for youth and young adults. We recommend that reports of intervention studies should follow commonly-accepted protocols for trial reporting, such as CONSORT and PEDro .There was wide variation in study settings, outcome measures, instruments, and intervention approaches across studies. Similarly, intervention methods were diverse: school curriculum on hearing conservation, distribution of printed materials , use of social media and the internet, face-to-face motivational activities, group learning, and technological approaches such as smartphone apps, computer training, and text message reminders.The role of technology in promoting many types of health behavior is of great interest. Among youth, several studies employing technology-based program have demonstrated high levels of efficacy with multiple health behaviors  –50. FurtWe reviewed the efficacy of hearing conservation interventions aimed at youth and young adults; analyzed the weaknesses and limitations of published studies, generated recommendations for future development of hearing conservation programs, including use of technology .The use of technology in promoting hearing conservation is a promising recent development in hearing conservation for youth. Although few studies to date have used technology in hearing conservation programs , 35, 38,


Mycoplasma hyopneumoniae, colonises ciliated epithelium and disrupts mucociliary function predisposing the upper respiratory tract to secondary pathogens. Alleviation of disease is reliant on antibiotics, vaccination, and sound animal husbandry, but none are effective at eliminating M. hyopneumoniae from large production systems. Sustainable pork production systems strive to lower reliance on antibiotics but lack of a detailed understanding of the pathobiology of M. hyopneumoniae has curtailed efforts to develop effective mitigation strategies. M. hyopneumoniae is considered an extracellular pathogen. Here we show that M. hyopneumoniae associates with integrin β1 on the surface of epithelial cells via interactions with surface-bound fibronectin and initiates signalling events that stimulate pathogen uptake into clathrin-coated vesicles (CCVs) and caveosomes. These early events allow M. hyopneumoniae to exploit an intracellular lifestyle by commandeering the endosomal pathway. Specifically, we show: (i) using a modified gentamicin protection assay that approximately 8% of M. hyopneumoniae cells reside intracellularly; (ii) integrin β1 expression specifically co-localises with the deposition of fibronectin precisely where M. hyopneumoniae cells assemble extracellularly; (iii) anti-integrin β1 antibodies block entry of M. hyopneumoniae into porcine cells; and (iv) M. hyopneumoniae survives phagolysosomal fusion, and resides within recycling endosomes that are trafficked to the cell membrane. Our data creates a paradigm shift by challenging the long-held view that M. hyopneumoniae is a strict extracellular pathogen and calls for in vivo studies to determine if M. hyopneumoniae can traffic to extrapulmonary sites in commercially-reared pigs.Enzootic pneumonia incurs major economic losses to pork production globally. The primary pathogen and causative agent,  Mycoplasma hyopneumoniae is the etiological agent of enzootic pneumonia and a primary pathogen in the porcine respiratory disease complex1. Globally, porcine enzootic pneumonia is widespread and inflicts significant economic losses to pork production. Losses are incurred via reduced growth rate and feed conversion efficiency, costs for treatment and vaccination, and excessive morbidity and mortalities resulting from the combined effects of multiple respiratory pathogens. M. hyopneumoniae influences the ciliary beat frequency, induces ciliostasis and causes epithelial cell death, culminating in a devastating assault on the mucociliary escalator and an excessive host immune response in the lungs5. M. hyopneumoniae colonises cilia that project into the luminal surface of epithelial cells of the respiratory tract and is rarely found associated with the epithelial cell body6. These observations suggest that M. hyopneumoniae recognises receptors expressed on the surface of cilia but are limited in their presentation on the cell body. M. hyopneumoniae attaches to cilia via highly expressed, structurally and functionally complex7 adhesins that are present on the cell surface of M. hyopneumoniae as a diverse combination of cleavage fragments that bind multiple host molecules including highly sulphated glycosaminoglycans, fibronectin and plasminogen21.M. hyopneumoniae include vaccination (predominantly with bacterin formulations); antibiotic therapy and herd management . Combinations of these strategies are effective, however, there remains a pressing need to lower reliance on antibiotics to control infection in intensively reared animal production systems25. Sound herd management is challenged by the need to identify pig production operations free of M. hyopneumoniae and it remains a challenge to identify subclinically-infected and carrier animals. Ultimately, further investigation into the survival mechanisms of this important porcine pathogen is required to aid in the development of future strategies to prevent and control transmission.Strategies that are implemented to control infection by 29, and although it has historically been characterised as a strict extracellular pathogen, M. hyopneumoniae has been cultured from the liver, spleen, kidneys and bronchial lymph nodes of pigs infected experimentally with M. hyopneumoniae32. However, it is not known if M. hyopneumoniae colonises tissue sites distal to the respiratory tract in commercially-reared herds. Interestingly, M. hyopneumoniae has been isolated in pure culture from both pericardial and synovial joint fluids in slaughter-age commercial pigs with fibrinous pericarditis33. It is not known how M. hyopneumoniae traffics to these sites nor is it known if M. hyopneumoniae can invade epithelial cells and trigger cellular uptake pathways.It is well known that numerous mycoplasma species can invade host cellsM. hyopneumoniae cells interact with integrin β1 via fibronectin and colocalise in a manner that promotes cellular uptake via caveosomes and clathrin-coated vesicles. We monitored the cellular events that depict M. hyopneumoniae trafficking via the endocytic pathway, and escaping phagolysosomal fusion, before residing free in the cytoplasm. Collectively, our data have significant implications for detecting animals infected with M. hyopneumoniae and for development of therapies to eliminate this difficult-to-control pathogen.In this study for the first time, we show that M. hyopneumoniae colonises host epithelial cell surfaces, scanning electron microscopy (SEM) was used to visualise the pattern of adherence to porcine kidney epithelial cells (PK-15) after 16 h. PK-15 cells have been used extensively as a model system for studying host-M. hyopneumoniae interactions36. SEM images showed both small clusters and individual M. hyopneumoniae cells associated intimately with the cell surface of PK-15 cell monolayers  M. hyopneumoniae cells that adhered to PK-15 cells reside intracellularly and are potentially invasive (data not shown). Although it is important to note that the gentamicin protection assay is a crude quantitative measure of cell invasion, we have routinely observed M. hyopneumoniae cells residing intracellularly using CLSM and 3D-SIM. Our observations presented here, are representative images of a minimum of 20 biological replicate experiments performed on different days over the course of several years.In order to gather insight into how ers Fig. . These aers Fig. , which oers Fig. . Using iAPI Fig. . In uninAPI Fig. . This coM. hyopneumoniae cells  images also depict PK-15 cell surface projections engulfing lls Fig. . Once inres Fig. , indicatasm Fig. . Notably39. We used confocal microscopy with monoclonal antibodies that recognise clathrin (mAbclath.) and caveolin-1 (using mAbcav) to show M. hyopneumoniae cells residing within clathrin-coated vesicles (CCVs) and caveosomes  for 2 h prior to infection with M. hyopneumoniae. Our modified gentamicin protection assay showed that the antibody blocked (~75% reduction in viable colonies) the ability of M. hyopneumoniae to become internalised within PK-15 cells  that were stained with F2P94-J antisera contained bacterial cells adhering along the ciliary border of the epithelium  PK-15 cells with anti-fibronectin (pAbFn) after membrane permeabilisation. It was found that M. hyopneumoniae cells associated both with CCVs  via clathrin- and caveolae-mediated endocytosis. Membrane bound M. hyopneumoniae captured via these mechanisms are transported intracellularly via early, recycling and late endosomes  reorganisation. Leukocytes are an established portal for the dissemination of many pathogens to distal tissue sites68 and provide a protective niche from the immune system. M. hyopneumoniae promotes the infiltration of leukocytes that stain with mAbβ-act to foci of infection. The inflammatory response, a hallmark of lung infection caused by M. hyopneumoniae, may provide the organism with a mechanism to invade and survive professional phagocytic cells for dissemination to the liver, spleen, kidneys and lymph nodes32. However, further studies are needed to interrogate this hypothesis.men Fig. 14,18. Sioci Fig.  at leveloci Fig. . CytoskeM. hyopneumoniae potentially hijacks the intrinsic eukaryotic α5β1-fibronectin recycling pathway to gain access to host cells where it can persist intracellularly. This has important implications for the trafficking of M. hyopneumoniae from the respiratory tract to distal tissue sites and potentially vice versa after extended periods of dormancy. This study paves the way for the development of therapeutic strategies that seek to interfere or block the ability of this pathogen to persist in its only known host Sus scrofa.Collectively, our data suggests that All data generated or analysed during this study are included in this published article (and its Supplementary Information files).Mycoplasma hyopneumoniae strain 232 was grown in modified Friis medium as described previously69. Broth cultures were incubated at 37 °C for approximately 16 h until they reached mid-exponential phase and the medium turned orange. At this stage, cultures contain approximately 5 × 105–1 × 106 CFU/mL. For growth on solid agar, cells were plated onto Friis agar using a method described previously70.5 cells/well) and incubated overnight. For infection studies, a 16 h culture of M. hyopneumoniae strain 232 