The Knee injury and Osteoarthritis Outcome Score–Anterior Cruciate Ligament (KOOS-ACL) is a short form version of the KOOS, developed to target populations of young active patients with ACL tears. The KOOS-ACL consists of 2 subscales: Function (8 items) and Sport (4 items). The KOOS-ACL was developed and validated using data from the Stability 1 study from baseline to postoperative 2 years.To validate the KOOS-ACL in an external sample of patients matching the outcome’s target population.Cohort study (diagnosis); Level of evidence, 1.The Multicenter Orthopaedic Outcomes Network group cohort of 839 patients aged 14 to 22 years who tore their ACLs while playing sports was used to assess internal consistency reliability, structural validity, convergent validity, responsiveness to change, and floor/ceiling effects of the KOOS-ACL at 4 time points: baseline and postoperative 2, 6, and 10 years. Detection of treatment effects between graft type (hamstring tendon vs bone–patellar tendon–bone) were also compared between the full-length KOOS and KOOS-ACL.d = 0.94 [Function] and d = 1.54 [Sport]). Stable scores and significant ceiling effects were seen from 2 to 10 years. No significant differences in KOOS or KOOS-ACL scores were detected between patients with different graft types.The KOOS-ACL demonstrated acceptable internal consistency reliability (α = .82-.89), structural validity , convergent validity , and responsiveness to change across time (large effect sizes from baseline to postoperative 2 years; The KOOS-ACL shows improved structural validity when compared with the full-length KOOS and adequate psychometric properties in a large external sample of high school and college athletes. This strengthens the argument to use the KOOS-ACL to assess young active patients with ACL tears in clinical research and practice. The intended structure of the KOOS— with items assigned to 1 of 5 subscales —was not supported in the sample, and it was evident that major restructuring and/or simplification would be required to establish adequate fit.The Knee injury and Osteoarthritis Outcome Score (KOOS) is commonly used in clinical research trials, registries, and clinical practice to assess high-functioning patients with anterior cruciate ligament (ACL) tears after ACL injury and reconstructive surgery. However, the KOOS is not specific to ACL injuries, is quite long (42 questions), and may not be as relevant to the young active patients with ACL tears whom we often wish to follow. Additionally, there is no evidence to support the structural validity of the KOOS in patient populations with ACL tears.7 A preliminary validation of the KOOS-ACL indicated adequate internal consistency reliability, structural validity, convergent validity, and responsiveness to change across time from baseline (preoperative) to postoperative 24 months. There were some remaining concerns surrounding measurement invariance and ceiling effects starting at 6 months, which, although not unexpected, may influence the ability to detect treatment effects between intervention groups in clinical research. Further studies may be required to identify the ability of the measure to detect change across various follow-up periods and between treatments.Therefore, a short form version of the KOOS was created with the goal of achieving structural validity, removing irrelevant items, and reducing responder burden, while still maintaining adequate psychometric properties. The development of the KOOS-ACL was conducted using data from the Stability 1 study, wherein patients were randomized to hamstring tendon (HT) autograft ACL reconstruction (ACLR) alone or with a lateral extra-articular tenodesis.These initial validation analyses were all conducted within the same data set from the Stability 1 study, which presents a lack of external validity and the potential to falsely declare adequate psychometric properties that may not hold outside of the utilized group of patients. Therefore, the purpose of this study was to externally validate the psychometric properties of the KOOS-ACL using the Multicenter Orthopaedic Outcomes Network (MOON) group data set of >800 young athletes  with ACL tears who underwent ACLR with HT or bone–patellar tendon–bone (BPTB) autografts and were assessed at baseline and postoperative 2, 6, and 10 years. It was hypothesized that the KOOS-ACL would show adequate internal consistency reliability, structural validity, convergent validity, and responsiveness to change in this data set of high-functioning patients with ACL tears.All abbreviations are defined in  This cohort study was approved by the Cleveland Clinic institutional review board (No. 3867). These patients completed the KOOS questionnaire before surgery (baseline) and at postoperative 2, 6, and 10 years. This sample represents the target population of the KOOS-ACL, young active patients undergoing ACLR, and is similar to that of the Stability 1 study, the sample in which the KOOS-ACL was developed and initially validated. However, the MOON cohort has a younger maximum age (22 vs 25 years) and younger mean age (17.7 vs 19.0 years), as well as longer follow-up time points (10 vs 2 years). Additionally, all patients in the MOON cohort were high school or college athletes, whereas patients in the Stability 1 study had to fit ≥2 of the following high-risk criteria: a pivot-shift grade ≥2, a desire to return to high-risk/pivoting sports, and generalized ligamentous laxity or knee hyperextension >10°.18 This meant that Stability 1 study participants could be eligible without sport participation, although this was rare.The MOON group cohort used in this validation study consisted of 839 patients aged 14 to 22 years who experienced an ACL rupture while playing sports and underwent a primary ACLR with HT or BPTB autografts.Appendix (available in the online version of this article). To conduct a rigorous external validation, methodology from the initial validation article was replicated using a data set from an external source (MOON group). Internal consistency reliability, structural validity, convergent validity, responsiveness, floor/ceiling effects, and detection of treatment effects were assessed for the KOOS-ACL, testing our hypotheses at each of the 4 MOON group cohort study time points, using the statistical methods and acceptable thresholds outlined in  Domain scores represent the sum of all items in the domain standardized to a score from 0 to 100 (worst to best). The IKDC is an 18-item knee-specific questionnaire querying symptoms, function, and sports activities. The items are summed and transformed to a score that ranges from 0 to 100 (worst to best). The WOMAC is a 21-item questionnaire widely used to assess symptoms and disability and was originally developed for individuals with hip and knee osteoarthritis. The function subscale contains 17 items and is scored out of 68. These outcomes measures have all been validated in patient populations of various knee conditions.The KOOS-ACL outcome measure is available in the lavaan, effsize, dplyr, and psych software packages.22All analyses were conducted in R Studio using the t tests were run on the main outcome measures to assess for systematic bias based on missingness, there were no significant differences in baseline KOOS-ACL scores—Function , Sport , or composite —between patients who were missing or not missing scores at postoperative 10 years.Patient characteristics are displayed in Acceptable Cronbach alpha values (.70 < α < .90) indicated reliable KOOS-ACL Function and Sport scores addressing the same underlying construct at all 4 time points .The 2-factor structure of the KOOS-ACL, with 8 Function items and 4 Sport items, was maintained in the MOON cohort as indicated by acceptable fit indices across time  . Equal fKOOS-ACL scores were strongly and significantly correlated with outcomes assessing similar constructs, including the IKDC  and WOMAC function , at all 4 time points . KOOS-ACF = 252.91; P < .001) and Sport  scores (d = 0.94) and Sport (d = 1.54) scores from baseline to 2 years, followed by stable mean scores (mean differences <1 point) from 2 to 10 years and negligible effect sizes . Floor effects were below the acceptable threshold for all scores and time points.F = 3.55; P = .03) and Sport , with consistently lower scores reported by patients who had a hamstring graft  but not the ADL subscale . The assumption of sphericity was violated for these analyses ; therefore, P values are reported with the Greenhouse-Geisser correction to reduce the likelihood of a type I error.There was a significant difference in baseline scores between groups (HT vs BPTB graft) detected in KOOS-ACL and full-length KOOS scores. When individual analyses of variance (ANOVAs) were conducted to compare groups at each postoperative time point, neither the KOOS-ACL nor the full-length KOOS detected a treatment effect between HT and BPTB autografts in the MOON patient cohort. However, repeated-measures mixed ANOVAs showed a significant interaction between time and graft type (HT or BPTB) in KOOS-ACL subscales, Function .4 A strict threshold for comparative fit (CFI and TLI) was used (0.95). Because the KOOS-ACL subscales contain relatively few items, especially the 4-item Sport scale, a more lenient threshold of absolute fit  was chosen (<0.08), as these fit indices are more strongly influenced by the number of degrees of freedom in a model. When all time points were combined, the CFI (0.937) was below the strict 0.95 threshold but still within the range of 0.9 to 0.95, which is generally deemed satisfactory for model fit.This external validation confirms the structural validity of the KOOS-ACL at extended time points of postoperative 6 and 10 years. This is important because the full-length KOOS does not show acceptable structural validity in populations with ACL tears, with significant overlap among subscales/constructs; therefore, we cannot be confident in interpreting the KOOS subscale scores for patients with ACL tears as they are labeled.P < .001). The larger MOON prospective cohort of 3202 patients undergoing ACLR also showed that younger age was a predictor of loss to follow-up, although this relationship was not significant (P = .065).Feasibility is an important psychometric property of patient-reported outcome measures, as evidenced by the diminishing proportions of patients with the required KOOS items available for analysis at subsequent time points (92% at baseline to 78% at postoperative 10 years). The KOOS-ACL is 71% shorter than the full-length KOOS (12 vs 42 items) and is estimated to take <5 minutes for patients to complete. A reduced length is likely to improve feasibility of completion and may help reduce the number of patients lost to follow-up. This may be especially helpful when working with youth athletes who may not be compliant in completing extensive batteries of patient-reported outcome measure questionnaires. In the Swedish knee ligament register, nonrespondents at 2-year follow-up were significantly younger  than respondents , Italian, Japanese, Korean, Latvian, Lithuanian, Norwegian, Persian, Polish, Portuguese, Russian, Singapore English, Slovakian, Slovenian, Spanish (Peru), Spanish (United States), Thai, Turkish, and Ukrainian.18 Interestingly, the MOON and Stability groups showed nearly identical scores for baseline Function (83 vs 82), 2-year Sport (95 vs 96), and 2-year Function (85 vs 85). The results of the 2 validation studies also show strikingly similar structural validity fit indices, correlations with IKDC scores, and Cronbach alpha values of internal consistency reliability. The likeness of scores and psychometric