Triclosan (TCS) is an organic substance showing antibacterial action, which is commonly used in many branches of industry, including, among others, cosmetics, pharmaceuticals and the food industry. TCS may penetrate into living organisms and negatively affect the health of humans and animals. The majority of previous investigations on TCS biomonitoring in humans have been performed on urine, but currently, studies on hair samples are becoming increasingly important. The aim of this study was to evaluate TCS concentration levels in residents of Olsztyn, a city in northeastern Poland, using a liquid chromatography-mass spectrometry technique. The presence of TCS was observed in 96.7% of samples tested, with concentration levels from 37.9 pg/mg to 3386.5 pg/mg. The mean concentration level of TCS in the present study was 402.6 (±803.6) pg/mg, and the median value was 103.3 pg/mg. Although there were some differences in TCS concentration levels between males and females, humans of various ages and humans with colored and natural hair had no statistically significant differences in TCS concentration levels. The obtained results have clearly indicated that people living in northeastern Poland are exposed to TCS to a large degree, and hair analysis, despite some limitations, is a suitable method for TCS biomonitoring in humans.  Triclosan   is an orPrevious studies have shown that TCS may penetrate into living organisms. Mainly, it penetrates the digestive tract and skin, but it can also enter the organism through the mucosal membrane of the oral cavity and lungs ,9,10,11.For years, TCS has been regarded as a neutral substance for the living organism. However, the latest observations have described that this substance adversely affects various internal organs and systems. Therefore, TCS is considered to be a potential endocrine disruptor. It is known that TCS affects the immune system, leading to changes in the synthesis of some cytokines ,26,27. IUntil recently, human exposure to organic xenobiotics that pollute the environment has been mainly determined by analyzing urine and/or blood samples, but in recent years, the analysis of hair samples has become more and more important ,41,42,43Nevertheless, it should be pointed out that the use of hair samples in the biomonitoring of xenobiotics is a relatively new method, and not all aspects connected with this issue are clear. According to the best knowledge of the authors, until now, only three investigations have described the presence of TCS in human hair samples. In one case, samples were collected in Germany , and in The main matrix in studies on human exposure to TCS are the urine samples. However, although urine analysis is invaluable in toxicology and well reflects TCS concentration levels in the organism, it does not explain all aspects connected with the evaluation of exposure to TCS and the distribution of this substance in the organism. This is because TCS levels in the urine change frequently due to daily excretion. It is known that orally administered TCS is rapidly distributed in the organism through the blood, but the major fraction of this substance is extracted within 24 h after administration, and even more than 80% of TCS is extracted during the first 4 days after exposure . TherefoTherefore, the aim of this study was to evaluate human exposure to TCS using hair samples collected from adult citizens of Olsztyn, a city in northeastern Poland, as well as correlating those samples with gender, age, hair coloring and TCS concentration levels. The study supplements knowledge both about human exposure to TCS in the world and hair analysis as a method used in xenobiotic toxicology.TCS and ammonium acetate (≥98%) were purchased from Sigma-Aldrich, methanol (LC-MS grade) from Honeywell Riedel de Haën , acetonitrile (LC-MS grade) from Fisher Chemical and phenobarbital-d5 from Isotec Inc. . Ultrapure water was produced by Merck’s Direct-Q 3UVwater purification system . Head hair samples (about 2 g) were collected in 2021 from 30 volunteers  who were residents of Olsztyn, a city with about 170,000 residents in northeast Poland. Volunteers were aged 22–67 (mean 39.2 ± 13.85) and did not report any chronic medical conditions and/or continued medication. Moreover, basic information, such as gender, age and if the hair was dyed or not, was collected from volunteers. Information about volunteers included in the study is shown in m/z ions for the monitoring of triclosan were 286.95, 289.00 and 291.00. The elution gradient program started at 15% solvent B (1 min), 95% solvent B (20 min) and finally at 15% solvent B for 4 min. Under the above chromatographic conditions, the retention time of TCS was 22.03 min.Hair samples were washed in ultrapure water (two times) and in methanol (two times) to remove external contamination. After washing, samples were dried and cut into small pieces. The extraction process was made in accordance with the method previously described ,46. In bDuring the present study, one measurement was made per hair sample. The analysis was performed with a liquid chromatography–mass spectrometry system , and the separation of TCS was made on a Discovery C18 column   at 30 °C. A gradient of 5 mM ammonium acetate (solvent A) and acetonitrile (solvent B) were chosen for the analysis at 0.6 mL/min. Atmospheric pressure chemical ionization (APCI) combined with a quadrapole mass filter in a selected ion monitoring (SIM) negative mode were used for monitoring the aforementioned substance. To ensure the efficacy of the method used in the present study, analytical parameters were checked. Standard solutions of TCS were performed in concentrations of 0, 50, 100, 250 and 500 ng/mL. Linearity of the standard solution responses was found to be 0.9998. Moreover, spiked samples were performed in concentrations of 0, 50, 100, 250 and 500 pg/mg, and linearity was found at 0.9937.Limit of detection (LOD) and limit of quantification (LOQ) were calculated using the signal to noise ratio. Recovery, accuracy and inter-day precision (%RSD) of the method were examined using 3 repeats of spiked samples (n = 3) for each of the above-mentioned tested concentrations  .p < 0.05.A non parametric Mann–Whitney test—GraphPad Prism version 9.2.0 —was used in the statistical analysis during the comparison of two groups of volunteers . The differences were considered statistically significant at TCS was found in 96.7% of samples tested. The concentration levels of this substance in samples from particular volunteers were very different and fluctuated from 37.9 pg/mg to 3386.5 pg/mg. The mean concentration level of TCS in the present study was 402.6 (± 803.6) pg/mg and the median was 103.3 pg/mg. Concentration levels of TCS noted in the present study are presented in During the present study, concentration levels of TCS in women and men were different. In women, the concentration levels of this substance fluctuated from 37.90 pg/mg to 3387 pg/mg (mean 643.6 ± 1075 pg/mg). In men, the values were lower and fluctuated from 51.80 pg/mg to 318.6 pg/mg (mean 144.4 ± 88.40 pg/mg) . HoweverSome differences in concentration levels of TCS were also visible between volunteers at the age of 22–35 and persons at the age of 45–67 . In the During the present study, the comparison of TCS concentration levels in volunteers with colored and non-colored  hair was also done. In persons with colored hair , the concentration levels of TCS fluctuated from 49.30 pg/mg to 3387 pg/mg, with a mean value of 997.9 ± 1290 pg/mg. In persons with natural hair, the values were smaller and ranged from 37.90 to 318.6 pg/mg, with a mean value of 134.7 ± 85.33 pg/mg. However, there were no statistically significant differences between these groups .During the present study, TCS was found in a high percentage 96.7%) of samples analyzed, which proves a high degree of human exposure to this substance. On one side, such exposure is connected with the common use of TCS in the production of personal care products, cosmetics, medicaments and numerous other everyday objects ,2. On th% of sampSuch visible differences between minimal and maximal TCS concentration levels in volunteers living in the same neighborhood strongly suggest that human exposure depends on various factors. In light of previous studies, the degree of human exposure to TCS depends not only on the scale of the use of cosmetics and personal care products and the industrialization of the region where the samples were collected, but also may be connected with many other factors, including diet, profession, type of clothing, medication use, lifestyle and even dental fillings and socioeconomic status ,2,56. InSuch comprehensive studies are also indispensable to clarify one more very important issue related to the presence of TCS in the human hair. Namely, it is about the possibility of bioavailability of TCS from the hair (through hair follicles) into the systemic blood circulation (and then into urine samples) and the associated risk of higher human exposure to this substance. If the degree of this bioavailability was established, then it would allow the updating of human biomonitoring assessment values  for TCS. There are two kinds of HBM values: the HBM I value—at which and below which there is no risk of adverse health effects—and the HBM II value—at which and above which adverse health effects are possible . CurrentThe exact determination of the source of human exposure to TCS in Poland poses considerable difficulties due to the fact that research on the presence of this compound in everyday products, as well as in food or the Polish natural environment, is very limited. All previous studies concerning this issue in Poland are presented in Due to the fact that previous studies on human exposure to TCS in Poland have been performed on urine samples, it is difficult to compare the present results with previous observations. Moreover, between particular studies conducted in Poland on urine samples, clear differences have been visible . This isIt should be pointed out that the previous studies described TCS concentration levels mainly in the urine, and the hair sample analysis for the presence of TCS is a relatively new method. Until recently, only three previous publications have shown TCS in human hair samples . CompariThe third previous publication described TCS in human hair collected in Germany  and was During the present study, statistically significant differences in TCS concentration levels were not found between males and females, although the values of concentration levels in women were higher than those noted in men. Although it is generally believed that a higher concentration of TCS in women may be connected with the fact that they use different types of cosmetics and personal care products containing this substance more often and to a greater extent than men, the previous studies have often reported contradicting results. For example, Allmyr et al.  conducteIntragender differences between TCS concentration in humans are difficult to establish due to a wide range of factors having an impact on exposure to TCS. However, experiments on animals strongly suggest that intragender differences may be connected not only with more frequent use of cosmetics by women, but also with various metabolic processes in males and females. Namely, it has been shown that the absorption of TCS is greater in female than in male mice , and so A comparison between groups of younger (aged 22–35) and older (45–67) volunteers performed in the present study have shown that TCS concentration levels were higher in the second group, but the differences were not statistically significant. In light of previous studies, it is known that samples collected from adult humans have shown higher TCS concentration levels than samples from children in the urine ,64 and hThe analysis of human hair samples to evaluate endocrine disruptor concentration levels, despite its many advantages, has some limitations. One of them is the possible influence of hair coloring on the results. During the preparation of samples for analysis, a special emphasis is placed on removing any contamination from the hair surface. However, it is commonly known that hair dyes, which often contain oxidative substances, impact the hair structure, which may lead to a change in structural integrity and therefore easier penetration of substances from the environment into the hair ,67,68. PAnother limitation connected with hair sample analysis is that the chemicals penetrate the hair in two ways: internally from blood to the hair bulb and externally directly from the environment to the inside of the hair. During analysis, the total concentration level in the hair is determined and it is impossible to separate the internal and external penetration. Moreover, it should be remembered that substances accumulate in the hair and are in the hair until the hair falls out, contrary to blood serum and urine in which the levels of substances may change in quick succession. Therefore, hair analysis is a method suitable for studies on long-term exposure to environmental pollutants, whilst investigations on urine and blood samples are more appropriate for the evaluation of short-term changes in exposure.Of course, this study also has some other limitations and did not answer many questions related to the investigation of human exposure in Poland to TCS by hair sample analysis. For example, due to the relatively small number of volunteers and the fact that all volunteers included in the study did not declare chronic diseases, it is difficult to determine if TCS exposure levels found in this study constitutes a risk factor for human health. On the other hand, in light of numerous previous studies ,30,31,37To sum up, the present study has shown that humans in Poland are exposed to TCS to a significant degree and that hair analysis may be used to bio-monitor this substance in humans. Large differences in TCS concentration levels between particular volunteers may suggest that exposure to this substance depends on various factors. Statistically significant correlations between TCS concentration levels and gender, age and/or hair coloring have not been observed. However, it cannot be clearly ruled out that hair coloring does not affect the concentration levels of TCS in the hair, which may limit the use of this method to some extent. To explain all aspects connected with the use of hair sample analysis to bio-monitor TCS in humans, further comprehensive studies on a larger population are necessary.