was pelleted, washed twice in PBS, and resuspended in 25 mM HEPES in DMEM containing 5% fetal bovine serum (infection medium) prior to incubation at 37 °C for 2 h. M. hyopneumoniae cells were added, so that each well received 0.5 mL representative volume (approximately 7.5 × 105 CFU/mL) of the initial culture, and allowed to adhere to PK-15 cells for 16 h at 37 °C/5% CO2. Non-adherent cells were removed by washing in PBS, followed by incubation in 300 µg ml−1 gentamicin in DMEM (filter sterilised through pore size 0.2 µm) for 4 h at 37 °C/5% CO2. Duplicate samples (positive controls) were not incubated with gentamicin. The gentamicin was removed and cells were washed 5 × in PBS to remove any residual antibiotic. The cell dissociation solution TrypLE  was added to each well and incubated at 37 °C for 20 minutes. Cells were then removed by gently pipetting the TrypLE 3–4 times. The cell suspension was serially diluted in Friis broth to 10−1 and 10−2. An aliquot (50 µl) of each dilution was pipetted onto 9 cm Friis agar plates, the liquid was allowed to air dry in a sterile environment for 10 minutes, and the plates were incubated at 37 °C/5% CO2 for at least 7 days. M. hyopneumoniae colonies were counted using a stereomicroscope (×40 objective). Inhibition experiments were performed by pre-incubating PK-15 cells with a 1:100 dilution of monoclonal antibodies against integrin β1  prior to the addition of M. hyopneumoniae cells.Porcine kidney epithelial-like monolayers (PK-15) were grown to semi-confluency and split into 12-well microtitre plates  was incubated for 1 h at RT. Cells were permeabilised in 0.5% (v/v) Triton X-100 in PBS for 5 min. Samples were then re-incubated with F2P94-J rabbit antisera at a dilution of 1:500 for 1 h at RT, followed by incubating with a 1:1000 dilution of anti-rabbit CF 568-labelled secondary antibody (Biotium) for 1 h at RT. DAPI (Roche) was then added for 5 minutes at RT to stain nucleic acids. Samples were then prepared as previously described35.Experiments were performed as described previouslyM. hyopneumoniae cells were labelled exclusively with DAPI and not F2P94-J antisera. Fibronectin was labelled prior to permeabilisation using polyclonal antisera raised in rabbit against fibronectin (pAbFn) as described previously14. Mouse monoclonal antibodies mAbs (Abcam) against clathrin , caveolin-1 , integrin β1 , RAB7  or LAMP1  were incubated post-permeabilisation overnight at 4 °C, and then incubated with a 1:1000 dilution of anti-mouse CF 488- or 568-conjugated secondary antibodies (Biotium) for 1 h at RT. Rabbit polyclonal antibodies (Abcam) against RAB5 (1:2000), EEA1 (1:1000) were incubated post-permeabilisation for 1 h at room temperature, and then incubated with a 1:1000 dilution of anti-mouse CF 488- or 568-conjugated secondary antibodies (Biotium) for 1 h at room temperature.Experiments were performed identically to those described above except that intracellular 71, using a Nikon A1 Confocal Laser Scanning Microscope and a V3 DeltaVision OMX 3D-SIM Imaging System .Samples were imaged as previously describedMaximum Intensity Projection (MIP) and slices images.Images captured with the Nikon A1 Confocal microscope and those generated by the DeltaVision OMX 3D-SIM were processed using Bitplane, Imaris Scientific 3D/4D image processing software to create M. hyopneumoniae-infected PK-cells and uninfected controls were labelled with integrin β1 (described above) and 10 random fields of view were captured using an Olympus BX51 Upright Epi Fluorescence Microscope at 20 × magnification and a constant exposure. The mean fluorescence of integrin β1-stained samples was calculated, after thresholding and binary conversion, using ImageJ. GraphPad Prism 7 was used to plot the data, including the standard error of the mean, and to perform the statistical analyses (unpaired t-test).35.Experiments were performed as described previously with no modificationsPreparation of tracheal sections and infected monolayers for transmission electron microscopy Samples were fixed in a fixation solution containing 5% formaldehyde and 2% glutaraldehyde in cacodylate buffer  and washed with cacodylate buffer. Samples were then osmificated with 1% aqueous osmium for 1 h at room temperature, washed and pellets of the samples were embedded in 2% water agar and cut into small cubes. Dehydration was achieved with a graded series of acetone  for 30 min on ice followed by contrasting with 2% uranyl acetate in 70% acetone for overnight at 4 °C and further dehydrated with 90% and 100% acetone. Samples in 100% acetone were allowed to reach room temperature and were infiltrated with epoxy resin according to Spurr’s formular for a medium resin72; 1 part 100% acetone/1 part resin for overnight, 1 part 100% acetone/2 parts resin for 8 h, pure resin for overnight and several changes the following 2 days. Samples were then transferred to resin filled gelatine capsules and polymerized for 8 h at 75 °C. Ultrathin sections were cut with a diamond knife, picked up with formvar-coated copper grids (300 mesh) and counterstained with 2% aqueous uranyl acetate and lead citrate. After air-drying samples were examined in a Zeiss transmission electron microscope TEM910 at an acceleration voltage of 80 kV. Images were recorded digitally with a Slow-Scan CCD-Camera  with ITEM-Software . Brightness and contrast were adjusted with Adobe Photoshop CS4.38 with a minor modification. Briefly, 19 male weaner pigs, sourced from a commercial herd, were infected endotracheally, with M. hyopneumoniae strain Hillcrest and euthanized 6 weeks post-infection. Serial sections of lung lesions from pigs, as well as healthy lung tissue from control pigs were examined for actin distribution by staining with 1:200 mAbβ-act for 1 hr at RT.All animal procedures were approved by the Animal Ethics Committee at the Elizabeth Macarthur Agricultural Institute and were in accordance with the Australian Code of Practice for the Care and Use of Animals for Scientific Purposes. Experiments were performed as previously describedSupplementary Figures


Iron and steel factory workers in Tanzania are likely to develop noise-induced hearing loss (NIHL) due to exposure to high sound levels. Studies on hearing status in this population are lacking. The aims of this study were to determine prevalence of NIHL among iron and steel workers and compare hearing thresholds at different frequencies with a control group. We conducted a cross-sectional study among 221 iron and steel workers exposed to average noise level of 92 dB(A), compared with 107 primary school teachers recruited as controls and exposed to average noise level of 79.7 dB(A). We used a questionnaire-based interview to collect information on socio demographic characteristics and other confounding variables. Hearing loss was defined as hearing threshold levels ≥25 dB hearing loss in either ear at 3000, 4000 or 6000 Hz. The prevalence of hearing loss was significantly higher among the exposed group than among the controls, i.e. 48% and 31%, respectively. There were significant differences in hearing thresholds between the exposed and control groups at 3000, 4000, 6000, and 8000 Hz. Hearing loss was more frequent among workers exposed to higher noise levels than among the controls suggesting that iron and steel workers run a higher risk of developing hearing loss. Noise-induced hearing loss (NIHL) is an underestimated public health concern ,2. GlobaStudies highlight noise exposure as one major risk factor contributing to hearing loss ,4,5. OthStudies conducted in Sub-Saharan Africa (SSA) have mainly focused on the mining sector and indicate, for instance, that the prevalence of hearing loss in this industry was 37% in Zimbabwe and 47% in Tanzania ,16. DespIn Tanzania, like in other SSA countries, investments in the manufacturing industries, including iron and steel industries, create jobs for a significant large number of employees. Globally the demand for steel is increasing, and this sector has provided employment for 50 million people . The conLEX,8h) for these workers of 92.0 dB(A) [This cross-sectional study was conducted from June 2016 until June 2017 and involved permanent male workers from four iron and steel factories in Tanzania exposed to noise. Characteristics and details of noise-exposure assessments in these factories have been presented elsewhere [Controls were male teachers from 34 public primary schools in Tanzania. This control group was chosen because they were expected to be exposed to low levels of occupational noise ,21,22,23The sample size calculation was based on the estimated prevalence of hearing loss among workers exposed to loud noise at work. Since there was no available information about hearing loss among noise exposed workers or among the general population in Tanzania, the sample size was calculated based on a community baseline survey conducted in Uganda that found the prevalence of hearing loss among adults to be 12% . In our A total of 376 permanent workers (253 from four iron and steel factories and 123 teachers from 34 public primary schools) were randomly selected by using a table of random numbers from the provided list of workers and were invited to participate in the study . WorkersWe used information collected through a structured interview with both exposed participants and controls to exclude the following categories of workers from the data analysis: those with congenital hearing loss, history of otitis media during childhood and those reported to have worked in noisy job among the controls. In addition, we excluded those who reported using long-term medication because the participants did not have adequate knowledge of the type of medication they used. Thus, we ended with a total of 328 (221 exposed and 107 controls) persons taking audiometric measurements .The participants were instructed to avoid areas with high level of noise for a minimum of 12 h prior to audiometric examinations to minimize the possibility of temporary threshold shift (TTS). The duration since last occupational noise exposure (free noise exposure) was recorded before the audiometric test was administered ,26.A structured interview questionnaire was used to collect information on demographic characteristics and factors that may influence hearing loss. These included age (in years), number of years of employment, history of noise exposure at work (yes/no), current smoking (yes/no), present use of long-term medication (yes/no), exposure to chemicals/organic solvents (yes/no), use of hearing protection while working in noisy areas (yes/no), ear infections