Background and Introduction: Idiopathic sudden sensorineural hearing loss (ISSNHL) is characterized by rapid onset, typically unilateral presentation, and variable recovery. This case-control observational study aimed to improve patient counseling by objectively characterizing long-term hearing loss progression following ISSNHL, using sequential audiometry in the largest-to-date cohort of patients with ISSNHL.Methods: Patients diagnosed with ISSNHL at a tertiary referral hospital from 1994 through 2018 with sequential audiometry were studied. Case controls with sensorineural hearing loss (SNHL) were matched by age, sex, baseline hearing status, and frequency of sequential audiometry. Hearing loss progression was quantified using Kaplan–Meier (K–M) analysis to account for variable follow-up duration. A subgroup analysis was performed by age, sex, preexisting comorbidities, ISSNHL-associated symptoms, ISSNHL treatment, and degree of post-ISSNHL hearing recovery.Results: A total of 660 patients were identified with ISSNHL. In patients with post-ISSNHL recovery to good hearing [pure tone average (PTA) <30 dB and word recognition score (WRS) > 70%], median time to progression to non-serviceable (PTA > 50 dB or WRS <50%) SNHL was 16.4 years. In patients with incomplete post-ISSNHL hearing recovery, contralateral ears were also at significantly higher risk of SNHL progression over the following 12-year period. Male sex was associated with increased risk of SNHL progression [odds ratio (OR) 3.45 male vs. female] at 5-year follow up. No other subgroup factors influenced the likelihood of SNHL progression.Discussion and Conclusion: Patients should be counseled on continued risk to long-term hearing after stabilization of hearing post-ISSNHL, with particular emphasis on greater risk to the contralateral ear in those with incomplete ipsilateral recovery. Idiopathic sudden sensorineural hearing loss (ISSNHL) occurs most often without any obvious triggering cause, reaches maximum deficit within an acute time period, and demonstrates variable recovery . AccordiThe etiology for ISSNHL is poorly understood and likely multifactorial. Previous research has supported a wide range of underlying comorbidities as being positively associated with ISSNHL, such as vascular, metabolic, allergic, autoimmune, and depressive disorders –11. EnviThe studies to date that have looked into recovery after ISSNHL are generally single-center, retrospective analyses performed on small patient populations , 18. WhiNowhere in the current literature exists a study of patients with ISSNHL that combines both a large enough patient population and comprehensive audiometric data to be capable of definitively characterizing patient demographics and comorbidities that influence the hearing loss progression after recovery form ISSNHL. As such, the long-term risk for continued rapid progression of hearing loss in patients post-ISSNHL is not well-understood.In this study, we propose that in patients with partial-to-complete recovery in hearing following ISSNHL, progression of hearing loss in affected ears would occur at a significantly faster rate than would otherwise be expected. A better understanding of hearing loss progression in ISSNHL-affected ears can allow for improved expectations management for these patients, and allow clinical otologists to think more proactively in terms of future possible interventions to improve or restore hearing. The impact of patient demographics, comorbidities, severity of initial hearing loss, and initial ISSNHL treatment strategies can furthermore aid in setting expectations for progression of future hearing loss and its management, and can lead also to improved understanding of disease etiology, potentially informing the development of novel treatments for ISSNHL.The Massachusetts Eye and Ear (MEE) is a tertiary referral center serving New England and adjacent regions in the Northeastern United States. The hospital has maintained comprehensive medical records for all patients during treatment at MEE, such as all audiometric evaluations performed at the facility dating back to 1994. For the purposes of this analysis, we identified patients who had received the diagnosis of ISSNHL per review of ICD9 and ICD10 billing codes from January 1, 1994 to September 30, 2018.Assessment of hearing loss was performed by comparing the affected ear to prior audiometry if available, and otherwise by comparison with the contralateral, unaffected ear, in line with AAO-HNS guidelines . AudiogrPatient charts and audiograms were reviewed to confirm ISSNHL, absence of known underlying etiology for sudden hearing loss, timing of initial diagnosis, treatment provided at time of initial diagnosis, associated symptoms at time of initial diagnosis, and baseline patient comorbidities, such as history of diabetes mellitus (DM), hypertension (HTN), hyperlipidemia (HLD), coronary artery disease (CAD), or cerebrovascular accident (CVA). All patients completed either MRI or auditory brainstem response testing to rule out retrocochlear pathology, and during initial workup did not have other associated symptoms or history to suggest any other than idiopathic etiology. Further audiometric analysis was performed to segment patients by degree of post-ISSNHL recovery as determined by best audiometry within 90 days following initial ISSNHL diagnosis, or next available audiometry if none completed within 90 days.After defining the ISSNHL study population, an age- and sex-matched case control population was selected at 3:1 ratio from the general population at MEE; patients in the case control population were selected by screening the MEE audiometry database for patients with diagnosis code of either SNHL or mixed hearing loss, however without any other known diagnosis of any etiology for sensorineural hearing loss other than presbycusis. When more than three case control candidates were available, priority for inclusion was given to patients for whom frequency of audiometric testing most closely matched their assigned patient with ISSNHL. Hearing in case controls was matched to the study population by both four-tone pure tone average (PTA) and WRS as per the AAO-HNS Hearing Classification guidelines . SpecifiA separate cohort of case controls were additionally matched by three-tone PTA  or word recognition criteria alone, with the remainder of screening for occult otologic disease performed in the same way as for those matched by AAO-HNS criteria. The analyses of hearing loss progression by isolated threshold and word recognition criteria could then be performed as secondary endpoints. As an isolated metric, a three-tone PTA was chosen for threshold analysis because this metric is most closely associated with speech reception thresholds and thus of highest clinical utility –27.Patients with ISSNHL were stratified by baseline audiometry; audiologic measurements included speech audiometry (WRS%) and threshold audiometry (dB HL). WRS was calculated using a standardized word list of monosyllables, measured as a percentage of correctly recognized words after listening to a recorded word list in quiet, typically at 70 dB or at the level at which the patient's speech intelligibility curve plateaus. Pure tone average (PTA) threshold was calculated as a four-tone average of bone conduction thresholds across 500 Hz, 1, 2, and 3 kHz frequencies as the primary threshold-based hearing metric. Good hearing was defined as PTA ≤ 30 dB and WRS ≥ 70%, equivalent to AAO-HNS Class A hearing, with hearing otherwise defined as poor. If a patient's initial audiometry was performed in the setting of sudden SNHL, however subsequent audiometry demonstrated return to good hearing within 90 days, then baseline hearing was considered to be good for purposes of baseline hearing assignment. For purpose of secondary endpoint analysis by isolated threshold and word recognition criteria, normal baseline hearing was defined as WRS ≥ 92% and a PTA threshold of ≤ 25 dB.Progression of audiometric change was assessed both ipsilateral and contralateral to initial ISSNHL event. For patients with good hearing after recovery, the primary assigned endpoint was defined as progression to non-serviceable hearing—either increase in thresholds >50 dB or decline in WRS <50%, equivalent to AAO-HNS Class C or worse hearing. For secondary endpoint analysis, a threshold endpoint was assigned as either moderate hearing loss (PTA > 40 dB) or moderately severe hearing loss (PTA > 55 dB) in patients with baseline normal hearing, per definition by the American Speech-Language-Hearing Association. Patients with baseline mild hearing loss (PTA > 25–40 dB) were also assessed for progression to moderately severe hearing loss. In addition, for secondary endpoint analysis, WRS% endpoint was defined either as WRS <78%  or word recognition <60%  , 28. PatORs) were calculated for patient subgroups based on age, sex, comorbidities, associated symptoms, degree of initial hearing loss, contralateral hearing status, and presence or absence of known steroid treatment at time of initial presentation. Primary endpoint for subgroup analyses was determined to be at 5 years post-ISSNHL to maintain sufficiently large sample size across all groups. Patients with less than 5 years' follow up were not included in this portion of the analysis.Odds ratios  and Excel .Progression of hearing loss over time was evaluated using Kaplan–Meier (K–M) analysis, with change in hearing used to define endpoint. Institutional Review Board (IRB) approval was obtained from the Human Studies Committee at Massachusetts Eye and Ear and Massachusetts General Hospital (IRB 17-120H), Massachusetts, United States. A written informed consent was not necessary given the observational nature of the study and no expected risk to participants from study inclusion.Per review of billing codes, 7,396 patients with ISSNHL were identified. Audiometric review that included requirement for sequential audiometry after initial diagnosis yielded 730 patients age ≥ 18 years with audiometric patterns consistent with ISSNHL as described in Methods. The majority of excluded patients either had historical rather than contemporary diagnosis of sudden hearing loss, or somehow were assigned the diagnosis code despite lack of supporting audiometric data. Further chart review revealed that of these patients, 24 had rapidly progressive SNHL rather than true sudden hearing loss, while 46 had sudden SNHL due to known etiology; all 70 of these patients were excluded. Of those with known etiology for sudden SNHL, 17 were due to acoustic trauma, 13 were due to head trauma, five due to barotrauma, five due to autoimmune inner ear disease , two due to known Meniere's disease, two with recent history of ototoxic drug administration, one due to complications of chronic otitis media, and one with underlying mitochondrial disease. After exclusions noted above, a core population of 660 patients diagnosed with ISSNHL was identified as outlined in Clinical characteristics, demographics, associated otologic symptoms, and initial treatment of patients included in the study are given in In conjunction with onset of sudden hearing loss, the majority of patients reported new onset or worsening of tinnitus (64.2%), while new onset or worsening of either dizziness (23.0%) or vertigo (16.5%) were less common; female patients, however, were more likely to report either dizziness or vertigo compared with males. Among the male population 218 (66.3%) reported episodes of tinnitus vs. 206 (62.2%) in the female group, but this finding was not significant. Treatment for the majority of the patients consisted of isolated oral steroids (53.0%), while both PO and IT steroids (13.0%) and only IT steroids (7.2%) were prescribed less often. While a wide range of