properties reported from these 2 cohorts indicates consistency in KOOS-ACL assessment of high-functioning patients with ACL tears. The MOON group cohort did have a notably higher mean baseline sport score as compared with the Stability sample (48 vs 38). This may reflect the fact that all patients in the MOON group cohort were athletes and injured while playing sports. In the Stability 1 study, playing a competitive pivoting sport was only 1 of several possible high-risk inclusion criteria, but even those who did not meet this criterion were highly active. The correlation between the subscales (Function and Sport) appeared to increase at later time points (postoperative 2-10 years). This could reflect that individuals are participating in lower levels of sport with increasing age; therefore, the constructs of Function and Sport start to become more similar.Because the KOOS-ACL was designed for a very specific target population—young active patients with ACL deficiency—it was important that validation be conducted within a data set of patients matching this profile, such as the MOON group young athlete cohort. The Stability and MOON patient samples are very similar in terms of age  and sport participation.In addition to the necessary step of validating the KOOS-ACL in an external data set, this study introduces new follow-up time points (6 and 10 years after ACLR) and a new graft type for ACLR (BPTB), which widens the applicability of the KOOS-ACL within the target population. Prominent ceiling effects are evident at these later follow-up time points, which may limit the usefulness of the KOOS-ACL in detecting treatment effects between intervention groups. However, it is not surprising that a large proportion of high school and college athletes are doing functionally well years after ACLR, as this is the desired and expected outcome of surgical intervention. More sensitive outcome measures, such as difficult performance-based or functional tests, may be needed and more appropriate to detect intervention effects, if they exist, at later time points. Importantly, the internal consistency reliability, structural validity, and convergent validity of the KOOS-ACL did not degrade below any of the acceptable thresholds after the 2-year time point assessed in preliminary validation. KOOS-ACL scores showed stability from postoperative 2 to 10 years, as hypothesized for this patient population. It may make sense to transition from the KOOS-ACL to full-length KOOS at later time points (>10 postoperative years), when symptoms of osteoarthritis may appear and items from the full-length KOOS assessing lower levels of function may be relevant. On the basis of this external validation study, we believe that the KOOS-ACL can be appropriately utilized for high-functioning patients with ACL tears until postoperative 10 years. It also showed that patients may have been more likely to receive a BPTB graft if they were younger, had high-grade laxity, played organized high school or college sports, had a lower body mass index, or were Black. It is not clear whether these characteristics would be linked to higher baseline scores on the KOOS.Although the KOOS-ACL did not elucidate any treatment effects between graft types in this cohort (despite known differences in resultant graft failure risk), the fact that similar results in the ANOVAs were seen with either the KOOS-ACL or the full-length KOOS indicates criterion validity of the short form. Additionally, the KOOS-ACL Function subscale was able to detect a significant time × graft type interaction, which was not seen in the full-length KOOS ADL subscale. This interaction may be driven by the lower baseline scores in patients who had a hamstring autograft reconstruction. Graft choice in this patient cohort was based on patient/surgeon decision (not randomized). A previous study showed that of 17 participating surgeons, 4 almost always chose BPTB, 4 almost always chose HT, and the remaining 9 switched between graft choices depending on their personal algorithms.The external validation is an important step in creating a patient-reported outcome measure. The KOOS-ACL has now been assessed and validated in >1400 young active patients with ACL tears. However, both validation studies to date have included North American patient cohorts. Further validation would be beneficial to increase our confidence in using the KOOS-ACL worldwide for high-functioning patients with ACL tears. The KOOS-ACL can be tested in any data set that has full-length KOOS data available.The KOOS-ACL shows improved structural validity as compared with the full-length KOOS and adequate psychometric properties in a large external sample of high school and college athletes. This strengthens the argument to use the KOOS-ACL to assess young active patients with ACL tears in clinical research and practice.Click here for additional data file.Supplemental material, sj-pdf-1-ajs-10.1177_03635465231160726 for External Validation of the KOOS-ACL in the MOON Group Cohort of Young Athletes Followed for 10 Postoperative Years by Hana Marmura, Paul F. Tremblay, Dianne M. Bryant, Kurt P. Spindler, Laura J. Huston and Alan M.J. Getgood in The American Journal of Sports Medicine


Mucus is a viscoelastic gel that acts as a protective barrier for epithelial surfaces. The mucosal vehicles and adjuvants need to pass through the mucus layer to make drugs and vaccine delivery by mucosal routes possible. The mucoadhesion of polymer particle adjuvants significantly increases the contact time between vaccine formulations and the mucosa; then, the particles can penetrate the mucus layer and epithelium to reach mucosa-associated lymphoid tissues. This review presents the key findings that have aided in understanding mucoadhesion and mucopenetration while exploring the influence of physicochemical characteristics on mucus–polymer interactions. We describe polymer-based particles designed with mucoadhesive or mucopenetrating properties and discuss the impact of mucoadhesive polymers on local and systemic immune responses after mucosal immunization. In future research, more attention paid to the design and development of mucosal adjuvants could lead to more effective vaccines. The development of mucosal vaccines continues to be a priority in the fight against microorganisms whose entry is the mucosa. However, the mucus layer limits the passage of antigens across the epithelium to reach mucosa-inductive sites. This protective barrier facilitates the clearance of foreign pathogens and particles. The mucosal compartments have the epithelium and the mucosal immune system as barriers to defense. The epithelial barrier interconnected by tight junctions, the mucus layer, antimicrobial peptides, and immunoglobulin A (IgA) production prevent, as a whole, access to pathogenic microorganisms, foreign particles, and toxins ,2,3.Furthermore, the mucosal immune system, comprising a network of mucosa-associated lymphoid tissues (MALT), is responsible for initiating and establishing the antigen-specific innate and adaptive immune response following infection or vaccination . Both inConsidering the barrier properties of mucosal surfaces, extensive studies have been performed to develop strategies for prolonging the residence time of vaccines in epithelial tissues; one of the most relevant is the use of mucoadhesive formulations. This review provides an overview of mucus, mucin, and how the interactions between mucus and particles occur. We discuss the physicochemical characteristics of particles that lead to improved mucoadhesion and/or mucopenetration.The paper aims to show solid scientific evidence to re-evaluate the correlation between improving the mucoadhesive or mucopenetrating characteristics of polymer-based particles used as mucosal vaccine adjuvants and the increase in specific systemic and mucosal immune responses. Likewise, we address potential strategies for mucus penetration, highlighting the importance of incorporating them to design more effective mucosal vaccines. For these purposes, most of the included research papers are those in which the mucoadhesion test of polymer particles was published together with the assessment of adjuvanted capacity following mucosal vaccination.The mucosal surfaces cover a vast extension of the body surface area. Due to continuous environmental exposure, many pathogens, or antigens, as well as particles found in the air and toxins, have the mucosal surfaces as portals of entry to the body. Therefore, the mucosal tissues play a fundamental role in protecting from invasion by harmful microorganisms through physical and biological barriers. The characteristic induction of antigen-specific sIgA antibodies, both local and distant, as well as protective immunity in systemic and mucosal compartments, makes mucosal vaccination the best tool for reducing mortality and morbidity caused by infectious pathogens that enter the body through the mucosal surfaces ,15.However, most vaccines licensed for use in humans are currently administered parenterally. Although parenteral immunization successfully induces a protective systemic immune response, it hardly induces an effective mucosal immune response, and the cellular mechanisms underlying this response remain largely unknown ,17,18. Vibrio cholerae   CS or HTCC. HTCC polymers have different degrees of quaternization or extent of positive charge . Zhang eCarboxymethyl chitosan (CMCS) is another water-soluble CS derivative with an improved degradation rate, a desired characteristic for its use in vivo . Recentl65CM50CS) were used in this work. The evaluation of swelling showed that the degree and rate of swelling of the TM65CM50CS-coated MPs were higher than those uncoated, both in HCL pH 1.2 and in PBS pH 7.4. Similarly, the in vitro mucoadhesion study using the everted gut sac with porcine jejunum showed that, while the adherence percentages of calcium alginate MPs and YMP MPs were low , the coating with TM65CM50CS of both particles increased mucosal adhesion during the first hour . Oral immunization resulted in significantly higher IgG and IgA levels in mice receiving OVA-loaded TM65CM50CS-coated MPs, which again confirms the role of mucoadhesive polymers in immunogenicity [For their part, methyl CS has been studied for diverse biological activities, including as tissue regeneration activator, absorption enhancer, and mucoadhesive . Suksamrgenicity .The ζ-potential of the CS-based vaccines significantly influences the induction of an immune response affecting more than one mechanism. Jesus et al. demonstrated that. after the intranasal administration of polycaprolactone/CS (PCL/CS) NPs in C57BL/6 mice, the lowest dose of adsorbed antigen (1.5 μg HBsAg) induced antibody titers comparable to the dose containing six times more adsorbed antigen (10 μg HBsAg). Furthermore, this group had the highest number of responding animals. However, serum IgG titers were significantly low compared to previous studies with the same dose of antigen (1.5 μg HBsAg), so the authors suggested that the decrease in ζ-potential (CS: +30 mV) to values close to neutrality generated by antigen interaction  leads to a reduced uptake in the epithelial barrier. These observations were independent of the mucoadhesive behavior of the particles without antigen evaluated in vitro. Therefore, the authors suggested that the antigen on the particle’s surface reduces the ζ-potential and hinders the interaction with mucin in vivo, avoiding particle–cell interactions and ultimately impacting the immune response . AlthougCarboxymethylcellulose (CMC), an anionic and water-soluble cellulose derivative , has beeSaccharomyces