An expert review panel (ERP) reviewed and updated the PhenX Toolkit Speech and Hearing domain to improve the precision and consistency of speech, language, and hearing disorder phenotypes. A three‐member ERP convened in August 2018 to review the measurement protocols in the PhenX Speech and Hearing domain. Aided by three additional experts in voice assessment, vertigo, and stuttering, the ERP updated the 28 protocols to reflect the latest science and technology. ERP recommendations include six new protocols, five updated protocols (from the same source), and one retired protocol. New additions include two voice‐related, three hearing‐related, and two speech‐related protocols. Additions reflect new phone/tablet applications for hearing and language, and clinical evaluations of voice. “Language” was added to the domain name, which is now “Speech, Language, and Hearing,” to represent language‐related protocols. These protocols can facilitate the assessment of speech, language, and hearing in clinical and population research. Common data elements  used by geneticists, otolaryngologists, audiologists, speech‐language pathologists, and in other disciplines can lead to cross‐study data integration and increased statistical power when studies are combined.The PhenX Toolkit ( PhenX, which began in 2006, is funded by the National Human Genome Research Institute (NHGRI), with additional funding from other National Institutes of Health (NIH) Institutes and Centers. During the earlier PhenX process, subject‐matter experts achieved consensus on well‐established measurement protocols that are made available to the public via the PhenX Toolkit (2https://www.phenxtoolkit.org/domains/view/200000). To address voice, vertigo, and stuttering, the ERP engaged additional outside experts for recommendations. Members of the ERP presented their assessments and recommendations to the group during six webinars. The ERP solicited feedback on their recommendation via email outreach to more than 2600 PhenX Toolkit users and through personalized communications to investigators in the field. Outreach included information on proposed changes to the protocols.The ERP was led by a hearing research expert and comprised of speech, language, and hearing experts  to the database of Genotypes and Phenotypes (dbGaP), which will facilitate study implementation and/or analyses. All PhenX protocols are also available in Research Electronic Data Capture (REDCap).3.1The hearScreen USA phone/tablet application is based on the Digits in Noise (DIN) test, which is a protocol that can be useful in identifying age‐related hearing loss , includes sustained phonation of two vowels (/a/ and /i/), reading of six sentences, and spontaneous speaking. The degree of roughness, breathiness, strain, pitch, loudness, and resonance—in addition to the overall severity of voice aberrance—is rated for each speech task using a visual analog scale. The CAPE‐V is designed to take 5 min and can be scored in real time; no instrumentation is required. The protocol is recommended by the American Speech‐Language‐Hearing Association (ASHA) for clinicians and speech‐language pathologists to identify the presence, severity, and characteristics of voice impairments , a tool to determine how accurately a child or adolescent can repeat nonwords. This screen is suitable for children with or without speech disorders  for adults aged 65 and older was added to the Voice Impairments measure. This protocol includes questions about how older individuals’ voice affects their daily activities . This protocol is an interviewer‐administered test for ages 5−70+ years, in which the respondent reads a short passage to identify a missing key word . This protocol assesses the respondent's ability to read real words and nonwords accurately and quickly  to identify specific categories of speech impairment  was updated to CELF‐5 2013). These age‐specific protocols assess the respondent's knowledge of grammar (. These a3.4https://www.phenxtoolkit.org/domains/view/200000). The Nonword Repetition measure was replaced with the SRT, which is thought to be a more useful assessment because it is easier to score and can be administered even to children who have speech sound disorders.The Nonword Repetition measure, which is part of the Comprehensive Test of Phonological Processing (CTOPP) and assesses phonological awareness, was “retired” but remains available via an archived protocols tab on the PhenX Toolkit website  and Oral Mechanism/Cranial Nerve Examination for Young Children. To date, no peer‐reviewed literature has been found to fill these measurement voids.Many different disciplines are involved in communications research. What should researchers interested in inherited factors know about these clinical tests and disorders? Conversely, what should clinicians know about inherited factors, and how can this knowledge be leveraged in clinical practice? A recent survey of over 500 audiologists and speech‐language pathologists showed that knowledge of genetics is considered very important, but actual training and competency in genetics is lacking , or the U.S. Government.Deborah Maiese, Tabitha Hendershot, and Carol M. Hamilton provided project guidance and oversight. Michael Phillips and Jennifer Schoden provided project management. CCM, BP, MP, and DM wrote the paper with contributions from all authors.


Fasciola hepatica (liver fluke), is a global disease of farm animals and a neglected disease of humans. Infection arises from the ingestion of resistant metacercariae that contaminate vegetation. Within the intestine, the parasite excysts as an active larvae, the newly excysted juvenile (NEJ), that borrows through the intestinal wall to infect the host and migrates to the liver. NEJ release, tissue penetration and migration are facilitated by enzymes secreted by the parasite, namely, cathepsin B1 (FhCB1), cathepsin B2 (FhCB2), cathepsin B3 (FhCB3) and cathepsin L3 (FhCL3). While our knowledge of these enzymes is growing, we have yet to understand why the parasites require all four of them to invade the host. In this study, we produced functional recombinant forms of these enzymes and demonstrated that they vary greatly in terms of activity, optimal pH and substrate specificity, suggesting that, combined, these enzymes provide the parasite with an efficient digestion system for different host tissues and molecules. We also identified several compounds that inhibited the activity of these enzymes, but did not affect the ability of the larvae to excyst or survive. However, this does not exclude these enzymes as targets for development of drugs or vaccines.Fasciolosis, caused by the worm parasite Fasciola hepatica is a major global disease of livestock and an important neglected helminthiasis of humans. Infection arises when encysted metacercariae are ingested by the mammalian host. Within the intestine, the parasite excysts as a newly excysted juvenile (NEJ) that penetrates the intestinal wall and migrates to the liver. NEJ excystment and tissue penetration are facilitated by the secretion of cysteine peptidases, namely, cathepsin B1 (FhCB1), cathepsin B2 (FhCB2), cathepsin B3 (FhCB3) and cathepsin L3 (FhCL3). While our knowledge of these peptidases is growing, we have yet to understand why multiple enzymes are required for parasite invasion. Here, we produced functional recombinant forms of these four peptidases and compared their physio-biochemical characteristics. Our studies show great variation of their pH optima for activity, substrate specificity and inhibitory profile. Carboxy-dipeptidase activity was exhibited exclusively by FhCB1. Our studies suggest that, combined, these peptidases create a powerful hydrolytic cocktail capable of digesting the various host tissues, cells and macromolecules. Although we found several inhibitors of these enzymes, they did not show potent inhibition of metacercarial excystment or NEJ viability in vitro. However, this does not exclude these peptidases as targets for future drug or vaccine development.Fasciolosis caused by  Fasciola hepatica, or liver fluke, is a food- and water-borne disease of humans and their livestock. The parasite has a global distribution and results in major losses to the agricultural community, conservatively placed at EUR 2.5 billion each year, due to the reduced productivity of infected sheep, cattle, water buffalo and goats  is the concentration of the formed product, [S] is the substrate concentration, Vmax is the maximum rate achieved by the system and KM is the Michaelis–Menten constant, which represents the concentration of substrate at which the reaction has a rate equal to half of Vmax.t] is the enzyme concentration and Kcat is the catalytic constant, representing the measure of substrate molecules converted to product by enzyme per second.To determine the kinetic constants, Ki) against the recombinant FhCBs and FhCL3. All the assays were carried out in 0.1 M C-P buffer with 1 mM DTT and 0.01% Brij® L23, at the pH corresponding to the optimum for each enzyme in a total 200 µL reaction volume. The peptidases (140 nM) were pre-incubated with the inhibitor (1000-15 nM) at room temperature, for 30 min, before starting the reaction by the addition of the substrate that was optimal for each enzyme . Substrate concentrations were equal to KM (shown in Kiapp) values were calculated with GraphPad Prism 5, using the Morrison equation for tight binding inhibition (Equation (3)). For competitive inhibitors, such as those used in this study, Kiapp was fitted to a second equation (Equation (4)) from which Ki can be determined [Commercially available inhibitors of cathepsin B (Ca074 and Ca074-OMe), cathepsin L , cathepsin K  and cathepsin S were purchased from Merck Millipore , human cathepsin B (HsCB) and cathepsin L (HsCL1) (positive controls). The exopeptidase activity was examined using two fluorescent substrates, Abz-Phe-Arg-Ala-Lys(Dnp)-OH and Abz-Phe-Arg-Ala-Lys(Dnp)-NH2, where Abz is 2-aminobenzoyl and Dnp is 2,4-dinitrophenyl . The peptidases were assayed using a similar protocol to that described above for measuring endopeptidase activity. The reactions were carried out in 0.1 M C-P buffer, pH 5.0, with a final substrate concentration of 20 μM. The enzymatic activity of the peptidases was monitored as RFU, in a PolarStar Omega Spectrophotometer, with readings taken at excitation and emission wavelengths of 355 nm and 460 nm, respectively.The exo-carboxypeptidase activity of the FhCBs and FhCL3 was compared with the activity of the F. hepatica metacercariae  were prepared for excystment as previously described by Robinson et al. [2, for 3 h to allow metacercarial excystment. Using a light microscope (25× magnification), the number of excysted NEJ was determined.n et al. . To veri2 at 37 °C for 24 h. After incubation, the number of moribund/dead parasites were counted using a light microscope (25× magnification). The criteria used to assess the NEJ viability were loss of motility and peristaltic movement and any obvious damage to the tegument or internal structures, as previously standardized by [NEJ were washed three times in a culturing medium  and their viability assessed by culturing the parasites in 1 mL of culture medium containing 100 μM of the inhibitor at 5% COdized by . The expP. pastoris system, the recombinant forms of FhCB1, FhCB2, FhCB3 and FhCL3 were