protocols for treatment was used, the most common form of oral steroids involved a prednisone burst and taper , or a series of 3–4 intratympanic injections of dexamethasone 10 mg/ml, typically with 0.5–0.8 ml volume per successful injection. Combination oral and IT steroid therapy usually involved initial prednisone treatment followed by salvage IT injections if less than complete recovery was achieved with prednisone alone. An additional 6.9% of the patients received other treatment, such as antiviral drugs, hyperbaric oxygen therapy (HBOT), or antibiotics. No treatment was provided in 6.5% of patients, and in 13.2% of patients, treatment was unable to be determined based on the medical record.p < 0.0001) associated with hearing status in the contralateral ear.p < 0.0001). In these recovered ears, median time to reach endpoint of non-serviceable hearing (either PTA > 50 dB or WRS <50%) was 16.4 years. Average follow-up interval between audiograms was 1.3 years for these patients, and also for selected case controls.Results of K–M survival analysis in p = 0.0035) is driven primarily by events earlier in follow-up. Average follow-up interval between audiograms was 1.3 years for these patients, and also for selected case controls.Further analyses seek to classify patients by using baseline hearing in terms of isolated threshold or word recognition criteria, rather than combined classifications. General trends are similar and demonstrate robustness of findings across a range of hearing loss endpoints, and can be found in the To our knowledge, this study represents the largest-to-date assessment of long-term hearing loss progression in patients with audiometrically-confirmed ISSNHL, and shows that continued progression of hearing loss following ISSNHL is faster than otherwise expected. In patients with incomplete ipsilateral recovery in hearing post-ISSNHL, an increased risk to long-term hearing was seen in contralateral ears as well. Subgroup analyses demonstrated that male sex was associated with significantly greater odds of hearing loss progression at 5 years post-ISSNHL recovery, both in ISSNHL-ipsilateral and contralateral ears. Other patient factors, such as age, comorbidities, drug therapy, and opposite ear hearing, were not significantly associated with odds of hearing loss progression.A minority of patients in our study demonstrated recovery to good hearing post-ISSNHL. Progression of hearing loss was later observed in these recovered ears, with median risk of progression to non-serviceable hearing by 16.4 years. Even contralateral ears, unaffected by initial sudden hearing loss, were shown to be at increased risk of hearing loss progression in patients with incomplete recovery in ISSNHL-ipsilateral hearing. The greater risk of hearing loss progression ipsilaterally may be due to continued presence of systemic risk factors or repeated exposure to the cause of the initial insult, or additionally due to the recovered ear maintaining a certain level of residual “hidden” hearing loss that is not detected by conventional audiometry; in this case, the loss and recovery of hearing with ISSNHL would be considered as similar to the temporary threshold shift seen in acoustic trauma . IncreasRecent studies have demonstrated the role of secreted factors in causing hearing loss in pathologies such as vestibular schwannoma, which can present with sudden SNHL and for which the audiometric hallmark is disproportionate loss in word recognition relative to threshold shift; undiagnosed Meniere's and autoimmune ear disease may also increase risk to bilateral hearing , 30, 31.Post-ISSNHL, all patients should thus be counseled of the persistent risk to hearing, both ipsi- and contralaterally. While the greatest risk seen is the accelerated ipsilateral hearing loss progression for patients with recovery to good hearing, contralateral ears continue to be at risk as well, particularly in those patients with incomplete recovery in ipsilateral hearing.These findings do not demonstrate significance of patient comorbidities or risk factors that have previously been shown to increase risk for sudden hearing loss, such as vasculopathic and cardiovascular risk factors , 11. FigOf note, males are actually over-represented in the patient group with recovery to normal hearing bilaterally—this finding could be explained by a greater degree of reversibility in mechanisms that are predominantly common in the male population, such as the cardiovascular risk factors noted above, while other mechanisms that are relatively more prevalent in the female population are less likely to be reversible . While sNo correlation was found among steroid treatment, route of steroid administration, or degree of initial hearing loss with risk of long-term hearing loss progression, and regardless of hearing loss endpoint evaluated—this may be because treatment choice is often confounded by other factors, such as patient steroid tolerance, and because the broad acceptance of steroids for sudden hearing loss led to very few patients not receiving some form of steroid therapy. Lack of effect seen with regard to initial degree of threshold shift is particularly interesting to note, given that a key component to the AAO-HNS guidelines defining ISSNHL is loss of ≥30 dB across three sequential frequencies, with previous research demonstrating a “floor effect” for improvement in hearing with small thresholds losses , 35. On Regarding relative risk due to symptoms associated with initial hearing loss, no significant associations were found between symptoms and risk of hearing loss progression. This is somewhat surprising given that tinnitus at least has previously been shown to be associated with worse prognosis in sudden hearing loss , 11.Alternate classification of hearing by isolated threshold or word recognition metrics showed that directionality and significance of results were consistent regardless of which metrics were used and how endpoints were defined. The details of these analyses can be found in the Due to the retrospective study design, availability of records was incomplete for some patients, and the interval and duration of follow-up audiometry inconsistent as well—K–M survival analysis was able to account for the effect of variable follow-up by matching average follow-up audiometry intervals between study patients and case controls, however cannot account for differences in total duration of follow-up among the individual patients. In many patients first seen at our institution for sudden hearing loss, no prior audiometry was available to assess baseline hearing, thus necessitating comparison with the contralateral ear to assess degree of initial hearing loss—as a result, it is possible that some cases of bilateral sudden hearing loss were excluded from the analysis. Mass Eye and Ear is a tertiary referral center—many patients whose initial diagnosis of ISSNHL was based on outside audiometry may not have had index audiometric data included in our audiology database, and thus would not have passed audiometric screening criteria to confirm the diagnosis and be included for analysis. Many patients who were referred to Mass Eye and Ear for evaluation likely returned to outside care after consultation for ISSNHL, and this undoubtedly contributed to the high initial screen out rate for patients without sequential audiometry after initial diagnosis. Additionally, certain patient factors, such as lack of steroid treatment, or history of CAD or CVA, were not present at sufficient frequency within the population to assess impact on long-term hearing outcomes with an appropriate level of confidence.Our analysis focuses on hearing loss progression only in ears with good hearing post-ISSNHL, either ipsi- or contralaterally. Progression of hearing loss in ears with incomplete recovery in hearing (either PTA > 30 dB or WRS <70%) was not assessed due to lower clinical relevance, since the vast majority of these patients already did not have useful hearing in the relevant ear.Of particular interest to this analysis is the selection of appropriate case controls—patients with SNHL but no other known sensorineural otologic history and thus presumed presbycusis, as well as a similar interval for follow-up audiometry compared with the study population. A potential selection bias could be present within the case control group if patients with occult otologic history were included, and if this history were not evident based on superficial review of diagnostic codes alone. To minimize this bias, we performed detailed chart review on any case control patients who demonstrated significant change in hearing over time, and replaced those with other otologic histories with “clean” controls for whom non-ISSNHL otologic disease burden was equivalent to the study population, as detailed in the Methods section. This finding reinforces the continued importance of detailed chart review in the context of large database analyses, since over-reliance on the “face value” of historical diagnosis codes can easily overlook key components to the patient history.These findings represent to date the largest study tracking long-term bilateral hearing outcomes in patients with ISSNHL. Our findings definitively demonstrate an increased risk of hearing loss progression in these patients, both in ISSNHL-ipsilateral and contralateral ears. Male sex was the only significant patient factor associated with the increased risk of hearing loss progression, however, this effect was confounded by greater incidence of HTN, HLD, diabetes, and cardiovascular disease in the male patient population. All ISSNHL patients should be counseled on the increased risk of accelerated long-term hearing loss progression, with primary concern for hearing in the ISSNHL-ipsilateral ear in patients with full recovery and additionally for the contralateral ear in patients with incomplete recovery. Additional care should be taken to avoid other otologic insults, such as acoustic trauma or potentially ototoxic medications in these patients, and guidance should be provided for optimal management of modifiable risk factors such as HTN and diabetes.Previous research indicates that secreted factors may play a role in accelerated hearing loss progression in patients with vestibular schwannoma (VS), with potential for long-distance percolation through CSF or blood given effects seen in VS-contralateral ears . Immune Improved understanding of causes and treatments for ISSNHL will require better understanding of disease pathophysiology, which is complicated by the likely multifactorial nature of the disease . We builkstankovic@stanford.edu.The primary dataset includes service dates for certain procedures which are considered protected under HIPAA guidelines, and as such cannot be shared publicly. Datasets can be made available upon request to interested parties. Requests to access these datasets should be directed to This study involving human participants was reviewed and approved by Human Studies Committee at Massachusetts Eye and Ear and Massachusetts General Hospital (IRB 17-120H). Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.KS conceived the project and supervised all the work. SE and KS designed the study. SE and JV collected data from patient charts. SE wrote code for data analysis. Primary analysis and interpretation were then performed by SE and JV, with input from all authors. SE, JV, and KS wrote the manuscript. All authors edited the manuscript and approved the final version.This work was supported by the National Institute on Deafness and Other Communication Disorders (R01DC015824), the Nancy Sayles Day Foundation, the Zwanziger Foundation, the Barnes Foundation, and the Sheldon and Dorothea Buckler (KS).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.