cerevisiae [Actinobacillus pleuropneumoniae loaded in the MPs groups. The findings also demonstrated that the microspheres reached the lungs and secondary lymphoid tissues and induced systemic IgG and secretory IgA responses to the ApxIIA in bronchoalveolar lavage (BAL) and nasal and vaginal washes. Although the immunostimulatory role of mannosylation in enhancing immunogenicity has been reported [Similar results have also been achieved using the dual immunostimulant and mucoadhesive capacity of mannan isolated from the cell wall of revisiae . Mannansreported ,134, in Bacillus anthracis protective antigen [Vyas and his team (2014) assessed the coating of CS MPs with alginate (A-CSMp). In contrast to most of the works reviewed up to this point, where the positive surface charge plays a fundamental role in adhesion to mucin, alginate as an anionic polyelectrolyte changes the ζ-potential of the particle to an electronegative value (−29.7 mV). FITC-BSA was rapidly washed from rat jejunal tissues; however, the in vitro retention time in the mucosa was prolonged when FITC-BSA was associated with A-CSMp. In the same way, in the in vivo assays, only A-CSM loaded with FITC-BSA successfully generated uptake by M cells in Peyer’s patches. When evaluating the efficacy of the particulate system in an oral anthrax vaccination model, high-titer anti-PA serum IgA and IgG antibodies were observed in animals receiving particles loaded with antigens compared to the free  antigen .Similarly, Saraf et al. loaded alginate-coated CS NPs (ACNPs) with HBsAg anchored to E. coli EH-100 lipopolysaccharide (LPS) (LPS-HBs-ACNPs) as an adjuvant for oral administration. As expected, the alginate coating changed the ζ-potential of the NPs from +45.2 mV (0.5% CS-0.1% TPP) to −26.2 mV  due to the negatively charged -COO- electrostatic interaction of the alginate on the positively charged -NH3 of the CS. Despite the ζ-potential’s more negative values, in vitro mucoadhesion studies showed that alginate-coated NPs were more mucoadhesive than CS NPs alone. Although anti-HBsAg serum IgG titers were higher for HB-ACNPs after oral administration, sIgA antibody titers in mucosal secretions were higher for LPS-HBs-ACNPs. The anchoring of LPS targeted the NPs to M cells, conferring immunogenicity to the system  independOn the other hand, sodium alginate protects the NPs from the hostile environment of the gastrointestinal tract, the same as the introduction of hydrophilic groups, such as hydroxyalkyls, carboxyalkyls, succinyls, and thiols, or polymer grafts, such as PEG. In this way, Amin and Boateng (2022) proposed a system based on OVA-loaded CS NPs coated with sodium alginate or PEG for oral vaccine administration. Both sodium alginate and PEG coatings increased the stability of NPs upon exposure to gastric fluids with the protection of the encapsulated protein , compared to uncoated NPs (<30 min). After transfer into simulated intestinal fluid, both coatings showed stability for 120 h, although with different release profiles of OVA. Increased alginate concentrations were related to a higher level of mucin binding. According to the authors, the alginate coating ensures stability, allows a higher antigen load to reach the site for mucosal immune response, improves mucoadhesive properties, and enhances the sustained release of antigen-loaded NPs .Quillaja saponins to vaccine formulations against brucellosis. While Brucella LPS was weakly immunogenic, when B. abortus LPS-loaded XG NPs were administered nasally to Balb/c mice, higher systemic and mucosal IgG antibody levels and mucosal IgA were induced. Increased immunogenicity was associated with a greater mucoadhesion force of the XG and the LPS-XG NPs compared to the LPS alone, as well as the ex vivo retention of LPS-XG NPs over 24 h in goat mucosa [Xyloglucan (XG), a non-anionic polysaccharide and the main hemicellulose component, has been applied with t mucosa .As in the case of CS, XG has been previously used in pharmaceutical applications in different formulations and by different routes, including mucosal, transdermal, and intraperitoneal, due to its biodegradability, cost-effectiveness, and non-toxicity. Some authors have suggested that the XG molecular structure, “mucin-like,” is responsible for mucoadhesive properties, including swelling capacity and increasing concentration-dependent viscosity ,143. All®, a highly cross-linked hydrophilic polymer, which provides it with mucoadhesive and viscoelastic properties. Coucke et al. used spray-dried powders of amylopectin (Amioca®) with polyacrylic acid (Carbopol® 974P) in different proportions  for the intranasal administration of H3N2-inactivated influenza virus and in combination with the LTR192G adjuvant in rabbits. The formulation SD25/75 induced the highest serum response of IgG anti-haemagglutinin compared to the formulation SD100/0, thus highlighting the importance of polyacrylic acid. Despite this, neither SD25/75 nor SD0/100 induce a local mucosal response. The immune response was directly related to the negative ζ-potential of Carbopol® 974P and the mucoadhesive properties of the formulations. The reticulated, predominantly elastic, or highly structured characteristics of SD25/75 (G’ >> G’’) increased the residence time in the nasal cavity. In contrast, the lowest viscosity and cross-linking of SD100/0 were associated with a low mucosal retention [Poly (acrylic acid) and its derivatives have excellent mucoadhesive capacity compared to cellulose, polycarbophil, chitosan, and pectin ,146,147.etention .The poly-γ-glutamic acid (γ-PGA)-based vaccine adjuvant, an anionic biopolymer, was used for the intranasal delivery of the influenza fusion protein sM2HA2 and OVA, co-administered with 3-O-desacyl-4′-monophosphoryl lipid A (MPL) and QS21 in a system denominated γ-PGA/MPL/QS21(CA-PMQ). Using in vivo single-positron-emission computed tomography imaging, it was possible to determine that γ-PGA increased the OVA residence time by up to 12 h in the nasal cavity. This signal decreased at 6 h when OVA was administered alone. This result is correlated with the higher serum IgG, IgG1, and IgG2a antibody responses in the groups vaccinated with OVA/CA-PMQ and sM2HA2/CA-PMQ compared to the groups that received OVA and sM2HA2 alone, as well as being superior to that induced by the cholera toxin used as a mucosal adjuvant. Likewise, animals vaccinated with the antigen/CA-PMQ induced more IL-4 and IFN-γ–secreting cell populations in the spleens stimulated with OVA, sM2, and HA2 protein than mice immunized with proteins alone or the control group. Additionally, the CA-PMQ induced high titers of sM2HA2-specific IgA antibodies at the administration and distal sites, along with an increased survival time (80–100%) following the challenge with influenza A subtypes and cleared pulmonary viral titers . The preThe anionic model (−35.5 mV) of Kurosaki et al. with benzalkonium chloride (BK) and γ-PGA NPs in a complex with OVA (OVA/BK/γ-PGA) was used for pulmonary administration. They observed an increased fluorescence intensity in the lung (Alexa647-OVA/BK/γ-PGA) indicative of lung deposition compared to Alexa647-OVA. OVA/BK/γ-PGA increased the levels of specific IgG antibodies, while in the animals that received OVA or the vehicle (BK/γ-PGA), anti-OVA IgG was not detected. The induction of immune responses at the mucosal site was also significantly higher in the OVA/BK/γ-PGA group . Their sThe previously reviewed polymer-based adjuvants could be thiolated to improve mucoadhesion. In the past two decades, important research has been conducted using thiolated polymers or so-call “thiomers”, mainly in excipients for drug delivery. Thiomers can interact with mucin through disulfide bonds with the cysteine-rich subdomains of mucus glycoproteins . These cUsing a tensile test and rotating cylinder method to obtain compressed tablets, Roldo et al. demonstrated that increasing the number of thiol groups covalently attached to chitosan-4-thio-butyl-amidine conjugated significantly improves mucoadhesion compared to unmodified CS. Thiolation increased the total adhesion work  up to 100 times . SimilarIn a recent study, Sinani et al. immunized Balb/c mice with BSA-loaded NPs prepared using aminated CS (aCS) and aminated and thiolated CS (atCS) polymers; mice were nasally immunized at 14-day intervals. At the end of the experiment (day 253), the nanoparticles (aCS and atCS) induced a more robust systemic response, resulting in an almost two orders of magnitude higher systemic IgG titer than the BSA/CpG ODN control, with atCS being the best. These results are correlated with the increased mucoadhesion observed in the aCS and the atCS. Both aCS and atCS modulated the Th2 immune response and enabled immune response at distal mucosal sites .Cellulose acetate phthalate (CAP) is widely used as an enteric coating for pharmaceutical dosage forms due to its solubility at pH values above 6 (such as in the intestines) but poor water solubility at a low pH (such as in the stomach). After exposure to intestinal fluids, the polymer swells, with the subsequent softening or complete dissolution of the phthalate, allowing the release of the biologically active compounds . Lee et For cationic thiomers such as atCS, the interactions are predominantly driven by electrostatic forces. In contrast, for anionic thiomers, such as T-CAP, interaction with the mucus occurs through hydrogen bonds, van der Waals interactions, and chain entanglement. In both cases, the bioavailability is improved by the extension of the residence time . NotablyFurther evidence has shown that thiomers are susceptible to thiol oxidation at pH ≥ 5, with their effectiveness being reduced following oral administration. Typically, thiol groups (R-SH) can form disulfide bonds with mercaptopyridine substructures, whereby thiol groups are stabilized against oxidation and increase their reactivity. S-protected thiomers, so-called “pre-activated”, have shown greater mucoadhesion than unprotected thiomers, according to Iqbal et al. (2012) . In thisAlthough mucoadhesive molecules improve the bioavailability of drugs and antigens administered via the mucosa, the mucus layer still limits passage into the epithelium. The transit time of particles in the mucosa is determined by the physiological renewal time of the secreted mucus layer . Mucus tTherefore, polymer-based adjuvants are expected to adhere to the mucous layer, penetrate the epithelium, and reach the inductive sites for mucosal immune responses before being removed. Hence, this section briefly describes the strategies to facilitate mucus barrier penetration and improve the permeability of polymer-based adjuvants once they are in the mucosa.Particles with a low adhesion and small size, thus with few steric hindrances to the mucin network, are often referred to as mucus-penetrating particles. Unlike mucoadhesive particles, mucus-penetrating particles seek to minimize the strength of electrostatic and hydrophobic interactions with the mucin. Polymers with neutral or low positive charges are generally included in the design of mucus-penetrating formulations. Several studies have reported the surface coating of particles with PEG. PEG is used as an adhesion promoter acting at the interface to improve adhesion. Hence, PEG chains tethered or grafted are covalently attached at one end on the polymer surface while the other is free, allowing PEG to diffuse from the polymer network to the mucus and enhancing interpenetration . Wang etDespite its widespread use in over-the-counter drugs and vaccines, recent approaches suggest that PEG is not immunologically inert ,166,167.Some works have also raised doubts about coating particles with PEG due to surface modifications that can alter the linked polymers’ physical and biological properties. Bamberger et al. evaluated the effects on APC response after functionalizing spermine NPs with acetylated dextran (Sp-Ac-DEX) through a process called DEXylation and PEGylation. The average particle size was considerably increased by DEXylation, with subsequent aggregation. PEGylation and DEXylation decreased the primary amines and, therefore, the ζ-potential. This was reflected in the 20% reduction in the cell viability of bone-marrow-derived dendritic cells and macrophages treated with DEXylated NPs, whereas PEGylation treatment increased viability by 10–20% compared to unmodified NPs. However, the binding and cellular uptake of surface-modified NPs was lower in PEGylated particles .