A male runner  had been running with suspected insertional Achilles tendinopathy (AT) for ~2 years when the pain reached a threshold that prevented running. Diagnostic ultrasound (US), prior to a high-volume stripping injection, confirmed right-sided medial insertional AT. The athlete failed to respond to injection therapy and ceased running for a period of 5 weeks. At the beginning of this period, the runner completed the Victoria institute of sports assessment–Achilles questionnaire (VISA-A), the foot and ankle disability index (FADI), and FADI sport prior to undergoing an assessment of bi-lateral gastrocnemius medialis (GM) muscle architecture (muscle thickness (MT) and pennation angle (PA); US), muscle contractile properties  and contraction time (Tc); Tensiomyography (TMG)) and calf endurance (40 raises/min). VISA-A and FADI scores were 59%/100% and 102/136 respectively. Compared to the left leg, the right GM had a lower MT (1.60 cm vs. 1.74 cm), a similar PA (22.0° vs. 21.0°), a lower Dm (1.2 mm vs. 2.0 mm) and Tc (16.5 ms vs. 17.7 ms). Calf endurance was higher in the right leg compared to the left (48 vs. 43 raises). The athlete began a metronome-guided (15 BPM), 12-week progressive eccentric training protocol using a weighted vest (1.5 kg increments per week), while receiving six sessions of shockwave therapy concurrently (within 5 weeks). On returning to running, the athlete kept daily pain  and running scores (miles*rate of perceived exertion (RPE)). Foot and ankle function improved according to scores recorded on the VISA-A (59% vs. 97%) and FADI (102 vs. 127/136). Improvements in MT (1.60 cm vs. 1.76 cm) and PA (22.0° vs. 24.8°) were recorded via US. Improvements in Dm (1.15 mm vs. 1.69 mm) and Tc (16.5 ms vs. 15.4 ms) were recorded via TMG. Calf endurance was lower in both legs and the asymmetry between legs remained . Pain intensity (mean weekly NRS scores) decreased between week 1 and week 12 (6.6 vs. 2.9), while running scores increased (20 vs. 38) during the same period. The program was maintained up to week 16 at which point mean weekly NRS was 2.2 and running score was 47.  Achilles tendinopathy (AT) is the 2nd most common running injury ,2. It isThe majority of running injuries are characterized by repetitive overuse and are usually to tissues ill-equipped for load absorption  . The insThe strongest evidence for the long-term resolution for Achilles tendon pain comes from exercise protocols . VariatiThe reasons pain and function respond positively to this form of training are multi-factorial . To the The patient provided written informed consent for the publication of this case report. A male runner  had been running with suspected insertional AT for 110 weeks (~2 years) having had the symptoms for the previous 5 years. The average miles per week for the 2-year period was 30. The average miles per week during phases of race preparation was ~40 and the maximum miles per week was 50. During this period, pain  and or stiffness was present every morning during the first steps on walking. Pain usually subsided as running progressed and returned after periods of prolonged rest. The runner used a 4-day-per-week running schedule and a 3-day-per-week cross-training schedule to manage running loads applied to the tendon and to lower the risk of other injury. Occasionally, pain increased  and was relieved using heavy double-legged isometric exercise (~85 kg) and reducing running from four days to three. No structured Achilles tendon rehabilitation was undertaken other than calf raise exercises on days the runner attended the gym (maximum 2*per week). The above summarises the training and pain status of the runner for much of the 2 years prior to cessation. This period included personal bests over 5 km (16:10), 10 km (33:46) and half-marathon (78:00).Nine weeks before cessation week 101/110), the runner began to increase the volume and sometimes intensity of one weekly run in an attempt to train for a marathon. This elevated daily pain to 7/10 and to a point where conservative self-management (as above) was no longer effective. The runner attended a sports medicine physician for an ultrasound guided high-volume stripping injection as prescribed in Humphreys et al. [1/110, thThe runner reported to the laboratory  at LeedsVISA-A and FADI scores were 59%/100% and 102/136, respectively. Compared to the left leg, the right GM had a lower MT (1.60 cm vs. 1.74 cm), a similar PA (22.0° vs. 21.0°), a lower Dm (1.2 mm vs. 2.0 mm) and Tc (16.5 ms vs. 17.7 ms). Calf endurance was higher in the injured leg (right) compared to the left (48 vs. 43 raises).The athlete ceased running for 5 weeks and began a metronome guided (15-BPM), 12-week progressive eccentric training protocol  on a flat surface, using a weighted-vest (1.5 kg increments per week). The exercise was performed by bilaterally plantar flexing both legs and lowering unilaterally on the affected leg to a count of 4 s (15 BPM). The athlete also received six sessions of shockwave therapy  concurrently within the first 5 weeks. The athlete recorded daily pain scores (NRS)  throughoThe athlete had a compliance of 80.4% (135/168 possible sessions) to the 12-week eccentric training protocol. At the 12-week point, foot and ankle function improved according to scores recorded on the VISA-A (59% vs. 97%) and FADI (102 vs. 127/136). In the injured leg, improvements in MT (1.60 cm vs. 1.76 cm) and PA (22.0° vs. 24.8°) were recorded via US. Improvements in Dm (1.15 mm vs. 1.69 mm) and Tc (16.5 ms vs. 15.4 ms) were recorded via TMG. Calf endurance was lower in both legs and the asymmetry between legs remained . Pain intensity (mean weekly NRS scores) decreased between week 1 and week 12 (6.6 vs. 2.9), while running scores increased (20 vs. 38) during the same period. The program was maintained up to week 16 at which point mean weekly NRS was 2.2 and running score was 47 .The runner in this case report ran for ~2 years with persistent pain and achieved several personal bests. This demonstrates, in this athlete at least, that pain can be managed in a way that allows a high level of athletic function. This underscores the title of this paper which suggests that pain and function are not as closely linked in chronic pain as they may be in pain resulting from traumatic insult. In fact, baseline results of the calf endurance test appear to demonstrate a compensatory mechanism whereby calf endurance in the injured leg was higher than that of the non-injured . Calf enThis athlete did not respond to a high-volume stripping injection, which may suggest that this type of intervention is not as effective in insertional tendinopathy as it appeared to be for mid-portion tendinopathy . It may In the injured leg (right), there were improvements in gastrocnemius medialis MT (~10%) and PA (~12.7%), as measured by US, which appeared to be reflected by the relative improvement in Dm (~47%) and, to a lesser extent, Tc (~6.7%). These findings were expected, as they mirror improvements in muscle size and function in response to resistance exercise . StrengtThis interpretation of an increase in Dm representing an increase in stiffness is somewhat at odds with researchers in the field who suggest a lower Dm to be associated with a greater muscle stiffness . These iThe modest reduction in Tc might be expected from an intervention that did not challenge the speed of muscle contraction. The improvements reported are likely due to an increase in the number of sarcomeres in series  and the There are several limitations we would like to acknowledge related to the interpretation of this case report. Firstly, although tendon structural abnormalities are present on ultrasound, this cannot be inferred to represent pathology . In factThis athlete ran at a high level for 2 years with persistent Achilles pain. Even after an evidence-based rehabilitation programme and a gradual return to running, this athlete was never entirely pain-free. The findings in this runner appear to underscore the need for greater awareness among clinicians about the dissociation between pain and function in Achilles tendinopathy. Pain education and limiting fear avoidance may be as important as physical rehabilitation. The neuromuscular adaptations in this study were substantial and the physiology which underpins them is multifactorial. To what extent these neuromuscular adaptations contribute to the reduction in pain and increase in running specific function is unknown and it will perhaps always be an almost impossible research question to answer. However, it is likely that the combination of neuromuscular adaptations and structural adaptations within the tendon itself help to better balance stress on biological tissue preventing overload of a specific portion of tendon.