Taken with the growing importance of cathepsin-mediated substrate proteolysis in tumor biology and progression, the focus and emphasis placed on therapeutic design and development is coming into fruition. Underpinning this approach is the invariable progression from the direction of fully characterizing cathepsin protease members and their substrate targets, towards targeting such an interaction with tangible therapeutics. The two groups of such substrates that have gained much attention over the years are the pro- and anti- apoptotic protein intermediates from the extrinsic and intrinsic signaling arms of the apoptosis pathway. As proteins that are central to determining cellular fate, some of them present themselves as very favorable candidates for therapeutic targeting. However, considering that both anti- and pro- apoptotic signaling intermediates have been reported to be downstream substrates for certain activated cathepsin proteases, therapeutic targeting approaches based on greater selectivity do need to be given greater consideration. Herein, we review the relationships shared by the cathepsin proteases and the Bcl-2 homology domain proteins, in the context of how the topical approach of adopting ‘BH3-mimetics’ can be explored further in modulating the relationship between the anti- and pro- apoptotic signaling intermediates from the intrinsic apoptosis pathway and their upstream cathepsin protease regulators. Based on this, we highlight important future considerations for improved therapeutic design. Over the last 10 years, great strides have been taken in identifying cathepsin protease-specific substrates, which have revealed a number of interesting regulatory paradigms based on the growing pleiotropic nature of these enzymes. While they were originally isolated as lysosomal proteases involved in the proteolysis of intracellular proteins, their role in determining the fate of cells as active components arising from the lysosomal ‘suicide bag’ have seen their importance evolve . At the When taken with the ability of cathepsins to reside in the cytoplasm, the potential for them to cleave and modulate a number of biochemically significant signaling pathway intermediates central to determining cell viability has taken on heightened importance. One such process that the cathepsins have been identified to regulate is apoptosis . WhiGenerally, apoptosis is the end-point of two main regulatory pathways . FirstlyThroughout this process, and of central importance from a regulatory standpoint, is the expression of the Bcl-2 family of proteins , the memTherapeutically, both extrinsic and intrinsic pathway intermediates have been the subject of intense scrutiny from a therapeutic targeting standpoint, helped and aided by the elucidation of the crystal structure of many signaling intermediates derived from these pathways . ClearlyMechanistically, the cathepsin proteases have been linked to regulating both of the intrinsic and extrinsic signaling cascades, and which forms the basis of this review article by addressing what recent evidence supports their input into this regulatory step of apoptosis. Herein, we outline the rationale for therapeutically targeting certain cathepsin proteases, in the context of either selectively abrogating their activity for the breakdown (and destabilization of the pro-apoptotic Bcl-2 proteins), or their activity to be selectively maintained for the breakdown of the anti-apoptotic Bcl-2 proteins. Clearly, a favorable endpoint is effectively altering the balance of active pro-apoptotic Bcl-2 proteins levels in relation to their anti-apoptotic counterparts, with a view to tipping the balance of these proteins, in order to favor apoptosis in the context of killing cancer cells and halting tumor progression . FinallyBAX gene frame-shift mutations can contribute to colon [BAX expression can give rise to accelerated mammary tumor development in mice models [BAX-knockout mice studies, the observations of abnormal B- and T-cell numbers and compromised spermatogenesis [Mammalian cells mainly encode three key pro-apoptotic proteins of importance, namely BAK, BAK and BOK, and each of which uniquely and directly induce MOMP based on their subcellular localization ,48,49. Wto colon , lung, ato colon ,58 and te models . From BAogenesis  have beeogenesis . Moreoveogenesis , double-ogenesis ,63,64. Cogenesis ,66, the ogenesis ,68,69. Hogenesis ,71,72.Based on such collectively strong evidence, whether such proteins are amenable to cathepsin-mediated digestion and deactivation has emerged as a key area of investigation, particularly in light of how deregulated cathepsin over-expression may also give cancer cells a survival and proliferative advantage during disease progression . MechaniWhile such BH3-mimetics do hold great therapeutic potential, their effects on cathepsin proteases, as key regulators for the cleavage of Bcl-2 family members and intrinsic pathway activation, remain largely unexplored. Similarly, the broader effects of cathepsin protease-directed inhibitors on the activation status of Bcl-2 family members, also remain poorly understood. This point is clearly highlighted through pepstatin A-mediated inhibition of cathepsin D, which delayed the onset of apoptosis, had the effect of altering the conformation of BAX, and which rendered it active independently of upstream BID cleavage-mediated activation . Being mIn summary, while a number of studies have looked at the intermolecular relationships between the cathepsins and Bcl-2 family of protein intermediates from the intrinsic pathway, one preferential cathepsin that directs BAX cleavage is cathepsin S (in vitro and in intact cells). Additionally, while other cysteine cathepsin proteases -B, -L, -S, and -K have been demonstrated to cleave BAK, no cathepsin proteases have been connected with BOK proteolysis. Moreover, progression towards therapeutically targeting this relationship, in order to spare enough pro-apoptotic monomeric protein for activation-mediated oligomerization, is indeed possible (as seen with the novel inhibitor CS-PEP1), and which offers a strong rationale for pursuing the BAK or BOK proteins as potential cathepsin substrates in a similar manner.−/− mice, abnormal death of lymphocytes had been reported [−/− mice possessed abnormalities in the demise of neurons and erythroid progenitor cells [−/− mouse studies reported the death of early stage embryos and mice with conditional deletions experienced a rapid loss of mature lymphocytes or hematopoietic stem cells [The anti-apoptotic Bcl-2 proteins have also gained increasing importance for therapeutic targeting, from their abilities to drive cancer progression and the most extensive studies of which have described the targeting of Bcl-2, Bcl-xL, and Mcl-1 ,82. Mechreported , while Bor cells . Additioem cells ,94.Based on the above evidence, the consequential role played by the cathepsin proteases during anti-apoptotic protein proteolysis has taken on significant importance. This was addressed by Droga-Mazovec et al. (2008), who reported the cleavage of proteins Bcl-2, Bcl-xL, and Mcl-1 upon the treatment of a variety of cell lines with the lysosomorphic agent, LeuLeuOMe . More spAs very promising candidates for targeted cancer therapy, early studies identifying novel therapeutics targeting the Bcl-2 subfamily of anti-apoptosis proteins involved the screening of natural compounds and which yielded little success . ThroughAs seen from our recent studies outlining an alternative approach for developing a peptide inhibitor to target cathepsin S-specific Bcl-2 family intermediates of the intrinsic pathway, effective peptide efficacy (and specificity) may also be guided by the intracellular pH of cancer cells, as a key-determining factor. In this context, we observed that cathepsin S could cleave Bcl-xL in vitro better at pH 7 than at pH 5 and that this reaction could be inhibited better at pH 7 (than at pH 5), by the novel peptide inhibitor CS-PEP1. Such a pH-sensitive cleavage reaction appeared fortuitously to favor the inhibition of Bcl-xL cleavage (by CS-PEP1) at a relatively higher pH, in relation to the lower pH that can be prevalent during tumor development and apoptosis. Favorably, cathepsin S-mediated cleavage of BAX (and the inhibition of this reaction by CS-PEP1) was observed to be effective at both pH 5 and 7 . CollectFrom the family of multiple BH3-only proteins, the two most characterized members, in the context of being cleaved by cathepsin proteases, are BID and Bim ,115,116.−/− mice failed to release mitochondrial cytochrome c upon their treatment with lysosomal extracts , thus linking lysosomal-derived, cathepsin-mediated cleavage (and the activation) of BID, with MOMP [Functionally, the main role of BID appears to link the death receptor pathway to MOMP ,122, folith MOMP ,128. Morith MOMP . From usith MOMP .In confirming such observations, from our recent studies, purified BID could be cleaved by cathepsin S in vitro or in mammalian cells, and this cleavage effect could be reversed upon the co-expression of BID with catalytically-null cathepsin S . While sc-myc transgenic mice developing B-cell leukemia [In the instance of the Bim protein (Bcl-2 interacting mediator of cell death), its importance in cell death is highlighted through its negative selection of B- and T-cell precursors and regulation of peripheral lymphocyte homeostasis . In suppleukemia ; or, 2) leukemia . The sigleukemia ,134,135.leukemia .As in the case of BAX and BAK, targeting the cathepsin(s) responsible for BH3-only protein degradation may hold good therapeutic value. In this context very little is known about how currently available anti-cathepsin therapeutics affect the activation (or proteolysis) of many other BH3-only proteins, including Bim.While a number of validated therapeutics directed at the Bcl-2 anti-apoptotic proteins have been described herein, further questions do arise to address what effects such therapeutics may have on the activity of upstream modulators, such as the cathepsin proteases. For example, during the inhibition of BAX-Bcl-2 protein binding, are BAX (or Bcl-2) protein molecules presented as good substrates for their cognate cathepsin proteases in the presence of BH3-mimetics? Based on recent developments (reported herein), this could indeed have an impact on the overall net stability of certain Bcl-2 proteins and how well they may fulfill their native roles as key apoptotic regulators.As a novel and alternative ‘semi-rational’ approach for therapeutic development, we have defined a peptide therapeutic based on its ability to potentially disrupt the BAX BH3-Bcl-xL hydrophobic groove interaction, while at the same time taking on properties of an allosteric inhibitor directed at the catalytic activity of cathepsin S. Of relevance, may also be the subcellular compartmentalization shared by the cathepsin and Bcl-2 proteins, and the penetrability of any arising therapeutics. In this context, while targeting the lysosome may be achieved with relative ease, other extra-lysosomal compartments, such as the unidentified and membranous perinuclear compartment in which enriched cathepsin S and BAX-derived proteins have been uniquely seen to co-localize  may present additional challenges . This isWhile one aspect of good therapeutic design directed at the Bcl-2 proteins appears to have taken shape and is yielding tangible effects against tumor progression and cancer in the form of BH3-mimetics , this reFrom our recent findings, one possible novel approach that could fulfill this is through identifying peptide sequences present in the cognate cathepsin protease inhibitor proteins (such as the cystatins), with a view to evaluating their abilities in modulating the pro- and anti-apoptotic protein interactions that the BH3-mimetics are designed to disrupt. Moreover, such therapeutics may also have the potential to interfere negatively with the cathepsin protease activity directed at certain pro-apoptotic protein members from the intrinsic apoptosis pathway. The net effect of such an approach may have greater benefits attached to them as such efforts may yield the development of a new generation of therapeutics that offer greater efficacy and selectivity. With this in mind, the resulting therapeutics may also have greater tolerance to operate at a fluctuating intracellular pH based upon the broad range at which the cysteine cathepsin protease are active, either within the tumor microenvironment or intracellularly in cells that have been chemotherapeutically treated. However, what effects such pH changes may have on cathepsin protease-substrate specificity, based upon substrate protein conformational changes and alterations in their sub-cellular localization, are also factors that need to be given greater consideration while defining the efficacy of any therapeutics arising from this novel approach.


