Innate immunity constitutes the main defense mechanism in fish and shellfish, which is the most antique and efficient system to protect organisms against microbial infections. In addition, lower vertebrates have less evolved adaptive immune responses compared to higher vertebrates, and therefore, rely on innate immune defenses for the control of pathogens . NonetheThe objective of this Research Topic is to reflect the significant advances in the study of the immune response against viral infections in fish. This topic contributes to identifying new mechanisms of response to infections and new effector molecules against viral pathogens. In addition, these studies provide a further understanding of the resistance of lower vertebrates to pathogens infection as well as to the prevention of exacerbated immune responses.The contributing articles collected in this Research Topic are grouped into the following proposed sections related to the antiviral defense in fish:1The first mechanism of response to infection is the innate immune response which comprises a set of cells and molecules that are involved in the body’s defense against infections.Krasnov et al. were able to distinguish between pathogen-specific responses from stress and general inflammation. They found that viral responsive genes were similar to different stressors such as DNA or RNA viruses, to the injection of bacterial DNA (plasmid), and the exposure to PAMPs (CpG and gardiquimod), but they responded to inflammation and bacteria with quite a low sensitivity.In this section, we can find the advances in the understanding of the defense strategies in the Atlantic salmon against viral infections through a meta-analysis based on microarray transcriptomic analysis. Vallejos-Vidal et al. revised the immune response mechanisms of one of the most pathogenic viruses, which affects farmed Atlantic salmon, the piscine orthoreovirus (PRV). They revised how an experimental infection of PRV triggers the antiviral immune response in salmonid RBCs. In addition, they showed how PRV elicited a Th1-type response and a positive regulatory effect on the CTL-mediated immune responses. The immune mechanisms related to the phenotype of permanently PRV-infected fish and their effects on the progression of secondary infection were also reviewed.Malik et al. showed the problem around melanized focal changes “black spots” in the white skeletal muscle of farmed Atlantic salmons infected with PRV-1. The first symptoms observed, which were the “red spots”, were featured by hemorrhages and acute inflammation that progressed into chronic inflammation developing the “black spots” in the muscle of salmons. Although the etiology of the spots is unknown, in this study, the authors found in both the red and black spots the presence of macrophages and melano-macrophages associated with abundant PRV-1. Here the authors studied the polarization status of the macrophages and cell-mediated immune responses in both PRV-1-infected and non-infected fish. Altogether, in this study, the authors suggested that PRV-1 could contribute to the pro-inflammatory environment that is essential for the pathogenesis of the spots.Jia et al. shed light on the mechanisms related to the immune response involved in both sensitivity and resistance against cyprinid herpesvirus-3 (CyHV3) in common carp. The authors demonstrated, with a transcriptomic analysis, that specific innate immune mechanisms which included phagocytosis, autophagy, cytotoxicity, and virus blockage by lectins and mucins like mucin 3 (MUC3) were the cause of a marked resistance to CyHV-3 infection in a breeding carp strain.Finally, in this section, 2Pathogens are able to trigger the immune response in the host through the activation of a huge variety of determinants or molecules responsible for the defense mechanisms. In this section, the authors identified two antiviral molecules and studied their role in the immune response of fish.Mou et al. identified different antiviral mechanisms for two viperin homologs (Cgviperin-A and Cgviperin-B) in the auto-allo-hexaploid gibel carp strain against crucian carp (Carasius auratus) herpesvirus (CaHV). The C-terminal domain of CgViperin-A and CgViperin-B were found to interact with a negative herpesvirus regulator of host interferon (IFN) production, the CaHV open reading frame 46 right (ORF46R) protein, and to develop the proteasomal degradation of ORF46R via reducing K63-linked ubiquitination. CgViperin-B was also found to mediate ORF46R degradation through autophagosomes. Their findings also clarified the different antiviral mechanisms of the duplicated viperin homologs in polyploid fish, which elucidated the evolution of teleost duplicated genes.Liu et al. showed how the toll-interleukin receptor (TIR)-domain-containing adapter-inducing interferon-β (TRIF), an essential adaptor downstream of Toll-like receptor signaling, played an important role in the innate immune response. The authors showed how common carp TRIF inhibited the replication of spring viremia carp virus (SVCV) in epithelioma papulosum cyprini (EPC) cells. In addition, the authors showed that TRIF increased under Aeromonas hydrophila and poly (I:C) stimulation in vivo and under poly (I:C), lipopolysaccharide, flagellin, peptidoglycan, and Pam3CSK4 stimulation in vitro. In summary, this study indicated that TRIF plays an important role in the innate immune responses of common carp against viral and bacterial infections.3A