produced as zymogens that could be purified to homogeneity using affinity chromatography at a yield from 5 to 15 milligrams per litre. The peptidases were resolved in SDS-PAGE, producing bands in the expected molecular weight of ~37 kDa , which exhibited a value at least eight times greater than that obtained with FhCB3 (Kcat/KM of 1708 M−1S−1).The hydrolysis of Z-Phe-Arg-AMC was observed to be the most efficient for FhCB1  and more efficient than that observed for FhCB3 . For both enzymes, Z-Val-Ile-Arg-AMC was the best peptide substrate.The catalytic efficiency of FhCB2 against Z-Gly-Pro-Arg-AMC and Z-Val-Ile-Arg-AMC was similar , designed to span the S2-S2′ subsites of the active peptidases [F. hepatica cathepsin peptidases assessed, only the recombinant FhCB1 cleaved Exo-OH and Exo-NH2; however, this activity was similar to that observed for SmCB1, but both were relatively low compared with the positive human control, HsCB -OH (Exo-OH) and Abz-Phe-Arg-Ala-Lys(Dnp)-NHptidases . Of the ol, HsCB .A panel of 12 commercially available cysteine peptidase inhibitors that preferentially target mammalian cathepsin B (Ca074 and Ca074-OMe), cathepsin L  cathepsin S (S) and cathepsin K  were screened against the FhCBs, FhCL3 and, as controls, HsCB and HsCL1. The broad-range cysteine peptidase inhibitor Z-Phe-Ala-FMK was also included .F. hepatica peptidase displayed a distinct profile against the inhibitors tested. However, it is noteworthy that the epoxysuccinyl cathepsin B inhibitors, Ca074 and Ca074-OMe, which we confirmed to be selective for HsCB compared to HsCL1, were poor inhibitors of all three FhCBs and of FhCL3 (Ki) were relatively high for these inhibitors against all FhCBs (Ki = 345 nM). The compounds KII and KIII abrogated the activity of FhCB3, whilst they were only weak inhibitors of FhCB1 and had no effect against FhCB2.Each of FhCL3 . Our stull FhCBs . HoweverKi = 0.7 nM). In addition, Phe-Ala-FMK, LII and S also strongly inhibited FhCL3 .Compared to the FhCBs, the activity of FhCL3 peptidase was potently abrogated by the inhibitor KII  (The excystment of f 100 μM . In the ≤ 0.005) A.p ≤ 0.001). None of the other inhibitors tested significantly influenced the survival of this stage of the parasite  more than 20 times higher than that of FhCB3, while this substrate was not cleaved by FhCB2. On the other hand, both FhCB2 and FhCB3 preferably cleaved Z-Val-Ile-Arg-AMC over all other substrates examined, whereas this was a poor substrate for FhCB1. Collectively, these data indicate major difference in the substrate specificity of the three F. hepatica cathepsin B peptidases.The S2 subsite of the active site of cathepsin B peptidases are of prime importance for holding substrates in position; these are largely conserved in cathepsin B peptidases . Of partS. mansoni mature cathepsin B1 enzyme numbering in Jílková et al. [Cathepsin B peptidases are uniquely characterised by having a flexible loop structure, termed the occluding loop, which is located at the apical region of the active site cleft and confers these enzymes with unique carboxy-peptidyl-dipeptidase activity. This activity brings about the removal of two amino acid residues from the carboxy-terminus of protein substrates ,35,46,47á et al. ; SupplemF. gigantica cathepsin B5, which possesses both His110 and His111, exhibits exopeptidase activity [F. hepatica cathepsins B.Sequence alignments show that FhCB1, FhCB2 and FhCB3 possess an occluding loop, but, while these retain His110, they each lack His 111 . Absenceactivity . Our stuactivity , were veF. hepatica and S. mansoni cathepsins B were not highly conserved. This region only shares 11–23% sequence identity between the FhCBs, whereas it is generally conserved in cathepsins B from mammalian species [F. hepatica cathepsins B and/or could confer unique, so far undisclosed, exopeptidase activities.Interestingly, we found that the occluding loops of the  species . This unF. hepatica peptidases but this compound also inhibited human cathepsin L and B demonstrating a potent but less specific interaction of this inhibitor with the parasite and host enzymes.A screen of commercially available cathepsin inhibitors revealed that the three FhCBs and FhCL3 exhibited distinct inhibition profiles. While the five cathepsin L inhibitors  showed specificity for HsCL1 over HsCBs, they exhibited high variability against the FhCBs and FhCL3, again emphasising the difference between the parasite enzymes and their mammalian hosts. For example, the epoxysuccinyl peptide inhibitor, L, which did not inhibit FhCB1 or FhCB2, significantly reduced FhCB3 and FhCL3 activity, whereas L1, containing a Phe-Phe peptide, was the most effective at blocking FhCB1 and FhCB3 activity. LII, which contains the dipeptide Phe-Tyr, was the most effective at reducing the enzymatic activity of all The mammalian cathepsin K inhibitors  were generally poor inhibitors of FhCB1 and FhCB2 but reduced FhCB3 and FhCL3 activity. This observation is in agreement with our earlier finding that FhCL3 exhibits cathepsin K-like collagenase activity  and furtKi values in the low nM range, <15 nM. The cathepsin S inhibitor, S, a Phe-Leu dipeptide with a keto-aldehyde (COCHO) group, which has reported Ki values for inhibition of human cathepsins S and B of 0.185 nM and 76 nM, respectively, ref. [Ki values of 14 nM and 35 nM for FhCB1 and FhCL3, respectively).Detailed kinetics studies were performed with six inhibitors and confirmed the poor inhibition of all four parasite enzymes with the inhibitors Ca074 and Ca074-OMe. In general, FhCB1 and FhCL3 were more susceptible to inhibition by cysteine peptidase inhibitors than FhCB2 and FhCB3. Nevertheless, we found that the cathepsin L inhibitor, LII and the broad range inhibitors Z-Phe-Ala-FMK were extremely potent inhibitors of FhCB1 and FhCL3 with ly, ref.  was the Our NEJ excystment experiments revealed that the majority of cysteine protease inhibitors examined at a 100 μM final concentration had little effect on the ability of the fluke to emerge from the cysts. The only compound to show significant inhibition of excystment was Z-Phe-Ala-FMK. Previous studies from our laboratory have shown that a final concentration of 1 mM of this compound completely prevented metacercariae excystment  that theWhen NEJ were cultured for 24 h with each inhibitor, we observed that most of the cysteine peptidase inhibitors did not elicit a significant effect on parasite viability. Our data differ from those of Beckham et al. , who repFasciola infection have shown that NEJ proteinases catalyse the hydrolysis of gelatin [In summary, we undertook a series of experiments on enzyme–substrate/inhibitor kinetics and revealed distinct differences in the specificity of the NEJ FhCB1, FhCB2, FhCB3 and FhCL3. This variation provides evidence for distinct and over-lapping functional roles of the peptidases that would allow the parasite to degrade a multitude of macromolecular substrates. Investigations into the digestion of protein substrates from mammalian hosts susceptible to  gelatin ,55, coll gelatin ,44,56, f gelatin , serum a gelatin , immunog gelatin  and haem gelatin . Collect gelatin . Hence,


Leishmania major (Kinetoplastida: Trypanosomatidae), which is transmitted by the sand fly Phlebotomus papatasi. In Tunisia, the fat sand rat Psammomys obesus and the desert jird Meriones shawi are the principal reservoir hosts of L. major. The presence of the P. papatasi vector of the L. major etiologic agent of ZCL was assessed in the vicinity of villages in endemic areas of Central Tunisia. The study was performed from September through October 2019, a period corresponding to the main peak of activity of P. papatasi. Sand flies were collected from rodent burrows located at the ecotone level, which is the transition zone between the natural environment and human settlement. Sand flies were identified to species level and tested for the presence of L. major by PCR. Our entomological survey showed that P. papatasi is the most abundant sand fly species associated with rodent burrows, and this abundance is even higher in ecotones primarily occupied by P. obesus in comparison to ecotones occupied by M. shawi. Infections with Leishmania major were detected only in P. papatasi, with an overall minimum infection rate (MIR) of 2.64%. No significant difference was observed between the MIRs in ecotones of P. obesus and of M. shawi. Incidence of ZCL in the studied areas ranged from 200 to 700 cases per 100,000 inhabitants, with a mean incidence of 385.41 per 100,000. Higher ZCL incidence was identified in ecotones of M. shawi compared to ecotones of P. obesus. ZCL cases are positively correlated with the MIRs. Considering the short flight range of P. papatasi, increases in its densities associated with burrows of P. obesus or M. shawi at the ecotone level expand the overlap of infected vectors with communities and subsequently increase ZCL incidence. Therefore, control measures should target P. papatasi populations at the ecotones.Zoonotic cutaneous leishmaniasis (ZCL), endemic in Central and Southern Tunisia, is caused by  Leishmania major and is transmitted by infected bites of the sand fly species Phlebotomus papatasi  × 100) . A Wilcop-values were significant at a value of 0.05. Relationships between factors were tested with a Pearson’s correlation. All tests were computed using R v. 3.6.0.All data from clinically confirmed cases during the ZCL epidemiological season starting in July 2019 until June 2020 were obtained from the Regional Health Department of Sidi Bouzid. Numbers of ZCL cases were given by delegation. The ZCL incidence between ecotones was analyzed with a Wilcoxon rank sum test; P. papatasi was the most abundant species (73.29%) followed by Sergentomyia minuta (18.70%), Sergentomyia fallax (5.72%), P. longicuspis (0.55%), Sergentomyia christophersi (0.55%), Sergentomyia deryfussi (0.06%), and Sergentomyia antennata (1.1%) . The abundances of P. papatasi in the ecotones of M. shawi (256) and of P. obesus (806) were significantly different .A total of 885 male and 564 female sand flies were collected from all investigated sites. Overall, a (1.1%) . The abuP. papatasi were screened for Leishmania infection by nested PCR. Eleven pools were found to be infected with Leishmania DNA. Only pools of P. papatasi were infected with Leishmania DNA. Thus, the overall minimum infection rate of P. papatasi with Leishmania DNA was 2.64% (11/417) . MIR medians were not significantly different among delegations .A total of 24 pools of female (11/417) . The meaM. shawi and of P. obesus biotopes were 504.76 ± 123.72 and 266.06 ± 19.93, respectively. ZCL incidence medians differed significantly between ecotones .The incidence of ZCL varied from 200 to 700 cases per 100,000 inhabitants among studied sites . The meap-value = 0.06687, A positive correlation (0.78) between the MIR and the number of clinical cases was observed, with results approaching significance . All L. major sequences clustered together with other L. major strains from different North African and Middle Eastern countries, separately from L. tropica and L. infantum, which appear in two different clusters   . PhylogeL. major are clustered at the margin of some villages where the gerbil reservoir hosts (Meriones species) are widespread [Sand flies were collected in sites located at the edge of rural villages