Thedonor/acceptor weight ratio is crucial for photovoltaic performanceof organic solar cells (OSCs). Here, we systematically investigatethe photovoltaic behaviors of PM6:Y6 solar cells with different stoichiometries.It is found that the photovoltaic performance is tolerant to PM6 contentsranging from 10 to 60 wt %. Especially an impressive efficiency over10% has been achieved in dilute donor solar cells with 10 wt % PM6enabled by efficient charge generation, electron/hole transport, slowcharge recombination, and field-insensitive extraction. This raisesthe question about the origin of efficient hole transport in suchdilute donor structure. By investigating hole mobilities of PM6 dilutedin Y6 and insulators, we find that effective hole transport pathwayis mainly through PM6 phase in PM6:Y6 blends despite with low PM6content. The results indicate that a low fraction of polymer donorscombines with near-infrared nonfullerene acceptors could achieve highphotovoltaic performance, which might be a candidate for semitransparentwindows. It has been demonstratedthat efficient charge transport and suppressed recombination lossesare largely related to balanced electron and hole mobility in OSCs.14 Generally, a continuous interpenetrating network of donor and acceptoris considered to be significant for charge transport.15 Therefore, the donor/acceptor (D/A) blend composition isimportant for charge transport and device performance. Typically,in dilute donor heterojunctions where the average distance betweenindividual donor domain enlarges, that may reduce hopping rate ofhole. The unbalanced charge transport can induce a space-charge effect,causing more recombination loss and poor performance.16 Interestingly, some previous studies have reported thatfullerene based OSCs with very low donor contents could maintain efficienthole transport.18 It was proposed that hole tunnelingcould occur between isolated donors,20 another possiblechannel is that fullerene acceptors such as PCBM can act as an ambipolarconductor for both electrons and holes in dilute donor solar cells.22Organic solar cells (OSCs) basedon nonfullerene acceptors (NFAs) have achieved rapid development inrecent years due to the tunable energy levels and absorption spectraof NFAs.Jsc exceeding 25 mA cm–2 with a low voltageloss (∼0.48 eV) can be achieved simultaneously.23 A report with single crystal analysis and molecular simulation hashighlighted the unique packing in Y6 films can lead to strong electroniccoupling between adjacent Y6 molecules and efficient 3-dimensionalambipolar transport networks.24 These interestingphenomena raise an important question about hole transport mechanismsin bulk heterojunctions (BHJs) with this emerging family of NFAs,which could contribute to high device performance. Dilute donor solarcells are interesting models to understand the role of acceptors oncharge generation, transport and extraction. Therefore, it will beinformative to investigate the correlation between photovoltaic performanceand donor/acceptor blend composition in NFA-OSCs.Compared to the clarificationon the exclusive role of fullereneacceptors in dilute donor solar cells, charge generation, transport,and recombination in dilute donor devices with NFAs are rarely investigated.Recently, NFA Y6 with the A–D–A′–D–Aconfiguration has received great attention. In the optimized solarcells based on PM6:Y6 , a high Jsc of 18.5 mAcm–2 and FF of 0.66. The results of femtosecondtransient absorption (TA) spectroscopy indicate efficient charge transferin dilute donor blend films. Moreover, slow charge recombination andalmost field-independent charge extraction in the dilute donor deviceswere observed in dilute donor solar cells. Importantly, it is foundthat PM6:Y6 blends even with 10 wt % PM6 exhibit efficient hole mobilityand balanced charge transport. By comparing the hole mobility of PM6diluted at 10 wt % in Y6 and insulating polymers, we conclude that10 wt % PM6 in PM6:Y6 blends can still retain efficient hole transportpathway through the active layer.In this work,we studied the photovoltaic behaviors of PM6:Y6 solarcells with various PM6 contents. The OSCs with 10 wt % PM6 yield ahigh PCE of 10.3% with J–V curves  are summarized in Table S1. As shownin Jsc of 18.5 mA cm–2 and FF of 0.66. It is worth noting that dilute donor devices showmuch better performance than the devices with high donor contents,especially Jsc and FF. To understand theorigin of high performance of the OSCs with such low donor contents,charge generation, transport, and recombination were investigated.To study the photovoltaicperformance, PM6:Y6 solar cells withvarious PM6 contents were fabricated with a standard structure ofITO/PEDOT:PSS/active layer/PDINO/Al. The photovoltaic parameters extractedfrom V curves S1 are suV curves S1 are suFigure S2). Compared to the PL of pure Y6 films, the largedegree of PL quenching in 10 wt % PM6 BHJ indicates efficient interfacialcharge transfer. Hole transfer dynamics from Y6 to PM6 was studiedwith femtosecond TA spectroscopy. In Figure S3 and Figure S4, a 750 nm pump laser was used to selectivelyexcited Y6. The rising kinetics of PM6 ground state bleach in theblend at ∼605 nm reflects the hole transfer process from Y6to PM6, since photoexcited Y6 shows no TA signal there . The hole transfer process exhibitsa biphasic behavior with an ultrafast interfacial hole transfer processand a relatively slower diffusion mediated process.27 We fit thehole transfer kinetics (Table S2) by abiexponential function1 andτ2 and prefactors of A1 and A2.28 The initial ultrafast charge transfer process (1 (<250 fs) can be attributed to quantum coherence, whichplays a role in the formation of charge carriers29 and was experimentally observed in OSCs.30 The exciton diffusion process characterized by A2 and τ2 becomes more dominantwith the decreasing of PM6 contents. Typically, large amounts of smallmolecules tend to aggregate,25 there wouldbe expected larger domains in 10 wt % PM6 BHJ films. The larger root-mean-square(RMS) of 10 wt % PM6:Y6 films  also indicates strong molecular aggregation, which could lead tooversized domains.31 Our previous workhas shown that excitons will be created away from the interface andthus spend longer time to diffuse to the interfaces because of thelarger domain in BHJ films with low donor contents.25 Similarly, the exciton diffusion mediated hole transferlifetime τ2 in PM6:Y6 blends decreases with the amountof donor, as shown in Table S3. ATHE of 83% in 10 wt % PM6:Y6 blend reveals efficient hole transfer.In addition, the decay signals in Charge transfer between PM6 and Y6 was investigated by measuringsteady-state photoluminescence (PL) spectroscopy S2. CompaFigure S6 and –4 cm2 V–1 s–1 and a hole mobility of 6.8× 10–5 cm2 V–1 s–1 (Table S4), indicatingefficient charge transport in such dilute donor structure. However,the electron mobilities show an obvious decrease when PM6 contentsis over 60 wt %, the unbalanced charge transport results in a decreaseddevice performance (–4 cm2 V–1 s–1) and hole mobility (1.8× 10–4 cm2 V–1 s–1), indicating Y6 is an ambipolar conductorfor electron and hole.To investigate the charge transport, charge carrier mobilitieswere determined from space charge limited current (SCLC) in single-carrierdevices, as shown in Jsc under different lightintensities. As shown in 32Voc dependence on light intensity, the slopes(n) of PM6:Y6 devices with 10 and 45 wt % donor arearound 1.3, indicating that the recombination is primarily via thebimolecular pathways with a minor role of trap-assisted recombination.34 The slope of the dilute acceptor device (90 wt % PM6) is very closeto unity, which suggests that photogenerated carriers are purely lostvia bimolecular recombination. Moreover, transient photovoltage (TPV)measurements were performed on PM6:Y6 solar cells, as shown in To study the charge recombination in device operational conditions,we examined Figure S7); charge extraction time versus bias were obtained fromTPC with a monoexponential decay model. In In the case of extremely unbalanced D/A ratios,a discontinuousinterpenetrating network of donor and acceptor might be an importantissue for charge transport. Also, a reduced contact area of anode/donoror cathode/acceptor could lead to a low charge extraction efficiency.Here we investigated charge extraction processes by using bias-dependenttransient photocurrent (TPC) measurements S7; charg14 Bias-dependent EQE spectra of 10, 45, and 90wt % PM6-based devices were recorded and are shown in Figure S8. The EQE spectra of devices with 10and 45 wt % donor contents are almost insensitive to bias, which isconsistent with the bias-TPC results. This feature can arise fromthe suppressed recombination loss at low internal fields, so thatphotogenerated carriers can be collected more efficiently, yieldinghigh Jsc and FF in the OSCs with 10 wt% PM6.Our previous work demonstrated that charge extraction correlateswith bias dependence of the external quantum efficiency (EQE) of OSCs.19 (ii) Holes can transport directly in donor phase.(iii) Y6 may facilitate the hole transport because it is an ambipolarconductor.Above results demonstrate that the OSCs with 10 wt %PM6 show highand balanced charge transport, which enables slow charge recombinationand fast charge extraction. As for the origin of efficient hole transportin dilute donor devices, we assume following hole transport pathways:(i) There is possibility for holes tunneling between isolated donors.–8 cm2 V–1 s–1. As forthe results shown in Figure S9 and Figure S10), the (100) diffractionpeak for PM6 at 0.32 Å–1 in in-plane was alsoobserved in 10 wt % PM6:Y6 films, which was most probably inducedby the face-oriented Y6 molecules.35 Thisphenomenon reveals that, although in the dilute donor blend, the donorexhibits ordered structure, which can account for high hole mobilityin dilute donor devices.To clarity the hole transport mechanism in PM6:Y6solar cells with10 wt % PM6, we compared the hole mobilities of PM6:Y6 with PM6:PS(polystyrene) with the same PM6 content because PS is an insulatorwith very low hole mobility in the order of 10J–V curvesof hole-only devices based on pristine Y6 exhibit a trap-filling process,but this phenomenon does not exist in PM6:Y6, PM6:PS, and PM6:PMMAblends . In addition,hole transfer from donor to acceptor is generally unfavored, becausethe highest occupied molecular orbital (HOMO) level of Y6 (−5.7eV) is deeper than that of PM6 (−5.48 eV), which was confirmedby cyclic voltammetry (CV) measurements . Therefore, it is plausible that NFA Y6 haslittle contribution to hole transport in the dilute donor structurewith 10 wt % PM6 (iii), even though Y6 has ambipolar transport characteristics.Given the discussions above, in the dilute donor devices, 10% of PM6can form efficient hole transport pathways, enabling well-balancedcharge transport with sufficiently high mobilities.It is obvious that the dark To conclude,the performance of PM6:Y6 solar cells with varyingstoichiometries was comparatively studied. A high PCE exceeding 10%was achieved in the dilute donor devices (10 wt % PM6) synergizedby efficient hole transfer, charge transport, slow charge recombination,and field-insensitive extraction proved with results of TA, TPV, TPC,and bias-dependent EQE. Surprisingly, balanced electron and hole mobilitiesare achieved in the 10 wt % PM6 OSCs evaluated with SCLC in singlecarrier devices. By comparing the hole mobility of 10 wt % PM6 mixedwith those of Y6, PS, or PMMA, we propose that the dominant hole transportin dilute PM6:Y6 blends is via the PM6 phase. Good performance ofdilute donor heterojunctions may open new applications such as fornear-infrared photodetectors or semitransparent photovoltaic panels.