®. These block copolymers consist of hydrophilic poly (ethylene oxide) (PEO), and hydrophobic block-poly (propylene oxide) (PPO) ordered in an A-B-A triblock structure: PEO-PPO-PEO [Brucella spp., along with the reduced excretion of Brucella ovis [® (PF127) and Gantrez® AN119 thermally sensitive hydrogel for intranasal vaccine delivery since the hydrogel increases the residence time of the antigens in the nasal epithelium, allowing their penetration into the deep skin layers of the nose thus reaching the submucosa, where they can trigger an immune response [Other polymers with mucopenetration ability are poloxamers, also known as Pluronic-PPO-PEO ,173,174.lla ovis . Pastor response .® mucopenetrates by the collapse of the mucus mesh [Another type of mucopenetrant includes nanoemulsions (NEs). Di Cola et al. evaluated PEG-coated O/W NEs with emulsified, added CS as a proposal for the nasal administration of drugs or vaccines. They observed that CS-added NE led to a local shrinking of the mucin gel network, forming larger pores between the mucin bundles. This phenomenon does not occur in the absence of CS. The SAXS  monitoring of the penetration of solute CS-added NE into the PGM showed a higher diffusion over time (20 min) through the mucus mesh. SANS (neutron scattering) confirmed that, unlike the steric hindrance caused by the pore-like size of mucus caused by mucoinert NPs, the CS-added NE based on Solutolcus mesh .Coating dextran particles with mucopenetration properties have also been explored to improve drug administration performance ,180,181 Helicobacter pylori, coated with a cell-penetrating peptide, and coated with PEG derivative. NPs were transported transepithelially, improving the systemic and mucosal antibody response and the protection against H. pylori after the challenge [An additional consideration for the design of polymer-based particles is passing through the second barrier, the epithelial cell membrane. The permeability of peptides, proteins, and drugs is often deficient. In this sense, absorption enhancers have been developed, which, in addition to preventing enzymatic degradation, facilitate the opening of the epithelial barrier and improve absorption through intracellular or paracellular mechanisms . Absorpthallenge . It willFor several decades, many polymer-based particles have shown promise as potential human mucosal vaccine adjuvants due to their biodegradability, biocompatibility, and nontoxicity characteristics. Added to this is the extensive study of the adjuvant mechanisms of particulate systems. However, in the mucosa, the mucin networks that cover the compartments are often considered a barrier for the particles, so the mucoadhesive and mucopenetrating capacity of the polymer-based particles often defines their adjuvant mechanism of action.The search for polymers with better mucoadhesive properties, regardless of the polymer’s source, but focusing on the physicochemical characterization of polymeric particles and the contribution of these properties to mucoadhesion, will allow the rational design of mucosal vaccines. However, it is not an easy task because, on the one hand, the smallest nanoparticles are the most mucopenetrating. Still, on the other, there is a lack of studies that suggest an ideal surface charge or a hydrophobicity that favors adhesion. At the same time, it cannot be ignored that there are multiple other cellular mechanisms to elicit the immune response triggered by the polymeric particles, i.e., enhanced antigen uptake, immune cell presentation and recruitment, and traffic to lymph nodes.Studies demonstrating the correlation between the observed immune response, the physicochemical characteristics, the mucoadhesion, and the mucopenetration ability are scarce. More studies that examine all these factors simultaneously are required to position mucoadhesion as another immune response mechanism necessary for designing more efficient polymer-based particulate adjuvants.The COVID-19 pandemic highlighted the need for mucosal vaccination as an effective strategy to eradicate infectious diseases that have the mucosa as a natural route of infection. Mucoadhesion is probably the most important feature to improve local and systemic immune responses since, by prolonging the residence time of particulate polymers in mucosal tissues, the absorption and sometimes penetration through the mucosal epithelia are allowed and improved. In this sense, studying the physicochemical characteristics of the polymeric particles used as mucosal vaccine adjuvants and how they affect mucoadhesion is crucial to developing new mucosal vaccines.


Anti-TAA antibodies were not generally related to ANA, a common marker of systemic autoimmunity. Associations of ANA with particular antigens inducing autoimmunity prior to breast cancer warrant further investigation.Autoantibodies to tumor-associated antigens (anti-TAA) are potential biomarkers for breast cancer, but their relationship systemic autoimmunity as ascertained though antinuclear antibodies (ANA) is unknown and warrants consideration given the common occurrence of autoimmunity and autoimmune diseases among women. The relationship between anti-TAAs and ANA among women who were later diagnosed with breast cancer and others who remained cancer free in the Women’s Health Initiative cohort. The study sample included 145 post-menopausal women with baseline ANA data. A total of 37 ANA-positive women who developed breast cancer  were matched to a random sample of 36 ANA-negative cases by age and time to diagnosis. An age-matched control sample was selected including 35 ANA-positive and 37 ANA-negative women who did not develop breast cancer . Baseline sera were assessed for Immunoglobulin G (IgG) antibodies, measured by custom microarray for 171 breast and other cancer-associated TAA. We used linear regression to estimate cross-sectional associations of ANA with log-transformed anti-TAA among cases and controls. Most anti-TAA did not vary by ANA status. Two anti-TAA were elevated in ANA-positive compared to ANA-negative cases: anti-PGM3 ( A growing body of research has sought to identify pre-clinical cancer-associated autoantibodies as potential biomarkers . A diverAntinuclear antibodies (ANA) are a common marker of systemic autoimmunity found in 15–20% of women in the general population . ANA areafter serum collection), and in age and ANA-matched “controls” who did not develop breast cancer during follow-up. Serum specimens, obtained from a prior study of women with self-reported rheumatoid arthritis screened for ANA, were tested against a custom array of anti-TAA identified in prior studies [Using data and samples from the WHI, we explored the association between ANA and anti-TAA antibodies in baseline sera obtained from women who later developed breast cancer  at enrollment or follow-up  and time to diagnosis (±1 year). Sample eligibility required complete data on hormone therapy, parity, estrogen-receptor tumor status and adequate serum volume (<200 μL). Incident breast cancer diagnoses were adjudicated on 96% of cases. Three cases with a positive ANA had a cancer history at baseline . Other cancers identified at follow-up  were equally distributed in ANA-positive and ANA-negative cases.To evaluate the relationship of ANA with anti-TAA in women without known breast cancer, a random sample was selected of 35 ANA-positive  and 37 ANA-negative women who did not develop breast or any other cancers during follow-up , matched by age (±1 year) and in a time-forward manner, to ensure controls had at least as much follow-up time as cases.The study was approved by IRB boards at the National Institute of Environmental Health Science and Fred Hutchinson Cancer Research Center. Existing samples and deidentified data were obtained from the WHI, which obtained informed consent from all participants.Antinuclear antibodies (ANA) were measured on all specimens as previously described . In brieAutoantibodies to tumor-associated antigens (anti-TAA) were assessed quantitatively using a custom array, including recombinant proteins that have previously elicited an autoantibody response to breast and other common cancers ,19,20,21Sixteen samples were hybridized on each slide at a dilution of 1:150, and a secondary antibody reaction with a fluorescent anti-human IgG  was performed. Then, the protein arrays were scanned with a GenePix scanner  using a red laser (635 nm), and local background-subtracted median spot intensities were generated using GenePix Pro 7 Software. Normalized values subtracted the log transformed anti-TAA (Log2TAA) median array intensities from individual intensities  to account for variability between arrays. Valid levels were detected for most antigens, with low detects reflected by a Log2TAA value of zero. There was no significant difference in zero values by case and ANA status . Correlat-test p-value of 0.05 for the mean value and a median ratio value of <0.90 or >1.10. A total list of all anti-TAA means ratios, p-values, and median ratios, are shown in p = 0.0058 for statistical significance was determined by the number of total comparisons (0.05 divided by 171).Differences in anti-TAA by ANA status were examined separately in women diagnosed with breast cancer during follow-up (cases) and in women who were not diagnosed with breast or other cancers during the same time (controls). Initial descriptive analyses visually explored the average of median Log2TAA values for each woman, stratified by ANA and case status. Ratio values comparing median anti-TAA values in ANA-positive versus ANA-negative women, stratified by case status, were calculated, and screened against the null  using a Using linear regression, we examined ANA as a predictor of anti-TAA intensities for those antigens that met the screening threshold . Models were run separately in controls and cases, and in cases, models were also stratified by time from serum collection to diagnosis (<7 vs. 7+ years) in cases. We first adjusted for age (Model 1) and then added EBNA, RF, and current hormone replacement therapy . Sensitivity analyses excluded women who were currently using DMARDS, cases with a history of other cancer at enrollment or during follow-up, or reported a diagnosis of systemic lupus erythematosus, and limited to women ER-positive cancers. Because age is an important determinant of both ANA and cancer risk, we explored the consistency of our main findings in models stratified by age (younger than 65 years and 65 and older). Zero values were included in regression models of continuously distributed Log2TAA. In sensitivity analyses, we imputed values as half the lowest detected value. We did not test for statistical interactions in these exploratory models.Visual inspection of overall anti-TAA antibody reactivity  showed a broad distributed that did not appear to