balanced immune response can protect the organism from pathogens, but an exacerbated response can impair the immune homeostasis, leading to uncontrolled inflammation or pathogen invasion. In this section, the molecular regulation of some innate immune responses against viral infection was reviewed.Miiuy croaker antimicrobial immunity. Gao et al. showed that pathogens such as rhabdovirus and bacteria up-regulated the expression of miRNAs, showing that an up-regulation of miR-2187 was able to reduce the production of antiviral genes and inflammatory factors through targeting TNF receptor-associated factor 6 (TRAF6), therefore, avoiding an extreme inflammatory response. On the other hand, Sun et al. showed that miR-181b-2 and miR-21-1 modulated antiviral and antibacterial immunity by means of the TRIF-mediated nuclear factor-κB (NF-κB) and interferon regulatory factor 3 (IRF3) signaling pathways.MicroRNAs (miRNAs) are molecules that are extensively involved in the regulatory systems of inflammation and immune responses in mammals. However, the regulatory pathway of miRNA-mediated immune responses is not well understood in lower vertebrates. In this section, the authors showed that different miRNAs could play an adverse role in the Li et al. described the role of miR-124 in the cellular immune response of Epinephelus coioides by Singapore grouper iridovirus (SGIV) infection (entry and replication). The authors showed a significant up-regulation of the miR-124 expression after SGIV infection. Their results suggested that E. coioides miR-124 increased viral replication and negatively modulated the host immune response by targeting Jun N-terminal kinases 3 (JNK3)/p38α MAPKinasa. Their results broaden the knowledge of the host immune interactions with viruses.In the same way, He et al. performed metabolome studies of Grass carp infected with GCRV and showed that, after viral infection, most metabolites increased in three-year-old fish and decreased in five-month-old fish. In addition, those differentially expressed metabolites presented antiviral effects both in vivo and in vitro. In summary, the authors concluded that the age-dependent viral susceptibility in grass carp depended on the immune system and metabolism of the host.Metabolites are also known to regulate the immune response and the susceptibility to pathogen infections. In summary, this Research Topic compiles recent developments and research in relation to the importance of innate immunity in antiviral defense in fish. Our objective was to increase the interest of research communities in these areas of research and direct this knowledge into new perspectives and strategies related to antiviral defense across aquatic organisms.VC and MO-V wrote the manuscript with contributions from other authors. VC, MO-V and LM contributed to conception of the Research Topic. All authors contributed to the article and approved the submitted version.


The kidney has a sophisticated vascular structure that performs the unique function of filtering blood and managing blood pressure. Tubuloglomerular feedback is an intra-nephron negative feedback mechanism stabilizing single-nephron blood flow, glomerular filtration rate, and tubular flow rate, which is exhibited as self-sustained oscillations in single-nephron blood flow. We report the application of multi-scale laser speckle imaging to monitor global blood flow changes across the kidney surface (low zoom) and local changes in individual microvessels (high zoom) in normotensive and spontaneously hypertensive rats in vivo. We reveal significant differences in the parameters of TGF-mediated hemodynamics and patterns of synchronization. Furthermore, systemic infusion of a glucagon-like-peptide-1 receptor agonist, a potential renoprotective agent, induces vasodilation in both groups but only alters the magnitude of the TGF in Sprague Dawleys, although the underlying mechanisms remain unclear. In investigating the influence of hypertension on the kidney, accumulating evidence suggests that microcirculatory hemodynamic changes occur in the pathogenesis of hypertension3. Two major mechanisms of renal autoregulation have been thoroughly investigated in connection to hypertension: myogenic response (MR) and tubuloglomerular feedback (TGF), which modulate the resistance of afferent arterioles periodically at 5. While the MR activates in response to increased afferent arteriolar transmural pressure, the TGF responds to the luminal Cl- concentration increase at the distal end of the thick ascending loop of Henle via the macula densa7. Notably, a comparison of the dynamic features of the TGF between normotensive Sprague Dawleys (SD) and spontaneously hypertensive rats (SHR) showed that while the prior had more stable and periodic TGF oscillations, the latter was found to be highly irregular11. The traditional dogma that views the TGF as a single nephron event has been shifting: studies have shown that nephrons can form synchronized nephrovascular ensembles via TGF signal propagation along the common artery’s arteriolar branches17. This urges an investigation into nephron ensembles of SDs and SHRs.Renal autoregulation is a crucial aspect of microcirculation that allows the kidney to maintain a stable renal blood flow and glomerular filtration rate despite the variations in blood pressure. Nephrons accomplish this by modulating the blood flow into the glomerular capillaries18. Conventional strategies such as