LPAI H9N2 infection among the poultry population in Egypt constitutes an additional risk factor in the poultry industry. This study aimed to determine the prevalence of H9N2 avian influenza virus (AIV) in commercial and backyard chickens in Egypt. A 2-year survey of H9N2 AIV in chickens in farms and backyards was carried out in 2015 and 2016.Direct detection of H9N2 AIV was performed by detecting the virus in tracheal and cloacal swabs using real-time polymerase chain reaction assays. A total of 20,421 samples were collected from chickens in farms and backyards in 26 Egyptian governorates.In 2015, cases positive for H9N2 AIV numbered 388 (3.9%) out of 10,016 examined cases. However, in 2016, the total positive cases numbered 447 (4.3%) out of 10,405 examined cases. The prevalence of H9N2 AIV among chickens on commercial farms was 4.6% out of the 16,666 chickens examined. The rates of positive cases in 2015 and 2016 were 4.4% (349/7884) and 4.7% (417/8782), respectively. The prevalence of H9N2 AIV in backyard chickens was 1.8% (69/3755). The rates of positive cases in backyard chickens were 1.8% (39/2132) in 2015 and again 1.8% (30/1623) in 2016. The highest positivity rate of H9N2 in chicken farms was in Beni-Suef (61.5%) (8/13), whereas the highest positivity rate in backyard chickens was in Fayoum (8.2%) (8/97).The analysis of H9N2 infections among chicken farms and in backyard chickens in the different governorates of Egypt over 2 years indicated widespread infection throughout the country. Thus, continuous surveillance and implementation of control programs are warranted. Since the first detection of H9N2 avian influenza virus (AIV) in poultry species (in turkeys) in the USA in 1966 . A wide Zoonotic transmission of H9N2 AIVs have been documented causing mild or asymptomatic infections in humans . Since tSeveral outbreaks of H9N2 AIV have been documented with the occurrence of single infection and coinfection with other bacterial and viral pathogens ,14. The In Egypt, H9N2 AIV was first isolated from a commercial quail farm in 2011 . Since tThe circulation of H9N2 AIV in Egypt increases the need to study the distribution pattern of H9N2 AIV in poultry through extensive surveillance and monitoring of field cases in commercial and backyard chickens throughout the country. Here, surveillance activities were conducted to determine the prevalence of H9N2 AIV in commercial and backyard chickens in Egypt.Ethical approval for this study was obtained from Animal Health Research Institute of Egypt.Molecular surveillance of H9N2 AIV was conducted in the Reference Laboratory for Veterinary Quality Control on Poultry Production (RLQP). A 2-year survey of the H9N2 AIV in chicken farms and the household chicken were carried out in 2015 and 2016.As shown in The samples were collected as five tracheal and five cloacal swabs from each collection site. All swabs from each farm or backyard were pooled as a single sample. The swabs were collected in sterile phosphate-buffered saline. Samples were transported under freezing conditions with minimal delay to the Laboratory of Virology, RLQP, Animal Health Research Institute (AHRI), Ministry of Agriculture, Egypt.Total RNA was extracted from the pooled samples using a QiaAmp Viral RNA Mini Kit , in accordance with the manufacturer’s instructions. Extracted RNA was subjected to real-time RT-PCR for detection of the H9 subtype of influenza A virus using Qiagen Quantitect probe real-time RT-PCR kit , using a Stratagene Mx3005p real-time PCR system , and the primer sequences and protocol reported previously : H9F priDifferences between groups were analyzed for statistical significance using Chi-squared test with “Statistical Package for Social Sciences, SPSS version 24.0” .On the farms, symptoms including reduced water consumption and respiratory manifestations were observed in broilers, while in layers there was reduced egg production with respiratory distress. At other farms, no apparent symptoms were identified. Mortality reached up to 30%. A few cases were vaccinated against H9 AIV (31 cases), while some cases were vaccinated against H5 AIV (482 cases).As shown in Fayoum Governorate had the highest rates of positive H9N2 detection in chicken farms among the governorates in Upper Egypt, with a 2-year prevalence of 23.6% (34.5% in 2015 and 16.5% in 2016). In contrast, H9N2 infection was not recorded at all in Matrouh Governorate . The difThe results shown in -2016, while it was detected in farms in this region only in 2016  was higher than that in Lower Egypt (3.8%), while in 2016, Lower Egypt (5.0%) had a higher positivity rate than Upper Egypt (4.1%). The prevalence of infection in East Egypt was 1.6% in 2015 and 2.6% in 2016 for farms and it was 1.8% in 2015 and 4.1% in 2016 for backyards. The infection was not detected in West Egypt in backyards in 2015 in 2016 . StatistAs shown in The endemicity and widespread of H9N2 AIV infection among the poultry population in Egypt have made it an additional risk factor in the poultry industry. H9N2 AIV also has the potential to be transmitted to humans, especially with the cocirculation of other influenza subtypes in Egypt ,24. AvaiIn this study, H9N2 AIV was surveyed among chickens in commercial farms and backyards in 26 governorates throughout Egypt. The data on the geographical prevalence of H9 subtype-positive cases revealed that the infection was recorded in 21/26 governorates in 2015 and 22/26 governorates in 2016. The spread of the virus showed no particular geographical pattern as there were no distinct barriers between most of the Egyptian governorates. In 2015, H9N2 infection was recorded in Upper Egypt, Lower Egypt, and East Egypt, while it was absent in West Egypt. Meanwhile, in 2016, the infection was recorded in all the four regions of Egypt. These results confirm the endemicity of H9N2 AIV infection among chicken populations in Egypt. Our results showed that the total prevalence of chicken infections with H9N2 was 3.9% in 2015 and 4.3% in 2016, which was higher than that reported in the previous work (3%) . The disOur results revealed that H9 infection was recorded in almost all governorates included in this study during 2015 and 2016, with the exception of Matrouh. This geographically widespread of H9N2 AIV infection in chickens in Egypt might be attributable to several factors, such as the low pathogenicity of the virus, poor biosecurity measures, and lack of a scheduled vaccination program. Moreover, household breeding is common in Egypt, where the birds are raised in a free-range set-up or at homes without biosecurity measures or vaccinations. Besides, the birds in backyards are regularly in close contact with carrier species such as aquatic birds, and with free-flying birds such as migratory birds, pigeons, and quails, which could be infected or act as carriers for different influenza subtypes . Other fet al. [The results of this surveillance indicated that the positivity rate in the commercial poultry sector (4.6%) was higher than that in the household poultry sector (1.8%), which matches the findings in previous work documenting a positivity rate in the commercial sector of 3.3%, but one in the backyard sector of 0.74% . In addiet al.  that theThe prevention and control programs of influenza viruses are focused on H5 AIVs subtype. However, the significance and economic impacts of H9N2 AIV should be considered when establishing programs to prevent and control H9 AIVs subtype. Although our findings indicated that the spread of H9N2 AIV among backyard birds occurred at a low rate, the backyard system is considered a major problem as it provides the opportunity for more poultry species to be reared together. In Egypt, particularly in rural regions, chickens, ducks, geese, turkeys, and pigeons are often reared in close contact with other animals and humans in the same household. The birds are mostly reared in primitive rooftops, cages, or as scavengers without the application of biosecurity measures. They move or graze through the streets or fields. These birds may thus come into contact with either domestic poultry or migratory birds . Thus, t-2012 [In this study, the incidence of H9N2 infection increased during the winter and spring months, while it decreased throughout the summer and fall, when the temperature increased. The climatic patterns of H9N2 infection in 2015 and 2016 were similar, with the infection rate gradually decreasing from April to June, and then infection being detected at a low rate from July to November. Finally, the infection rate increased from December to April. An increase in the rate of positive H9N2 cases in the spring months indicated that the epidemiology of H9N2 in chickens in Egypt in 2015 and 2016 changed over time, with outbreaks now occurring in warmer months. In 2015, the highest recorded numbers of cases were in January, February, and April, while in 2016 they were in April, March, and May. The results of this study agree with the previous work that reported H9N2 AIV infection throughout the year, with a particular association with winter months in Egypt during 2011-2012 . In addi-2012  that fou-human interface and contributes to the emergence of new reassortants, which could potentially produce a pandemic [Avian influenza of the H9N2 subtype could be naturally transmitted and humans contacts with infected birds . Seropospandemic . Therefopandemic .et al. [H9N2 AIVs are continuing to circulate through inter-species and intra-species transmission among various host species and subsequent zoonotic transmission has been reported by Rahimi Rad et al. . GeneticThe high prevalence of H9N2 infections among chicken farms and backyards in Egypt over 2 years indicated that such infections are widespread throughout the country. Thus, there is a need to encourage adequate measures to control H9N2 AIV in poultry in Egypt to minimize the risk factors to the poultry industry and the possibility of novel pandemic variants arising.MME drafted the manuscript, AIY revised the manuscript, analyzed the data, ASA designated the work, MME and MA collected the samples, and performed the laboratory work. All authors read and approved the final manuscript.