vary by ANA status in either cases or controls . Comparip < 0.05; p = 0.004; TTN, p = 0.005. Similar, non-significant elevations of these anti-TAA with ANA were seen in controls. Two other anti-TAA were higher in ANA-positive women (DUSP26 in cases and PRKD1 and UCLH1 in controls) and three were lower , only in cases.Of all anti-TAA screened . In age-stratified models, the association of TTN levels with ANA was most apparent in younger  but not older cases . A sensitivity analysis with imputed values for two Log2TAA zero values (PRKD1 and UCHL1) showed no substantial differences  and with breast cancer. Although a few differences in specific anti-TAA levels were seen, depending on ANA status among controls (PRKD1 and UHL1), no significant differences remained after adjusting for multiple comparisons. Among cases, however, significantly higher levels of two specific anti-TAA (PGM3 and TTN) were observed in ANA-positive compared to ANA-negative women , and in The association of ANA with anti-TTN TAA levels was stronger in women whose cancers were diagnosed within seven years, which suggests a temporal relationship of these autoantibodies in the development of breast cancer. Titin (encoded by the TTN gene) is a very large protein, which plays an important structural, developmental and regulatory role in skeletal and cardiac muscle ,24. NotaOur initial screening comparisons also showed that ANA was positively associated with anti-PGM3 TAA levels in cases, and an elevated but non-statistically significant association was seen for anti-PGM3 and ANA in controls. Subsequent adjusted models suggested modest associations of similar strength in both cases and controls. Phosphoacetylglucosamine mutase (PGM3) is one of several paralogues  that control glucose metabolism in most cells , and defp = 0.0058; however, some findings were suggestive. For example, ANA was associated with elevated levels of anti-PRKD1 TAA in controls, but negatively associated in cases at least 7 years prior to diagnosis. Protein Kinase D1 (PRKD1) is a serine/threonine kinase expressed in normal breast tissue, where it is responsible for maintaining the epithelial phenotype [In our overall analyses, no other associations reached statistical significance at the Bonferroni threshold of henotype . Methylahenotype . It is pPrior studies suggested levels of breast cancer-specific anti-TAA may be lower in current users of hormone therapy, especially estrogen , but we In addition to the example described for TTN, myasthenia gravis, and thyoma described above, cancer-associated autoimmunity has been found to occur across a range of diseases, such as myositis and scleroderma, considered to be a paraneoplastic phenomenon . Recent Cancer immunoediting has been proposed to play a role in immune surveillance and control of growing tumors . In the Antinuclear autoantibodies associated with clinical autoimmune diseases may have different characteristics, causes and consequences, compared to those seen in the general population. Autoimmune patients tend to have higher titers or disease-related autoantibodies to complexes of proteins with DNA or RNA , and arep-value adjusted for multiple comparisons (0.0058). Because analyses of smaller sub-samples can introduce bias through effect size magnification, we did not consider all 171 anti-TAA with respect to age or other potential modifiers. Our results support the development of larger studies to provide adequate power for stratified analyses.This study has limitations, especially given the relatively small sample, which limited sub-analyses. However, the focused design allowed us to address our research question overall and in the context of future breast cancer. Our TAA assay included many specific antigens, increasing the number of comparisons and the possibility that associations could arise due to chance. Therefore, we highlighted only those results that would have been statistically significant at a Our study sample was drawn from a larger sample of women who reported rheumatoid arthritis (RA) who had existing data on ANA. The proportion of clinical RA and lupus in our study sample is likely to be low, given the poor specificity of self-report ,45 and lThe current study was not intended to investigate the utility of anti-TAA antibodies in breast cancer screening, given the extended time between serum collection and diagnosis (average of 7 years). Results may have been different for samples taken closer to the time of diagnosis. While there is some evidence of an association of ANA with breast cancer in clinical studies , this qIn conclusion, our findings that ANA positivity was not broadly associated with higher TAA reactivity suggest that ANA are unlikely to confound studies of tumor autoantibodies and cancer. However, observed associations of ANA with anti-TAA in women who later developed cancer suggest that cancer-associated adaptive immune responses may exist to years prior to the development of clinically apparent tumors, and support the development of larger prospective studies with repeated measures to investigate autoimmunity and pre-clinical cancer.


Stunting has been a major concern in sub-Saharan Africa. However, littleevidence exists on the spatiotemporal variations in under-five stuntingwithin a national context.This paper examines the spatiotemporal variations in under-five stunting anddeterminants using data from the Ethiopia Demographic and Health Surveys(2000-2016).Spatial autocorrelation and multilevel logistic regression models were usedto conduct the analyses.The stunting prevalence has decreased from 51% to 37%, while the prevalenceof severe stunting has decreased by more than half (from 28% to 12%). Wideregional variations in stunting have been consistently observed over theyears, which exhibited a higher level of stunting in Tigray (48%), Afar(42%), and Amhara (42%). The results show considerable local and regionalvariations in under-five stunting levels with diverse patterns ofimprovements in regional stunting levels over time. Stunting levels wereassociated with child-level factors such as the sex of a child, birth size,age of a child, birth order, preceding birth interval, and place of birth.Maternal educational attainment, nutritional status, household wealth,toilet facility type, and place of residence were linked to under-fivestunting. The regional-level infant mortality rate was associated withunder-five stunting.Specially tailored policies and interventions should be devised to addresspersistent spatial inequalities in stunting by focusing on higher riskpopulations. Child stunting levels have shown steady improvement globally, except inAfrica where improvement has slowed in recent years.2 Consequently, the World HealthOrganization’s (WHO) global target of a 40% reduction in the number of stuntedchildren by 2025 is unlikely to be met. In Ethiopia, stunting cases among under-five childrenhave decreased from 58% in 2000 to 38% in 2016, a reduction of about a third, which leaves more work to be done.Childhood stunting (low height-for-age) adversely affects the survival, growth, anddevelopment of children, impacting 149 million (21.9%) under-five children worldwide. To achieve the 2025 global target, WHO suggested that situational analysis beconducted to uncover geographic variation in childhood stunting and its underlyingcontext-specific causes. Multiple studies have shown spatial and temporal differences in childhoodundernutrition and its determinants in sub-Saharan Africa.7 Substantial spatiotemporalvariations in chronic malnutrition among under-five children have been found in Ghana. Additionally, regions within the country have been identified as high-riskareas for chronic malnutrition. In Cote d’Ivoire, temporal changes in stunting among under-three childrenbetween 1994 and 2011 have been documented, along with spatial differences at thenational and subnational levels. These previous studies have indicated that age, sex, household wealth,maternal education, and maternal body mass index are significantly associated withunder-five stunting,68 butregional-level factors were not examined.Several long-term adverse effects including poor health, limited cognitive, physical,economic, and reproductive performance among others, have been widely linked tochildhood stunting.810 These studiesdid not consider the trends of stunting prevalence at the local and regional levelsover time, as they were based on only one survey year while another focused on a specific part of the country and, therefore, hadno national representation. Besides, none of these studies examined individual-level factors incombination with aggregate-level regional variables across regions and overtime.Few previous studies have, however, considered the spatial aspect of stunting amongchildren in Ethiopia.This study fills these literature gaps by drawing on nationally representative datato conduct a spatiotemporal analysis of under-five stunting in Ethiopia. The studyis guided by 2 main research questions: (1) Are there spatial variations in childstunting in Ethiopia and do they change over time? (2) Are spatial and temporalvariations in child stunting explained in part or in full by individual and regionalcontextual factors? The spatiotemporal analysis is relevant because it may have anadditional advantage over separately analyzing spatial or temporal trends. Theanalysis identifies local areas of high risk, illuminates any unusual patterns andpersistence of undernutrition disparities or clustering over time, and ultimatelyhelps to direct tailored programs and interventions to address stunting disparities.The statistical analysis also provides a better understanding of theindividual-level and regional contextual factors associated with spatiotemporaldisparities in under-five stunting in Ethiopia.1113 These data sets arecomparative and provide a reliable source of maternal and child health dataincluding height-for-age (stunting) data at the individual level. EthiopianDemographic and Health Surveys are nationally representative data that used amultistage sampling procedure where respondents were selected from households inrandomly selected clusters in both rural and urban settings in the country. TheNational Ethics Review Committee of Ethiopia reviewed and approved the EDHSprotocol. The study sample comprised 8890, 4363, 10 222, and 9341 children under5 years of age for the years 2000, 2005, 2011, and 2016, respectively, for atotal sample of 32 816.This study draws on pooled cross-sectional data from the Ethiopian Demographicand Health Surveys (EDHS) (2000-2016) which are part of the global demographicand health survey (DHS) series that are collected at regular periods. We use the Z-scores of height-for-age in the dataset to create adichotomous measure  based on these standards.The outcome variable is stunting as determined by height-for-age among childrenunder the age of 5. Height-for-age is a measure of linear growth retardation andcumulative growth deficits. Based on the WHO growth standards, children whoseheight-for-age Z-score is negative 2 standard deviations or more (−2 SD) fromthe median of the reference population are considered stunted.1518 Thus, variable selectionand measurements are mainly supported by the existing literature. The sex of achild  was included in the study, while the age of a child wasmeasured in months as < 12, 12-23, 24-35, 36-47, and 48-59. Child size atbirth was measured in 3 categories such as small, average, and large. Thepreceding birth interval was measured as < 2 years, 2-4 years, and > 4years, while birth order number was measured as 1 to 2, 2 to 4, and 5+. Child’splace of delivery was measured as