IntroductionOur objective was to determine the efficacy of ketoconazole (KTZ) 2% cream for the treatment of mild adult female acne (AFA).Materials and methodsThis placebo-controlled trial was conducted in District Headquarters (DHQ) Teaching Hospital, Sahiwal, Pakistan. The study was completed in a period of January 2019-June 2020. A total of 60 females of age > 25 years having mild AFA were included. In Group I, the patients were advised to apply 2.0% KTZ cream covering the whole skin area twice daily for a period of eight weeks. In Group II (placebo group) patients, a topical cream containing propylene glycol was applied for similar period. After eight weeks, the cream was discontinued and participants were advised to use routine skincare products, and follow-up was done after two weeks. The main study outcome was reduction in the total count of acne lesions including both inflammatory and non-inflammatory lesions and overall success rate of treatment.ResultsMean age was 36.2 ± 6.3 years in KTZ group versus 35.4 ± 6.5 years in control group. Mean duration of acne was 14.3 ± 7.3 years in KTZ group versus 15.1 ± 6.9 years in control group. Improvement in facial adult female acne scoring tool (AFAST) scale (AFAST-F) was observed in 13 (43.3%) patients in KTZ group and in only 4 (13.3%) patients in control group . Improvement in submandibular AFAST (AFAST-S) was observed in 12 (40.0%) patients in KTZ group and in eight (26.7%) patients in control group. The overall success rate of treatment was 14 (46.7%) in KTZ group versus 4 (13.3%) in control group .ConclusionsIn our study, we found significant improvement in reduction of acne lesions as well as complete recovery using 2.0% ketoconazole for the treatment of mild AFA. So KTZ can be used as a preferred treatment option for these patients. Acne vulgaris is a very common skin disease particularly affecting the young females. It has significant psychological impact and impairs the daily living of affected ones . It has Hormonal, genetic, and environmental factors have been shown to be the main players in the pathogenesis of AFA. Among the environmental factors, stress, disturbed sleep, diet, excessive face washing, use of certain cosmetics, and exposure to ultraviolet radiation have been reported to be the triggering factors of AFA . AFA can be in continuous or intermittent form and has reported to have high recurrence rates. The intermittent and recurrence nature of AFA has resulted in higher prevalence of antibiotic resistance. Therefore, long-term and continuous treatment is need is some cases that may continue even for years ,7.Malassezia dysbiosis. Ketoconazole (KTZ) cream has been shown to be effective for the treatment of AFA. KTZ is a broad spectrum azole antifungal, which was originally approved in 1981 for the treatment of systemic fungal infections. It has also been shown to have anti-inflammatory and anti-androgenic activity [Cutibacterium acnes; it is also effective against resistant isolates [The commonly used treatment options for the management of AFA are benzoyl peroxide and adapalene . These disolates .Keeping in view the above-mentioned benefits of KTZ, the aim of the present study is to determine the efficacy of KTZ 2% cream for the treatment of mild AFA.This placebo-controlled trial was conducted in District Headquarters (DHQ) Teaching Hospital, Sahiwal, Pakistan. The study was completed in a period of January 2019-June 2020. The ethical clearance approval for this study was taken from Ethical Review Committee (ERC) of hospital. The patients were informed about study protocol before taking informed consent. A total of 60 females of age > 25 years having mild AFA  were included. Exclusion criteria were patients taking topic or systemic treatment for AFA for at least last two weeks, other facial skin lesions that can effect acne assessment, or pregnant females.Patients were allocated to two equal groups using draw randomization. In Group I, the patients were advised to apply 2.0% KTZ cream covering the whole skin area twice daily for the period of eight weeks. In Group II (placebo group) patients, a topical cream containing propylene glycol was applied for similar period. After eight weeks, the cream was discontinued and participants were advised to use routine skincare products, and follow-up was done two weeks later. All the participants were advised to use skin-cleansing product based on nonmedical based ingredients such as sodium lauryl sulfate formulation before applying the cream. The follow-up was performed at two, four, six, and eight weeks; the final assessment was done at eighth week. A senior dermatologist who was unaware of the study protocol was asked to do final assessment of all patients.The main study outcome was reduction in the total count of acne lesions including both inflammatory and non-inflammatory lesions and overall success rate of treatment. We used adult female acne scoring tool (AFAST) to evaluate face and submandibular areas separately. Acne improvement was defined as an improvement in acne severity score at least 1 point AFAST scale reduction from the baseline value, while the success rate was defined as achievement of 0 (clear) or 1  AFAST score at the end of follow-up.Data analysis was done using SPSS v25 software . Acne improvement and success rate were compared using chi-square test between the KTZ and control group by taking p value ≤ 0.05 as a significant difference.There was no significant difference regarding mean age, duration of acne, and family history between the group. Mean age was 36.2 ± 6.3 years in KTZ group versus 35.4 ± 6.5 years in control group. Mean duration of acne was 14.3 ± 7.3 years in KTZ group versus 15.1 ± 6.9 years in control group. There were 29 (96.6%) patients in KTZ group having persistent acne, while 27 (90.0%) patients in control group had persistent acne . The aggravating factors of acne were also similar between the groups  was observed in 13 (43.3%) patients in KTZ group and in only four (13.3%) patients in control group . Improvement in submandibular AFAST (AFAST-S) was observed in 12 (40.0%) patients in KTZ group and in eight (26.7%) patients in control group. The overall success rate of treatment was 14 (46.7%) in KTZ group versus 4 (13.3%) in control group  (Table In this placebo-controlled randomized trial we determined the efficacy of KTZ 2.0% cream for the treatment of mild AFA. We found significant improvement in the severity of AFA using KTZ, with overall success rate of 46.7% as compared to only 13.3% in control group.Chottawornsak et al. conducted a similar study on the efficacy of KTZ for the treatment of AFA. The authors reported overall success rate of 45.0% in KTZ group and only 14.3% in control group. The study reported improvement in AFAST-F scale in 42.9% KTZ group patients and in 9.5% control group patients, and improvement in AFAST-S scale was reported in 35.0% KTZ group patients and in 33.3% control group patients .In the present study, improvement in AFAST-F was observed in 43.3% patients in KTZ group and in 13.3% patients in control group, while improvement in AFAST-F scale was observed in 40.0% patients in KTZ group and in 26.7% patients in control group.We chose the AFAST scale for evaluation of response rate of KTZ cream as this scale is more reliable for determining the improvements in acne severity ,13. MoreC. acnes; it produces inflammation and follicular hyperplasia in infected individuals [Along with antifungal and anti-inflammatory properties, the other possible mechanisms of KTZ effectiveness may be due to its antilipase and anti-androgenic activity. Lipase is the most important enzyme for virulence activity of ividuals . Inhibitividuals . So thisividuals .Malassezia spp., which also has lipase activity more than that of C. acnes [Malassezia spp. is responsible for recurrent acne and other skin inflammatory lesions such as seborrheic dermatitis [KTZ also exhibits its antilipase activity against C. acnes . Some rermatitis . As KTZ The major limitation of present study is that we only included patients having mild degree of AFA; so the results of present study cannot be generalized to treat patients having moderate to severe acne. There is a need to conduct studies with large sample size including patients having moderate and severe AFA so that the efficacy of KTZ treatment can be determined in these patients as well.In our study, we found significant improvement in reduction of acne lesions as well as complete recovery using 2.0% ketoconazole for the treatment of mild AFA. So KTZ can be used as a preferred treatment option for these patients.


Background: An antibody panel is needed to definitively differentiate between adenocarcinoma (AC) and squamous cell carcinoma (SCC) in order to meet more stringent requirements for the histologic classification of lung cancers. Staining of desmosomal plaque-related proteins may be useful in the diagnosis of lung SCC.Materials and methods: We compared the usefulness of six conventional  and three novel  markers to distinguish between lung SCC and AC in 85 small biopsy specimens (41 ACs and 44 SCCs). Correlations were examined between expression of the markers and patients’ histologic and clinical data.Results: The specificity for SCC of membrane staining for PKP1, KRT15, and DSG3 was 97.4%, 94.6%, and 100%, respectively, and it was 100% when the markers were used together and in combination with the conventional markers . In a stepwise multivariate logistic regression model, the combination of CK5/6, p63, and PKP1 in membrane was the optimal panel to differentiate between SCC and AC, with a percentage correct classification of 96.2% overall (94.6% of ACs and 97.6% of SCCs). PKP1 and DSG3 are related to the prognosis.Conclusions: PKP1, KRT15, and DSG3 are highly specific for SCC, but they were more useful to differentiate between SCC and AC when used together and in combination with conventional markers. PKP1 and DSG3 expressions may have prognostic value. EML4–ALK (echinoderm microtubule-associated protein-like 4–anaplastic lymphoma kinase) or EGFR gene-activating mutations can respond to the respective tyrosine–kinase inhibitors  family is required to form functional desmosomes . PKP1 isDesmoglein 3 (DSG3) is one of seven desmosomal cadherins. Desmosomal proteins act as tumour suppressors and are downregulated in epithelial–mesenchymal transition and in tumour cell invasion and metastasis. However, some studies have shown the upregulation of several desmosomal components in cancer, including DSG3, and overexpression of these proteins has been related to the prognosis. Therefore, desmosomal proteins can potentially serve as diagnostic and prognostic markers . KeratinImmunohistochemistry is widely used for the subtyping of lung carcinomas. Thyroid transcription factor 1 (TTF1)  and NapsIn this study, we compared the usefulness of six conventional and three novel markers for the differential diagnosis of lung SCC and AC in small biopsy specimens. We also explored correlations between the expression of these markers and the histologic and clinical data of the patients.Specimens used in this study were from 87 patients who underwent surgical resection for lung cancer. All patients were stages I (54 patients), II (20 patients), or III (10 patients), except for three patients in stage IV, and were selected for surgery with no previous chemotherapy or radiotherapy treatments. Only cases in which the histological diagnosis of the resected specimen was invasive AC or SCC were included, representing the majority of the series (85 patients). The remaining two cases, which corresponded to sarcomatoid carcinoma, were excluded from the study.Histological diagnosis of the surgical specimens followed the recommendations of the WHO 2015 classification . The othThe 85 incisional biopsies were analyzed by two pathologists (authors MGM and MCM) and classified according to the WHO 2015 classification . Pathological evaluation of resected specimens was carried out using the 1997 revision of the International System for Staging Lung Cancer  and revia4), and p40 .Sections of 4-μm thickness were taken from the paraffin blocks, mounted on pre-treated slices, and stained for PKP1 , KRT15 , and DSG3 . Normal human skin was used as positive control. After testing several dilutions, PKP1 and DSG3 were diluted at 1:200 and KRT15 at 1:500. Prediluted monoclonal