Automated speech recognition (ASR) systems have become increasingly sophisticated, accurate, and deployable on many digital devices, including on a smartphone. This pilot study aims to examine the speech recognition performance of ASR apps using audiological speech tests. In addition, we compare ASR speech recognition performance to normal hearing and hearing impaired listeners and evaluate if standard clinical audiological tests are a meaningful and quick measure of the performance of ASR apps.Four apps have been tested on a smartphone, respectively AVA, Earfy, Live Transcribe, and Speechy. The Dutch audiological speech tests performed were speech audiometry in quiet (Dutch CNC-test), Digits-in-Noise (DIN)-test with steady-state speech-shaped noise, sentences in quiet and in averaged long-term speech-shaped spectrum noise (Plomp-test). For comparison, the app's ability to transcribe a spoken dialogue (Dutch and English) was tested.All apps scored at least 50% phonemes correct on the Dutch CNC-test for a conversational speech intensity level (65 dB SPL) and achieved 90–100% phoneme recognition at higher intensity levels. On the DIN-test, AVA and Live Transcribe had the lowest (best) signal-to-noise ratio +8 dB. The lowest signal-to-noise measured with the Plomp-test was +8 to 9 dB for Earfy (Android) and Live Transcribe (Android). Overall, the word error rate for the dialogue in English (19–34%) was lower (better) than for the Dutch dialogue (25–66%).The performance of the apps was limited on audiological tests that provide little linguistic context or use low signal to noise levels. For Dutch audiological speech tests in quiet, ASR apps performed similarly to a person with a moderate hearing loss. In noise, the ASR apps performed more poorly than most profoundly deaf people using a hearing aid or cochlear implant. Adding new performance metrics including the semantic difference as a function of SNR and reverberation time could help to monitor and further improve ASR performance. Automated Speech Recognition (ASR) has become increasingly sophisticated and accurate as a result of advances in deep learning, cloud computing, and the availability of large training sets , 2. The Since 2017, several ASR systems have claimed speech recognition performance close to that of normally hearing humans . A lowerIn order to obtain estimates of human speech recognition performance that could be used for comparison with ASR, some researchers have determined the WER among professional transcribers of speech from the SwitchBoard and CallHome corpora. Saon et al.  estimateFor some commonly-used ASR systems, WERs of 5.1% (Microsoft) and 5.5% (IBM) have been reported using the SwitchBoard corpus , which iFor people with hearing impairments, there are specific user needs to consider when developing ASR apps. For example, these listeners might use both speechreading  and textThe settings where an ASR is used may also differ between individuals with impaired or normal hearing. For example, the settings where people with hearing loss use ASR may be more often in a more homely atmosphere between family members that might use more colloquial language or slang. That situation may be similar to closed caption for video series. The most common complaint of people with hearing loss is the reduced speech perception in complex listening environments including cocktail parties, restaurants, in conversations with their doctor, and family gatherings , 16. AdvA standardized task that fully captures the skills of humans to recognize speech does not yet exist, to our knowledge. Such a task would need to account for factors as background noise, reverberation, accent, and speech impairment. This is needed to verify claims that ASR speech recognition performance is close to humans , 2 and sThis pilot study aimed to examine the speech recognition performance of ASR apps using audiological speech tests. We normally administer clinical audiology tests in patients from normal hearing to profound hearing loss to assess speech recognition. We tested the hypothesis that our clinical tests might thus provide objective metrics for performance of ASR systems for people with hearing loss, helping us to determine what range of hearing losses could benefit from ASR apps. In addition, we compared ASR results to normal hearing and hearing impaired listeners and evaluated if standard clinical audiological tests provide a meaningful and quick measure of the performance of ASR apps.Four different apps on two smartphones, with various operating systems, were tested on their ability to transcribe speech. For this project, the iOS operating apps were tested using an iPhone 6, and for the Android operating apps, a Samsung A3 was used. Both smartphone devices are widely used. We decided to select inexpensive ASR apps (<$10) for a user-license since they would be most widely used by our patients while the cost for ASR apps is not reimbursed in the Netherlands. The four apps tested were Ava and Earfy that both run on iOS and Android, Speechy iOS only, and Live Transcribe Android only. The tested apps were chosen by searching on the Internet on November 18th, 2019, for the best-known speech recognition apps for the hearing impaired and deaf as well as good reviews on the different app-stores. Also, the apps needed to be suitable to convert English and Dutch speech into text.The apps were evaluated in similar test conditions used to assess speech reception in human listeners in Dutch Audiology Centers according to best local clinical practice. The smartphones were placed one meter in front of a speaker in a sound treated room compliant with ISO 8253-1 . StandarFirst, the apps were tested on speech recognition in quiet by converting a list of single words into text. The standard Dutch speech recognition test for this purpose is the Dutch CNC-test, which consists of phonetically balanced lists of twelve monosyllabic Dutch words in quiet . The woSecond, the Plomp-test (Dutch sentences in noise) was presented . The tesThird, a DIN-test (Digits-in-Noise) was performed. Digit triplets  were presented in long-term average speech-spectrum noise via a 1-up, 1-down adaptive SNR procedure. SRT was expressed in dB SNR, where a listener can on average recognize 50% of the digit triplets correctly. A test series consisted of 24 triplets. The first four triples were not used to determine the test outcome. The noise level was set at a fixed level of 60 dB with an initial positive SNR of 6 dB. The stepsize to adjust the level of the triplets was 2 dB. The DIN-test has a measurement error in humans of 0.7 dB .https://www.youtube.com/watch?v=zBJBD1-ePRw). For the English dialogue, part of an advanced English tutorial was played. In this video, a conversation could be heard between a male and female voice (https://www.youtube.com/watch?v=JtMgw2rCYSo&t=1s). The Dutch dialogue consisted of 256 words, while the English dialogue consisted of 248 words. After the whole dialogue was played, scoring was performed on the transcript outputted by the app. The number of missing, wrong, and inserted words was counted and expressed in the WER.Fourth, a fragment of dialogue in Dutch and English at 72.2 dB(A) was presented through the speaker to recreate a more realistic listening setting. The Dutch dialogue was an introduction video of the Radboudumc with a female voice, talking clearly and at a normal pace  was calculated for these results. No retest was performed for the dialogue.The results for all apps on the Dutch CNC-test (words in quiet) are shown in The speech-in-noise results are shown in In The test-retest reliability of the CNC-test can be seen in None of the ASR apps achieved performance close to normal hearing listeners on audiological tests. In quiet, ASR apps performed similarly to listeners with a moderate hearing loss. When transcribing speech-in-noise, the ASR apps performed in the performance range of CI recipients. Sentences-in-noise provided a quick test to assess ASR performance since that test material provided more linguistic cues than digits-in-noise or lists of CNC words.The performance of the ASR apps on speech-in-quiet tests seems comparable to listeners with a moderate conductive hearing loss (30–35 dB threshold shift), which is known as disabling for certain activities in daily life . In compThe performance of the ASR apps on the Digits-in-Noise test was very limited compared to humans. Normal hearing listeners achieve on the DIN-test, monaurally using headphones, an SNR of −8.8 dB . CI reciThe performance of ASR apps on sentences in noise (Plomp-test) was very limited and much poorer than in people with a moderate hearing loss . Normal Sentences with and without noise (Plomp-test) could be considered as a performance metric for ASR apps in difficult listening conditions. Possibly with more natural sentences to provide even more linguistic cues. Testing through a loudspeaker has the advantage that it takes the effect of room acoustics into account, making the test condition more realistic. Instead of a sound booth, a room with more representative acoustics for daily situations  would provide even more representative results. The current scoring procedure of the Plomp-test, based on fully correct sentences, leads to very high SNRs that may underestimate the practical value of ASR for hearing impaired persons. For instance, if an ASR in a conversation under noisy conditions provides keywords, it may already benefit the person with hearing impairment. One could easily adopt the Plomp-test by determining the WER score on a fixed SNR level to simulate above example. Or alternatively, accept a higher number of mistakes (compared to none) in the adaptive test by using keywords . BesidesIn longer dialogues, all tested apps provided a running English transcript with a WER around 19–34%. This roughly corresponds to 60–80% correct word (~1-WER) scores and this is in the same range as for persons with profound hearing loss who use a cochlear implant  and bettA possible explanation for the poor performance at low levels could be the smartphone's microphone gain settings and limited dynamic range rendering soft sounds undetectable . We chosIn English, there is not much difference between the apps or between the operating platforms. Therefore, we do not expect differences in the Dutch version to stem from hardware differences between the smartphones  but from the implementation of the Dutch language in the specific app or the used training data. The difference between iOS and Android was only visible in Dutch. In Dutch, Earfy (iOS) and Ava (iOS) score significantly poorer.There was no consistent difference favoring either iOS or Android versions of the apps. Earfy performed better on Android, while AVA performed better on iOS. For prospective users, the performance of the app depends on language, and may depend on the platform.The administered tests did not include the effect of accents or speech impairments . ThWhen measuring performance in noise, an adaptive SNR procedure was used. The effect of noise could be more extensively studied by evaluating ASR by determining the Word Recognition Score (the convention in the field of audiology) or the Word Error Rate (the convention in the field of ASR research) on several fixed SNR levels  that correspond to realistic listening conditions for people using a hearing aid . For ecoIn this study, only  speech-to-text performance of the apps was measured. The usability, processing speed, effect on speechreading, and readability of the transcript were not evaluated. Other researchers looked into requirements for speed and user interface and concluded that those are important factors to improve usability . We expeThe number of apps tested in this study is limited. We did not perform a standardized procedures for literature review  to find and include ASR apps for this pilot study. In English, more apps may be available than in Dutch and we did not include expensive state-of-the-art  ASR systems.Other factors to consider not included in this pilot study are the distance between speaker and listener, especially in these times of social distancing and the effect of face masks on a speaker's voice and intelligibility . FeedbacInstead of the quick audiological tests we performed here, a more conventional and elaborate evaluation method would be to record several hours of conversations with hearing impaired users  via a smartphone while the screen is oriented such that the user can read the transcript. Subsequently, one could create transcripts of the recordings by human transcribers as ground truth, pass the recordings through several ASR apps and determine a performance rating based on WER and other automated metrics such as the semantic distance between the ASR transcript and ground truth .ASR may benefit from domain-specific evaluation tools and have domain-specific applications. For instance, Miner et al.  developeMetrics as, for example, the WER , or semantic difference , as functions of signal-to-noise ratio and reverberation time (RT) can provide more ecologically valid estimates of the benefits ASR apps could provide in daily life. Representative SNR values could include −5, +10, +30 (quiet) dB SNR. For ecological valid measures, realistic fluctuating noise maskers should be used , 38. RevASR apps can provide benefits in conversations between patients and their audiologists . In addiA factor of variability in rating procedures is that in many speech-in-noise tests, the test is made easier for CI recipients by only scoring correct keywords rather than full sentences , 30. AltNone of the ASR apps achieved performance close to normal hearing listeners on audiological tests. No app stood out from the others on performance level. On audiological speech tests in quiet, ASR apps performed similarly to listeners with a moderate hearing loss. When transcribing speech-in-noise, the ASR apps performed in the performance range of CI recipients. Sentences-in-noise provided a quick test to assess ASR performance. Additional performance measures are needed to evaluate ASR apps. Besides the speech material, also type of noise and the presentation mode audio-only vs. audiovisual need to be considered. Adding new performance metrics including the semantic difference as a function of SNR and reverberation time can help to monitor and further improve ASR performance. Clinicians can use benchmarks based on such metrics to counsel prospective users and may benefit from automated procedures. Several hearing impaired listeners, especially CI recipients, report that they benefit from the apps in certain situations , which iThe raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.LP, J-WW, and PH conceptualized the study. LP, PH, and DG collected the data. LP and J-WW took the lead in drafting the manuscript. All authors contributed to the data interpretation, reviewed the results, and edited the 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.