where cultivated plains overlap with fields of chenopods or jujube trees, the natural habitat of the rodent reservoirs forming an ecotone, which is the interface between human settlements and natural ecosystems . Previoudespread . Thus, tdespread . As the P. obesus and M. shawi with L. major varying from 40 to 70% [L. major infection prevalence in P. papatasi associated with rodent burrows and its association with ZCL. In the present study, we reported that P. papatasi is the most abundant sand fly species associated with rodent burrows in all investigated sites. In addition, L. major is the only Leishmania species infecting P. papatasi associated with rodent burrows. Phylogenetic analysis based on a partial region of the ITS-rDNA is in favor of the co-circulation of two different strains, phylogenetically identified as a sub-cluster. Further studies are needed to investigate the epidemiological impact of the circulating L. major strains on the incidence of ZCL. The infection prevalence of P. papatasi with L. major did not differ significantly between the ecotones of P. obesus and of M. shawi. Our results provide further evidence that P. papatasi is indeed the main vector of L. major in ZCL endemic areas.While several studies performed in Tunisia reported high infection rates of 0 to 70% ,7, few sP. papatasi is the main sand fly species associated with rodent burrows in dumpsites and in chenopod field located at the edge of these villages, with an overall minimum infection rate of 5% [P. papatasi is the most abundant sand fly species associated with burrows of the great gerbil Rhombomys opimus [L. major is the most frequent Leishmania species circulating among populations of P. papatasi associated with rodent burrows.Previous study performed by our group in Hichria and Ouled Mhamed from the delegation of Souk Jedid showed that te of 5% . Our ress opimus ,28,29,30P. papatasi associated with burrows of the great gerbil varied from 0.18% [P. papatasi is the most abundant sand fly species (94.4%) associated with burrows of P. obesus [L. major was detected in populations of P. papatasi associated with rodent burrows, with an MIR of 2.5% [P. papatasi was associated with burrows of P. obesus, and the infection rate with L. major was 2% [The infection prevalence of om 0.18%  to 4% [2om 0.18% . Similar. obesus . In addi of 2.5% . In Jordr was 2% .P. papatasi with L. major did not differ significantly between ecotones, ZCL incidence is significantly higher in the M. shawi ecotones compared to those of P. obesus. This finding could be explained by the high infection prevalence of M. shawi with L. major (reaching 53% in autumn) [P. obesus’ sedentary reservoir, living in small colonies as the result of fragmented chenopod fields, maintains a sylvatic cycle of for L. major. Considering that the flight range of P. papatasi is around 0.75 km [L. major-infected P. papatasi in the ecotone of M. shawi expands the overlap of the infected ZCL vector with human habitations and communities, contributing to the emergence of epidemics among naïve human populations.While the minimum infection rates of  autumn) , and by  autumn) . By comp 0.75 km , increasP. papatasi with L. major peaks in September–October, as have been reported in Israel [During the study period, the mean ZCL incidence among delegation was 385.41 ± 77.29. Similar results concerning the period from 1999 to 2004 were reported, showing that the average annual incidence rate of ZCL was 666.7/100,000 inhabitants in the governorate of Sidi Bouzid, and the dynamics of ZCL incidence are significantly heterogenic, occurring in outbreaks and clustering in space and time . Qualitan Israel . A positP. papatasi with P. obesus and M. shawi are intertwined and play critical roles regarding the origin of ZCL emergence in rural communities. Our data suggest that for the P. obesus ecotone, the balance that exists between rodent and sand fly populations allied with the behavior of this rodent possibly creates a veiled buffer zone for disease emergence. Nevertheless, L. major-infected sand flies can still find their way from the burrows of P. obesus and transmit ZCL to humans. In contrast, M. shawi, due to its invasive and its migratory behavior, likely plays a greater role in disseminating ZCL into the peridomicile [Based on our entomological and epidemiological findings, we provided strong evidence that the two clearly distinct ecotones associated with the interaction of domicile .L. major between reservoir hosts and P. papatasi should occur at the ecotone level. Environmental management measures through the destruction of the burrows of P. obesus by deep plowing of chenopod fields have at best led to some arguable but generally not sustainable success [M. shawi, subsequently exacerbating ZCL transmission. The practice of poisoning rodent reservoir hosts has reduced the incidence of ZCL [M. shawi burrows. However, the impact of insecticide-treated baits on ZCL incidence remains to be determined [Breaking the tenuous transmission cycle of  success , and wit success . In addie of ZCL ,36, but termined . From ou


Aeq,8h 85–105 dB) workplace, while 155 women did not experience noise. Age categories were determined according to the World Health Organization . Hearing thresholds were measured at frequencies of 0.125–16 kHz. Higher hearing thresholds were found in the youngest age groups (18–29 and 30–44 years) among those exposed to noise, as compared to those who were not. The difference in hearing thresholds between the exposed and unexposed groups increased with age, as well as with the frequencies. The highest difference in hearing thresholds for these age categories was measured at 11.25 kHz. The oldest age group (45–63 years) exposed to noise showed lower hearing thresholds than the unexposed group at all frequencies from 4 kHz to 16 kHz. High-frequency audiometry can be used for the early detection of increased hearing thresholds at high frequencies. High-frequency audiometry could be included in preventive programs, especially for younger people exposed to noise, in order to enable earlier detection of noise-induced hearing loss.For this study, high-frequency audiometry was used to compare the hearing thresholds, with respect to age, among women exposed to noise in their working environment, as well as those not exposed to such noise. The cohort comprised 243 women (average age 36.2 years), of which 88 women were employed in a noisy (L At present, conventional pure-tone audiometry is one of the most widely used examination methods for diagnosing hearing defects. In pure-tone audiometry, the hearing of subjects is tested in an acoustically soundproofed chamber, using an audiometer that generates clear tones over the frequency range of 0.125 kHz to 8 kHz, according to current legislative standards. However, the human auditory range is much wider than the frequency range covered by conventional pure-tone audiometry. The human ear is capable of perceiving tones up to 20 kHz. Therefore, high-frequency audiometry is required to examine the hearing threshold from 8 kHz to 20 kHz [High-frequency audiometry has been studied for several decades, but the lack of commercially available equipment and the standardization of calibration recommendations have long limited its use ,4,5. At The presumed age-related hearing loss at frequencies ranging from 0.125 kHz to 8 kHz can be calculated using the valid ISO standard: “Acoustics—Statistical distribution of hearing thresholds as a function of age” . Thus, tHistorically, more men have worked in noisy work environments than women. Even today, far more men than women work in places with high-noise risk; however, the proportion of women in such occupations is on the rise. This is mainly due to changes in production technology and, more generally, in the labour sector. Men tend to use hearing protective devices (HPD) much longer, such that the use of HPD is often automatic for them; to the contrary, women often do not use HPD, although they generally take more care in terms of their own health. Some studies have even stated that the prevalence of HPD use in women is up to three times lower than in men . The NatAeq,8h reaches or exceeds 85 dB, or an impulse noise whose A-weighted equivalent sound pressure level LAeq,8h is equal to or greater than 85 dB and whose peak sound pressure level C LCpeak reaches or exceeds 140 dB [Aeq,8h 85 dB). Exposures at or above this value are considered hazardous [Current legislation defines a noise level in the working environment as hazardous if a steady or variable noise whose A-weighted equivalent sound pressure level Ls 140 dB . There iazardous .Noise-induced hearing loss (NIHL) is a form of sensorineural hearing loss (SNHL), which initially manifests itself as a loss at 4 kHz. As NIHL progresses, the damage spreads to cover all high frequencies. This is tested for by extending the range of frequencies for tone audiometry measurements to values higher than 8 kHz, mostly between 14 kHz and 16 kHz. Hearing loss is usually bilateral and symmetrical; however, when the exposure is acute, damage to only one ear may also be found. There is also a frequent association between tinnitus and otalgia following noise exposure .The aim of this study is to compare the hearing threshold as a function of age in respondents exposed or unexposed to workplace noise at frequencies from 0.125 kHz to 16 kHz using high-frequency audiometry.The data were collected between 2018 and 2020 in a specialized clinic. The company where the study cohort was employed gave its consent to conduct the study, provided that all applicable ethical standards would be complied with. All respondents completed and signed an informed consent form to be included in the study.Aeq,T ranged from 87.0 dB to 90.2 dB. The average A-weighted equivalent sound pressure level LAeq,T, measured by a specialized laboratory throughout the 8-hour shift (430 min of working time), was 88.7 dB and the measurement uncertainty was 1.6 dB. As it was higher than the hygienic limit of A-weighted equivalent sound pressure level LAeq,T (85 dB), the workplace was considered hazardous in terms of noise. The maximum of peak sound pressure level C LCpeak measured was 110.4 dB. The noise in the workplace was variable. The maximum working time was 2 h, and there was a break after every 2 h, comprising a total of 50 min of breaks per shift. The analysed company produced various types of filters, transmissions, and other components for the automotive industry, and mainly employed women for manual work. The workplace with exposure to noise was an air-conditioned hall, while the filters were manufactured in a separate room. The room had dimensions of 13.2 × 5.9 × 3.2 m, the floor was made of concrete, the ceiling was coffered, and the walls were lined with tiles. Respondents exposed to a noisy working environment were recruited for this study between 2018 and 2020, and their hearing threshold was evaluated. The employer had a long experience with protecting workers from hazards in their working environment. Occupational hygiene inspections were carried out regularly, and the only risk factor in the workplace was noise. Hearing protection was meticulously observed and strictly enforced, in accordance with Czech legislation and the European standard  [The cohort comprised respondents employed in a noisy workplace, where