antibodies from Master Diagnostica were used for the remaining determinations: TTF1 , Napsin A , CK7 , CK5/6 , p63  using the Master Polymer Plus Detection System (MAD-000327QK Master Diagnostic) and following the manufacturer’s instructions. Diaminobenzidine was used as chromogen, and sections were counterstained with haematoxylin. Immunohistochemical staining was evaluated independently by two pathologists (authors MGM and MCM), who assigned a score based on the extent and intensity of immunoreactivity. Agreement between pathologists was >90%, and any differences in interpreting results were resolved by consensus. Staining for PKP1, CK15, and DSG3 was evaluated semiquantitatively in nucleus, cytoplasm, and membrane as negative , positive 1+ (6–25% cells stained), positive 2+ (26–50% cells stained), or positive 3+ (>50% cells stained). Staining intensity at the same localizations was scored semiquantitatively from 0 to 3. Immunostaining results were scored as the sum of the extent and intensity of immunoreactivity, considering a score ≥3 positive and a score <3 negative. Only cases with nuclear staining were considered positive for TTF1, p63, and p40, while those with cytoplasmic/membranous staining were considered positive for CK5/6 and CK7, and those with characteristic granular and cytoplasmic staining were considered positive for Napsin.2) or Fisher’s exact tests. Odds ratios with 95% confidence intervals (CIs) were calculated for 2 × 2 tables. Accuracy measures were calculated for each marker with the corresponding 95% CIs. p ≤ 0.05 was considered statistically significant. Survival analyses were performed with R 3.4.0 software, using the ‘survival’ (v2.42–3) and ‘survminer’ (v0.4.2) packages. Clinical information and normalized gene expression data were downloaded from Firebrowse  for the TCGA-LUSC project (n = 504). Patients with SCC from the TCGA-LUSC cohort were divided into two groups for analysis of the relationship between survival outcomes and gene expression: ‘high’ (above the median for the specific gene) and ‘low’ (below the median). Kaplan–Meier curves were plotted for the raw survival data, and Cox proportional hazards models were then constructed, accounting for tumour stage and for patient age and sex.IBM SPSS20 statistical package was used for data analyses unless otherwise specified. The relationship of clinical-pathological characteristics with marker expression was evaluated using the chi-square , followed by CK5/6 93.2%) and p40 (88.6%), whereas p40 was the most specific (95.1%), followed by CK5/6 (92.7%) and p63 (85.4%). Other statistical analyses evaluating agreement and reproducibility are displayed in % and p40We studied three novel markers for SCC  that may contribute to the differential diagnosis of SCC and AC, also analyzing their localization . Immunohistochemical staining for PKP1 was mainly detected in SCC, with a heterogeneous distribution and intensity among the different tumours analyzed, and in different areas of the same neoplasm. PKP1 most frequently stained cellular membranes, marking intercellular junctions, followed by staining of the cytoplasm. Membranous staining was mainly observed in well to moderately differentiated areas of SCC. Nuclear staining was less often observed and was more frequent in cells with a more immature appearance . MembranStaining for DSG3 was not observed in ACs, with the exception of only a few nuclei in a small number of cases. As occurred with PKP1 and KRT15, DSG3 stained SCCs with irregular distribution and was more often observed in cell membranes of better-differentiated areas and to a lesser degree in cytoplasms. Nuclear staining was much less frequent and mainly observed in poorly differentiated areas .In our cohort, the most sensitive marker was PKP1 in nucleus (86%), followed by KRT15 in membrane (75.6%) and DSG3 in membrane (64.3%). However, DSG3 in membrane was the most specific (100%), followed by PKP1 in membrane (97.4%) and KRT15 in membrane (94.6%). Other statistical analyses evaluating agreement and reproducibility are displayed in According to these results, membranous staining with the novel markers  was specific for ≥94% of the SCC samples, obtaining ≥94% true positives in our cohort of patients.The antibodies were also tested in combination. For SCC samples, membranous PKP1 staining showed a specificity of 97.4% and sensitivity of 76.7% and membranous DSG3 staining a specificity of 100% and sensitivity of 64.3% when separately evaluated. When considered together, membranous staining with these two markers achieved a specificity of 100% (95% CI = 90.6%–100.0%) but sensitivity of 52.4% (95% CI = 37.7%–66.6%). Positive TTF1 staining provided a specificity of 97.7% and a sensitivity of 75% for AC. When all three markers were considered together in Panel 1 SCC (positive membrane staining for PKP1 and DSG3 and negative staining for TTF1), the specificity for SCC samples was increased to 100.0% (95% CI = 90.6%–100.0%), but the sensitivity was 52.4% (95% CI = 37.7%–66.6%), with an AUC of 0.7619. Membranous KRT15 staining yielded a specificity of 97.4% and a sensitivity of 75.6% for SCC samples, and Napsin A staining a specificity of 93.2% and a sensitivity of 80.5% for AC samples when evaluated separately. When all five markers were considered together in Panel 2 SCC , the specificity for SCC samples was increased to 100.0% (95% CI = 89.6%–100.0%), but the sensitivity was 47.5% (95% CI = 32.9%–62.5%), with an AUC of 0.7375. For AC samples, the combined evaluation of negative membranous staining for PKP1 and DSG3 and positive TTF1, in Panel 1 AC, increased the specificity for AC samples to 100.0% (95% CI = 91.6%–100.0%), but the sensitivity was 24.3% (95% CI = 13.4%–40.1%), with an AUC of 0.8552. When negative membranous staining for PKP1, DSG3, and KRT15 and positive staining for TTF1 and Napsin A were considered together in Panel 2 AC, the specificity for AC samples increased to 100.0% (95% CI = 91.2%–100.0%), and the sensitivity was 63.6% (95% CI = 46.6–77.8%), with an AUC of 0.8088 .The area under the ROC curve (or AUC) represents an optimal summary statistic for comparing the sensitivity and specificity of the nine markers . For the cohort of 85 primary malignant lung neoplasms (41 ACs and 44 SCCs), including all histological grades, p63 (0.965) had the largest AUC, followed by CK5/6 (0.960), and p40 (0.938), with AUC values >0.9, while AUC values for the remaining markers were between 0.8 and 0.9. AUC values are displayed in A stepwise multivariate logistic regression model was constructed to determine the optimal immunohistochemical marker panel for differentiating between SCC and AC. The best result included the combination of markers CK5/6, p63, and PKP1 in membrane, giving a percentage correct classification of 96.2% overall (94.6% of ACs and 97.6% of SCCs).p = 0.19), possibly due to the limited sample size. There was also a non-significant trend for longer survival in SCC versus AC patients (log-rank p = 0.47). In order to overcome this sample size limitation and remove tumour subtype as a confounder, we also studied 495 patients in the TCGA-LUSC cohort (The Cancer Genome Atlas) for whom survival and gene expression data were available . In this cohort, a significant relationship was found between higher PKP1 expression and better overall survival (p = 0.036) in a Cox proportional hazards model after adjusting for confounding variables.Data on 80 patients were available for this analysis. The Kaplan–Meier survival curve depicted in survival , obtainiSupplemental Figures 1 and 2, available online). In the multivariate analyses, we applied Cox proportional hazard models for each marker, accounting for patient age, sex, and tumour stage, in order to assess the relationship of expression at mRNA level with patient survival.We also performed survival analyses for all conventional and novel SCC biomarkers in the TCGA cohort under the same conditions as the PKP1 analysis. For the univariate analyses, we stratified the patients as ‘high’ or ‘low’ for each marker based on the expression being above or below the median value, and we plotted Kaplan–Meier curves , PKP1 , and CK5 . In addition, the association of DSG3 with survival was close to statistical significance . Expression of KRT15 or p40 at the mRNA level was not associated with patient survival.In the multivariate analyses, the high expression of three SCC markers was associated with better survival: p63  for a detailed evaluation and compared them with six conventional markers that are well described in the literature and are routinely used by many histopathology laboratories . Given tStudies in whole-tissue sections indicate that SCCs have a relatively precise immunophenotype, i.e. negativity for TTF1 and positivity for p63, CK5/6, and 34βE12. In contrast, ACs are much more heterogeneous, and only diffuse positivity for TTF1 is considered characteristic, given that a proportion of ACs also express markers considered typical of SCCs. Hence, albeit useful, no SCC marker is wholly specific .According to the present results, among the expressions of the conventional markers CK7, TTF1, and Napsin A, characteristic of AC, and CK5/6, p40, and p63, characteristic of SCC, only TTF1 in AC and p40 in SCC were specific in ≥94% of samples in the present cohort, with ≥94% true positives. Membrane staining with the novel markers  was specific for ≥94% of the SCC samples, with ≥94% true positives in the present cohort. The antibody panel studied was most effective for the classification of SCC and AC when the antibodies were applied successively in a stepwise manner. We found that the specificity was higher (100%) when the antibodies were used in combination rather than individually. For SCC, the most effective panels were: Panel 1 SCC  and Panel 2 SCC . For AC, the most effective panels were: Panel 1 AC  and Panel 2 AC .The novel markers described here showed a heterogeneous staining of SCC, which was observed in the membranes and cytoplasm of more differentiated cells, marking the intercellular junctions, with staining of nuclei more frequently detected in areas of more immature appearance. In AC samples, focal staining with these novel markers was detected in nucleus and cytoplasm, but never in membrane. In addition to their role in cell adhesion, plakophilins, including PKP1, have been reported to localize to the cytoplasm and nucleus, where they are thought to have several functions that are not completely understood . This exIn our cohort of patients, the relationship of PKP1 staining with better survival of SCC and AC patients did not reach statistical significance; however, we were able to confirm the relationship between high PKP1 mRNA expression and better overall survival in an additional analysis of 495 SCC patients from TCGA. In addition, our TCGA analysis revealed a close-to-significant association between high DSG3 mRNA expression and improved overall survival. This agrees with previous reports that associated positive DSG3 staining with longer survival in lung cancer patients of all histologic subtypes . Taken tAccording to our findings, evaluation of CK15 and especially of DSG3 and PKP1 improves the differential diagnosis of SCC and AC. However, a potential limitation of these SCC markers is that membrane-specific positivity is found in better-differentiated samples that are more readily identifiable under the microscope. Nevertheless, they may be especially useful in samples affected by a major artefact, because they offer a more objective parameter for evaluating the degree of differentiation. A further advantage of these markers is the complete absence of staining in poorly differentiated ACs, observing cytoplasmic and nuclear positivity only in moderately or well-differentiated ACs. These antibodies could possibly be used in a cocktail, with the consequent saving of histological sections. This is an important issue, given the need for genetic as well as immunohistochemical analyses in tissue from small biopsies in patients with lung carcinoma.Despite the small number of samples analyzed, the stepwise multivariate logistic regression model showed that the combination of markers CK5/6, p63, and PKP1 in membrane gave a percentage correct classification of 96.2% overall (94.6% of AC and 97.6% of SCC), being the best immunohistochemical marker panel to distinguish between SCC and AC.