a home or health facility.The study examined numerous individual-level sociodemographic predictors thatwere selected based on their significant association with under-five stunting inthe extant empirical literature.Several maternal factors were as well included in the study. Maternal age atbirth was measured as < 20 years and 20 years or more, and maternaleducational attainment was measured as no education, primary education, andsecondary education or higher. Maternal employment status was recoded asemployed or not employed, while maternal nutritional status was recoded asunderweight or not underweight, and maternal breastfeeding status was measuredas ever breastfed or never breastfed. Household wealth status was based on awealth index collapsed into poor, middle, and rich, while the source of drinkingwater and type of toilet were both measured as improved or unimproved. Ruralversus urban places of residence were also included in the study.Also, some regional aggregate level factors were included in the study to assessregional effects on stunting at the child level. We considered the medianduration of exclusive breastfeeding by women in the region, the percentage ofchildren with diarrhea who received an oral rehydration solution or recommendedhomemade fluids treatment, the percentage of married women with an unmet needfor family planning, the region’s infant mortality rate, and the percentage ofhouseholds in the region with electricity. These aggregate-level variables weregenerated from the DHS for aggregate-level analysis and are all included ascontinuous measures. Two main analyses were performed. The first level comprised the analysisof the background characteristics of respondents and the spatialtemporalvariations in stunting using geospatial methods. In the spatial data analysis,DHS global positioning system (GPS) data points were used to generate spatialweights at the primary sampling unit (cluster) level using the 3-nearestneighbors rule for each of the survey years. Voronoi polygons were used tocreate buffers around neighboring GPS points. We used the 3-nearest neighborrule as it provided the best clustering patterns during the exploratoryanalysis. The spatial weights were attached to the study data to calculateMoran’s I statistic to assess potential spatial autocorrelation of stunting atthe cluster level. The Moran’s I analysis generated positive results indicatingspatial clustering of stunting. Local Moran’s I analysis was further conductedto generate Z-scores whereby positive values represent “high” clustering whilenegative values represent “low” clustering of stunting. The results were thenattached to shapefiles and plotted as cluster maps showing hot spots and coldspots of stunting for all the years. A logistic regression model was used toproduce regional estimates of stunting which were used to create regional mapsand geometric lines to visualize the spatial and temporal trends ofstunting.The R statistical package (version 3.6.3) was used for all data processing and analysis.For the second analysis, multilevel logistic regression was used to examine theassociation between individual and region-level predictors on under-fivestunting. We specified the models at two levels, where children at theindividual level were nested within regions (higher level hierarchy) over time,taking into consideration both individual and aggregate-level predictors. Thiscan be mathematically specified as:j represents the region of residence of the children,while t. The Year term is theoverall time trend within the country while the Where Three models were estimated. As a baseline model, Model 1 was specified toexamine the temporal effect on stunting among the regions by using only timerandom slopes and the regional level random intercepts without the individualand region-level predictors. Model 2 adjusted for individual-levelsociodemographic factors, while Model 3 further controlled for regionalaggregate-level predictors. Due to the multistage sampling procedure used forselecting respondents, the analysis was weighted using complex survey weightscreated by nesting the primary sampling unit (clusters) and the strata(residence) in the regions to address potential over-sampling and under-samplingin the study sample and reduce biased estimates. The model parameters were usedto calculate odds ratios (OR) and 95% confidence intervals (CI).Children of higher birth order , children with a precedingbirth interval of less than 2 years (48.6%), and children born at home (43.7%)had significantly higher levels of stunting compared to their respectivecounterparts. There were also significantly higher stunting levels amongchildren whose mothers had no education (44.8%), were underweight (45.0%), everbreastfed (41.7%), and lived in a poor household (44.4%). Children in householdswith unimproved drinking water (44.8%) and toilet facility (42.7%) also hadconsiderably higher stunting levels than their counterparts with improvedconditions . Additionally, children who residedin rural areas (43.1%) were found to have higher stunting levels than theirurban counterparts. High levels of regional variation in stunting are observed,with the highest levels of stunting found among children residing in the Amhara(49.4%), Tigray (44.0%), and the Southern Nations, Nationalities, and Peoples’Region (44.0%), and the lowest prevalence (17.1%) found in Addis Ababa.P <.0001), 0.47 (P < .0001), 0.48 (P <.0001), and 0.49 (P < .0001) for the years 2000, 2005, 2010,and 2016, respectively, suggesting significant spatial clustering of stunting.The analysis generated Moran’s I statistics of 0.46  more likely to bestunted compared to female children. Average-sized and larger-sized children at birth  hadsignificantly lower odds of being stunted compared to the smaller size childrenat birth. The odds of stunting increased significantly with the age of thechild, with the odds of stunting being 6.83 times among older children ages36-47 months compared to children aged < 12 months (95% CI: 6.06-7.71). Theresults also showed that the odds of stunting increased significantly for higherbirth order children, such as order 3-4  and order5+  compared to first or second-born children,while the odds of stunting significantly decreased with longer preceding birthintervals  compared to the shortest birth interval (< 2 years). Relativeto home-delivered children, children delivered at a health facility had 10%lower odds of stunting (95% CI: 0.83-0.97).Compared to children whose mothers had no education, children whose mothers hadhigher education levels with primary education or a secondary level of educationhad a least 10%  or 44%  lower odds of being stunted, respectively. Children of underweightmothers had 13% higher odds of being stunted compared to children of mothers who were not underweight. Children from themiddle-income  and rich households  had lower odds of being stunted compared to children from poorhouseholds. Similarly, children from households with unimproved toiletfacilities had higher odds  of being stuntedcompared to households with improved toilet facilities. Children in urban areashad at least 18% lower odds  of stunting comparedto children who reside in rural areas. At the regional level, only infantmortality rate was associated with under-five stunting, with the odds beingsignificantly higher in regions that had above-average infant mortality rates inthe country over the study period . Papua New Guinea, India, and Ghana. Generally, there is notable evidence of a steady decline in the regionallevels of under-five stunting for the 15-year study period although the levelsremain high. Diverse patterns of improvement in the stunting prevalence occurred inthe various regions over time. Despite the observed temporal improvements, there wasevidence of a north-south divide in stunting levels which mainly disadvantaged thenorthern states  and to the advantage of thesouthern and the central part of the country, particularly the Addis Ababa Regionwhich is also the capital city of the country.The current study presents findings of a spatial and temporal analysis of under-fivestunting using a nationally representative sample of children under 5 years inEthiopia. We found several clusters of both high and low stunting values at thelocal cluster level over time, suggesting considerable disparities in stunting withmost of the hot spots existing in the northern part of the country. This may beexplained by the recurrent drought and poor sociocultural feeding habits in theseregions. Evidence of spatial variations in under-five stunting has also beenestablished in various countries including Uganda, According to the joint child malnutrition estimates, when the prevalence ofstunting becomes 30% or more, it is considered very high or critical. Therefore, our observed prevalence of stunting in these regions  has been in a serious range over the studyperiod (2000-2016). A study indicated that conflict-driven displacements in Afar andTigray regions have been aggravating hunger and malnutrition rates up over the years. Also, intensified conflict on the Tigray-Afar border in recent days isexpected to force more communities from their homes and deeper into hunger. The same study also indicated that there is a higher shortage of safe andadequate water supply and limited access to health services in these regions inregions of Tigray and Afar regions, which ultimately contributes to the higher level of stunting among under-fivechildren in these regions. A study has linked stunting to food diversity and thenumber of meals a child ate per day and being underweight in the Amhara region. Consequently, child nutrition intervention strategies should seriouslyconsider food security, and dietary diversity, and be specifically targeted toresidential locations.The prevalence of stunting in Ethiopia was estimated to be 37% in 2019; in the sameyear, the percentage of stunting was higher in Amhara (41%), Benshangul-Gumuz (41%),Tigray (48%), and Afar (42%) compared to the national estimate. A recent study by UNICEF in the Benshangul-Gumuz region discovered thatdespite the decline in food poverty from 55% in 1999/2000 to 24% in 2015/2016, only1.1% of rural households were in the Productive Safety Net Programme compared to 11%of households at the national level. There is also some evidence of an effect of poor dietary patterns andpractices (in younger age groups), and the absence of good water and sanitationpractices and facilities in the Benshangul-Gumuz that contribute significantly tochildhood stunting. The 2019 EDHS data also shows that only 2% of households use improved (notshared) sanitation facilities in Benishangul-Gumuz.In the Benshangul-Gumuz region, there has been evidence of a high multidimensionalchild deprivation index in which 89% of children were found to be deprived in 3 to 6dimensions as well as a high prevalence of food poverty (24%). Older children (ages 24-59 months) were the most at-risk age-group forstunting, as previously reported by various studies.31 Thus, timely initiation ofsupplementary feeding to infants is important to meet their changing nutritionalrequirements to avert any adverse nutritional status later in life. Also, childrenwith smaller birth sizes were more likely to be stunted before age 5 than average-and larger-sized children similar to findings by several studies in Nepal, Nigeria, and Tanzania, which may be driven by a lack of nutritional supplements during pregnancy andlater after birth. Additionally, higher birth order was associated with higher oddsof stunting,35 which may follow from an additional birth placing an economicstrain on households, leading to adverse