the A-weighted equivalent sound pressure level Larmuffs) . This EuAeq,8h < 80 dB), selected randomly from the database of registrations in the outpatient clinic. The control group was the general population, with exposure to noise in public areas, but without exposure to workplace noise. This group was chosen to fulfil the condition requiring the comparison of average hearing thresholds in respondents exposed and unexposed to workplace noise in individual age categories. The history of noise exposure in public areas and leisure activities  was not investigated in this work. Both groups lived in the same area and came from similar social groups. Exposure to workplace noise in the current and former workplace was monitored in the respondents. To allow for comparison of our results with those from other surveys, the respondents were divided into age categories according to the World Health Organization (WHO) [The relative number of women and men employed in the noisy workplace depended on the designation and the company workload. The control group consisted of respondents aged 18–63 years who worked in a job without noise exposure  .The exclusion criteria were disagreement with inclusion in the study and age outside the range of 18–63 years. Another exclusion criterion was male gender, as the designation in the risk category was associated with female employees. The respondents with differences between right and left ears at frequencies from 0.125 kHz to 4 kHz higher than 10 dB were excluded from the study.Before beginning the examination, the participants were briefed about the process and the principle of the audiometric measurement. Data collection began with taking a brief case history and other data needed for further processing. The respondents with exposition to a noisy working environment were not exposed to noise on the day and the day before the examination. The condition for the respondents before the audiometric examination itself was 24 h of work rest and no noise exposure. This was followed by tympanometry , conventional pure-tone audiometry, and high-frequency audiometry , performed according to the standards EN ISO 8253-1:2010 Acoustics—Audiometric test methods and EN ISO 266:1997 Acoustics—Preferred frequencies ,17. ThesThe respondents were fitted with headphones, through which tones of different intensities and frequencies were played, first into one ear, then into the other ear. The measurement started with the ear that the respondent identified as the one with better hearing. If the respondent did not perceive a subjective difference of hearing between the ears, the measurement began in the left ear. The hearing threshold was measured at 0.125, 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 6, 8, 9, 10, 11.25, 12.5, 14, and 16 kHz. The result was plotted as an audiogram, where the hearing loss was expressed in dB (decibel hearing level dB HL) for each frequency and separately for each ear. The average values of hearing loss for individual frequencies were calculated and the average threshold curves for individual age categories were compiled. They were statistically compared in order to determine the difference in hearing threshold (in dB), as well as its statistical significance.t-test, the Wilcoxon paired test, and the Mann–Whitney test. Statistical significance was analyzed using the Stata version 13 software  and the online program Openepi version 3.01 . The significance level for testing was set at 5%.The results recorded in the audiogram were exported to Microsoft Office Excel 2017  for calculation of statistics, following which tables and graphs were drawn. Statistical comparison of data was performed using basic descriptive statistics, the chi-square test, the Student’s p < 0.001). The group was then divided into WHO age categories: 18–29 years, 30–44 years, 45–59 years, and 60–74 years. The age category of 60–74 years was merged with the 45–59 category, as the number in this category was too low; thus, the oldest age category was 45–63 years. This category (45–63 years) was the least represented, with a total of 61 women. The second most represented age category was 30–44 years with 87 women, while the most numerous category was the youngest (18–29 years), with a total of 95 individuals (the sizes of groups with and without noise exposure in the workplace are given in p = 0.0004). The average working time at risk of noise was 5.69 years . The youngest age category (18–29 years) worked in a noisy workplace for an average of 3.15 years . The second age category (30–44 years) worked in a noisy workplace for an average of 5.08 years , while the oldest age category (45–63 years) worked in a noisy workplace for an average of 8.26 years . The group was then divided, for further analysis, into categories according to noise exposure: exposure less than 5 years , exposure 5–10 years , and exposure more than 10 years .A total of 88 respondents exposed to a noisy working environment were recruited over the period from 2018 to 2020. The exposed group comprised a total of 88 women employed in the noisy working environment, while the control group included 155 women without exposure to workplace noise. A total of 243 women were examined. The average age in the whole group was 36.2 years . Respondents not exposed to workplace noise were more than 5 years younger, on average , compared to those in the exposed group . This age difference between the exposed and unexposed groups of women was statistically significant . Nevertheless, the average losses at frequencies of 0.125–16 kHz did not exceed 20 dB in both the exposed and unexposed groups. For exposed respondents, the highest average loss was 20 dB at 16 kHz. The difference in the calculated average values of the hearing threshold in this age category ranged from 0 dB to 6 dB. The largest difference (6 dB) was at 11.25 kHz, where the average hearing loss was calculated to be 11 dB for the exposed group and 5 dB for the unexposed group. A statistically significant difference was found between the average hearing thresholds at 0.75 kHz, 6 kHz, 11.25 kHz, and 12.5 k Hz frequencies for average hearing losses in dB. No statistically significant differences were found for the other frequencies. At the frequencies of 0.5 kHz and 9 kHz, no statistically significant differences were found, but the calculated In the 30–44 years age group, the average hearing losses did not exceed 20 dB in either the exposed or the unexposed group at any measured frequency up to 11.25 kHz. At higher frequencies—above 12.5 kHz—the average threshold curves gradually sloped upward, with higher values being reached for the exposed group. The highest differences (4 dB) between the exposed and unexposed groups were at the frequencies 9–12.5 kHz. Statistically significant differences were found at 2 kHz, 3 kHz, 8 kHz, 9 kHz, 11.25 kHz, and 12.5 kHz. No statistically significant difference at 6 kHz was found, but the calculated value approached 0.05 .p = 0.042). Average values, standard deviations, and minimum and maximum hearing losses for the individual frequencies in the exposed and unexposed groups are given in In the oldest age group (45 to 63 years), the differences in average hearing losses were the highest, ranging from 0 dB to 7 dB. As in the other age groups, the highest differences in average hearing losses between the individual groups were at 11.25 kHz and 14 kHz. The hearing loss at 11.25 kHz was 23 dB for exposed respondents and 30 dB for unexposed respondents, such that the difference in these values was as large as 7 dB. The same difference of average hearing loss was observed at 14 kHz: 50 dB for exposed respondents and 57 dB for unexposed respondents. Statistically significant differences were found between the average hearing loss of exposed and unexposed respondents at 1.5 kHz  and unexposed group are given in hed 0.05 . Figure High-frequency audiometry is used mainly for the early detection of hearing loss. For instance, it is used to test people working in a noisy environment, undergoing treatment with ototoxic drugs, or when examining the hearing threshold in patients with tinnitus, in whom a hearing impairment has not been detected by conventional pure-tone audiometry. It serves mainly for the early detection of hearing impairment, manifested as lowered hearing threshold at high frequencies. The hearing loss becomes first apparent at high frequencies, before it spreads to frequencies essential for communication . The method for high-frequency audiometric examination does not differ from that used in conventional pure-tone audiometry. For example, in treatments using ototoxic drugs, high frequencies are the first to become damaged. When monitoring hearing threshold damage with ototoxic drugs, the hearing thresholds are measured for each person separately and compared individually with an initial hearing threshold. Therefore, this method can be employed successfully, even though normative hearing thresholds have not been specified ,18,19.p-values approached the statistically significant value of 0.05 . Furthermore, in this study, we compared the hearing threshold of unexposed persons with persons exposed to noise at work for less than 5 years, 5–10 years, and more than 10 years. Statistically significant differences were not found between the average hearing threshold of those exposed for less than 5 years and unexposed persons only at the frequencies 0.125 kHz, 0.25 kHz, 3 kHz, 4 kHz, 14 kHz, and 16 kHz. In the group exposed to noise for 5–10 years, statistically significant hearing thresholds were found at the frequencies 0.75 kHz, 3 kHz, 6 kHz, 8 kHz, 9 kHz, 11.25 kHz, and 12.5 kHz; while those for the group exposed to workplace noise for more than 10 years were observed at the frequencies from 0.75 kHz to 3 kHz and 9 kHz to 12.5 kHz. At a few frequencies, statistically significant differences were not found, but the calculated p-values approached 0.05 .In our study, the average hearing thresholds from 0.125 kHz to 16 kHz were compared, depending on age and length of exposure to noise at work (number of years at risk of noise at work) for women exposed and unexposed to workplace noise. Statistically significant differences were found between the average hearing thresholds of exposed and unexposed respondents at the frequencies of 0.75 kHz, 6 kHz, 11.25 kHz, and 12.5 kHz in the 18–29 years age group; at 2 kHz, 3 kHz, 8 kHz, 9 kHz, 11.25 kHz, and 12.5 kHz in the 30–44 years age group; and at 1.5 kHz in the 45–63 years age group. No statistically significant difference was found for a few frequencies, but the corresponding calculated The standard EN ISO 1999: 2013 provides procedures for estimating the hearing loss due to noise exposure of populations free from auditory impairment, other than that due to noise , or of unscreened populations whose hearing capability has been measured or estimated. Persons regularly exposed to noise can develop hearing loss of varying severity. This International Standard can be applied to calculate the risk of sustaining hearing loss due to regular occupational noise exposure, or due to any daily repeated noise exposure. Consequently, this International Standard does not stipulate a specific formula for assessment of the risk of impairment, but specifies uniform methods for the prediction of hearing loss, which can be used for the assessment