The accuracy of continuous glucose monitoring (CGM) sensors may be significantly impacted by exercise. We evaluated the impact of three different types of exercise on the accuracy of the Dexcom G6 sensor. Twenty-four adults with type 1 diabetes on multiple daily injections wore a G6 sensor. Participants were randomized to aerobic, resistance, or high intensity interval training (HIIT) exercise. Each participant completed two in-clinic 30-min exercise sessions. The sensors were applied on average 5.3 days prior to the in-clinic visits (range 0.6–9.9). Capillary blood glucose (CBG) measurements with a Contour Next meter were performed before and after exercise as well as every 10 min during exercise. No CGM calibrations were performed. The median absolute relative difference (MARD) and median relative difference (MRD) of the CGM as compared with the reference CBG did not differ significantly from the start of exercise to the end exercise across all exercise types . The accuracy of the no-calibration Dexcom G6 CGM was not significantly impacted by aerobic, resistance, or HIIT exercise. Regular physical activity can help people with type 1 diabetes improve glycemic control, decrease total daily insulin needs, achieve a healthier body mass index, and reduce risk of cardiovascular and peripheral neuropathy complications ,3. DespiThe emergence of continuous glucose monitoring (CGM) has transformed how type 1 diabetes is managed and enabled better monitoring of glucose during exercise ,12. The While several studies have assessed the accuracy of CGM systems during exercise ,27,28,29With diverse weekly physical activity being recommended for all adults living with diabetes , it is chttps://clinicaltrials.gov under the identifier NCT03443713. All participants gave informed consent prior to participating in this trial.This work is a planned secondary analysis of a single center randomized prospective study involving 24 adults with type 1 diabetes using multiple daily injection therapy. This outpatient study aimed to assess the impact of weekly physician-driven insulin setting changes on participants with elevated HbA1c levels of 7.0% or more. Physician’s recommendations were used to improve and compare the performance of an artificial-intelligence-based decision support algorithm as previously published . This clParticipants wore a commercial Dexcom G6 CGM  on their abdomen for 10 days at a time, for a total of 28 days, and were instructed not to perform calibrations. Daytime physical activity was captured using an Apple Watch . Participants used the InPen device  for administration of insulin aspart and were instructed to use the InPen bolus calculator on a smartphone. They used either insulin glargine or insulin degludec for long-acting insulin. Participants logged all food and drinks consumed during the study using a mobile application developed at OHSU. Each participant was asked to exercise at home four times and to complete two exercise sessions in clinic. The first in-clinic session was scheduled between day 9 and 14 of the study, and the second between day 16 and 20. Both at-home and in-clinic exercise sessions followed standardized videos. Participants were randomly assigned to either an aerobic, resistance, or HIIT exercise video. The 30-min aerobic exercise video was designed for steady aerobic workout and consisted of 45 s of activity interrupted by 15 s of instructional transition. The 30-min resistance exercise video was designed to work all muscle groups and made use of resistance bands with self-selected resistance level. The shorter 20-min HIIT video consisted of 40-s activities followed by 80 s of rest.During the in-clinic exercise sessions, capillary blood glucose (CBG) was measured using a Contour Next meter , which is among the most accurate commercially available glucose meters (mean absolute relative difference (MARD) of 5.6 ± 6.4%) . CapillaExercise was only initiated once the participant’s glucose was above 80 mg/dL (4.4 mmol/L) with carbohydrates administered for values below 120 mg/dL (6.7 mmol/L) at the start of exercise, or below 70 mg/dL (3.9 mmol/L) thereafter. Serum ketone levels were checked for glucose values above 250 mg/dL (13.9 mmol/L) and the exercise session was rescheduled if the ketone levels were above 0.6 mmol/L.Capillary glucose measurements from the Contour Next meter were used as reference glucose and were paired with the temporally nearest CGM values within 2.5 min. Only CBG values between 40 and 400 mg/dL (2.2–22.3 mmol/L) were considered, excluding 3 out of 272 m values.The accuracy of the Dexcom G6 system during exercise was evaluated using median absolute relative difference (ARD) and relative difference (RD), comparing subsequent measures to baseline. Because the paired differences of these measures did not follow a normal distribution, Wilcoxon signed-rank tests were used to determine the statistical significance of the presented results, at a 95% confidence level  .p-values from the Wald test of model coefficients and 95% confidence intervals (CI). The gamma model for ARD was estimated using a generalized linear mixed model with a log link. Because mean ARD did not follow a normal distribution, its effects are reported as ratios. Lastly, aerobic exercise was selected as reference in both models, as it represented the greatest change in glucose when compared with resistance and HIIT exercises. In a preliminary analysis, we investigated whether the time since application of the CGM influenced accuracy. We did not find that sensor wear-time was statistically related to accuracy (p = 0.201), and since the study was not powered to detect differences in accuracy across wear times, we did not include this variable in the final mixed effects analysis.Next, mixed-effects regression models were used to estimate the effects of four independent variables on mean ARD and mean RD. The mean RD was found to reasonably fit a normal distribution. However, the mean ARD data were strictly positive and right-skewed, thus better fitting a gamma distribution. As such, mean RD was estimated using a linear regression model while mean ARD was estimated using a gamma regression model. The independent variables were selected based on hypothesized relationships to the endpoints and include (1) the exercise type , (2) the reference CBG value, centered at the mean, (3) the change in CBG over the previous 10-min interval (glucose drop), and (4) the time since exercise start in 10-min increments, starting at 10 min. These variables were then scaled for interpretability. Higher-order terms and splines were considered and tested for some of the models based on exploratory analyses, but these failed to provide a better fit and were not used in the final analysis. Additionally, both regression models also included nested random intercepts for the study participant and the exercise session. Robust  variance estimators were used to calculate 2 test [The Clarke error grid analysis (EGA) was then used to visualize the data and quantify the clinical accuracy of the system . This an2 test ,40. Next2 test . This le2 test ,43.A total of 25 participants were enrolled, of which one withdrew prior to any exercise visits due to scheduling conflicts. All remaining 24 participants completed two exercise sessions in clinic and were included in the analysis. Eight persons were randomly assigned to each exercise group  for a total of 48 exercise visits and 16 sessions per exercise type. The baseline characteristics and demographics of these participants are reported in In total, 272 paired Dexcom G6 CGM and reference Contour Next CBG values were analyzed: 96 pairs for aerobic, 96 for resistance exercise, and 80 for HIIT. As expected, the sharpest drop in median CBG from the start to the end of exercise was observed during aerobic exercise (−47.5 mg/dL or −2.6 mmol/L), followed closely by resistance exercise (−36.5 mg/dL or −2.0 mmol/L) and HIIT (−26.0 mg/dL or −1.4 mmol/L). Interstitial glucose results from the G6 sensor showed a similar trend with the largest glucose drop during aerobic exercise (−46.5 mg/dL or −2.6 mmol/L), followed by resistance (−20.5 mg/dL or −1.1 mmol/L) and HIIT (−15.5 mg/dL or −0.9 mmol/L). From the start to the end of exercise, the majority (69%) of participants experienced a drop in blood glucose.p = 0.03). For resistance exercise, the RD at time 30 min was statistically different from baseline (p = 0.01), but the negative bias at time 30 min was smaller than baseline. Relative to the start of exercise, the median RD for all other time points did not show any significant changes.Median ARD did not differ significantly from the start of exercise to the end of exercise across all exercise types, as illustrated in p = 0.96), meaning that there was no substantial bias in the CGM when controlling for other variables (95% CI −7.2 to 6.8). When evaluating the influence of exercise type on RD, we found that the mean RD was significantly larger in the negative direction during HIIT with a difference of −11.3% compared to aerobic exercise . In comparison, resistance exercise was found to be somewhat closer to aerobic exercise, with a difference of −4.6% . Mean RD was also found to be strongly linked with capillary glucose levels with a change of −0.74% for every 10 mg/dL (0.5 mmol/L) increase in the reference blood glucose . Lastly, mean RD and glucose drop were strongly correlated with a 3.1% change in mean RD for every 10 mg/dL of glucose drop, meaning that as glucose dropped faster, we observed a more positive RD . Observations from exercise sessions were correlated with an intraclass correlation coefficient (ICC) of 0.78, while the ICC from participants was 0.43.Three independent variables including exercise type, reference CBG value and glucose drop were significantly associated with RD in the mixed-effects linear regression model . Time frp = 0.40; 95% CI 0.97 to 1.1), meaning that a 0.3% increase in mean ARD is predicted for every 10 min of exercise. The intercept, or mean ARD when other variables are null, was significantly different from zero at 10.85% . Mean ARD during HIIT was nearly the same as during aerobic exercise, with a ratio of 0.98 . Resistance exercise was somewhat lower at 0.93 . In practice, these ratios represent a predicted mean ARD of 13.3% (CI 8.4 to 18.2) for aerobic exercise, followed by 13.0% (CI 9.3 to 16.7) for HIIT and 12.4% (CI 8.8 to 15.9) for resistance exercise. ARD changed only fractionally for every 10 mg/dL increase in the reference blood glucose . This represents a small decrease in mean ARD as CBG increases, with predicted ARD ranging from 13.3% (CI 9.3 to 17.3) at 70 mg/dL (3.9 mmol/L) down to 12.6% (CI 9.2 to 16) for glucose levels near 250 mg/dL (13.9 mmol/L). ARD changed little for every 10 mg/dL of glucose drop . That is, mean ARD is predicted to rise from 12.7% (CI 10.5 to 14.8) at constant glucose levels up to 14.0% (CI 11.1 to 17.0) for glucose drops of 30 mg/dL every 10 min.On the other hand, the regression model for mean ARD did not link changes with any of the independent variables at a statistically significant level. Time from start of exercise was not significantly associated with ARD when meter value and glucose drop were included in the regression model. The estimated effect was 1.03 on the ratio scale for every 10 min (p = 0.04), the bias in the clinically safe zone (A+B) was comparable across exercise types. Additionally, no notable differences were observed in these metrics when comparing baseline values (before exercise) to those at different time points during exercise.The Clarke error grid analysis (EGA) results for the different types of exercises are shown in Lastly, the Clarke EGA shown in The lead-lag analysis for all exercises combined resulted in a median estimated lag of 13 min (interquartile range (IQR): −25.5 to 29.5 min). Lag appeared to be greater for resistance and HIIT exercise with a median of 18 min (IQR −26.5 to 30 min) and 19 min (IQR 4.5 to 29 min), respectively, compared to a 1-min (IQR: −26.5 to 21 min) estimated lag for aerobic exercise. Visually, however, lag was not apparent in many of the individual glucose traces.Throughout this study, a total of three adverse events occurred that were deemed unrelated to the study. One additional adverse event originated from a 3 cm redness near the CGM application site. No serious adverse events occurred.p-value of 0.02 for both CGM systems) [Overall, the data presented here demonstrate that the accuracy of the Dexcom G6 sensor was not significantly impacted by different exercise types. While a statistically significant decrease in accuracy (ARD and sensor bias) had been reported for the Dexcom G4 Platinum during moderate aerobic exercise , the Dexsystems) . Similarsystems) . Recentlsystems) .Across most time points, the Dexcom G6 CGM demonstrated a negative bias as compared to the meter, indicating an under-estimation of glucose levels. There is a clear advantage to under-estimating glucose levels as compared to over-estimating glucose levels, as the latter could be a safety concern due to the risk of undetected hypoglycemia. This is particularly true during aerobic exercise where glucose levels can drop rapidly. This CGM under-estimation trend is similar to past observations made on the Medtronic Gold, Dexcom G4 Platinum and Medtronic Paradigm Veo Enlite CGM devices ,45,47. AOn the other hand, the absolute error showed no significant impact from the different types of exercise, capillary glucose values, time since exercise start, or magnitude of the glucose drop. The observed trend in With mean ARD ranging from 11.2 to 15.9% during aerobic, resistance and HIIT exercises, this study observed somewhat reduced numeric accuracy compared to recent at-rest results of the G6 system in pregnant women with diabetes for example (mean ARD 8.7–11.5%) . Many vaA few limitations to this study should be considered. Although the overall change in glucose followed an expected trend, the magnitude of the glucose drop during exercise (maximum median drop of 2.6 mmol/L) was modest in part likely due to the participants ingesting carbohydrates prior to exercise to prevent hypoglycemia ,51. As sThis study is one of the first comprehensive evaluations of the Dexcom G6 accuracy under different types of exercises. Ultimately, our findings support the use of the Dexcom G6 system to reliably and safely assess glucose levels during 30-min exercise sessions. This is vital information with the recent FDA-approval of Control-IQ, which modulates insulin delivery based on real-time Dexcom G6 CGM data .