as a child or adult (yes/no), head injury/trauma (yes/no), tinnitus (yes/no). In addition, information about otitis in childhood (yes/no), known congenital hearing loss (yes/no), any relatives with hearing loss and any history of ear-related medical condition (diabetes and hypertension) was collected. This information was collected before the audiometry day and used to exclude participants before audiometry .Prior to otoscopy, but on the same day as the audiometry, participants were interviewed using a checklist indicating whether they had symptoms of upper respiratory infections  (yes/no), ear discharge (yes/no), time and date they left work (hours) and the most recent time they were exposed to high noise at a level that made it difficult to communicate. Afterwards, otoscopy was performed by an occupational physician; in circumstances when the ear canal was completely obstructed with wax or cerumen, the latter were removed, and a new appointment was scheduled for audiometry. This also applied to participants with upper respiratory infections; the test was postponed until they were asymptomatic.max) in the booth was 51 dB at 31.5 Hz. For best results, audiometry was conducted in the morning before any work exposure. In addition, the city is less noisy in the morning compared to other times of the day when the participants could potentially be exposed to a higher level of environmental noise. Pure tone audiometry was conducted using an Interacoustics AD226  with Amplivox Audiocup earphones having lower test limit of −10 dB. The equipment was pre-calibrated. Test frequencies were 250–8000 Hz in the order starting with 1000, 2000, 3000, 4000, 6000, 8000, 500, 250 and finish at 1000 Hz [Audiometric measurements (pure tone audiometry) were conducted in an ear-screening locally-constructed booth in a quiet room at the headquarters of the Occupational Safety and Health Authority (OSHA) in Tanzania. The same technical personnel conducted all audiometric tests using a standardized protocol. Background noise in the test booth was monitored by a calibrated hand-held Sound Level Meter , and checked for conformity with ISO 8253-1:2010 standard . The hig 1000 Hz . A manua 1000 Hz ,28.t-tests were used to compare categorical and continuous descriptive variables, respectively. NIHL was defined as hearing threshold level ≥25 dB hearing loss in either ear at 3000, 4000 or 6000 Hz [Descriptive statistics were presented as mean and standard deviation or percentage. Chi-square and independent samples  6000 Hz .Potential determinants of hearing loss were identified. Age was categorized into three age groups (tertiles) based on the age distribution among the controls. Duration of work was categorized arbitrarily into three groups . History of ear-related medical conditions  was combined into a dichotomized variable (yes/no), current smoking (yes/no), relatives with hearing impairment (yes/no), tinnitus (yes/no) and previous noisy work (yes/no). A chi-square test was used to explore the relationship between these variables and hearing loss in exposed participants compared with controls. The intercorrelation between participant’s age and duration of work determinants was tested with the Pearson correlation test. In multiple regression analyses, we chose the determinant that contributed most to the hearing loss. We used log binomial regression models with a 95% confidence interval (CI) to ascertain differences in hearing loss (yes/no) between exposed and controls within each age strata and within the total group of workers while adjusting for the significant determinants selected from Chi-square analyses and the correlation test.We computed the mean hearing threshold for the different test frequencies for both exposed participants and controls, as well as for the three age groups within the main exposure groups. For each test frequency, multiple linear regression was used to analyze for differences between exposed and controls, while adjusting for age as a continuous variable, previous noisy work and history of ear-related medical condition.Aeq8h of 92 dB(A) [d) (in years), and the square of noise exposure level (LAeq8h), with frequency specific constants u, v, and L0  and a L92 dB(A) . Within decibels :NIPTS = p < 0.05 as statistical significance. NIPTS was estimated using Microsoft Excel .We used IBM SPSS Statistics, Version 25  for data analysis and set a parameter of We obtained ethical clearance from The Regional Committee of Medical and Health Research Ethics (REK-VEST) in Norway (number 2016/635/REK sør-øst dated 20 May 2016); and later from The Muhimbili University of Health and Allied Sciences (MUHAS) Ethics Committee in Tanzania number 2016-06-24/AEC/Vol. XI/38 dated 24 June 2016. Each iron and steel factory and primary school administration was contacted individually, and all of them granted permission to conduct the study. Individual participants were contacted and informed about the research objectives and activities to be conducted and gave written consent. Information that was collected was treated as confidential and was not accessed by unauthorized parties. We used participants’ identification instead of names in data collection, processing and analysis.t-test; p < 0.001) (p < 0.001) but not for the other variables i.e., current smoking, tinnitus, relative with hearing impairment and history of ear-related medical condition (p < 0.05). The overall prevalence of hearing loss was significantly higher  among exposed workers (48%) than among the controls (31%) (The participation rate was 87% for both the exposed and controls. The exposed group was significantly younger than the controls (independent sample < 0.001) . There wondition . Among tondition . In addils (31%) . p 1 = 0.002; p 2 = 0.013) but not for the older age group  had the highest prevalence ratio (2.5), although it was not statistically significant .t-test, p < 0.05)  . In line < 0.05) . In anal < 0.05) .The mean hearing threshold among the participants in the 18–35 age group was similar to the predicted NIPTS according to ISO 1999 at the lower frequencies , while it was about 1 dB higher than ISO 1999 for the higher frequencies (4000 and 6000 Hz) a. For thWe found a higher prevalence of hearing loss among Tanzanian iron and steel factory workers compared to controls i.e., 48% vs. 31% respectively. In addition, a comparison of hearing thresholds between the two groups for the frequencies 4000 and 6000Hz revealed significant differences. To our knowledge, this is the first study in SSA to document the prevalence of hearing loss among workers exposed to noise in iron and steel factories.In the present study, we found a significantly higher prevalence of NIHL among iron and steel workers than the controls. The noise exposed workers were exposed to a mean noise level of 92 dB(A), without using hearing protection devices . At thisAge is one of the main factors for the development of hearing loss. To adjust for age can be difficult in statistical analyses. In the present study, we stratified the working population into three age groups and found a borderline increased risk for hearing loss among the younger age group 18–35 years), and significant differences between exposed and controls in hearing thresholds for the frequencies of 4000 and 6000Hz. The significant difference in the dip for the 4000 and 6000 Hz frequencies is a sign indicating hearing loss due to noise exposure in this age stratum ,18,38,39–35 yearsIn this study we found higher estimates of NIPTS than predicted by ISO 1999 standard for 18–35 years at frequencies of 4000 and 6000 Hz. These frequencies are likely to be affected by noise exposure . HoweverThe control group in our study had a significant lower prevalence of hearing loss compared to the exposed workers at higher frequencies. The measured hearing loss in this group was lower than that recorded among the controls in South African miners study for the higher test frequencies i.e., 31% versus 46% respectively . The conStrengths of this study are the high response rate among the participants, the use of a control group from workplaces with low sound levels, and the use of standardized methods for audiometry. In addition, it was possible to control for the effect of age in hearing thresholds by stratification of age groups while adjusting for age as a continuous variable within the age strata. The statistical analyses made it possible to adjust for potential confounding factors related to hearing loss, such as current smoking, previous noise exposure, tinnitus, history of ear-related medical conditions, duration of work and relatives with hearing impairments. The use of calibrated research equipment and devices together with adherence to the novel procedure related to audiometry testing and ISO 8253-1:2010 standard for ambient noise improved the findings. Our study had some limitations; The design of the study was cross-sectional, and this reduces the possibility to conclude regarding the causal relationship between noise at work and hearing loss. Still, this study indicates that the sound levels are of importance to the registered hearing losses in this working population, as the frequencies involved are in the upper frequency area and the sound levels measured were above 85 dB(A). A longitudinal study would have provided a better exposure-effect association. Information collected through interview questionnaire might introduced recall bias. To minimize this bias, we used the same trained research personnel and method for both the exposed and the controls. In addition, in many societies today, people listen to music at high volume levels, and this may affect their hearing ability. We have limited information about leisure time exposure to noise among our study participants, but we have no reason to believe that the workers in iron and steel factories are more exposed to leisure time sound than are the control workers. In addition, iron and steel workers spent most of their time during the day at work. Lastly, it was impractical to monitor workers at their homes before audiometry.Based on these findings, this study should be a wake-up call for stakeholders in the establishment and should serve to encourage the implementation of noise control measures such as the use of hearing protection devices in these workplaces. The information we found on the high prevalence of hearing loss may be used by policy and decision-makers in awareness creation programmes aimed at noise control such as establishing hearing conservation programmes and preventive services among working populations exposed to noise .