of impairment, according to the formula desired or stipulated in a specific country. As noise-induced hearing loss is the result not only of occupational noise exposure but also of the total noise exposure of the population, it may be important to take the non-occupational exposure of individuals  into account. Only if this non-occupational exposure is negligible—compared with the occupational exposure—does this International Standard allow for prediction of the occurrence of hearing loss due to occupational noise exposure. Otherwise, it should be used to calculate the hearing loss to be expected from the combined  total daily noise exposure. The contribution of the occupational noise exposure to the total hearing loss can then be estimated, if desired. All sound pressure levels stated in this International Standard do not consider the effect of hearing protectors, which would reduce the effective exposure levels and modify the spectrum at the ear. In our study, both employers and workers followed the rules for hearing protection from noise in the workplace. Apart from noise and age, there were no other risks affecting their hearing threshold in the workplace. Possible noise hearing loss was not evaluated, but the current noise hearing loss in the population exposed to work noise and unexposed population living in the same conditions was evaluated, with respect to high-frequency audiometry [Kovalová et al. monitored and compared the average hearing thresholds of 2698 women exposed and unexposed to workplace noise, of whom 479 were exposed to noise in the work environment and the remaining 2219 women formed a control group without noise exposure. They found no statistically significant differences between the average hearing loss, at all examined frequencies, for women under the age of 44. From the graphs presented in their study, it is clear that the average hearing loss in the 18–29 years and 30–44 years age groups for exposed and unexposed subjects did not exceed 20 dB; thus, they were normograms in both monitored groups [Silva et al. reported statistically significant differences in hearing loss, as measured by conventional tonal audiometry, between young adults and older adults, even among non-hearing-impaired respondents. The respondents in that study were divided into only two age groups: 25–35 years and 45–55 years . The samSilva et al. found that, for the 25–35 years age group, the average hearing loss at high frequencies did not exceed 20 dB; however, in the older age group (45–55 years), the average hearing loss reached up to 50 dB. The highest losses were calculated from measurements performed at 12.5 kHz . These vMaccà et al. compared average hearing losses at frequencies up to 18 kHz, with respect to age and noise exposure. They monitored 113 individuals exposed to noise and 148 people without noise exposure. The results of their study confirmed the dependence of high-frequency hearing thresholds on age, but also found higher high-frequency hearing thresholds in people exposed to noise, compared to the average hearing loss in people without noise exposure. The largest differences in average hearing threshold values between exposed and unexposed groups were at 4 kHz, 6 kHz, and 14 kHz. They also found more pronounced differences between high-frequency hearing thresholds at and above the age of 30. Greater differences in the average hearing loss at high frequencies among the 30–44 and 45–65 years age groups were not found, compared to that in the 18–29 years age group. Differences in the average hearing loss at high frequencies ranged from 1 to 3 dB (in the 18–29 years age group), 0 to 3 dB (in the 30–44 years age group), and 1 to 3 dB (in the 45–65 years age group). Maccà et al. also observed an average working time at risk of noise of 17.62 years . In our study was observed an average working time at risk of noise 5.69 years  .A Scottish study of hearing loss in female weavers in 1965, cited by Kovalová et al. , also toIn this study, a shorter average time of noise exposure was found, but there was an evident trend between time spent exposed to noise risk, increasing age, and increase in the hearing threshold, especially at high frequencies. The threshold of respondents increased with age and exposure to noise in the workplace. All respondents with exposition to workplace noise used HPDs throughout all the time with noise exposure at work. However, the correctness of effective use by the respondents and the wear of HPDs has not been checked by the authors. The wear of HPDs could explain the increase in hearing threshold levels in women with exposition to workplace noise. The employer should check the correct use of the HPDs as rigorously as the noise in the workplace itself.A multivariate analysis by Van Wieringen et al. showed that age was the primary predictor of change in hearing thresholds in the high frequency range (10–18 kHz), followed by noise exposure. This conclusion was also supported by our results, where the highest differences in the average hearing threshold among exposed respondents in the youngest age group (18–29 years) were found at the frequencies of 11.25 kHz and 12.5 kHz (6 dB). The analysis also showed the opposite for the conventional 0.25–8 kHz frequency range, where noise exposure was the primary predictor. The results of this study suggest that high-frequency audiometry could be used as an early indicator of hearing loss due to noise and acoustic trauma, rather than audiometry at the conventional frequency of 4 kHz, especially for younger groups .The effect of age and noise exposure on the hearing threshold at high frequencies has also been investigated by Ahmed et al. in a cross-sectional study with 187 respondents exposed to noise and 52 respondents without noise exposure at work. The hearing thresholds of all monitored subjects were measured at frequencies ranging from 0.25 kHz to 18 kHz [The average hearing loss, in the study by Ahmed et al., increased with age at high frequencies. Exposed subjects had significantly higher hearing thresholds than those not exposed at all the high frequencies tested, where the difference between the two groups was greatest at 14 kHz. Higher hearing thresholds in the unexposed group were found only for the oldest group at frequencies over 4 kHz .Other observed values related to the tendency of the thresholds to deteriorate with increasing frequency in both groups. This finding, presenting a descending configuration of the audiometric curve, can be explained by the fact that the human ear requires more acoustic energy to detect higher frequencies. However, a significant deterioration at 14 and 16 kHz indicates a further effect on cochlear function, which significantly increases the hearing thresholds at these frequencies in the tinnitus group . A similRocha et al. concluded that noise in the work environment interferes with high frequency thresholds. All mean hearing threshold values found in the experimental group exposed to noise were higher than in the control group . They suggested that these data reinforce the importance of high-frequency studies, even in conventional pure-tone audiometry, for the early detection of noise-induced hearing loss. A group of 92 firefighters aged 30–49 years were included in this study. Although in this case they were all men, the results correlated with the results of other studies . TherefoIn a study by Garcia et al., hearing loss was observed at higher frequencies compared to conventional pure-tone audiometry, with the range from 9 kHz to 11.25 kHz being the most affected. These results indicate that high-frequency audiometry could be useful in the early diagnosis of acoustic trauma in those who have not yet been affected in conventionally investigated frequencies . These cIn some cases, high-frequency audiometry can be used to distinguish between hearing loss caused by noise and presbycusis. In presbycusis, the auditory threshold progressively increases with the higher frequency range. With noise-induced hearing loss, the hearing threshold usually increases first in the 4–6 kHz range, while there is a marked improvement at 10, 12.5, or 14 kHz [Sheppard et al. have stated that the prescribed regulations for the protection of hearing from adverse noise in the work environment have not been appropriately designed to protect the entire population working in noise. Rather, they are designed to protect a substantial portion of professional noise-induced hearing loss while maintaining a feasible cost–benefit ratio. There is also a growing awareness in the population that noise exposure, although mild, can contribute to a number of other auditory and non-auditory consequences, which may not be identified in hearing protection monitoring programs . The devThe standardization of normal hearing losses at high frequencies, considering the results of a number of studies published so far, remains controversial. It is, therefore, necessary that studies are performed using the same methodology so better data comparison and standard thresholds can be derived. Given that many of these are variable , the methodology needs to be the same so that a reliable comparison of studies can be made .At present, conventional pure-tone audiometry is widely employed to measure hearing thresholds. This method measures hearing loss at frequencies from 0.125 kHz to 8 kHz, even though the human auditory field is much wider—ranging from 0.016 kHz to 20 kHz. High-frequency audiometry is used to measure hearing loss at frequencies from 8 kHz to 20 kHz. As there exist no high-frequency hearing loss standards, high frequency audiometry is not currently used to diagnose hearing impairment, and is mainly used only for the early detection of increased hearing thresholds at high frequencies.In this work, the average hearing thresholds from 0.125 kHz to 16 kHz were compared between female respondents exposed and unexposed to workplace noise. Hearing loss was higher in exposed respondents compared to the unexposed respondents in the younger age groups (18–44 years). This comparison was made for each age group separately, as the hearing threshold at high frequencies increases with age. This should be taken into account in the future when setting normative values, which should, therefore, be set separately for each age group. The results of this work also confirmed that the hearing threshold increases, depending on the duration of noise exposure.In the future, it would be desirable to expand the possibilities of using high-frequency audiometry by setting normative standards. In the meantime, when monitoring noise damage, it is necessary to measure the auditory thresholds for each person separately and compare them individually with the initial hearing threshold. Hearing loss due to workplace noise can be prevented relatively simply. The health and safety of employees in the workplace is regulated by legislation, and observance of these regulations, their enforcement, and early diagnosis are necessary and can be highly effective. Especially with the increasing age of workers, early detection of and compensation for any hearing loss is critical. One of the possibilities could be to employ high-frequency audiometry for preventive examinations in people exposed to noise in order to enable the early detection of damage or hearing loss as a result of such noise exposure.