To compare the repeatability and reproducibility of axial and lateral retinal measurements using handheld optical coherence tomography (OCT) systems and a tabletop OCT system.Graders measured central foveal thickness (CFT), optic nerve-to-fovea distance (OFD), and retinal nerve fiber layer (RNFL) thickness on OCT scans of the right eye of 10 healthy adults. Three OCT systems were used: handheld Leica Envisu, investigational handheld swept-source OCT (UC3), and Heidelberg Spectralis tabletop system. All eyes were imaged five times with each OCT system by each of two imagers. A components of variance analysis provided estimates of repeatability (variation due to random error) and reproducibility  expressed as standard deviation and (coefficient of variation %).Repeatability of CFT (µm) for Envisu, UC3, and Spectralis was 5.9 (2.6%), 6.9 (2.9%), and 4.7 (2.1%), and the reproducibility was 6.1 (2.7%), 7.3 (3.1%), and 4.7 (2.1%), respectively. The repeatability of OFD (mm) was 0.13 (2.9%), 0.10 (2.3%), and 0.07 (1.6%), and the reproducibility was 0.13 (3.0%), 0.10 (2.3%), and 0.07  respectively. The repeatability for RNFL thickness (µm) for Envisu, UC3, and Spectralis was 4.3 (7.8%), 2.7 (5.4%), and 2.9 (4.9%), and the reproducibility was 4.5 (8.3%), 2.9 (5.8%), and 2.9 (4.9%), respectively.All three OCT systems had good repeatability and reproducibility with coefficients of variation of less than 3.5% for CFT and OFD measurements, and less than 8.5% for RNFL thickness.Our findings inform the repeatability and reproducibility of retinal axial and lateral measurements on handheld OCT and are useful for both clinical research and patient care. CFT is an axial measurement, OFD is a lateral measurement, and RNFL thickness depends on both to obtain thickness at specific lateral locations from the optic nerve head. Factors that could influence real-world CFT measurements include image quality, image tilt, foveal frame selection, variable retinal layer reflectance from different OCT systems, and segmentation error. Lateral measurements on OCT are further affected by subject eye length, scan size, image rotation (variable dependence on fast and slow axis), imager hand motion, and subject body position (sitting or supine).Our group has previously adapted a commercial handheld OCT system for infant use  and has subsequently developed our current research handheld OCT system (UC3). These handheld OCT systems have been used in our ongoing infant retinal imaging study . We are unable to conduct rigorous reproducibility studies with multiple repetitions and multiple research systems in the pediatric vulnerable population. Our goal of the current study was to evaluate the repeatability and reproducibility of these handheld OCT systems compared with a tabletop OCT system  in healthy adult volunteers. In addition, we evaluated the variance induced by body position (sitting vs. supine) of handheld OCT measurements in adult volunteers.This study was approved by the Duke University Health System institutional review board and adhered to the tenets of the Declaration of Helsinki. Informed consent was obtained from the subjects after explanation of the nature, risks, and benefits of the study. We conducted a prospective imaging study in 10 healthy adult volunteers with minimal refractive error  to compare the repeatability and reproducibility of OCT measurements of the two handheld systems  to the tabletop system . For each volunteer, two imagers (DT and XC) obtained five sets of volume scans in the right eye with each of the three imaging systems. Each set of volume scans had two volumes per set in the Envisu, five volumes per set in UC3, and one volume per set in Spectralis. For each system, 10 volume scans (five volumes for each imager) containing both the optic disc and fovea were selected for CFT, OFD, and RNFL thickness measurements . Envisu To evaluate the variance induced by positional changes (sitting vs. supine), one imager (DT) obtained five sets of volume scans in the left eye of three healthy adult volunteers in sitting position and five sets of volume scans in supine position with Envisu and UC3. Scans for CFT and OFD measurements were selected as noted earlier.Automatic segmentation of each selected volume was performed using custom MATLAB  code (DOCTRAP).Following the principles of reliability analysis,The 10 participants had an average age (SD) of 37.9 (10.4) years with minimal refractive error . Eight participants were women. All were healthy adult volunteers with no known eye issues or history of ocular surgeries.Out of the 300 scans (100 scans with 3 OCT systems) that were graded independently by 2 graders, 13 scans  were ungradable due to image quality or an image being cropped and 587 scans (97.8%) were analyzed. The mean (SD) of CFT across all three OCT systems was 227.3 (16.1) µm. For Envisu, UC3, and Spectralis, the mean (SD) of CFT was 223.0 (14.7), 236.0 (15.8), and 223.0 (13.9) µm, respectively. For CFT overall, repeatability was 6.2 µm (2.72%) and reproducibility was 9.7 µm (4.28%) . The repImager variation was the smallest for UC3  compared with Envisu  and Spectralis . Grader variation only occurred in UC3  for gradable scans . The calOut of the 300 scans (100 scans with 3 OCT systems) that were graded independently by 3 graders, 34 scans  were ungradable due to optic disc and fovea not well-identified or not both included and 866 scans (96.2%) were analyzed. The mean (SD) of OFD across all three OCT systems was 4.37 (0.37) mm. For Envisu, UC3, and Spectralis, the mean (SD) was 4.41 (0.40), 4.33 (0.38), and 4.38 (0.34) mm, respectively. For OFD overall, repeatability was 0.13 mm (2.9%) and reproducibility was 0.14 mm (3.1%) . The repImager variation was the smallest for Spectralis  and UC3 , and larger for Envisu . Grader variation was the smallest for Envisu  and Spectralis , and slightly larger for UC3  for gradable scans . The calTo evaluate the variance induced by positional change, handheld OCT imaging was performed in the left eye of three adult volunteers using Envisu and UC3. For CFT, out of the 60 scans graded by 2 graders, 9 scans  were ungradable due to image quality and unable to determine CFT and 111 scans (92.5%) were analyzed. The Envisu mean (SD) was 234.0 (24.0) µm at supine position and 229.0 (15.5) µm at sitting position, and the UC3 mean (SD) was 247.0 (20.5) µm at supine position and 245.0 (20.3) µm at sitting position. The position variation (CV) was larger for Envisu  than UC3  .For OFD, out of 60 scans graded by 3 graders, 14 scans  were ungradable due to image quality or unable to determine both fovea and optic nerve and 166 scans (92.2%) were analyzed. The Envisu mean (SD) was 4.30 (0.20) mm at supine position and 4.36 (0.23) mm at sitting position, and the UC3 mean (SD) was 4.28 (0.20) mm at supine position and 4.25 (0.21) mm at sitting position. UC3 again yielded smaller position variation (CV) of 0.00 mm (0.00%) compared with 0.03 mm (0.68%) for Envisu . We atteOut of the 300 scans graded by 1 grader, 16 scans  were ungradable due to optic disc and fovea not well-identified or not both included, poor image quality, image crop, or incomplete coverage of the area of RNFL measurement and 284 scans (94.7%) were analyzed. The mean (SD) of RNFL thickness across all three OCT systems was 54.8 (11.2) µm. For Envisu, UC3, and Spectralis, the mean (SD) was 54.3 (12.2), 50.3 (9.9), and 59.6 (9.5) µm, respectively. For RNFL thickness overall, repeatability was 3.8 µm (7.00%) and reproducibility was 6.2 µm (11.40%) . The repOur current study examined the repeatability and reproducibility of axial and lateral retinal measurements of two handheld OCT systems (Envisu and UC3) compared with a tabletop OCT system . We found that for both CFT and OFD measurements, all systems yielded a repeatability and reproducibility coefficient of variation of less than 3.5%. When compared with the handheld systems, the tabletop Spectralis system did yield a smaller variance for both repeatability and reproducibility of each measure. For all systems, RNFL thickness exhibited more variability than either CFT or OFD, as evident by the higher value of residual coefficient of variation, as it was dependent on variations from both axial and lateral measurements. For RNFL thickness, Spectralis again yielded a smaller variance, whereas all systems yielded a coefficient of variation of less than 8.5% for both repeatability and reproducibility. Both CFT and RNFL thickness are primary outcomes for the ongoing BabySTEPS study. For all three values , all three OCT systems had strong repeatability as represented by ICC values above 0.80, which reflected the small variation due to other components and an ability to detect changes between subjects.Compared with tabletop systems, handheld OCT has more potential sources of variance. First, handheld systems are often used on infants or children who do not fixate. It is frequent to observe artifacts due to eye movement, saccades, and hand movement. Second, it can be difficult to maintain a particular posture through the duration of a volume acquisition, which may be ameliorated by increasing the speed of acquisition. Third, there is a higher likelihood of image rotation as it is more difficult to determine the eye alignment. Despite this, a recent retrospective study from our group noted similar reproducibility in handheld OCT in infants compared with tabletop OCT in adult volunteers.Axial measurement is a property of the laser and is less affected by hand/patient eye movement during the acquisition process. However, it is plausible that the wavelength difference  of CFT measured by UC3 compared with Envisu and Spectralis.,Lateral measurements, however, depend on the axial length of the eye and the reference arm length, as well as the inherent eye model length of the respective OCT device.RNFL thicknesses have added complexity as they rely on both axial and lateral OCT measurements, especially in infant and pediatric eyes that are changing in size. To longitudinally follow the RNFL thickness, one may consider a different approach of measuring RNFL thicknesses: rather than a fixed distance from the optic disc, a proportion of distance between optic disc and the fovea may be considered. This may ameliorate the need to model the change in axial length and its related change in scan size.The results from our study needs to be interpreted with limited generalizability, as it was conducted in healthy, compliant adult volunteers with minimal refractive error, and not the typical patient population in which handheld OCTs are commonly used . Infants and children, with limited ability of fixation and varying axial length through different stages of development, will have inherently more variability in OCT measurements. Limitations of our study include the small sample size that failed to compare the difference of OCT measurements with positional changes and the number of observers selected for each retinal measurement. As the goal of the study was to evaluate the handheld OCT devices compared with tabletop device and their utilization in BabySTEPS, the limited number of observers may limit its inference to retinal measurements of these devices in the general population by any observer.Our study informs us that handheld OCT systems are repeatable and reproducible instruments to measure CFT, OFD, and RNFL thicknesses in healthy adult volunteers. This result will be applicable to current and future research studies and clinical practice using handheld OCT systems to evaluate changes in retinal measurements.