In the present study, two experiments were carried out for studying the pathogenicity of H9N2 avian influenza virus (AIV) in broiler chickens after vaccination with different live respiratory viral vaccines.One-day-old specific pathogen-free (SPF) chicks were divided into four groups in each experiment. In experiment 1, Groups 1 and 2 were inoculated with H9N2 AIV through nasal route in 1 day old, Groups 1 and 3 were vaccinated with live infectious bronchitis coronavirus (IBV) vaccine in 5 days old, and Group 4 was left as a negative control. In experiment 2, Groups 5 and 6 were inoculated with AIV subtype H9N2 through nasal route in 1 day old, Group 5 was vaccinated with live IBV vaccine and live Newcastle disease virus (NDV) vaccine in 5 and 18 days old, respectively, Groups 6 and 7 were vaccinated with live NDV vaccine in 18 days old, and Group 8 was left as a negative control. Chicks were kept in isolators for 18 days in the first experiment and 35 days in the second experiment. Tracheal and cloacal swabs were collected from 3, 5, 7, 10, 12, and 15 day’s old chicks from all groups in experiment 1 and 21, 23, 25, and 28 days old from all groups in experiment 2. Quantitative real-time reverse-transcriptase polymerase chain reaction (rRT-PCR) was applied on the collected tracheal swabs for detecting RNA copies of H9N2 AIV. Cloacal swabs and the positive rRT-PCR tracheal swabs were inoculated in 10-day-old SPF embryonated chicken eggs (ECE) to confirm rRT-PCR results. Internal organs  from all chicken groups were collected weekly for histopathological examination to determine severity of the lesions. Serum samples were collected on a weekly basis for the detection of humoral immune response against H9N2, NDV, and IBV from all chicken groups.rRT-PCR results with virus titration in ECEs revealed a significant increase in H9N2 AIV titer with extension in the period of viral shedding in Groups 1 and 5. Severe lesion score was observed for Groups 1 and 5. The humoral immune response against H9N2 AIV, NDV, and IBV revealed a significant increase in H9N2 AIV titer in Groups 1 and 5, NDV titer showed a significant increase in Group 7, and IBV titer increased in Groups 1, 3, and 5.Results demonstrated the increase in pathogenicity of H9N2 AIV, especially when H9N2-infected chicks vaccinated with live IBV vaccine. Avian influenza (AI) is a respiratory disease in poultry of zoonotic importance caused by influenza A viruses of the family Orthomyxoviridae. Type A influenza viruses are classified into different subtypes according to the two major surface glycoprotein: Hemagglutinin (H) and neuraminidase (N) . Avian iIn Egypt, serological evidence of H9 spread throughout Egypt was recorded on 2009-2010 before isolation of the virus in 2011 .The infections with H9N2 viruses in Egypt are higher in chicken than other species, mostly in apparently healthy broilers, and recorded in layers and breeders .In the present study, two experiments were carried out for studying the pathogenicity of H9N2 avian influenza virus (AIV) in broiler chickens after vaccination with different live respiratory viral vaccines.All applicable institutional guidelines for the care and use of animals were followed in the Central Laboratory for Evaluation of veterinary Biologics (CLEVB).50 was calculated  and 2.319×10 [ct 27.58]) than Group 2 (1.89 [ct 30.73] and 1.19 [ct 31.91]) observed clearly at 7 and 10 days, respectively. Virus shedding extended in Group 1 up to 15 days versus Group 2 only up to 10 days.In experiment 2, there was a significant increase in RNA copies in Group 5 with copies ranged between 2.319×10 (ct 27.80) at 21 days and 1.86 (ct 31.16) at 28 days than Group 6 with copies ranged between 2.017×10 (ct 27.81) at 21 days and 1.36 (ct 31.68) at 28 days .Figure-Titration of H9N2 AIV from tracheal swabs revealed that:In experiment 1, there was a significant increase in Group 1 (3) than Group 2 (1) observed clearly at 7 days old with virus shedding extended in Group 1 up to 15 days versus in Group 2 only up to 7 days.In experiment 2, there was a significant increase in Group 5  than Group 6  at 3, 5, and 7 days postvaccination, respectively, with virus shedding extended in Group 5 up to 10 days versus Group 6 only up to 7 days .Figure-H9N2 AIV titration from cloacal swabs revealed that:In experiment 1, there was a significant increase in virus shedding in Group 1 extended up to 15 days versus Group 2 only up to 5 days.In experiment 2, there was a significant increase in Group 5  than Group 6  at 3, 5, and 7 days old with virus shedding extended up to 10 days in Group 5 versus Group 6 only up to 7 days .Figure-Staphylococcus aureus and Haemophilus paragallinarum activated HA of the virus and increased its replication and spread [Escherichia coli has a vital role in viral spread and virulence of H9N2 AIV [H9N2 AIV causes great economic losses in broiler chickens although the virus is classified as low pathogenic virus ,13,18,19d spread . FurtherH9N2 AIV .On the other hand, it is well known that the hemagglutinin glycoprotein for influenza viruses is produced as a precursor, HA0, which requires cleavage by host proteases before it is functional and virus particles are infectious . The HA0In the present study, lesions of internal organs  of the group coinfected with H9N2 AIV and live IBV vaccines were significantly higher than the H9N2 AIV-infected group in experiment 1; histopathological changes in the kidney were as the results of direct influence of H9N2 AIV replication in renal tubules -33. TherTaken together, results of rRT-PCR assay and virus titration revealed that the vaccination with live IBV vaccine postinfection of the chicks with H9N2 AIV has a great effect on enhancement of the virulence of H9N2 AIV, extend the period of viral shedding, and increase the pathogenicity versus H9N2 AIV-infected group in the experiment ,8. Such It has been demonstrated that live IBV vaccine, H120 strain, exacerbates the manifestation of experimental H9N2 AIV infection in broiler chickens ,38 as IBIn the present study, the results of experiment 1 revealed that HI titer of H9N2 AIV increased in the group infected with H9N2 AIV and vaccinated with live IBV vaccine versus H9N2 AIV-infected group, and these results indicated that IBV could enhance the propagation of H9N2AIV and consequently an increase in H9N2 AIV HI titer ,8. FurthThe results of HI titer of H9N2 AIV in experiment 2 revealed that H9N2 AIV HI titer was reduced in the group infected with H9N2AIV and vaccinated with live NDV vaccine versus the group infected with H9N2 AIV and vaccinated with both live IBV and NDV vaccines, indicating that live NDV vaccine interfered with H9N2 AIV replication and this is in accordance with others -45. InteHistopathological studies in experiment 2 revealed that the infection with H9N2 AIV in birds vaccinated with live NDV alone induces mild lesions. Meanwhile, infection with H9N2 AIV in birds vaccinated with both live IBV and NDV vaccines induced severe lesions in all collected internal organs and these results indicated that live NDV vaccine has no significant effect on the severity of H9N2 AIV . Resultsin vitro using cell cultures or chicken embryos, and interference between these viruses has been reported, with one virus inhibiting the growth of the other [Based on the obtained results, there was no effect of live NDV vaccine on the severity of lesions caused due to H9N2 AIV.Coinfections of H9N2 AIV and NDV have been previously studied he other -50. Virahe other . Viral ihe other . NDV andhe other .Collectively, the overall results of the present study demonstrated that live IBV vaccine postinfection with AIV subtype H9N2 increases its pathogenicity and this may be one of the factors causing economic losses observed in the past 5 years in broiler chickens in Egypt.ZMI conducted the experiment and drafted the manuscript, HAH and AHE designed and followed up the experiment and critically reviewed the manuscript, and MME participated in designing and followed up the practical work. This work is part of the master degree of ZI supervised by HAH, A EL-Deeb, and M EL-Safty. All authors read and approved the final manuscript.