TCOF1, POLR1D, POLR1C and POLR1B genes. Genetically, the TCOF1 gene contains 27 exons which encodes the Treacle protein. In TCOF1, over 200 pathogenic variants have been identified, of which most are deletions leading to a frame-shift, that result in the formation of a termination codon. In the presented article, we review the genetics and phenotype of TCS as well as the management and surgical procedures utilized for treatment.Treacher Collins syndrome (TCS) is associated with abnormal differentiation of the first and second pharyngeal arches, occurring during fetal development. Features of TCS include microtia with conductive hearing loss, slanting palpebral fissures with possibly coloboma of the lateral part of lower eyelids, midface hypoplasia, micrognathia as well as sporadically cleft palate and choanal atresia or stenosis. TCS occurs in the general population at a frequency of 1 in 50,000 live births. Four subtypes of Treacher Collins syndrome exist. TCS can be caused by pathogenic variants in the  Treacher Collins syndrome , also referred to as Franceschetti syndrome or mandibulofacial dysostosis (MFD1), is a rare developmental disorder. The disorder has been described by Berry  and FranTCOF1 gene; Treacher Collins syndromes 2: TCS2 is caused by pathogenic variants in the POLR1D gene; and Treacher Collins syndromes 3: TCS3 caused by pathogenic variants in the POLR1C gene and lately identified POLR1B gene as a new gene responsible for a novel Treacher Collins syndromes 4: TCS4 [TCS syndrome is genetically heterogeneous . The lit 4: TCS4  /protein kinase C (PKC) phosphorylation sites . TreacleTCOF1 coding treacle can cause apoptosis of neural crest cells during embryogenesis by influence on crucial apoptotic regulators [Treacle is involved in the transport of proteins and ribosomal subunits between the nucleolus and cytoplasm . This prgulators .Loss of treacle protein function in neuroblastoma cells affects the expression of genes that are involved in proliferation, apoptosis and cell cycle.TCOF1 complexes. They have also discovered TCOF1 as a DDR (DNA damage response) factor, that recruits NBS1 to nucleoli after DNA damage. DNA damage due to oxidative stress occurs during embryonic development [Ciccia et al.  identifielopment .POLR1C and POLR1D genes are involved in TCS syndrome by detecting deletion and pathogenic variants in the POLR1D and POLR1C genes in patients with TCS [POLR1C and POLR1D genes were discovered more than 10 years after the discovery of the TCOF1 gene. The POLR1D gene is encoded by 3 exons, POLR1C by 9 exons, while the POLR1B gene consists of 15 exons. The structure of TCS-associated genes, their exons, and differences in length are presented  in TCS patients, while a report by Sanchez et al. [POLR1B.Dauwerse et al.  detectedz et al.  identifiIn the case of Treacher Collins syndrome patients, no correlation is observed between the type of the pathogenic variants and the phenotype.external and middle ear abnormalities including microtia with conductive hearing loss attributed most commonly to malformation of the ossicles,bilateral and symmetric downslanting palpebral fissures,coloboma or notching of the lateral part of lower eyelids with medial absence or sparse of the eyelashes and tear ducts defect,hypoplasia of the facial bones with micrognathia and retrognathia as well as zygomatic bones. Hypoplasia of the zygomatic bones and mandible can cause significant feeding and respiratory difficulties.Symptoms of TCS are clinically very variable and include following mean malformations/features:Other common manifestations include cleft palate, unilateral or bilateral choanal stenosis or atresia and pharyngeal hypoplasia. Most patients with TCS have unaffected intellect, and TCS occurs in both genders equally .The phenotypic features of TCS are observed bilaterally, and disorder intensity is determined by skeletal deformities. The lateral and lower orbital walls are shorter due to hypoplasia of the zygomatic bone, there is narrowing and flattening of the bony face, as well as displacement of the outer corner of the eye and symmetric downslanting palpebral fissures . A forwaIn 60% of children is noticed a poor quality of speech. Speech difficulties have resulted from hearing loss problems and speech acquisition and are associated with structural defects of the ears. Researchers noted that speech therapy may be more effective after structural normalization; however, it is also recommended before such normalization. Patients with TCS may use speech through expressions such as gestures, facial expressions, and vocalizations, which the speech pathologist should consider to contribute to social interactions. Children who have tracheostomies require special attention from speech pathologists.SF3B4 gene, which is located on chromosome 1q21.2. Patients with Nager syndrome are characterized by the following phenotype: downslanting palpebral fissures, hypoplasia of zygomatic bone, micrognathia, cleft lower eyelid, dysplastic ears associated with hearing loss, combined with predominantly radial limb defects such as lateral absence or hypoplastic of thumbs and hypoplasia of the radius with radioulnar synostosis. Miller syndrome is an autosomal recessive disorder caused by mutations in the DHODH gene, which is located on chromosome 16q22.2. Patients with Miller syndrome are characterized by malar hypoplasia, micrognathia, microstomia and a cleft lip and/or palate. The limb defects in Miller syndrome are distinctive, consisting of an absence or incomplete development of the 5th digital ray of all four limbs, and, frequently, forearm abnormalities . Pathogenic variants in the EFTUD2 gene are responsible for mandibulofacial dysostosis with microcephaly (MFDM), characterized by the following phenotypic features: microcephaly, asymmetry of facial features with unusual ears with preauricular tags, sensorineural deafness, a small jaw and major defects including choanal atresia, cleft palate, cardiac defects and even esophageal atresia as well as significant developmental delay and speech delay.Clinical diagnosis of patients with TCS may be difficult as they possess features similar to other disorders, such as oculoauriculovertebral spectrum (known as Goldenhar syndrome), Miller and Nager syndrome, as well as mandibulofacial dysostosis with microcephaly. These disorders are caused by abnormal development of the first and second branchial arches of embryonic development . In the Patients with TCS need specialized, multidisciplinary treatment involving pediatrics, clinical genetics, otolaryngologist, orthodontist, audiologist, psychologist and having major impact variable types of surgeons. Of significant importance is (computed tomography) CT scanning, used to document the anatomy of the head in order to prepare a plan for surgery, to recognize defects of hearing organs as well as to document progress in different stages of treatment . Neuro-oBirth to age 2 years—facilitation of elementary living functions: breathing, e.g., tracheostomy , feedingAge 3 to 12 years—speech therapy, integration into society, bone reconstructions including mandibular , which cAge 13 to 18—orthognathic therapies, maxillomandibular and nasal reconstruction as well as integration into society. Multistage reconstructive treatment forming face is possible in this age and makes noticeable improvements . HoweverTreatment can be divided into three main phases depending on the age of the patient.Treatment must be started as early as the first year of life in cases where there is a life-threatening risk, e.g., nocturnal apnea in children with underdevelopment of the mandible and hypertrophy of the tongue. In the most severe cases, this can lead to suffocation (apnea). In these cases, a mandibular distraction is most effective. The correction of macrostomia or cleft lip surgery is generally performed in infancy. Similarly, due to the need to protect the eyeball, coloboma surgery is generally carried out in the first year of life (even in the first months) if indicated.To determine the severity of mandibular underdevelopment, the Kaban classification is used (types I–III). For patients with TCS, surgical treatment is critical and often begins in the first year of life and continues until growth is complete. The shortening of the mandible and falling back of the tongue results in upper airway obstruction. If macrostomia is present, surgery is needed before the child is one year of age. The lip is sewn at the age of 6 months and the cleft palate between the first and second years of life .The cleft upper eyelid (coloboma) should be operated on before the end of the first year of life. This is necessary to create mechanical eye protection and to ensure proper corneal hydration. Misalignment of the eyelid crevices is corrected by lateral canthopexy, and eyelid defects and coloboma are corrected with local plasty eyelid flaps.Another phenotypic feature seen in children with TCS is symmetric downslanting palpebral fissures, surgery for which begins at preschool age. The procedure involves displacement of a long dermal-musculocutaneous flap from the upper eyelid to the lower eyelid and canthopexy . In caseOrbital cavity wall deformations are operated simultaneously with proper eyelid positioning. To improve facial appearance due to wide and long nose structure, horizontal chin osteotomy and rhinoplasty can be performed . OrbitalMolecular diagnosis plays a very significant role for patients with Treacher Collins syndrome in the prenatal and postnatal periods. Genetic counselling is very important for every family with a TCS child. Each patient with TCS is unique and treatment should be developed by a multidisciplinary team. As there are no clinical differences between the four subtypes of Treacher Collins syndrome, the surgical treatment also does not differ according to the type of pathogenic variants in the genes described. Apart from age, it is the appearance, location and severity of the deformity that determines treatment priorities.The complete etiology of TCS is not yet fully understood. In about 10% of cases, it remains unknown what implies the disorder. This may indicate a role of other genes or other genetic mechanisms in the pathogenesis of TCS .