IntroductionDysphagia is highly prevalent in patients with a history of recurrent acid peptic disease. Endoscopy is the mainstay of diagnostic workup of these patients to reach underlying cause and appropriate subsequent treatment. The objective of the study was to determine the frequency of various types of endoscopic findings in patients with dysphagia and the association of these findings with gender, age and duration of symptoms.MethodsThis cross-sectional study was carried out in the Department of Gastroenterology of a tertiary care hospital in Rawalpindi, Pakistan. A total of 137 patients who presented with a history of dysphagia for at least two weeks were enrolled in the study. Duration of symptoms was noted, and all patients underwent upper gastrointestinal endoscopy to find out the cause of dysphagia. Tissue biopsies were obtained, and further histopathological examination was performed to correlate the findings with symptoms of dysphagia. ResultsA total of 137 patients were enrolled for six months. The mean age of the patients was 56.9 ± 17.44 years, and the mean duration of symptoms was 15.96 ± 12.31 weeks. There were 65 (47.4%) males and 72 (52.6%) females in the study. Majority of them, 123 (89.8%), presented with a short duration of symptoms that varied between 2-24 weeks and were mainly middle-aged (31-60 years) and old-aged (61-80 years). The most commonly observed endoscopic findings were esophageal stricture in 25 (18.2%), achalasia cardia in 20 (14.6%), esophageal mass in 12 (8.8%) and reflux esophagitis in 7 (5%) patients. No association was seen between age, gender and duration of symptoms and findings on the endoscopy.ConclusionDysphagia is associated with many endoscopic findings that are not related to demographic variables and must be evaluated earlier to reduce further morbidity and mortality. Dysphagia is the symptom that describes the difficulty in swallowing due to any cause . NumerouThe most frequently performed procedure for initial diagnosis and evaluation of dysphagia of esophageal origin is upper gastrointestinal (GI) endoscopy . It helpThe findings on endoscopy in patients with dysphagia are well known. In one literature review, it was found that in patients who had dysphagia, upper GI endoscopy revealed that 32.5% patients had normal findings, 25% had Schatzki's ring, 22.5% had malignant esophageal stricture, 10% had reflux esophagitis, and 5% had a benign stricture of esophagus . In anotNumerous studies have been carried out on the findings of endoscopy with dysphagia; however, the prevalence of these findings have not been well documented in local databases of Pakistan. Therefore, the current study aimed to determine the frequency of endoscopic findings in patients with dysphagia and the association of these findings with age, gender and duration of symptoms.It was a cross-sectional study conducted at the Department of Gastroenterology and Hepatology, Pak Emirates Military Hospital, Rawalpindi, Pakistan for a period of six months from July 2018 to December 2018. A sample size of 137 was calculated using the Epi-Pen calculator based on the prevalence of dysphagia to be 9% in general population-based on previous studies (8). The patients were recruited using consecutive non-probability sampling. All patients presenting with dysphagia from age 18-80 years were included in the study. Patients with a previous history of upper gastrointestinal (GI) endoscopy and any past GI surgery were excluded from the study. The study was conducted following principles of ethical medical practice as laid down in the Declaration of Helsinki after obtaining its formal approval from the Ethical Committee of the same institution.Patients fulfilling the inclusion criteria were recruited in the study after obtaining informed written consent. The patients were assessed with adequate history, thorough examination and investigations and findings were noted down on a pre-designed proforma. Endoscopy was performed by the same consultant gastroenterologist (HR), and biopsies were taken where required and were sent for histopathological analysis. The results of the analysis were recorded on the proforma and were subjected to statistical analysis. Radiography was advised in all cases to see radiological findings in patients with dysphagia.The data were entered and analyzed through Statistical Package for Social Sciences . Quantitative data such as age and duration of symptoms were presented as mean and standard deviation. Qualitative data such as gender, age groups, duration of symptoms, physical findings, radiological findings, endoscopic findings and biopsy findings were calculated as frequencies and percentages. Chi-square test was used to compare qualitative data, whereas a student's t-test was used for quantitative variables. A p-value of ≤ 0.05 was considered as statistically significant.A total of 137 patients were enrolled in this study. The mean age of the patients was 56.9 ± 17.4 years, and the mean duration of symptoms was 16.0 ± 12.3 weeks. There were 65 (47.4%) males and 72 (52.6%) females in the study. Distribution of patients in various age groups and duration of dysphagia is shown in Table Various endoscopic findings were seen in 57 41.6%) patients in the elderly (61-80 years) as described below. The most common findings in this age group were achalasia cardia in 10 (7.3%), esophagitis in 8 (5.8%) and esophageal masses in 7 (5.1%). In middle age group (31-60 years), endoscopic findings were present in 50 (36.5%) patients and the most common findings were esophageal stricture 14 (10.2%), esophageal mass 5 (3.6%) and achalasia in 5 (3.6%). Among young age group (18-30 years), positive endoscopic findings were observed in 15 (11.0%) patients and the most common findings were of achalasia cardia in 5 (3.6%), corrosive stricture in 4 (2.9%) and esophageal stricture in 3 (2.2%). Despite variability in presentations at different age groups, there was no significant association found between age and endoscopic findings . The common results in males were esophageal stricture 14 (10.2%), achalasia cardia in 10 (7.3%), esophageal mass in 5 (3.6%) and gastro-oesophagal junctional (GEJ) growth in 4 (2.9%). In females, the typical findings were esophageal stricture 11 (8%), achalasia cardia 10 (7.3%), esophageal mass 7 (5.1%) and corrosive stricture in 4 (2.9%). There was no statistically significant association between gender and the findings on endoscopy in patients who presented with dysphagia.In patients who presented with short duration of dysphagia, i.e. from 2 to 24 weeks, the most common findings were esophageal stricture in 22 (16.1%) patients, achalasia cardia in 19 (13.9%), esophageal mass in 9 (6.6%), reflux esophagitis in 7 5.1%), GEJ growth in 5 (3.6%) and esophageal stricture, esophageal candidiasis, esophageal diverticulum and peptic stricture in 4 (2.9%) each. Among patients who presented with long duration of dysphagia, i.e. from 25-48 weeks, common endoscopic findings were of esophageal stricture in 3 (2.2%) and corrosive stricture in 2 (1.5%) patients. No endoscopic finding was found in patients who presented with too long duration dysphagia, i.e. > 48 weeks. No significant association was found between the duration of dysphagia and endoscopic findings as indicated by the p-value of >0.05. The results of tissue histopathological examination are presented below , achalasia cardia in 20 (14.6%), esophageal mass in 12 (8.8%), reflux esophagitis in 7 (5.1%), esophageal diverticulum in 4 (2.9%), corrosive stricture in 4 (2.9%), esophageal candidiasis in 4 (2.9%), esophageal web in 4 (2.9%) and peptic stricture in 4 (2.9%).Esophageal stricture was seen more in patients who were of middle age  followed by in old age patients above 60 years of age 8, 5.8%). It was more frequent in males  compared to females  and was more common in patients who presented with short duration of dysphagia (2-24 weeks) as compared to patients who presented with long term (> 48 weeks) . This implies that esophageal stricture is mainly found in middle-aged males with short duration of symptoms of dysphagia. In a study by Eslick et al. [.8%. It wThe second most common finding on endoscopy was of achalasia cardia. It was commonly seen in patients who had a short duration of symptoms (<24 weeks), i.e. in 19 (13.9%). Only one (0.7%) patient with a duration of symptoms of >24 weeks had achalasia cardia. Both males and females had similar findings of achalasia cardia, i.e. 10 (7.3%) each. It was more prevalent in the old age group of >60 years age in a frequency of 10 (7.6%), whereas young and middle-aged patients had similar rates of achalasia cardia, i.e. in 5 (3.6%) each. In one study, achalasia cardia was found to be the 4th most common cause of dysphagia and was seen in 9.4% patients, and the majority of the patients were of age <30 years, i.e. 43.8%, 11.1% were of age 30-50 years and no patient above 50 years had achalasia cardia [Thirdly, esophageal mass was found as a common finding. It was more common in old age people (61-80 years), i.e. in seven (5.1%) patients followed by in middle-aged patients 5 3.6%) patients. It was more commonly seen in females seven (5.1%) and was more frequent in patients with dysphagia of short duration, i.e. 9 (6.6%). Yahya et al. reported that in patients with dysphagia, esophageal mass was a common cause, and it was reported to occur in 28.4% patients . In our .6% patieA fourth common finding was that of reflux esophagitis in 7 5.1%) patients. Reflux esophagitis was slightly more frequent in females than males, i.e. four (2.9%) and three (2.2%) respectively. It was more frequent in patients with a short duration of symptoms, i.e. in 7.1% and was more frequent in middle-aged people, i.e. in 4 (2.9%). Findings from other studies revealed similar prevalence rates in terms of gender. However, other studies showed that reflux esophagitis was more common in old age patients with duration of dysphagia for >24 weeks -5,17..1% patieThis study had certain limitations. First of all, the study was carried out in a single-centre, so the results cannot be generalized to the general population with different dietary habits. Secondly, the sample size was small, so it cannot be taken as a representation of the whole population. Thirdly, endoscopy was only used for diagnosis, and the therapeutic use of the endoscopy was not evaluated. Lastly, the database did not provide thorough details for evaluation of causes of these findings and additional testing for further diagnostic confirmation was not done. Future studies on large sample size with subjects from various backgrounds and different dietary habits must be carried out in order to look for the association between demographics and findings on endoscopy as well as with other factors that can play an essential role in the pathogenesis of dysphagia.Based on results, the current study concluded that the most common findings on endoscopy in patients with dysphagia were esophageal stricture, achalasia cardia, esophageal mass and reflux esophagitis. No significant association was found between the endoscopic findings and age, gender and duration of dysphagia. Upper gastrointestinal endoscopy must be carried out in all patients who present with dysphagia in order to look for causes like esophageal stricture, achalasia cardia, esophageal mass and reflux esophagitis , which if diagnosed earlier and treated promptly can reduce the risk of further morbidity and mortality associated with dysphagia. Endoscopy of the upper gastrointestinal tract is a safe, convenient and effective procedure for establishing the diagnosis and providing information about different treatment strategy.