R. Soc. open sci.5, 171529. (Published 27 June 2018). (doi:10.1098/rsos.171529)This correction refers to tables


CLDN14, was reported to be responsible for human hereditary hearing loss, DFNB29. Until now, nine pathogenic variants have been reported, and most phenotypic features remain unclear. In the present study, genetic screening for 68 previously reported deafness causative genes was carried out to identify CLDN14 variants in a large series of Japanese hearing loss patients, and to clarify the prevalence and clinical characteristics of DFNB29 in the Japanese population. One patient had a homozygous novel variant (c.241C>T: p.Arg81Cys) (0.04%: 1/2549). The patient showed progressive bilateral hearing loss, with post-lingual onset. Pure-tone audiograms indicated a high-frequency hearing loss type, and the deterioration gradually spread to other frequencies. The patient showed normal vestibular function. Cochlear implantation improved the patient’s sound field threshold levels, but not speech discrimination scores. This report indicated that claudin-14 is essential for maintaining the inner ear environment and suggested the possible phenotypic expansion of DFNB29. This is the first report of a patient with a tight junction variant receiving a cochlear implantation.Tight junctions are cellular junctions that play a major role in the epithelial barrier function. In the inner ear, claudins, occludin, tricellulin, and angulins form the bicellular or tricellular binding of membrane proteins. In these, one type of claudin gene,  CLDN14 gene.Hearing loss (HL) is the most common sensory impairment and is diagnosed in approximately two in every 1000 children . The inh+ and low Na+ concentrations [+ concentration and EP) are indispensable for cochlear hair cells to transduce acoustic stimuli into electrical signals [LSR, angulin-2/ILDR1, and angulin-3/ILDR2) form the bicellular or tricellular binding of membrane proteins. Among these, claudin-14, tricellulin, and angulin-2 are reported to be responsible for human deafness DFNB29, DFNB49, and DFNB42, respectively [For our sound receiving process, it is necessary to convert sound vibration to nerve action potential (mechano-transduction) . The centrations . Furthertrations ,10. It i signals . The bar signals . Tight jectively ,14,15.Cldn9 mutant mice and Cldn14 mutant mice demonstrate hearing loss. However, these two animal models do not display reduced EP [The claudins are a family of proteins that play a major role in epithelial barrier function, especially in bicellular junctions. There are 24 claudins that have been identified in humans thus far ,16,17,18duced EP ,23. The CLDN14 [CLDN14 variants are a relatively common cause of recessive hearing loss, which is responsible for 2.25% of HL patients in a Pakistani study cohort [CLDN14 variants, the audiovestibular findings, such as progression and details of vestibular function, remain unclear. In addition, there are no reports on patients with cochlear implantation (CI), thus the outcome of CI is also unknown. In the present study, we used massively parallel DNA sequencing (MPS) to detect pathogenic variants in CLDN14 among a large series of Japanese HL patients. To date, nine different variants in CLDN14 ,27,28,29y cohort ,31. AlthCLDN14 gene identified from a group of non-consanguineous HL patients and discuss additional clinical features including CI outcomes. This is the first report of a diagnosis of hearing caused by CLDN14 in an east Asian population and in such a patient receiving a cochlear implant as intervention.Here, we present a patient with a novel variant in the CLDN14: NM_144492: c.241C>T, leading to p.Arg81Cys, was homozygously detected in one of 1577 probands with autosomal recessive HL , and almost all scores indicated “damaging” . The corThe proband was a 37-year-old female. The newborn hearing screening program had not yet started at birth and there were no particular complications in the perinatal period. The female proband also passed an elementary school health checkup at six years of age, but was suspected of hearing loss at age nine and was referred to an otolaryngology clinic at a local general hospital. Pure-tone audiograms showed steep high-frequency sensorineural HL (SNHL) with an average of 77 dBHL in both ears A. It appThe proband underwent a CI (MED-EL FLEX28) in the right ear at the age 35. Sound field threshold levels were improved from 77.5 dBSPL (with hearing aids) to 37.5 dBSPL (with the cochlear implant) A. To evaCLDN14 gene as a cause of progressive SNHL in a Japanese patient. To date, a total of nine pathogenic variants in CLDN14 have been reported  among the Japanese HL patients, and 0.06% (1/1577) among the families with autosomal recessive HL in the Japanese population. In clear contrast to this, variants of CLDN14 account for 2.25% of autosomal recessive HL in the Pakistani population. In this study, the patient was identified from a non-consanguineous family and the parents of the proband were from a geographically remote area, whereas almost all of the previous reports were from consanguineous families [CLDN14 variants were found in our cohort, but one HL patient with a homozygous CLDN14 variant was identified, it is necessary to consider such rare HL cases.The incidence of families ,27,28,29CLDN14 remains unclear, and further study including serial audiograms is required to clarify this. Until Pater et al. reported the progression of HL , the pheCldn14 knockout model mice has not been reported and the details remain unknown. This is the first report to mention in detail the vestibular function, including caloric testing, cVEMPs, and oVEMPs. Endolymph in cochlear is characterized by extremely high resting potential called EP, in addition to high K+ concentration. In contrast, in the vestibular endolymph, the K+ concentration is high, but no EP can be detected. Claudin-14 may be necessary for maintaining a barrier to resist EP, but not K+.We could not identify any dysfunction on the vestibular examinations. According to past reports, claudin-14 is also expressed in the vestibule of mice (such as in the sensory epithelia) . VestibuCLDN14-associated HL patient. A CI represents the most successful neural prosthesis in clinical cases [This is the first report showing the outcome for a CI for a al cases . For patal cases . The impal cases .CLDN14 was expressed at the same level in the organ of Corti and spiral ganglions [As mentioned above, the patient showed relatively poor improvement in speech discrimination scores, whereas good improvement in sound field threshold levels was observed. Since the cochlear implant directly stimulates the cochlear nerve, the influence of the inner ear should be excluded. Thus, it is likely that the unsatisfactory speech discrimination was due to deficiencies in the spiral ganglion or central auditory pathway. In our previous study, anglions . IncludiAll procedures were approved by the Shinshu University Ethical Committee (No. 387—4 September 2012 and No. 576—2 May 2017) as well as the respective Ethical Committees of the other participating institutions described elsewhere  and wereFor the genetic analysis for this proband, we performed target genome enrichment for 68 previously reported genetic causes for deafness and MPS analysis described elsewhere . In brieThe missense, nonsense, insertion/deletion, and splicing variants were selected from among the identified variants. Variants were further selected as less than 1% of (1) the ExAC ,43, (2) CLDN14 variant.The age of onset of HL, the incidence of progressive HL, and episodes of vertigo and dizziness were analyzed based on the medical charts of the probands with the Pure-tone audiometry was performed to evaluate HL. The pure-tone average (PTA) was calculated from the audiometric thresholds at four frequencies . If an individual did not respond to the maximum hearing level at a frequency, 5 dB was added to the maximum hearing level. The severity of HL was classified into mild (PTA: 20–40 dBHL), moderate (41–70 dBHL), severe (71–95 dBHL), and profound (>95 dBHL). The audiometric configurations were categorized as low-frequency, mid-frequency, high-frequency (gently or steeply sloping), flat, and deaf . The vestibular examination findings, including caloric testing and the measurement of cervical and ocular vestibular evoked myogenic potentials (c/oVEMPs), were analyzed. Caloric testing involved the measurement of the maximum slow phase velocity (SPV) by cold water irrigation . We defined a maximum SPV value below 10 deg/s as representing areflexia and a value between 10 and 20 deg/s as representing hyporeflexia. For cVEMPs testing, electromyography (EMG) was performed using a pair of surface electrodes mounted on the upper half and sternal head of the sternocleidomastoid muscle, respectively. The electrographic signal was recorded using a Neuropack evoked potential recorder . Clicks lasting for 0.1 ms at 105 dBnHL were presented through a headphone. The stimulation rate was 5 Hz, the bandpass filter intensity was 20 to 2000 Hz, and the analysis time was 50 ms. The responses to 100 stimuli were averaged twice. The oVEMPs testing was measured by bone-conductive vibration (BCV). The BCV was delivered in 4 ms tone bursts of 500 Hz vibration  by using a hand-held 4810 mini-shaker , which was placed on the midline (Fz) of the forehead. The active electrode was located over the inferior orbital margin and a reference electrode was placed 2 cm below the active electrode. The ground electrode was placed on the chin. The patients laid in a supine position on a bed and looked up at an angle of approximately 30 degrees above straight ahead during the recording. The signals were amplified and bandpass filtered between 20 and 2000 Hz. The stimulus intensity was 115 dB force level, 500 Hz with an analysis time of 40 ms, and 50 responses were averaged for each run. The VEMPs asymmetry was calculated as follows: asymmetry ratio (AR) =  × 100/. In this study, an asymmetry ratio of >30% was defined as a decreased reaction and no reaction in amplitude VEMPs as absent.Intervention for HL, including the use of hearing aids or cochlear implants, was investigated. To evaluate speech perception outcomes, speech discrimination scores (using the 67S Japanese monosyllable test) were used. The subjects sat 1 m away from the sound source facing zero-degree azimuth, and recorded monosyllable words in quiet were presented in the sound field at 65 dBSPL.CLDN14 gene identified from a non-consanguineous family. This is the first report of CLDN14-associated HL in an east Asian population. Serial audiograms indicated high-frequency hearing loss type, and the deterioration gradually spread to other frequencies, finally resulting in deafness. The patient showed normal vestibular function for caloric testing, cVEMPs, and oVEMPs. This is also the first report of an HL patient with a tight junction variant receiving a CI. The CI improved the proband’s sound field threshold levels, but not their speech discrimination scores. This information contributes to our understanding, diagnosis, and treatment of HL caused by TJ disorders.We present a patient with a novel variant in the