nutritional status.Some sociodemographic characteristics were found to be associated with the observedspatial and temporal disparities in stunting. Our findings have linked male childrento higher odds of stunting which was previously reported by a study conducted insub-Saharan Africa that suggests that boys are more likely to be stunted than girlsbecause male children may be more vulnerable to health inequalities than femalechildren in the same age-group.38 which may have adverse nutritional implications for the child. Adequately spaced children may have the necessary nutrition for growth anddevelopment and a strong immune system, thereby reducing the likelihood of childhood undernutrition. Home-born children were more likely to be stunted than those born at a healthfacility and, thus, access to a health facility may play a crucial role in reducingthe stunting disparities. Various studies have linked improved access to skilleddelivery care to enhanced maternal and child health42 likely because mothers may beprovided with the relevant information and an understanding of health practicesneeded to improve their nutritional status and that of their children. As with thepresent study, children of richer households38 and higher maternaleducational attainment4347 have been linked to reducedodds of stunting emphasizing the importance of higher socioeconomic characteristicsas a protective factor against adverse child nutritional outcomes. Also, children ofunderweight mothers were more likely to be stunted than children of normal oroverweight mothers, providing support for previous reports of an association betweenmaternal body mass index and odds of under-five stunting.4851Moreover, a short (less than 2 years) birth interval was associated with higher oddsof stunting, while unsanitary conditions, such as open defecation due to lack of access toimproved toilet facilities, have also been linked to childhood stunting.Our findings also highlight sanitation concerns, such as the use of unimproved toiletfacilities, which are linked to increased under-five stunting. Water supply andsanitation, given their direct impact on infectious diseases, especially diarrhea,have been considered important for preventing malnutrition and contrasts29 findings of previous studies. At the regional contextual level,it appeared that regions with above-average infant mortality rates were associatedwith higher stunting odds than below-average regions. Therefore, residing in regionswith higher infant mortality levels may offer a considerable stunting disadvantageto under-five children. Individual-level research has linked childhood stunting tounder-five mortality, while child undernutrition has in turn been linked to increased childmorbidity and mortality risks.56 Thus, providing a deeperunderstanding of the under-five and infant mortality nexus may provide crucialfuture program directions for improving under-five stunting levels in Ethiopia.Furthermore, we found notable residential disparities in under-five stunting aschildren living in rural settings were more likely to be stunted, which bothdirectly supportsThis study is not without potential limitations. The cross-sectional nature of thedata does not allow for causal inferences. Also, the study did not include someregional aggregate factors which are directly relevant for under-five nutritionalstatus, including food security and nutritional program coverage which were notcaptured by the survey data. Despite this, the use of a spatial and temporalanalytical approach has provided an invaluable contribution to the under-fivestunting literature.Although childhood stunting has improved in Ethiopia over time, substantial local andregional disparities in under-five stunting remain. Addis Ababa, the capital region,has seen consistently low prevalence, while northern regions have experienced higherlevels with slower and inconsistent declines. While Ethiopia may be on track toachieve the WHO’s target of reducing childhood stunting by 40% in 2025, regionsespecially in the north may not achieve this target.To ensure continuity across the country, policy decisions should bridge local andregional disparities by focusing on areas that have made slow or uneven progress.Programs and interventions can be targeted to hot spots and high-risk regions foundin this study. Moving forward, organizations can project what areas will be at highrisk for stunting by identifying regions with larger populations of older under-fivechildren and children from poor households, among other higher risk populations ofunder-five stunting. Policymakers and stakeholders should also commit resources toimprove socioeconomic inequalities among women, as well as household socioeconomicconditions since these underlying factors also impact under-five stunting.An in-depth investigation into the worsening under-five stunting levels observed inthe Dire Dawa region can help to better understand the underlying contextualfactors. Future research should consider additional regional-level factors onstunting, including seasonal food insecurity and climatic conditions on stunting.Also, future research can investigate changes in stunting patterns over time usingpoint estimation at smaller spatial scales in Ethiopia.


Concurrent chemoradiotherapy using weekly cisplatin remains standard of care for locally advanced cervical cancer in Sub-Saharan Africa. While cisplatin remains a popular cancer chemotherapeutic, it has an irreversible ototoxic effect on patients’ auditory system. However, there is a paucity of epidemiological information on its extent and severity during cervical cancer treatment. In a region with a high burden of cervical cancer, this has serious consequences for aural intervention and rehabilitation.2 body surface) at a tertiary level hospital in KwaZulu-Natal Province of South Africa, underwent audiological assessments at various intervals. We describe the temporal impact of cisplatin exposure on hearing loss, its combined effect with HIV-infection, and estimate ototoxicity incidence in this cohort. The median age was 52 years with Stages IIB (45%) and IIIB (35.4%) cancers being most common. Complaints of reduced hearing sensitivity increased significantly (p<0.0001). Bilateral, asymmetrical sensorineural hearing loss, with greater effect in the extended high-frequency range, was evident. Cisplatin dosage was significantly associated with ototoxicity severity at one- (p = 0.017), three- (p = 0.010), and six-month (p = 0.015) post-treatment follow-up. HIV-seropositivity (53.7%) was significantly associated with NCI-CTCAE Grading Scale at three- (p = 0.022) and six-months (p = 0.023) post-treatment. Multiple Tobit regression revealed a cumulative dose effect bilaterally, after adjustment for age and HIV status, evident from 9000Hz and above in the right ear, while a plateau effect was observed at 250mg/m2 in the left ear. The incidence was ototoxicity was 98% at a cumulative dose of 150mg/m2.Using a prospective cohort study design, 82 patients with incident cervical cancer, receiving weekly cisplatin chemotherapy (50 mg/mThe findings of this epidemiologic study highlight the temporal course and severity of ototoxicity experienced by cervical cancer patients treated with cisplatin, with greater impact in HIV-positive subgroup, thus underscores the need for audiological monitoring and timely interventions in this cohort. Cervical cancer continues to significantly burden the Sub-Saharan continent with the highest regional incidence . Based oWhile cisplatin is one of the most popular anti-cancer drugs due to its effectiveness, it is known to have a diverse toxicity profile affecting the gastro-intestinal, renal , hematolCisplatin-associated ototoxicity is characterized by a pattern of bilateral, progressive sensorineural hearing loss, with a greater influence on the high frequencies –16. The Hearing loss is also often a common occurrence of diabetes and hypertension, due to either the disease pathophysiology or the medication used in the treatment of these conditions. Diabetes-related-auditory manifestations are due to “spiral ganglion atrophy, degeneration of the vestibulocochlear myelin sheath, reduction of the number of spiral lamina nerve fibres, the thickening of the capillary walls of the stria vascularis and small arteries” . HyperteGiven the high incidence of cervical cancer in Sub-Saharan Africa, the associated comorbidities described above and the resulting poor quality of life in affected patients that may be further exacerbated by hearing loss, this prospective cohort study aimed to evaluate the extent and severity of cisplatin-associated ototoxicity experienced by cervical cancer patients in South Africa.2 body surface weekly for six cycles) in combination with dexamethasone (48 mg) and ondansetron (24 mg) offered as outpatient therapy. In some cases, the number of chemotherapy cycles was reduced due to some patients experiencing side effects i.e. severe gastrointestinal disturbances such as nausea, vomiting, and/or diarrhoea. Patients with cervix carcinoma were routinely tested for HIV during diagnostic work-up. All patients identified with reduced hearing ability were counseled regarding aural rehabilitation and referred for further interventions.Ethical clearance was obtained from the University of KwaZulu-Natal Biomedical and Research Ethics Committee (BE064/13) and the participating hospital, a tertiary level hospital, and one of the main referral centers for cancer patients housing an audiology department in KwaZulu-Natal (KZN), South Africa. Following written informed consent, 82 women diagnosed with histologically confirmed incident cervical cancer were prospectively enrolled into the study before treatment, within two years. Women presenting with profound hearing loss at baseline assessment, previous exposure to cisplatin chemotherapy, or with a history of tuberculosis, and/or malaria were excluded. Of the 82 participants, 80 attended all follow-up appointments. Attrition in the study was due to participant death. Chemotherapy consisted of cisplatin  staging system. All participants underwent audiological assessments before commencing cisplatin chemotherapy, with follow-up assessments at the beginning of the fourth cycle, and at one-, three-, and six-month post-treatment. These assessments included a case history, otoscopic examination, immittance audiometry , pure tone audiometry (including masking where required), extended high-frequency audiometry (up to 20000Hz), speech reception threshold (SRT) testing, word recognition score (WRS) testing and distortion product otoacoustic emission (DPOAE) testing, as prescribed for ototoxicity monitoring by the American Speech-Language and Hearing Association (ASHA) . The casPatient characteristics were summarized using frequencies and proportions for categorical and/or binary variables, whilst medians and interquartile ranges for asymmetric distributions. The distribution of participant’s pure-tone air conduction thresholds at the different frequencies was analysed separately for left and right ears, to avoid mixing frequencies, which have different upper cut-off limits on the audiometer. Data analysis is per ear and not per patient to prevent the effects of correlation between the ears, as ototoxic hearing loss is reported to be bilateral. For example, a higher pure tone threshold in the right ear may affect the threshold in the left ear and vice versa.Classification of hearing loss was based on frequencies in the range 125Hz– 8000Hz, as high-frequency audiometry does not have standardized results . The extThe Cochran-Armitage test for trend was used to test for trend in binomial