Inflammatory bowel disease (IBD) is today a global disease, the incidence of which is growing in the pediatric population. This prospective study aims to decipher IBD incidence and its trend in a pediatric population through 16 years in the South Moravian Region of the Czech Republic.We evaluated data concerning 358 pediatric patients with newly diagnosed IBD at University Hospital Brno, which is a gastroenterology center for the entire pediatric population (0–18 years) and cares for all pediatric IBD patients in the South Moravian Region .P < 0.001). The overall IBD incidence per 100,000/year was 9.8 . Constituent incidences per 100,000/year were CD 5.2 (95% CI: 4.5--6.0), UC 3.4 (95% CI: 2.8--4.0), and IBD-U 1.2 (95% CI: 0.9--1.6). IBD incidence was projected to reach 18.9 per 100,000/year in 2022.The study encompassed 3,488,907 children during 16 years. We diagnosed 192 children (53.6%) with Crohn disease (CD), 123 (34.4%) with ulcerative colitis (UC), and 43 (12.0%) with IBD-unclassified (IBD-U). The incidence of IBD increased from 3.8  per 100 000/year in 2002 to 14.7  per 100,000/year in 2017 (The overall incidence of pediatric IBD in the Czech Republic is increasing, and especially that of CD, whereas trends in UC and IBD-U appear to be constant. These data highlight the need to identify risk factors involved in the rising incidence of IBD. What Is KnownThe incidence of pediatric-onset inflammatory bowel disease shows geographical variability worldwide.The incidence and prevalence of childhood-onset inflammatory bowel disease have risen rapidly in recent decades.What Is NewThis research describes an increase in the incidence of inflammatory bowel disease, mainly because of a rise in the incidence of Crohn disease, between 2002 and 2017 in a population of Czech children.Prospectively evaluated incidence rates of pediatric inflammatory bowel disease and its subtypes in the Czech Republic are among the highest in the literature.Inflammatory bowel disease (IBD), consisting of Crohn disease (CD), ulcerative colitis (UC), and inflammatory bowel disease unclassified (IBD-U), is a chronic relapsing inflammatory disorder with a multifactorial etiology. Both genetic aspects and environmental factors are important for IBD pathogenesis ,2. ApproWe have had relatively limited data on the incidence of IBD in Czech children. Therefore, it would be highly desirable to have more precise data in this field, to describe the exact trends in current developments, and to compare these data with those from other regions, especially within Central Europe.The aim of our study was to determine the IBD incidence and trends in the Czech pediatric population. The study takes in a 16-year period in the South Moravian Region within the Czech Republic Fig. .2. The health system in the Czech Republic is tax-funded and offers universal access.We conducted the study in the South Moravian Region, one of the 14 regional administrative units in the Czech Republic. The region has 1,187,667 inhabitants , constituting 11% of the Czech Republic's total population. Children account for 19% of this number, or 229,375. The region covers an area of 7188 kmThe diagnosis and treatment of pediatric patients with IBD in the Czech Republic is limited only to highly specialized centers, which are strategically located and provide comprehensive care to IBD patients. In the South Moravian Region, all regional hospitals are referring pediatric patients for a comprehensive examination to 1 center in the regional capital, University Hospital in Brno. This referral pathway is rigid and has not changed throughout the study period.Diagnosis of patients under 18 years of age by gastroenterologists serving adults is very unlikely, as in the Czech Republic, this would not be reimbursed by health insurance. The only reason for potential drop-out from our study is diagnosis of a local inhabitant outside of our region. We consider this to be very unlikely, but it does constitute a potential bias in our study. We believe the absolute majority of pediatric IBD patients from the South Moravian region are referred to our center.The inclusion criteria consisted of children having been diagnosed with IBD according to relevant guidelines ,13  by magnetic resonance enterography. All children were 0–18 years of age at the time of diagnosis and were resident in the South Moravian Region. The newly diagnosed IBD patients were subdivided into 3 main clinical types: CD, UC, and IBD-U. If a differentiation between UC and CD could not be determined after a complete workup, these patients were designated as IBD-U ,13. The Standard statistical methods were adopted for data description. Count data are summarized using absolute and relative frequencies. Median and interquartile range of nonmissing observations are reported for continuous data. Kruskal-Wallis test was used to examine between-group differences for continuous data, whereas Fisher exact test and exact rate ratio test , assuminglm function from the built-in stats package.Age-gender adjusted incidence rates are expressed as newly diagnosed children per 100,000 pediatric population per year  and reported with 95% confidence intervals based on a gamma distribution. Data regarding the size of the pediatric population were obtained from the Czech Statistical Office, which counts the number of inhabitants either in actual years or accounts for all regions’ redistributions and adjusts past numbers with regard to the regions’ present sizes. For this study, we chose to work with figures related to actual years. Incidence trends and future projections were estimated using Poisson regression. All statistical significances were evaluated on a level of α = 0.05. The entire analysis was conducted in the statistical software R. Poisson models were estimated using the P > 0.05).The basic demographic characteristics of 358 children (<19 years of age) diagnosed for IBD between 2002 and 2017 in the South Moravian Region are shown in Table P > 0.05), this is not the case for IBD (P < 0.001) and CD (P = 0.003). The predominance of the CD diagnosis after 2009 is confirmed by its being significantly higher than the UC and IBD-U incidences  in 2017, but its incidence was only higher than that of IBD-U (P = 0.016) in 2002.Incidence rates per 100,000/year for the given period 2002 to 2017 are captured in Figure P < 0.001). The difference in CD versus UC incidences also was significant (P < 0.001).A detailed overview of overall incidences by diagnosis, age, and sex is provided in Table According to the Paris Classification , there wAmong 123 patients diagnosed with UC, there were 6 (4.9%) patients only with proctitis (E1), 16 (13.0%) with left-sided  colitis (E2), and 8 (6.5%) with extensive  colitis (E3). Pancolitis was present in 90 (73.2%) patients. In 3 patients (2.4%), the extent of the disease could not be validated.P < 0.001) incidence of 14.7 (95% CI: 13.1--16.3) occurred for children aged 7 years and older. Likewise CD, UC, and IBD-U incidences among young children (<7 years) were significantly lower  than were those for older children (≥ 7 years). No significant between-diagnoses differences were found for patients up to 6 years of age. For patients aged 7 and older, the CD incidence was found to be significantly greater than were those for UC and IBD-U , which was true also for incidences of UC vs. IBD-U (P < 0.001).The IBD incidence begins to grow quickly from the age of 8. The peak occurs at 17 years of age, with decrease thereafter. Of those diagnosed with IBD, only 26 (7.3%) were children under 7 years of age, whereas the majority of child patients were at least 7 years of age at diagnosis. Early presentation of CD, UC, and IBD-U at age 6 and earlier occurred in 14 (3.9%), 7 (2.0%), and 5 (1.4%) children, respectively. The overall IBD incidence per 100,000/year for children ages up to 6 years was 2.0 (95% CI: 1.3--3.0), whereas significantly higher (P = 0.606). No significant sex-related differences were found between incidences of CD, UC, and IBD-U . Among both boys and girls, CD and UC incidences were proven to be significantly higher compared with those for IBD-U  as well as CD vs. UC incidence .Of the 358 children diagnosed with IBD, 53.1% were boys . Overall male and female IBD incidences per 100,000/year were 10.1 (95% CI: 8.7--11.7) and 9.5 (95% CI: 8.1--11.1), respectively, with no statistically significant difference between the 2 (P < 0.001). The rising trend in IBD mainly reflects significant increase in newly diagnosed CD cases , whereas neither UC  nor IBD-U incidence  showed any significant changes. The IBD incidence is projected to reach 18.9 per 100,000/year in 2022 (14.2 for CD).Figure P < 0.001) than for boys . The difference is substantially smaller for the CD diagnosis, with girls facing relative risk of being diagnosed that is 1.2% higher than in the case of boys . There appeared a significant increase in incidence over the 16-year period of girls being diagnosed with UC .The relative risk per year of being diagnosed with IBD was 2.7% greater for girls (RR = 1.072, We provide a detailed longitudinal data set describing IBD incidence and its trends in pediatric patients through the 16 years between 2002 and 2017 within a well-defined geographical area of the Czech Republic. To our knowledge, this is the most recent and comprehensive study in this field. Our results provide important insights into the high incidence of IBD and its increasing trend, which are due mainly to the rise in rates of CD. The overall IBD incidence per 100,000/year shows IBD incidence in children within the Czech Republic currently to be among the highest in the world –6,16,17.