The median durations (interquartile range) for DIHL due to platinum compounds, sulfonamides (plain), interferons, antivirals for treatment of hepatitis C virus (HCV) infections, other aminoglycosides, carboxamide derivatives, macrolides, and pneumococcal vaccines were 25.5 (7.5–111.3), 80.5 (4.5–143.0), 64.0 (14.0–132.0), 53.0 (9.0–121.0), 11.0 (3.0–26.8), 1.5 (0.3–11.5), 3.5 (1.3–6.8), and 2.0 (1.0–4.5), respectively. Our results demonstrated potential risks associated with several drugs based on their RORs. We recommend to closely monitor patients treated with aminoglycosides for DIHL for at least two weeks. Moreover, individuals receiving platinum compounds, sulfonamides (plain), interferons, and antivirals for HCV infection therapy should be carefully observed for DIHL for at least several months.Many drugs can cause hearing loss, leading to sensorineural deafness. The aim of this study was to evaluate the risk of drug-induced hearing loss (DIHL) by using the Japanese Adverse Drug Event Report (JADER) database and to obtain profiles of DIHL onset in clinical settings. We relied on the Medical Dictionary for Regulatory Activities preferred terms and standardized queries, and calculated the reporting odds ratios (RORs). Furthermore, we applied multivariate logistic regression analysis, association rule mining, and time-to-onset analysis using Weibull proportional hazard models. Of 534688 reports recorded in the JADER database from April 2004 to June 2018, adverse event signals were detected for platinum compounds, sulfonamides (plain) (loop diuretics), interferons, ribavirin, other aminoglycosides, papillomavirus vaccines, drugs used in erectile dysfunction, vancomycin, erythromycin, and pancuronium by determining RORs. The RORs of other aminoglycosides, other quaternary ammonium compounds, drugs used in erectile dysfunction, and sulfonamides (plain) were 29.4 (22.4–38.6), 18.5 (11.2–30.6), 15.4 (10.6–22.5), and 12.6 (10.0–16.0), respectively. High  Around 466 million people worldwide have disabling hearing loss, and it is estimated that by 2050, over 900 million people will have disabling hearing loss (www.who.int/news-room/fact-sheets/detail/deafness-and-hearing-loss). Thus, hearing loss can have significant emotional and social impact.Hearing loss leads to a number of issues such as inability to recognize speech, depression, withdrawal, anger, loss of self-esteem, and poor quality of life (www.who.int/news-room/fact-sheets/detail/deafness-and-hearing-loss). More than 150 drugs such as platinum-based anticancer drugs and aminoglycosides are currently known to ototoxic  → Y , where X and Y are mutually exclusive sets of items [support, confidence, and lift values can be calculated to evaluate the correlation of this rule. Support determines how often a rule, which in this case is the combination of X and Y, is observed in the database. Support was measured using the following formula:Association rule mining is a common technique used to identify associations among numerous variables. Given a set of transactions of items . If the confidence of the association rule demonstrates the rule’s strength [Confidence was calculated by the following equation:The strength . ConfideLift represents the ratio of probability. Given a rule, X and Y occur together to the multiple of the two individual probabilities for X and Y:Lift evaluates the independence of X and Y, and higher lift values indicate stronger relationships. If X and Y are independent, lift equals 1. If X and Y are positively or negatively correlated, lift is > 1 or < 1, respectively. Association rule mining was performed utilizing the arules package of the R software (version 3.6.0). The parameter maxlen  is the maximum size of mined frequent itemsets. To extract association rules efficiently, the thresholds for the optimized support, confidence, and maxlen are defined depending on factors such as size of the data, number of items, and research purpose. In this study, we defined the minimum support and confidence thresholds as 0.00001 and 0.1, respectively; furthermore, maxlen was restricted to 3.Time-to-onset duration from the JADER database was calculated from the time of the patient’s first prescription to the occurrence of the AEs. The median duration, quartiles, and WSPs were used to evaluate the time-to-onset data. The scale parameter α of the Weibull distribution determines the scale of the distribution function. A larger-scale value (α) stretches the distribution, while a smaller scale value shrinks the data distribution. The shape parameter β of the Weibull distribution determines the shape of the distribution function. A larger shape value gives a left-skewed curve, whereas a smaller shape value gives a right-skewed curve. In the analysis of the SRS, the shape parameter β of the Weibull distribution was used to indicate hazards without a reference population as follows. When β was equal to 1, the hazard was estimated to be constant over time. If β was greater than 1 and the 95% CI of β excluded the value 1, the hazard was considered to increase over time. Finally, if β was less than 1 and the 95% CI of β excluded the value 1, the hazard was considered to decrease over time , 10–13. The JADER database contains 534688 reports submitted from April 2004 to June 2018, and we identified 1218 DIHL events. The top seven AEs were deafness , sudden hearing loss , hypoacusis , deafness neurosensory , deafness unilateral , auditory disorder , and deafness bilateral  . The lowUsing a stepwise logistic regression model, we examined and selected significant DIHL-related variables among the demographic factors (sex and age-stratified group) and reporting year. The results in the final model indicated significant contributions to DIHL of the ≤ 19 years group (p < 0.0001), the female and ≤ 19 years group (p < 0.0001), and the female and 40–59 years group (p = 0.0012). Contrarily, the reporting year did not significantly contribute to DIHL (data not shown). Adjusted RORs for the analyzed groups were as follows: 1.10 for the female group ; 2.03 for the ≤ 19 years group ; 1.13 for the 40–59 years group ; 4.32 for the female and ≤ 19 years group ; and 0.75 for the female and 40–59 years group . In the mosaic plot, papillomavirus vaccines or other quaternary ammonium compounds were associated with DIHL only in the ≤ 19 years group, causing DIHL in 12.0% and 7.4% of the cases, respectively .Furosemide, ribavirin, and cisplatin were the three drugs most frequently associated with DIHL . Therefolift value , interferons, antivirals for treatment of HCV infections, other aminoglycosides, drugs used in erectile dysfunction, glycopeptide antibacterials, and other quaternary ammonium compounds are stated in the many reports –37, whiclift and support values of {≤ 19 years, pancuronium} → {hearing loss} were high  and cisplatin (p = 0.1873) displayed a tendency of being low. However, there were no significant differences in the average doses between cases with and without DIHL.Time-to-onset profiles of DIHL have been systematically addressed using the JADER database. Our results suggest that attention should be paid to the possibility of DIHL onset with aminoglycosides and careful observation is recommended soon after the administration for at least 2 weeks. The median durations for DIHL due to platinum compounds and macrolides were 25.5 and 3.5, respectively, and these results support those reported previously [Cisplatin chemotherapy has been widely used in cancer treatment . FollowiIt is reported that high trough levels for aminoglycosides  and aging may be the risk factors for auditory toxicity . NeomyciThe ROR signals were detected for sulfonamides (plain) and vancomycin . It is sSeveral cases of hearing loss during interferon therapy or combination treatment with peginterferon and antiviral for HCV infections (ribavirin) have been reported –30. In oROR signals were detected for papillomavirus vaccines . To the Phosphodiesterase 5 (PDE5) inhibitors such as sildenafil are prescribed for erectile dysfunction . Case stlift value  and adopted the most recent case number to identify duplicate patient reports and exclude them from the analysis. However, there is no key code to identify duplicate reports in the JADER database, rendering their removal challenging. The PMDA has introduced an evaluation method based on a match score for duplicate detection in the JADER database [In the JADER database, duplicate data may be present due to follow-up reports on a case or different individuals disclosing the same patient case. It is recommended to identify duplicate patient reports originating from different sources and to exclude them from the analysis. For example, for the US Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS), we followed the agency’s recommendations  (loop diuretics), interferons, ribavirin, other aminoglycosides, papillomavirus vaccines, drugs used in erectile dysfunction, vancomycin, erythromycin, and pancuronium. Our finding indicated that patients with congenital diaphragmatic hernia should be monitored after pancuronium administration. Patients treated with aminoglycosides should be closely observed for DIHL for at least 2 weeks. After the administration of macrolides, pneumococcal vaccines, and carbamazepine, it may be necessary to monitor individuals for hearing loss for a few days. However, careful observation for at least several months is recommended after the administration of platinum compounds, sulfonamides (plain), interferons, and antivirals for treatment of HCV infections. Despite the inherent limitations associated with SRS data, we believe that our findings represent a valuable contribution to the clinical knowledge and will help improve the management of DIHL.