Mucormycosis, a rare fungal infection, mainly affects individuals with diabetes mellitus and those who were immunocompromised and has a high mortality rate. Its most common presentation is similar to that of acute bacterial sinusitis with symptoms of nasal congestion, headache, and fever. The involvement of multiple cranial nerves in mucormycosis was rarely reported in the literature and indicates severe disease. Herein, we report the case of a 56-year-old man who was referred to the ophthalmology outpatient clinic for facial nerve palsy. He was treated with systemic steroids for 10 days with no improvement. On examination, he had a loss of vision and a frozen orbit due to involvement of cranial nerves II, III, IV, V, VI, and VII. An extensive workup revealed a hemoglobin A1C of 10%. However, he was never diagnosed with diabetes mellitus previously and denied any of the classical symptoms of diabetes mellitus. He underwent ethmoidectomy, maxillectomy, and drainage of an intraorbital abscess after appropriate imaging studies. Histopathology confirmed the diagnosis of mucormycosis, and the patient was started on systemic amphotericin B. This case emphasizes the importance of screening for diabetes mellitus. Early recognition of underlying diabetes mellitus in this patient may have prevented the development of mucormycosis along with its devastating complications. According to reports from the United States, mucormycosis has a very low incidence of 1.7 in every million individuals. Diabetes mellitus is the most common risk factor for the development of mucormycosis across Asia, while hematological malignancies are the major risk factors across Western countries. Although mucormycosis is a severe complication of poorly controlled diabetes mellitus, it is very rare for diabetes mellitus to be diagnosed after the development of mucormycosis. Mucormycosis can involve many organ systems, including the cutaneous, pulmonary, and gastrointestinal systems, and in severe forms, it can spread to other organs. However, rhinocerebral involvement is the most frequent presentation. Involvement of cranial nerve VII along with the widespread involvement of other cranial nerves was rarely reported in the literature. Herein, we report a very unusual case of mucormycosis with extensive rhino-orbit cerebral involvement presenting with multiple cranial nerve palsies as a first presentation of diabetes mellitus in a previously healthy patient.Mucormycosis is a rare opportunistic angio-invasive infection caused by fungi of the order Mucorales. It mainly affects individuals with diabetes mellitus and patients who have underlying immunosuppression, with a mortality rate reaching up to 80%.A 56-year-old man was referred to our institution for a case of facial palsy. He was treated with systemic methylprednisolone for 10 days, without improvement. On examination, he had a visual acuity of 1.0 in the right eye and hand motion in the left. Motility examination revealed limitation of abduction of the right eye. The patient had a frozen orbit with a limitation of all gazes in the left eye. Fundus examination revealed a pale optic disk, afferent pupillary defect, and an ischemic retina, indicating central retinal artery occlusion. Examination findings were also consistent with oculomotor, trochlear, and abducens nerve paresis. Mucormycosis was a result of steroid therapy, as the patient was diagnosed 2 weeks previously with an isolated cranial nerve VII and was started with steroids in a peripheral ophthalmology clinic. At his first presentation in Jordan University Hospital, he had a frozen orbit with cranial nerve III, IV, and VI palsies.The patient was admitted, and the results of baseline investigations were normal, except for a high white blood cell count and hemoglobin A1C (HbA1c) of 10%. Systemic steroids were stopped, and the patient was started on insulin to correct his blood sugar. He was never diagnosed with diabetes mellitus before and denied any symptoms of diabetes mellitus. He was initially diagnosed with diabetes mellitus. He tested negative for ketone bodies and was treated in the ward. Results of other routine investigations, such as kidney function tests and complete blood count, were normal, except for high blood sugar and a high white blood cell count.The differential diagnosis of his current symptoms was cavernous sinus thrombosis, a space-occupying lesion in the cavernous sinus or orbital cellulitis.Computed tomography of the brain and orbit was performed and revealed extensive sinus disease with left complicated ethmoidal and maxillary sinusitis extending into the left orbit with abscess formation. There was also extensive orbital cellulitis and evidence of left cavernous sinus thrombosis secondary to infection .It is possible that the patient had undiagnosed diabetes mellitus as indicated by a high HbA1c and was worsened by steroid therapyThe patient was started on systemic vancomycin and ceftriaxone, and a swab from the nasal cavity was sent for evaluation. Nasal swab revealed methicillin-resistant staphylococcus and positive fungal staining. The patient was started on systemic amphotericin B at a loading dose and continued with the therapeutic dose after 2 days of his presentation to our institution. Nasal endoscopy revealed diffuse white filaments and necrosis of the bone from which a biopsy was taken. Sinuses were drained, followed by ethmoidectomy and maxillectomy. A biopsy from the necrotic tissue was sent, which showed the presence of non-septate hyphae branching at right angles consistent with angio-invasive mucormycosis. However, workup to identify the exact causative organism of mucormycosis was not performed as it did not affect further management.The patient was kept on daily irrigation of the sinuses along with administration of systemic amphotericin B. On follow-up magnetic resonance imaging, the patient had an intraorbital collection . Subsequ Even with early diagnosis and aggressive therapy, its prognosis remains poor. Diabetes mellitus is the most common risk factor for mucormycosis, particularly during ketoacidosis. Ketones induce the fungi to utilize and produce ketoreductase, which facilitates its growth through various mechanisms. Roden et al. reviewed the characteristics of 929 patients with mucormycosis, and of 929 patients, 36% had diabetes mellitus, 17% had an underlying malignancy, and 19% had no underlying conditions. In a review of literature of 851 patients, the median age was 51 ± 12 years, and 531 (63%) were men. Diabetes mellitus was the most common underlying condition  had documented ketoacidosis), followed by hematological malignancy  had acute myeloid leukemia) and solid organ transplantation  had received a kidney transplant). Of the predisposing factors, the use of corticosteroids at the time of presentation was the most common , followed by neutropenia  and trauma . Rhino-orbito-cerebral mucormycosis was the most commonly observed manifestation , followed by cutaneous  and pulmonary mucormycosis  to rule out diabetes mellitus as the underlying cause of his facial palsy and (2) to measure the baseline blood sugar before the initiation of steroid therapy.Mucormycosis is an opportunistic infection with a high mortality rate, reaching up to 80% without treatment. In advanced disease, symptoms include chemosis, ptosis, proptosis, ophthalmoplegia, blindness, and multiple cranial nerve palsies .4,5 The classical clinical presentations are facial pain, an irregular black eschar on the palatal or nasal mucosa, and pus discharge from the eye and nose. In a retrospective chart review of 48 patients with 49 cases of acute fulminant-invasive fungal sinusitis over 19 years, mucormycosis and aspergillosis were found in 22 and 27 cases, respectively. Orbital  and cranial nerve symptoms were seen at presentation more frequently in mucormycosis (6 [27%] and 9 [41%]) than in aspergillosis cases . Long-term orbital and cranial nerve sequelae were reported in 16 (72%) mucormycosis cases and 10 (37%) aspergillosis cases (p = 0.0210). These data suggest that the presence of orbital and neurological symptoms at presentation warrants more aggressive surgical intervention because of the likelihood of mucormycosis. In another review of mucormycosis cases, all six patients had uncontrolled diabetes mellitus with fungal-invasive disease and had multiple cranial nerve involvement, including cranial nerves II to VII. However, our patient had not been definitely diagnosed with diabetes mellitus before he had the fungal infection. That cohort chiefly complained of facial swelling, blood-stained nasal discharge, and crusting. All six patients were on prolonged antibiotic treatment with an incomplete response. Facial dysesthesia was present in all sides involved. Six patients had impaired vision, two had vision loss, and four only had a perception of light. These patients improved to finger counting during treatment. All patients had periorbital edema, ptosis, and 7th cranial nerve palsy of the lower motor type, and two patients had irritability and disoriented state and succumbed to the disease within 48 h of treatment. By contrast, our patient was fully oriented and had an initial visual acuity of hand motion with limited improvement.Clinically, mucormycosis is characterized by rhinitis with granular and purulent discharge, nasal ulceration, black spots of infarcted mucosa, and paranasal sinusitis. However, other common presenting symptoms include epistaxis, ophthalmoplegia with blindness, proptosis and orbital cellulitis, hemiplegia or stroke, and decreased mental function.A diagnosis of rhino-orbito-cerebral mucormycosis requires high clinical suspicion, and a definitive diagnosis requires a biopsy indicating the presence of the fungi. Furthermore, imaging is required to identify the degree of local involvement and spread across the rhino-orbital structures.Mucormycosis is treated with surgical debridement of necrotic tissues, use of intravenous antifungal drugs, and elimination of predisposing factors. In retrospective studies, the combination of surgery and antifungal drugs showed higher survival rates than medical treatment alone. Early detection and treatment of this patient's underlying diabetes mellitus may have prevented the development of mucormycosis, especially in a country like Jordan where the prevalence of diabetes mellitus among men aged >25 years is as high as 33%.Our case emphasizes the importance of screening for diabetes mellitus to prevent such a devastating complication. Our patient denied any of the classical symptoms of diabetes mellitus, and the only risk factor he had for diabetes mellitus was his age. Given the relatively long asymptomatic period of diabetes mellitus, the American Diabetes Association recommends screening for diabetes mellitus at age 45 years. We are unsure whether the involvement of cranial nerve VII in this patient was the first manifestation of mucormycosis or it was due to an independent pathology in which its treatment with glucocorticoids facilitated the development of mucormycosis. Furthermore, the widespread involvement of cranial nerves III, IV, and VI is also uncommon and indicates severe disease possibly due to cavernous sinus thrombosis.The involvement of cranial nerve VII in mucormycosis is very rare, and its pathogenesis is not well understood.Mucormycosis is a rare and possibly fatal opportunistic infection. It should be suspected in any patient who presents with symptoms of sinusitis along with the widespread involvement of the orbit and cranial nerves. Workup should be initiated for an underlying systemic disease, such as diabetes mellitus, in any patient diagnosed with mucormycosis. In the present case, initiation of steroid therapy may have influenced the development of fungal infection and its progression, and this highlights the importance of screening. It is critical to exclude diabetes mellitus before starting steroid treatment and to screen a susceptible population perhaps at a younger age than recommended. Treatment with systemic antifungals, surgical debridement of necrotic tissues, and correction of underlying hyperglycemia should be initiated as soon as possible in diabetic patients with mucormycosis. Although this patient was 56 years old, screening for diabetes mellitus in younger populations earlier than 45 years is recommended, especially in the presence of opportunistic infections.Ethical approval was obtained from the Jordan University Board. Informed consent was also obtained from the patient.All authors declare no conflict of interestThis work was not funded by any funding bodies, and all authors have read and approved the contents of this manuscript.