proportions of auditory complaints over time. ASHA criteria  was used≥ 20dB decrease at any one test frequency,≥10dB decrease at any two adjacent frequencies, orloss of response at three consecutive frequencies where responses were previously obtained. Repeated Measures Analysis of Variance (ANOVA) was used to determine if the difference in mean DPOAE amplitudes between 1500Hz - 6000Hz was statistically significant at each audiological evaluation.2 versus High, 250mg/m2 and 300mg/m2) and HIV seropositivity on the severity of ototoxicity. As a result of the right censoring in pure-tone frequency threshold measurements (outcome variable), due to the upper limits of the audiometer at the various frequencies (125Hz-14000Hz), Tobit regression was employed to examine the association between the outcome variable and cumulative cisplatin dosage (mg/m2) (predictor variable). This approach was key as the number of ‘No Responses’ in the extended high-frequency range increased at each successive audiological evaluation (2) of 0.15, α = 0.05, power = 80% and 3 predictors  resulted in a sample size of 77. All statistical tests were 2-sided with statistical significance defined as p-value ≤ 0.05. All analyses were conducted in Stata version 15 .The Fisher’s exact test was used to determine the association between cisplatin dosage  while the remaining participants all received either 150 mg/m2, 200 mg/m2 or 250 mg/m2 of cisplatin, respectively. At each case history ascertainment, all participants reported no ongoing exposure to high levels of recreational and occupational noise. Furthermore, a review of the medical file and patient reports revealed normal renal function in all participants.Participant demographic and medical characteristics are presented in A summary of the self-reported audiological symptoms is presented in Abnormal otoscopic findings (in this case tympanic membrane perforation) were evident in the right ear of one participant, and the left ear of another participant at all audiological evaluations. At all audiological evaluations, 99% of the participants presented with Type A tympanograms bilaterally based on Jerger (1970)  classifiThere was a significant increase in the number of individuals presenting with abnormal DPOAEs bilaterally at each successive audiological evaluation  . DPOAEs There was a consistent decrease in the DPOAE amplitude bilaterally at each follow-up, as evidenced by the median DPOAE profiles with the decrease being more pronounced at the higher frequencies for both the right  and leftSpeech audiometry revealed that more than 79 participants (96%) presented with good SRT-PTA correlation bilaterally at each time point; therefore, validating the pure tone audiometry test results. Word recognition scores testing revealed that 75 participants (>95%) presented with excellent word recognition ability within normal conversational levels (below 60 dB HL) bilaterally at each audiological evaluation.2, an incidence of ototoxicity of 98% was recorded in this cohort.The ASHA criteria were used to determine the presence of an ototoxic shift in the patient’s hearing at each audiological evaluation. Given that any of the three criteria need to be fulfilled in the assessment for ototoxicity, 2) and high (250 & 300 mg/m2), respectively is illustrated in 2. At subsequent follow-up assessments, it becomes evident that the severity of ototoxicity is associated with treatment doses as more participants in the higher dosage group begin to show greater ototoxic change. The temporal association between cisplatin dosage and ototoxicity severity is significant at one- (p = 0.017), three- (p = 0.009) and six-month (p = 0.015) post-cisplatin treatment. At the end of the 6-month follow-up, 15/47 (32%) of patients in the high-dose group were in the Grade 2/3 category compared to only 5/33 (15%) of the patients in the low-dose group.The deterioration in hearing thresholds during cisplatin-chemotherapy treatment and their ability to guide recommendations regarding hearing amplification was explored. The temporal association in the severity of ototoxicity  and cumulative cisplatin dosage, stratified into low  in both the right and left ear, after adjustment for the effect of participant age and HIV status. Correlates of pure tone threshold measurements for the right ear revealed that significant changes were evident from 1000 Hz at a cumulative dose of 150 mg/mβ-coefficient 10.89) in hearing threshold at 200 mg/m2 cisplatin dose (p ≤ 0.05), compared to a 4-unit increase (β-coefficient 4.29) among the HIV-negative participants exposed to the same dose , despite there being a steady increase in the number of these participants presenting with hearing loss at each successive audiological evaluation.This study highlighted the adverse audiological impact of cisplatin in the course of cervical cancer treatment, with HIV-positive participants exhibiting a greater severity in hearing loss than their HIV-negative counterparts. By the end of the six-month follow up period, all of the participants (100%) presented with an ASHA  significThe percentage of significant ototoxic change in the current study, being higher than previously reported in South Africa , and othHIV and the subsequent use of ARTs aggravate cisplatin-ototoxicity, with there being a significant association by the three and six-month post treatment evaluations. The synergistic effect of ARTs and cisplatin on the auditory system may also account for a lower cumulative cisplatin dosage resulting in a more rapid range in the hearing thresholds in the HIV-positive group than the HIV-negative group. A synergistic effect on hearing, implies that the resultant hearing loss is greater than the sum of the hearing losses that occur from any one co-morbidity or ototoxic medication alone . The ototh cranial nerve, opportunistic infections associated with hearing loss and ototoxicity as a result of highly active antiretroviral treatment (HAART), and medication administered in the treatment of opportunistic infections” (p. 37). Whilst conductive hearing loss and outer/middle ear infections are common in HIV-infected individuals, the low incidence of these otologic conditions in the current study  highlights the benefits of the use of ARTs in preventing upper respiratory tract infections, which predispose affected individuals to middle ear infections and consequent conductive hearing loss. Additionally, it may indicate a lower viral load in this cohort due to the use of ARTs for a minimum of one year, which has proven to improve the impaired functioning of the immune system. However, it was not possible to identify the participants’ viral load, CD4 count, and treatment adherence in the current study, as this information was not captured in the hospital medical file due to participants receiving their ART from other decentralized local clinics. Additionally, participants were not knowledgeable of this information. As neurological testing was not conducted in the current study, no comment can be made about the direct action of HIV on the auditory nerve contributing to hearing loss.van der Westhuizen et al.  reported2 and 236.84 mg/m2 were likely to result in ototoxic hearing loss for the ototoxicity-free and ototoxicity groups, respectively, our study revealed that increasing cumulative dosages from 150mg/m2 have increasing effects in the high and extended high-frequency region bilaterally, with a plateau effect seen in the left ear at 250mg/m2, with no further change in hearing thresholds at increased dosages.Our findings on cumulative cisplatin dosage effects concur with literature , 45, 46 Our findings on progressively worsening pure-tone hearing thresholds, evident in the extended frequency range, supports previous literature , 40, 47.Deterioration of hearing thresholds in the extended high-frequency range does not generally negatively affect word recognition scores, as is evident in the current study with more than 90% of the participants presenting with excellent word recognition scores within normal conversational levels (60 dBHL) at all audiological evaluations. These findings concur with Theunissen et al. , who repThese results also, therefore, concur with the large number of participants presenting with ‘No Change’ when the NCI-CTCAE grading scale is used at the mid-cycle audiological evaluation, as this scale reflects the functional impact the ototoxic change has on the patient’s communication . As timeComplaints of tinnitus increased during treatment and thereafter steadily decreased from the one-month post-treatment assessment; thus, indicating a resolution of tinnitus post-treatment. The resolution of tinnitus is in keeping with Bokemeyer, Berger , MelamedThe reported progression of hearing loss for six months after cessation of cisplatin chemotherapy in this study is an indication of cisplatin retention in the cochlear indefinitely  and, theFurthermore, the increasing number of abnormal DPOAE findings in each ear with subsequent audiological evaluations is congruent with sensorineural hearing loss. Surprisingly, despite there being a significant decrease in the DPOAE amplitudes at all frequencies bilaterally, the number of abnormal DPOAE results are lower than would be expected when considering the increasing number of participants presenting with ototoxic hearing loss. These findings concur with Teotia et al. , who alsFactors that have been overlooked in previous studies investigating the effects of cisplatin, namely co-morbidities such as diabetes, hypertension, HIV infection, and the subsequent exposure to other ototoxic medication, have been considered in the current study as many women in South Africa present with such comorbidities . Due to The strength of this study is evident by the selection of the study design, which allowed for the prospective observation of cisplatin’s effect on hearing in individuals within the cohort over time . The useThis prospective cohort study, being the first in South Africa and the African continent, has revealed cisplatin’s detrimental effect on hearing with an even lower cumulative dosage of the treatment, possibly due to the large prevalence of HIV and subsequent use of ARTs. To prevent or avoid further deterioration in the quality of life of women with cervical cancer at a time when they are probably living in both physical and mental distress, early and effective identification of ototoxicity is crucial. Audiological monitoring should, therefore, be an essential part of the holistic management of patients receiving cisplatin chemotherapy, irrespective of cancer type, particularly in South Africa and other countries, which have a high burden of HIV. For patients who experience asymptomatic hearing loss but require continuation of cisplatin treatment, intensive audiological monitoring needs to be implemented with possible considerations of utilising oto-protective medication intra-tympanically. The findings of this current study will increase healthcare personnel’s awareness of the ototoxic effect of cisplatin and the need for comprehensive audiological monitoring with the potential of improving the quality of life for oncology patients. The potential for further investigation into chemotherapeutic regimens with minimal or no ototoxic effects is also warranted.S1 File(TIF)Click here for additional data file.S2 File(TIF)Click here for additional data file.S3 File(TIF)Click here for additional data file.S4 File(TIF)Click here for additional data file.S5 File(TIF)Click here for additional data file.S6 File(TIF)Click here for additional data file.S7 File(TIF)Click here for additional data file.S8 File(TIF)Click here for additional data file.