Horizontal Gene Transfer (HGT) refers to the transfer of genetic materials between organisms through mechanisms other than parent-offspring inheritance. HGTs may affect human health through a large number of microorganisms, especially the gut microbiomes which the human body harbors. The transferred segments may lead to complicated local genome structural variations. Details of the local genome structure can elucidate the effects of the HGTs.In this work, we propose a graph-based method to reconstruct the local strains from the gut metagenomics data at the HGT sites. The method is implemented in a package named LEMON. The simulated results indicate that the method can identify transferred segments accurately on reference sequences of the microbiome. Simulation results illustrate that LEMON could recover local strains with complicated structure variation. Furthermore, the gene fusion points detected in real data near HGT breakpoints validate the accuracy of LEMON. Some strains reconstructed by LEMON have a replication time profile with lower standard error, which demonstrates HGT events recovered by LEMON is reliable.Through LEMON we could reconstruct the sequence structure of bacteria, which harbors HGT events. This helps us to study gene flow among different microbial species. Horizontal Gene Transfer , 2 is thHGTs may affect human health through a large number of human microbiota , includiIt is necessary to understand HGTs better. However, our current research is mainly focused on inferring ancient (lineage) HGTs from genomic sequences . While thttps://github.com/lichen2018/LEMON) takes use of existing shotgun NGS datasets to detect HGT breakpoints, identify the transferred segments, and reconstruct the local strain, which has complicated structure variation.LEMON [R, we utilize BWA[D and H respectively.Only the assembly results from multiple time-point metagenomics data of one individual can be used to discover the HGT events that exactly occurred between these time points. However, these samples are insufficient in published data, and this method cannot evaluate the difference in HGT events between different samples. To solve this problem, we construct a n (NCBI) . We selen (NCBI) . The refilize BWA to alignbreakpoints. We need to identify the breakpoints and segments involved in the local strains from D and H. The data are heterogeneous. According to studies on virus integration [The donor segments and receptor segments interweave in a local strain, separated by HGT egration  and studegration . Hence wA and B in Fig. Bx and By in Fig. B on reference X and Y respectively. Then pair B can be transformed to the point B′ with coordinates  in Fig. supports a cluster if the mapped positions belong to the cluster. Each cluster contains multiple breakpoint pair candidates.First, we map paired-end shotgun reads to reference genomes using BWA, here all references are indexed together to generate Burrows–Wheeler Transform (BWT) indexes. If the two sides of a paired-end read are mapped to two different references, we call such a pair a junction pair, such as junction pairs R1 and R2. Given a candidate genomic positions pair p1 and p2 which belongs to R1 and R2, respectively, we identify the split reads aligned to R1 where the alignment terminated at position p1. Denote the portions of such a split read s mapped to R1 and R2 as e1(s) and e2(s), respectively. Then, the score is defined according to the alignment qualities of e2(s) against R2 nearby p2. The alignment quality qs of e2(s) is calculated as m/l, where m is the number of matched positions and should be at least 15 bps, l is the length of e2(s). The quality score of p2 is calculated as p1. The candidate with the highest score is reported as the final breakpoint pair positions of the cluster.To find the exact positions, we use a scoring scheme to rank the candidate positions. The candidate with the highest score is reported as the final breakpoint pair position. The scoring scheme evaluates the split reads that support the cluster. Suppose that there are two references involved in the cluster, xu,xv∈{+,−} where + and − respectively indicates the positive and negative strands, and u,v∈V. We call such a pair a connected pair. A connected pair is directed; that is, E, the copy number of e∈E as c(e) w.r.t. R. It is easy to see that the segments and connected pairs specify a graph G as illustrated in Fig. Each breakpoint involves two segments. Denote the segment pair as mentclass2pt{minimk HGT events captured in G. Since each HGT event results in one local strain, there are k local strains to be constructed according to G. The first and last segments of each local strain are from the same receptor. Without loss of generality let all the first segments and the last segments of the local strains at the integration sites be denoted B={b1,...,bk}, and T={t1,...,tk}, respectively. Denote all the other segments involved in the HGTs as V={v1,...,vn} (excluding B and T). Next, we estimate the number of copies (copy number) of each segment v within R, and denote it as c(v), where v∈B∪T∪V. Factors such as species coverage are incorporated into the copy number estimation. For example, if segment vi is belong to a receptor r, the coverage of vi is cov(vi) and the average coverage of r is cov(r), then the initial copy number of vi is estimated as c(vi)=cov(vi)/cov(r).Assume that there are k local strains captured in G. B and T can be identified with the input data. We assume that the copy numbers of the first segment and the last segment are the same for each strain, that is, c(bi)=c(ti), without loss of generality.Our task is to identify the k local strains.We formulate the problem as a Eulerian circuit problem to find the c(bi)=c(ti),1≤i≤k. We insert a dummy edge c(bi)−1,1≤i≤k. The edges First, we insert dummy edges to transform the solution into a circuit. Without loss of generality, we assume v∈V, there should be a path that starts from a source node in S, passes through v, and ends up at some target in T. However, due to sequencing errors, edges or nodes can be missing or spuriously introduced. If no target and source can reach a node v, we remove v and its adjacent edges from G. If a path exists from some vertex s to v, but no path exists from v to a target t, we insert vertices and edges to form a path from v to t. The inserted edge candidates are taken from the reference sets D and S. If v belongs to the same reference as t, this would suffice to reconstruct the path. Otherwise, we add edges that connect v and some nodes on the reference of t. In both situations, we use the minimum number of edges required. All the introduced edges and vertex are assigned a copy number of 1.Second, we insert edges and nodes to ensure connectivity. In the ideal case, for each vertex u as in(u) and the outbound edges of u as out(u). That is, in(u)={u|,∈E,x∈{+,−}},out(u)={u|,∈E,x∈{+,−}}. The in-copy and and out-copy of a vertex v are defined as cin(u)=c(u)=cout(u), but this may be broken due to experimental and sequencing errors.Denote the set of inbound edges of u (respectively μ) to a target copy number t(u) (respectively t(μ)) according to Eq. We propose an integer linear programming (ILP) approach to optimize the degree balance property. First, assign each segment i be denoted as hi. We choose a solution in which each reconstructed strain i has hi as large as possible.It can be shown that a Eulerian circuit to the graph constructed as illustrated in Fig. To evaluate the performance of LEMON, we construct true local strains containing transferred segments and use LEMON to recover them. We propose two metrics Reconstruction Accuracy and Detection Rate to measure results.n is the number of receptors, and each true local strain {Hi} consists of mi segments, that is, hi is the reconstructed local strain which has the same receptor of Hi; if Hi doesn’t have hi in the result, we set hi=∅. The segments in hi are denoted Hi and hi. match_score and mismatch_score of Smith-Waterman algorithm are set as 1 and -1, respectively.We denote the true local strains as RAi of hi is defined in formula  is the alignment score of hi and Hi, while mi is the number of segments in Hi. When all segments are matched in hi and Hi, which means all transferred segments are recovered. So SW is equal to mi∗match_score, that is RAi=1.Here, N as 8 and define the mean value of reconstruction accuracy We set the number of repetitions nk denotes the number of non-empty hi in the j-th repetitions.Detection Rate is defined as the rate of the number of acceptable recovered transferred segments and the number of all transferred segments. The average detection rate is the average of 8 repetitions for each test.An acceptable detection of one transferred segment should have its breakpoint pair located within 20 bp  of the teps, which is the maximum radius of one cluster, and minsample, which is the minimum number of points in one cluster. In our paper, eps is set as the average insert size. minsample is set as 1.Most third-party tools used in this article are set default parameters, including BWA, LUMPY , iRep 2, and STATo simulate human gut microbiome with different complexity as mentioned in “In order to prove the performance of LEMON, we compared its performance with another popular breakpoint detection-based structural variant discovery software LUMPY.Figure s1 and s2 in Fig. In order to evaluate the performance of LEMON in HGT breakpoints detection as mentioned in “In this simulation, we set the number of species s to 2560 and the number of HGT events to 100. In order to simulate a complicated HGT event structure, we changed the number of transferred segments in each HGT event from 1 to 5. Simulated paired-end reads were generated from true local strains by using WGSIM at 5X, 10X, 20X, and 30X coverage. 4 repetitions were performed.