The Taiwanese national 23-valent pneumococcal polysaccharide vaccine (PPV23) program in adults ≥75 years of age and the 13-valent pneumococcal conjugate vaccine (PCV13) program for children were implemented in 2008 and 2013, respectively. In this study we evaluated PPV23 vaccine effectiveness (PPV23VE) in the elderly, with regard to both direct protection from the vaccine itself and the indirect protection conferred by PCV13 immunization in children.The incidence of invasive pneumococcal disease (IPD) in Taiwan from July 2008 to June 2016 was collected from IPD surveillance data. A comparison of IPD incidence with a nationwide vaccination registry allowed an estimation of PPV23VE by the screening and indirect cohort methods.The incidence of IPD in adults ≥75 years of age ranged from 13.9 per 100,000 inhabitants during the period July 2008–June 2013 to 10.4 per 100,000 inhabitants between July 2013 and June 2016 . According to the screening method, PPV23VE against death within 30 days of IPD onset, all IPD, and PPV23-serotype IPD was 32.5% (95% CI: 17.5–44.7%), 33.9% (95% CI: 25.2–41.5%) and 43.4% (95% CI: 34.4–51.2%), respectively. PPV23VE with the indirect cohort method was 39.0% (95% CI: 15.5–55.9%) for all PPV23 serotypes and 71.5% (95% CI: 44.2–85.4%) for 11 serotypes included in PPV23 but not in PCV13. During the period July 2008–June 2012, PPV23VE against PPV23-serotype IPD was 55.1% (95% CI: 27.2–72.3%).PPV23 is able to prevent IPD and 30-day fatality in adults 75 years of age and older due to a combination of direct effects from PPV23 and indirect effects from PCV13. It might confer higher protection against PPV23-serotype IPD before the introduction of PCV13 program in children.The online version contains supplementary material available at 10.1186/s12879-020-05721-0. S. pneumoniae from a normally sterile body site, such as the blood, cerebrospinal fluid or pleural effusion, poses significant health threats to young children (< 5 years of age), older adults, and individuals with chronic medical conditions [Invasive pneumococcal disease (IPD), defined as the isolation of nditions , 2. In tnditions . The pednditions . Howevernditions , 6. Age-nditions , 7, 8.National 23-valent pneumococcal polysaccharide vaccine (PPV23) and 13-valent PCV (PCV13) vaccination programs were started in Taiwan in 2008 and 2013, respectively , 9. ThusWhile evidence-based recommendations form the cornerstone of the vaccination policy of the World Health Organization (WHO) and of many developed countries, such data are scarce in Africa and Asia , 11, 12.A national PPV23 vaccination program aimed at the elderly (≥75 years), with vaccines donated by the Formosa Plastic Group, was implemented in 2008 . In addiData on IPD cases in patients of all ages in whom disease onset was between July 1, 2008, and June 30, 2016, were obtained from the national IPD surveillance system, a hospital laboratory- and case-based passive surveillance system for monitoring IPD established on October 15, 2007, by the Taiwan Centers for Disease Control . The IPDThe PPV23 vaccination date was obtained from the National Immunization Information System (NIIS) database, described in previous studies , 19. TheIncidence rates of IPD were calculated per 100,000 inhabitants, and specific incidence rates by age groups  and vaccine serotype. Incidence rate of serotype 19A-IPD was specifically characterized because it emerged after 7-valent PCV introduction and tends to result in more complicated pneumonia with empyema [Pc=the proportion of cases who have been vaccinated and Pp =the proportion of the target population who have been vaccinated. Using logistic regression models, VE obtained by the screening method could control for confounding variables of age group  and sex when data on the vaccination coverage in each subgroup was available.PPV23VE was calculated using two methods: the screening method and the indirect cohort (Broome) method. The screening method, described by Farrington , 20, 21,On the other hand, the indirect (Broome) cohort design used IPD cases caused by PPV23 vaccine types (VT) as the case group and IPD cases caused by non-PPV23 serotypes (ST) as the control group  , 23. TheThe Taiwan CDC approved the protocol of this study and waived the requirement for written informed consent because of the study’s retrospective design and the use of data from administrative databases, thus, involving minimal risk to study participants.p values for trend in each age group < 0.05). Before the implementation of national PCV13 vaccination programs in children in 2013, the highest incidence of IPD in children ≤5 years occurred in the year of July 2010–June 2011, and a decrease from a peak of 21.23 cases per 100, 000 inhabitants during July 2010–June 2011 to 6.14 per 100,000 inhabitants during July 2015–June 2016 was determined . Table p < 0.05), presented with more HRMCs (p < 0.05), and had a lower proportion of PPV23-serotype IPD (p < 0.001) than unvaccinated patients. As shown in Fig. p < 0.05).The incidence of IPD in adults ≥75 years of age varied from 16.11 per 100,000 inhabitants during July 2008–June 2009 to 9.17 per 100,000 inhabitants during July 2015–June 2016  within 5 years of vaccination and 15.5% (95% CI: − 47.1 to 51.4) thereafter  than in the period of July 2008–June 2016 . We speculated that as the indirect protection emerged following PCV13 introduction, PPV23 unvaccinated elderly could benefit from lowering their risk of acquiring IPD caused by 12 serotypes common to PCV13 and PPV23, which might bias the PPV23VE estimation to be lower. On the other hand, VE waning since time of vaccination should possibly be considered.In addition, in our study, PPV23VE was higher against IPD caused by 11 serotypes that included in PPV23 but not in PCV13 (PPV23-non PCV13 VT) than against PPV23 serotype-IPD, as also found in a Spanish study . Such fiPp and Pc to bias VE estimation [The validity of VE estimation using the screening method could be influenced by the completeness of PPV23 immunization recorded in the NIIS. Although the accurate PPV23 vaccination coverage for IPD cases and elderly population might be greater than 32.8 and 41.9%, respectively, it may not result in very low or very high timation , 31. PPVtimation , 23. UnlThe adjusted VE determined by the screening method was slightly higher than the estimated VE obtained using the Broome method, perhaps because the screening method does not allow for the control of confounding factors, which can result in an overestimation of VE . MoreoveFollowing the 2006 recommendation by the WHO of a routine PCV-based immunization program in children, the IPD incidence caused by PCV VT gradually declined not only in vaccinated (direct protection) but also in unvaccinated (indirect protection) age groups , 34, 35.Vaccine protection at the population level can be rapidly estimated using the screening method whereas the duration of PPV23 protection at an individual level could be estimated by a stratified analysis of time since vaccination using the indirect cohort method , 11, 21.PPV23 is a T-cell-independent vaccine that lacks a mechanism for long-term boosting of the immune response . While POur study had several limitations beyond its observational design and the residual confounders. First, although the coverage of hospitals enrolled in IPD surveillance in Taiwan should have been 100%, detection bias may have occurred for cases identified using this passive surveillance system, thereby affecting the representativeness of the data , 17. SecIn conclusion, our results demonstrated a decrease in IPD incidence among all age groups across eight seasons covering the pre- to post-national PCV13 immunization era in Taiwan, accompanied by an increased trend of non-PCV13 ST IPD. The decline in IPD among adults ≥ 75 years is likely due to a combination of direct effects from PPV23 and indirect effects from PCV13. Using the indirect cohort method, PPV23 did confer moderate protection against PPV23-serotype IPD in adults ≥75 years before national PCV13 immunization program implemented in children.Additional file 1: Supplementary Table 1. Serotype percentage of IPD in Taiwan from July 2008 to June 2016, stratified by age group.Additional file 2: Supplementary Table 2. Accumulated coverage of PPV23 in older adults, stratified by counties and age groups in Taiwan.Additional file 3: Supplementary Table 3. Effectiveness of PPV23 in patients ≥75 years of age, estimated by the indirect cohort (Broome) method, July 2008 to June 2012 (n = 581).