Many community-dwelling older adults develop activity of daily living (ADL) disability and subsequently regain function. Using data from the ASPirin in Reducing Events in the Elderly (ASPREE) clinical trial, we examined the relationship of gender, incident disability, and persistent disability 6 months after the incident disability. Walking, bathing, dressing, transferring, toileting, and eating were assessed as ADLs, at bi-annual interviews. ADL disability was defined as requiring help with or inability to do or severe difficulty with ≥1 ADL; persistent disability was an ADL loss at 6 months after a first (incident) ADL disability. Discrete time, multivariable Cox proportional hazards regression was utilized to estimate associations with developing incident ADL disability described as cause-specific hazard ratios, with death as a competing outcome. For persons with incident ADL disability, odds of developing persistent disability at 6 months as compared to recovery was determined using multivariable logistic regression. These analyses included 18,414 (51.6% women) ASPREE participants in the United States and Australia aged 70+ years (65+ years if U.S. ethnic minority) without ADL disability at trial entry. During a median follow-up of 4.7 years, 1,485 participants (63.2% women) developed an incident ADL disability, and, of those, 272 (57.0% women) met criteria for persistent disability at 6 months. Women had an increased risk  of developing incident ADL disability; however, women were less likely to have persistent disability versus recovery 6 months later . Why persistent disability development is lower in older women needs further exploration.


SARS-CoV-2–infected patients are reported to show immunocompromised behavior that gives rise to a wide variety of complications due to impaired innate immune response, cytokine storm, and thrombo-inflammation. Prolonged use of steroids, diabetes mellitus, and diabetic ketoacidosis (DKA) are some of the factors responsible for the growth of Mucorales in such immunocompromised patients and, thus, can lead to a life-threatening condition referred to as mucormycosis. Therefore, an early diagnosis and cell-based management cosis is the need of the hour to help affected patients overcome this severe condition. In addition, extended exposure to antifungal drugs/therapeutics is found to initiate hormonal and neurological complications. More recently, mesenchymal stem cells (MSCs) have been used to exhibit immunomodulatory function and proven to be beneficial in a clinical cell-based regenerative approach. The immunomodulation ability of MSCs in mucormycosis patient boosts the immunity by the release of chemotactic proteins. MSC-based therapy in mucormycosis along with the combination of short-term antifungal drugs can be utilized as a prospective approach for mucormycosis treatment with promising outcomes. However, preclinical and in mucormyIn mucormycosis, the hyphae of clinical trials are needed to establish the precise mechanism of MSCs in mucormycosis treatment. These cells differentiate into multiple tissues, such as bone, cartilage, connective tissue, muscle, and fat cells . The Intolecules . MSCs plolecules . MSCs arolecules . MSCs arolecules .C. albicans  conducted 950 successful clinical trials over 10,000 subjects performing promising MSC therapies . MSCs poalbicans .In SARS-CoV-2-recovered subjects, post COVID-19 sepsis is among the common complications. The post-COVID-19 irregularities include impaired innate immune response, cytokine storm, thrombo-inflammation, and eventual immune exhaustion. Mucormycosis contributes to life-threatening fungal infections with higher than 50% mortality rates even postsurgical debridement and after antifungal drugs . Recentlrhizopusarrhizus is the culprit fungus responsible for causing pulmonary, gastric, cutaneous, and rhino-orbital-cerebral diseases, especially in adults , orbit, CNS, skin, bones, kidney, joints, gastrointestinal tract, and mediastinum (invasive type). Studies show that giant cell invasion, eosinophilic necrosis, and thrombosis are the prime hallmarks of mucormycosis . These mRhizopus, Lichtheimia, Cunninghamella, Rhizomucor, and Apophysomyces are reported  that induces the NF-κβ pathway-related genes  but not at alkaline pH  and pleural effusions are presentable. In computerized tomography (CT) scanning, a reverse halo sign (RHS) confirms the presence of mucormycosis . In anotRhizomucor antibody is used for immunohistochemical analysis to diagnose Mucorales and Entomophtorales , DNA sequencing, melt curve analysis of PCR products and restriction fragment length polymorphism analyses RFLP; . These t species .Rhizopus, Mucor, and Cunninghamella species at the same time in a single specimen , and implication of antifungal agents. In early management of mucormycosis, polyene therapy within 5days of early diagnosis shows significant improvement in survival compared with the therapy of polyene administered after 6days of the diagnosis  if possible in such patients. Such an approach also includes aggressive glycemic control and normalization of the acid–base environment to prevent DKA.Surgical management in mucormycosis involves debridement of the necrotic tissues for complete elimination of mucormycosis. This approach is the last option to treat such infected patients when there is no or limited penetration of antifungal agents/drugs at the site of infection due to tissue necrosis and blood vessel thrombosis. In a previously published study, surgical debridement of the tissue was found to be on top when tested in a logistic regression model in patients with mucormycosis with promising outcomes . In anotPrimary antifungal therapy includes the use of amphotericin B, liposomal amphotericin B, and amphotericin B lipid complex. The dose of these antifungal agents in the treatment of mucormycosis is still not known or fixed under treatment guidelines. However, starting dosages of 1mg/kg/day of amphotericin B and 5–7.5mg/kg/day for liposomal amphotericin B and amphotericin B lipid complex are commonly prescribed in clinical practices in both children and adults. A previously conducted study proved that a higher dose of liposomal amphotericin B (>10mg/kg/day) does not show any significant pharmacokinetic advantage in patients with mucormycosis .Salvage therapy provides an additional advantage in the management of mucormycosis. Deferasirox or posaconazole are some of the offered choices for patients with mucormycosis and those who are intolerant to polyene therapy. Sufficient data for posaconazole is available from clinical trials regarding the efficacy and safety of this drug in the management of mucormycosis. However, if deferasirox must be added, it should be given only for 2–4weeks during the salvage therapy as, beyond 4weeks, it causes toxicity . For patIn the treatment of mucormycosis, antifungal drugs are the first-line treatment options, especially in prophylaxis and invasive fungal infections. These azoles have associated long-term side effects, especially in patients with hematologic malignancies. The long-term complications or side effects associated with the azole implication include hormone-related impairments, such as alopecia, impotence, oligospermia, decreased libido, gynecomastia, hyponatremia, hypokalemia, and rarely adrenal insufficiency, along with hepatotoxicity . In anotin vitro and therapeutic effect in C. albicans–infected mice.Multipotent stem cells exhibit immunomodulation characteristics that were found to be safe for the treatment of COVID-19 . The FDAMucormycosis is a life-threatening opportunistic infection that severely affects immunocompromised patients, especially SARS-CoV-2-infected patients. In a recently published case report, it is found that a patient with acute myeloid leukemia (AML) presented with pulmonary mucormycosis and was treated with the antifungal agent voriconazole preceded by a 1month transplant with peripheral blood stem cells followed by isavuconazole, and the patient showed a significant reduction in ground glass opacities in the CT scan . This stAspergillus fumigatus hyphal extract (Aspergillus fumigatus conidia that further lowers the expression of tumor necrosis factor-α (TNF-α) and increases the expression of IL-10  contribute as vital components of the innate immune system, assuming the first line of defense . In a muironment . In anot defense . Mucoral defense .with L. corymbifera, R. oryzae, R. delemar, and C. bertholletiae mediated through platelet-derived growth factor receptor B (PDGFRB) signaling in the infection phase  in lungs  were observed to interact on phase . Moreovein lungs . Polymorin lungs .−) when encountered with the hyphae of mucoralean species that, in turn, modulate the neutrophils  exhibit CD73+, CD90+, and CD105+ along with CD45-, CD34-, CD14-, CD11b-, CD79a-, and HLA class II . StudiesIn vitro and in vivo study demonstrates that IL-17+ MSCs inhibit the growth of Candida albicans. MSCs are proven to show a protective effect in asthma induced by Aspergillus fumigatus hyphal extract mediated by inhibition of Th17 signaling mechanism  was investigated on anti-Aspergillus CD4+ T cells, and it was found that anti-Aspergillus T cells initiate the IL-6 production of hMSCs .MSCs modulate the immune molecules of the immune system and are proven to be beneficial in the survival of patients suffering from immune-related complications. MSCs are known to prevent the overstimulation of the immune system as they contribute to the suppression of the immune functions mediated by stimulation of pro-inflammatory factors  having the azole functional group is the first choice in mucormycosis possess some limitation of having side effects on long-term use, MSCs can be an alternative cell-based treatment approach that can be used without conflict. MSCs exhibit an immunomodulatory role in the host and protect it from fungal infection by boosting the immune system through the release of various chemotactic proteins.Moreover, very limited studies are present explaining the mechanism of immunomodulation adopted by MSCs in mucormycosis. However, assumptions can be made for using MSCs as prospective therapy in the treatment of mucormycosis. No conclusive data of clinical trials are available to include MSCs in treatment of mucormycosis.Currently involved in COVID-19 testing duties; Prashant Tripathi: Nodal Officer, COVID-19 Testing Laboratory, GSVM Medical College, Kanpur; Richa Giri: Nodal Officer, Multidisciplinary Research Unit, GSVM Medical College, Kanpur.AR, SA, and G-BJ: conceptualization. AR, SA, G-BJ, KG, SB, QM, SR, and RT: formal analysis. KG, SB, SR, and RT: investigation. AR, SA, and G-BJ: methodology. PT, RG, SA, MS, and HK: supervision. PT, RG, SA, MS, and HK: visualization. AR, SA, G-BJ, and QM: writing – original draft. PT, RG, SA, MS, and HK: writing – review and editing. All authors have read and approved the manuscript and ensure that this is the case.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.