ΔRA=RALEMON−RALUMPY to measure the performance difference between LEMON and LUMPY. If ΔRA is positive, LEMON achieves a higher RA than LUMPY. As we can see from Fig. ΔRA are positive and ΔRA decreases as coverage increases. Figure RALEMON is better than RALUMPY. Figure Figure In Fig. We applied LEMON on a recently released metagenomic dataset  to reconAs we can see from Fig. We used iRep to estimate the replication timing profile of each bacterium in metagenomics data . iRep utWe applied iRep to evaluate two coverage distributions for each receptor. The first coverage distribution is evaluated based on the original reference of the receptor containing no HGT event. The second coverage distribution is evaluated based on the reconstructed reference of the receptor containing HGT events. According to the two coverage distributions, we estimated two replication timing profiles for each receptor. Since iRep utilizes the regression method to estimate replication timing profiles, we use Standard Error to measure the accuracy of the estimated replication timing profiles. Figure In order to verify the HGT breakpoints detected by LEMON, we analyzed the IBD (Inflammatory Bowel Disease) data set published by HMP (Human Microbiome Project) . In addiThe reads supporting the breakpoint, NZ_DS981501.1:4185-NZ_CP015401.2:963348, are shown in Additional file We identify 3 main reasons for discrepancies between STAR-Fusion-detected gene fusion breakpoints and our HGT breakpoints:1) The results of STAR-fusion are based on STAR aligner, while our algorithm is based on BWA. STAR aligner and BWA employ different alignment algorithm, giving rise to different breakpoints results;2) Limited sequencing data. The amount of metagennomics sequencing data in the IBD data set is around 5G per sample, and the amount of metatranscriptome data is 2G per sample. This is insufficient for the statistical significance required for finding all the breakpoints;3) Based on our statistics in Table In summary, it is reasonable to find only 3 matching breakpoints in 17 pairs of data.In this paper we present LEMON, a novel HGT discovery software that can detect HGT events and reconstruct strains containing multiple HGT events with complicated structural variation.Using LEMON to reconstruct the sequence structure of bacteria allows us to study the metagenomics problem from the sequence level, thus no longer subjected to the comparison of abundance. For example, since HGT is the fundamental mechanism for the spread of antibiotic resistance in bacteria, by utilizing LEMON we could detect transferred Antibiotic Resistance Genes (ARG), determiHowever, as the amount of sequencing data is generally insufficient for current metagenomics analysis, challenges remain in identifying the HGTs sensitively and accurately. This results in several shortcomings in LEMON. First, LEMON remains weak in finding HGT between the sequences that do not exist in the reference genome. Second, because we only consider the reads of unique mapping, the HGT on the repeat region cannot be identified.At present, our reference set only contains the genome of bacteria. However, human gut microbiome also contains other microorganisms such as fungi and viruses. Therefore, a reference library that contains sequences of bacterial, viral and fungal more completely would be highly desirable for HGT analysis of the microbiome.Additional file 1Detailed reads mapping result at HGT breakpoints NZ_DS981501.1:4185 - NZ_CP015401.2:963348. Top-left is upstream genome; top-right is downstream genome. STAR-Fusion determined genes around gene fusion points are annotated with border bar. Red lines represent breakpoints detected from metagenomics data with HGT algorithm. Blue lines represent breakpoints detected from metatranscriptome data with STAR-Fusion. In red rectangle, top sequence with base name is local strain constructed with HGT breakpoints information, and other color bars are metagenomics reads support breakpoints. In blue rectangle, top sequence with base name is local strain constructed with gene fusion breakpoints, other color bars are metatranscriptome reads support those breakpoints.


Slow-moving tropical cyclones (TCs) can cause heavy rain because of their duration of influence. Combined with expected increase in rain rates associated with TCs in a warmer climate, there is growing interest in TC translation speed in the past and future. Here we present that a slowdown trend of the translation speed is not simulated for the period 1951–2011 based on historical model simulations. We also find that the annual-mean translation speed could increase under global warming. Although previous studies show large uncertainties in the future projections of TC characteristics, our model simulations show that the average TC translation speed at higher latitudes becomes smaller in a warmer climate, but the relative frequency of TCs at higher latitudes increases. Since the translation speed is much larger in the extratropics, the increase in the relative frequency of TCs at higher latitudes compensates the reduction of the translation speed there, leading to a global mean increase in TC translation speed. How the translation speed of tropical cyclones has changed in recent decades and will change in the future has been the subject of debate. Model results show that on average, they have not slowed down in the past, but despite a slowing of tropical cyclones at higher latitudes, a poleward shift in their mean track location causes a general speed up under high greenhouse gas emissions. In relation to the anticipated increase in precipitation associated with TCs in the future4, how the translation speed of TCs has changed in recent decades and will change in a future warmer climate is an area of active research8. Kossin6  concluded, based on observational data , that the translation speed of TCs has decreased globally by 10% between 1949 and 2016, possibly due to global warming. The magnitude of the slowdown varies from basin to basin, with the largest slowdown being in the western North Pacific basin. However, Moon et al.7  cast doubts on K18’s conclusions, pointing out that such a slowdown trend arose mainly due to the lack of detection of weak and over-the-sea TCs, which have a relatively slow translation speed, in the pre-satellite era (1965 and before). Because it is impossible to reproduce the observational data during the pre-satellite era with the same quality as those in the post-satellite era, we propose to analyze results of numerical simulations for the current and future climates.Tropical cyclones (TCs) are one of the most intense weather systems in the world and thus it is of great importance to better understand how the behavior of TCs may change under global warming such as their frequency and intensity. Translation speed of TCs is one important feature because the slower TCs move, the longer is their influence time, and the greater is the impact of severe weather events associated with TCs such as heavy rain and strong windsHere the following two questions are at the center of discussion in this study. The first question is whether there exists a decreasing trend in the translation speed of TCs during the historical period, and the second is how the translation speed might change in the future warmer climate. Results from the model used in this study show that the global mean TC translation speed has not slowed down in the past, but under global warming, they will in general speed up due to a poleward shift in their mean track location, although those moving into the extratropics slow down.10 (see Methods). It is important that these simulations reproduce important aspects of the observed TC frequency and distribution under current climate conditions in order to address the above questions with some confidence. Yoshida et al.10  showed that the simulated annual TC genesis and TC distribution match the observations quite well . Comparisons of the simulation results between the current and future climates enable us to explore the second question: how the translation speed might change in a future warmer climate. Table Care must be taken in interpreting this result. In a future warmer climate, the westerly winds are expected to weaken due to the decrease in baroclinicity in the atmosphere. Since the westerly winds play an important role as the steering flow of TCs, these weakened westerly winds will lead to a reduction in the TC translation speed in the extratropics  and sea ice concentration from COBE-SST229 are used as the lower boundary conditions. The period of the simulations is 60 years (1951–2010) and the ensemble size is 100. In addition to using different initial conditions, small perturbations of SST based on its analysis error are adopted for the ensemble experiments. Global mean concentrations of greenhouse gases are set to the observational values for each year. A future climate scenario in which the global mean surface air temperature is assumed to be 4 K warmer than the pre-industrial climate is simulated, corresponding to the global warming around the end of the twenty-first century under the representative concentration pathway 8.5 (RCP8.5) scenario of CMIP5. The amplitude of the warming is kept constant throughout the simulations. The period of the simulations is 60 years and the ensemble size is 90. Six CMIP5 models are used for obtaining the warming pattern of the SST and each pattern is added to the observational SST of 1951–2010. For each warming pattern, 15-member ensemble experiments are conducted using different initial conditions and sea surface perturbations for each SST change.The large-ensemble simulations were conducted by an atmospheric global circulation model (AGCM). The AGCM used in this study is the Meteorological Research Institute AGCM, version 3.2 (MRI-AGCM3.2) with a 60 km horizontal grid spacing, which is exactly the same as MRI-AGCM3.2H listed in the CMIP5 archive28. Appropriate threshold values of these variables are chosen so that the global number of TC geneses detected in the current climate simulations by MRI-AGCM3.2H is close to that observed. Compared to other explicit detection methods, the TC tracking method in the present study does not show large differences in TC detection30.The TC tracking is based on the model outputs, including sea level pressure, 850 hPa relative vorticity, 850 hPa and surface wind speed, vertical wind shear between 850 and 300 hPa, warm core temperature, and duration periodThe TC tracking data include the central position of TCs every 6 h. For each ensemble member and each year, we calculated great-circle distance between two locations along TC tracks and computed the annual average translation speed of the ensemble member for the year. We repeated this computation for all ensemble members and then took an ensemble mean to obtain the annual-mean translation speed. The absolute frequency is calculated by counting the number of 6-hourly tracked TC central locations that meet the conditions of the latitudinal or translation speed bins.Supplementary Information