Background: Alzheimer’s disease (AD) is the most prevalent cause of dementia which affects a growing number of people worldwide. Early identification of people at risk to develop AD should be prioritized. Hearing loss is considered an independent potentially modifiable risk factor for accelerated cognitive decline and dementia in older adults. The main outcome of interest of this review is the alteration of Cortical Auditory Evoked Potential (CAEP) morphology in an AD or mild cognitive impairment (MCI) population with and without hearing loss.Methods: Two investigators independently and systematically searched publications regarding auditory processing on a cortical level in people with cognitive impairment (MCI or AD) with and without hearing loss. Only articles which mentioned at least one auditory elicited event-related potential (ERP) component and that were written in English or Dutch were included. Animal studies were excluded. No restrictions were imposed regarding publication date. The reference list of potential sources were screened for additional articles.Results: This systematic review found no eligible articles that met all inclusion criteria. Therefore, no results were included, resulting in an empty systematic review.Conclusion: In general, dysfunction – being either from cognitive or auditory origin – reduces CAEP amplitudes and prolongs latencies. Therefore, CAEPs may be a prognostic indicator in the early stages of cognitive decline. However, it remains unclear which CAEP component alteration is due to cognitive impairment, and which is due to hearing loss (or even both). In addition, vestibular dysfunction – associated with hearing loss, cognitive impairment and AD – may also alter CAEP responses. Further CAEP studies are warranted, integrating cognitive, hearing, and vestibular evaluations. As the world’s population increases in age, a growing number of people are confronted with cognitive impairment and dementia. The most common cause of dementia is Alzheimer’s disease (AD) which, according to the World Health Organization, accounts for up to seventy percent of all dementia cases .When people experience a greater-than-expected cognitive decline but are still able to perform their activities of daily life autonomously, the concept mild cognitive impairment  atrophy on brain Magnetic Resonance Imaging (MRI) scan, brain fluorodeoxyglucose (FDG) positron emission tomography (PET) (FDG-PET), cerebrospinal fluid (CSF) biomarkers and amyloid PET. With recent advances in AD biomarkers, earlier and more accurate identification of individuals who are in a prodromal stage of AD has become possible. Early identification of AD is important as therapeutic interventions work best if started at the early stages of cognitive decline .Regarding the early detection of cognitive decline and the evaluation of conversion from MCI to AD, other parameters may be of added value . One proAssessment of electroencephalography-derived CAEPs provide insight into the extent to which hearing function is preserved on a cortical level. However, CAEP morphology is modulated by dysfunction such as hearing loss and/or cognitive impairment. Hearing loss, affecting a third of the older population, is a potentially modifiable risk factor for dementia . Both heSince hearing loss is a potentially modifiable risk factor of cognitive decline within AD, and hearing loss and AD both alter CAEP morphology in a similar way, how are CAEPs affected in people with cognitive impairment (MCI or AD) with or without hearing loss?The Preferred Reporting Items for Systematic Review and Meta-Analysis (PRISMA) guidelines were adhered for conducting and reporting this systematic review . The proParticipants: Older adults with diagnosed MCI or AD will have to be subdivided in a group with and a group without hearing loss. If possible, but not mandatory, these groups will have to be compared with older adults with preserved cognition. Animal studies will be excluded.Concept: How are CAEPs affected in different populations: in particular in people with MCI or AD. Therefore, only articles which mention at least one auditory elicited ERP component will be included. Since hearing loss is a recognized potentially modifiable risk factor for increased cognitive decline within AD , the impTypes of sources: This systematic review will consider analytical observational studies including prospective and retrospective cohort studies, case-control studies, and analytical cross-sectional studies. In addition, this systematic review will consider descriptive observational study designs such as descriptive cross-sectional studies. Systematic reviews that meet the inclusion criteria will also be considered.The search strategy was defined and ran in the databases of PubMed, Cochrane, Web of Science, and Scopus. The following search string was used: [ OR (ERP) OR (CAEP)] AND [(Alzheimer’s disease) OR [“mild cognitive impairment”) OR (MCI)] AND (hearing). The search strategy, including all identified keywords and index terms, were adapted for each included database. The reference list of potential sources were screened for additional articles. Studies published in English and Dutch were included. No restrictions were imposed regarding publication date. The date of the last search was 9 August 2021.Following the search, all identified results were collated and uploaded into EndNote X9.2 . Duplicates were removed. Title and abstracts were then screened by two independent reviewers (HG and JB). Potentially relevant sources were retrieved in full and the full text of selected citations were assessed in detail against the inclusion criteria by two independent reviewers (HG and JB). Discrepancies were discussed until consensus was reached. All steps of the screening procedure are presented in A total of 70 records were identified through database searches. There were no records identified through other sources. After duplicates removal, 55 records were identified. Based on title and abstract, 52 records were excluded because they did not include at least one auditory elicited ERP component, and/or they did not include people with cognitive impairment (either MCI or AD), and/or they did not include people with and without hearing loss. Other reasons for exclusion based on title and abstract were protocols, animal studies, and non-English or non-Dutch articles. The full texts of the remaining three articles were obtained, but these records also did not meet all inclusion criteria. One article did not include people with and without hearing loss, and two articles did not include at least one auditory elicited ERP component nor include people with and without hearing loss. Thus, no results were included in this systematic review. A summary of the inclusion process can be observed in Even though no record met all inclusion criteria, they have useful information regarding auditory processing on a cortical level in people with cognitive impairment (MCI or AD), or in people with hearing loss.An electroencephalography-derived (EEG) auditory evoked potential (AEP) is elicited when acoustic stimuli are presented to a patient. This AEP consists of multiple peaks, each with a certain amplitude (μV), representing the strength of the response, and latency (ms), being the time after stimulus onset. The presence, amplitude, and latency of each peak are determined by the intactness and functionality of the auditory pathway, and properties of the acoustic stimulus . Based oHearing loss, gradually raising in prevalence and severity with age, has been recognized as one of the most important potentially modifiable risk factors for dementia . In ordePresent P100-N100-P200 peaks indicate intact sound detection because auditory processing at the auditory cortex is maintained. In contrast, the preconscious MMN and conscious N200 and P300 reflect whether the brain can discriminate between different speech stimuli . The MMNSensorineural hearing loss, hearing loss caused by damage to the inner ear or vestibulocochlear nerve, modulates obligatory P100-N100-P200 responses, resulting in prolonged latencies and reduced amplitudes . McClannIn general, latency measures are more sensitive in detecting changes in hearing function, whereas amplitudes are more dependent on the person’s attention as well as stimulus properties, such as stimulus intensity . This isWhen presenting acoustic stimuli to people with  hearing loss, attention must be paid to the sound level of these stimuli. In general, prolonged latencies and reduced amplitudes are found in subjects with hearing loss in comparison with people with normal hearing when using stimuli at equal sound pressure levels . A similAs dementia alters cognitive processing in the brain, the characteristics of certain higher-order CAEPs, may be affected. In particular, the P300, or P3, component is a positive wave generally occurring around 300 ms post-stimulus. It is elicited by an oddball paradigm, in which rare “target” or “deviant” stimuli are randomly embedded in a train of often-recurring “standard” stimuli. The P300 component is evoked only by deviant stimuli and requires attention towards the stimuli, suggesting a top-down process, as depicted in Thus, both cognitive impairment (MCI and AD) and hearing loss may alter CAEP’s waveform morphology and should be integrated in future research when assessing CAEP measurements. Research that incorporates the role of hearing function on CAEP potentials in determining the risk of developing dementia is currently lacking.As previously stated, hearing loss is a potentially modifiable risk factor for cognitive decline and dementia, and impacts CAEP morphology. However, the other sensory organ in the inner ear, being the vestibular apparatus, may also be associated with cognitive decline and may also alter CAEP morphology.This study aimed to systematically review the literature on how CAEPs are affected in people with cognitive impairment (MCI or AD) with and without hearing loss. Unfortunately, the search resulted in no included studies reporting both the evaluation of hearing and cognitive status in CAEPs, resulting in an empty systematic review. Nevertheless, CAEPs can be a valuable measurement in the assessment of both hearing loss and cognitive decline.Because of their close anatomical relationship, there is a high occurrence of hearing loss in patients with vestibular dysfunction and vice versa. Early in evolution, otolith organs encompassed vestibular function, as well as sound detection. As vertebrates evolved, the cochlea became mainly responsible for sound detection . NeverthVestibular function is important in the reflexive coordination of eye, head, and body movement in a three-dimensional space, voluntary movement and balance, and higher cognitive functions such as navigation and spatial memory. Typical symptoms of vestibular dysfunction include unsteadiness, dizziness, disorientation, and vertigo. Although intact vestibular functioning is crucial for normal movement and overall quality of life, it is one of the most often overlooked sensory systems in EEG research .Besides hearing loss, vestibular dysfunction, affecting one in every five older adults, is also associated with cognitive impairment and AD and may even contribute to its onset . For a rTo the best of our knowledge, only one recent study evaluated CAEP recordings in individuals with peripheral vestibular dysfunction. In conclusion, vestibular dysfunction may alter CAEP’s waveform morphology and is associated with cognitive impairment and AD. However, literature that incorporates vestibular evaluation, CAEP potentials and dementia is currently unavailable.In general, decreased CAEP amplitudes and prolonged latencies indicate malfunctioning, which may involve a peripheral problem such as hearing loss, or a central problem such as cognitive impairment. Hence, people with AD or hearing loss may show a similar pattern of distorted CAEP waveform morphologies. It is therefore important to incorporate hearing assessments when evaluating CAEP measurements in people with AD, and likewise incorporate cognitive assessments when evaluating CAEP measurements in people with hearing loss. That way, the malfunctioning  associated with distorted CAEP waveform morphologies can be assessed more accurately.In addition to integrating hearing and cognitive assessments when evaluating CAEP measurements, it is also important to include vestibular function tests. Not only are the auditory and vestibular end organs anatomically nearby and closely intertwined, there is a high occurrence of vestibular loss in people with hearing loss and vice versa. Additionally, vestibular dysfunction has demonstrated to prolong CAEP latencies, despite not reaching statistical significance . FurtherTo the best of our knowledge, this is the first attempt to systematically review how CAEP morphology is affected in people with cognitive impairment (MCI or AD) with or without hearing loss. The results of our systematic search for literature concerning both the impact of hearing and cognitive status on CAEPs revealed that there are currently no published studies exploring this specific topic. Therefore, this systematic review increases the awareness of an “evidence gap” in this field of research, especially because hearing loss is a potentially modifiable risk factor for an accelerated cognitive decline and modulates CAEP responses.In conclusion, distorted waveform morphologies of CAEPs may serve as (early) prognostic indicators of cognitive decline and disease progression along the AD continuum. CAEPs have the advantage of being non-invasive, objective, and applicable in case of language barrier . Early iThe original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.VVR, HG, and JB conceived and designed the study. HG and JB drafted the manuscript. All authors critically revised the manuscript for important intellectual content, read, and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.