angiotensin-converting-enzyme (ACE) inhibitors and angiotensin-II-receptor blockers (ARBs) are renoprotective and reduce the risk of end-stage renal disease (ERSD)19. In treating diabetes, exenatide, a GLP-1 receptor agonist (GLP-1RA), is a promising pharmacological agent for reducing hyperglycemia and blood pressure, but it has also shown signs of renoprotection20. The decline in renal function threatens diabetic patients, with symptoms manifesting in the form of hypertension, sodium retention, albuminuria, glomerular hyperfiltration, and microvascular damage, to name a few21. In human and animal models, studies have shown that GLP-1RAs mitigate some of these risk factors through renal vasodilation, diuresis, natriuresis, and blood pressure reduction25. In Wistar Kyoto rats, acute exenatide infusion induced vasodilation in the kidney and increased the glomerular filtration rate (GFR)26. Moreover, infusing GLP-1 in rats caused natriuresis that is linked to the inhibition of tubular sodium reabsorption, which could potentially enhance the TGF-mediated vascular response via increased sodium chloride delivery to the macula densa27. Although these studies allude to how GLP-1RAs might affect renal hemodynamics, the real-time effects of GLP1-RA on the TGF-induced hemodynamic changes in vivo remain largely unexplored.An imbalance in the RAAS system and upregulated SGLT2 activity are major drivers of glomerular hyperfiltration and tubular hyper-reabsorption in the progression of CKD in diabetes28. Yet, Ronn et al. demonstrated that the intrarenal infusion of GLP-1 increased the renal blood flow and urinary flow rate in SHRs29. Savignano et al. observed diminished diuresis and natriuresis with no renal hemodynamic changes in response to systemic GLP-1 administration in SHRs compared to SDs30. These studies indicate that the TGF dynamic in response to GLP-1RA may differ between the SDs and SHRs. However, a study under the same experimental condition and mode of administration is required to compare the effect of GLP-1RA on the TGF-induced hemodynamics between SDs and SHR. Unfortunately, experimental approaches for evaluating TGF-mediated oscillations in vivo remain limited. Micropuncture, albeit the current gold standard, is only suitable for measuring [NaCl], single nephron GFR, and tubular flow and pressure changes elicited by the TGF in three nephrons maximally. Particularly for the TGF system, which is highly sensitive to physiological and experimental variations, a prolonged, non-contact recording method is optimal.Compared to SDs, the SHRs have reduced GLP-1R expression in the pre-glomerular vasculature, where TGF and MR modulations occur32. The LSCI technique can assess blood flow in microvessels (arising from the efferent arterioles) or the whole kidney perfusion, depending on the magnification: this is critical when the hemodynamic across the kidney surface is heterogeneous13. Such a system has been employed in the past to investigate the effect of drug distribution on the renal surface34 and to demonstrate that nitric oxide affects the TGF and myogenic-driven hemodynamics16. Recently, Postnov et al. introduced high-resolution laser speckle imaging of renal blood flow. They estimated the frequency and phase differences in rat kidney microcirculation: the angiotensin II infusion promoted stronger synchronization while acetylcholine infusion induced complete desynchronization12. Previously, we proposed a multi-scale LSCI (msLSCI) system suitable for evaluating tissue circulation at two different imaging scales simultaneously: high-zoom  and low-zoom (large field-of-view)35. Using the high-zoom component of the system, we demonstrated the TGF-induced hemodynamic changes in a population of microvessels associated with nephrons during systemic infusions of a Na-K-2Cl inhibitor (furosemide) and a sodium-glucose co-transporter inhibitor (phlorizin) in Sprague Dawleys36.Laser speckle contrast imaging (LSCI) is a robust, minimally invasive system for monitoring surface blood flow in various tissues, such as the skin, brain, and kidney, in real-time in vivo37, we aimed to discern the dynamical features of TGF-mediated vascular responses and synchronization patterns in the two rat strains. Furthermore, we evaluated the TGF-induced hemodynamic responses in vivo to the systemic administration of GLP-1R agonist (exenatide) in the presence (SDs) and low expression (SHRs) of GLP-1Rs in the preglomerular vascular smooth muscle cells.In this study, we employed the multi-scale LSCI to simultaneously assess the renal microcirculation over the kidney surface and in individual micro-vessels of SDs and SHRs. By applying the time-frequency super-resolution (SLT) analysisAnimals were weighed prior to the experiment. The average weights were 344Across all animals in SDs and SHR groups, we observed TGF-mediated changes in the blood flow dynamics across the kidney surface with the low zoom settings of the msLSCI. Scalograms obtained with SLT analysis of the blood flow clearly distinguished the TGF-mediated changes within the TGF frequency range 0.015–0.04 Hz) in both SDs and SHRs  and the intervention period. GLP-1RA induced no change in the blood pressure in the SD group from 122After evaluating the differences between SDs and SHRs during baseline measurement, we assessed how the exenatide could perturb renal microcirculatory hemodynamics. Exenatide infusion increased the BFI  and noisier (SNR) in SHR compared to SD , the SDs and SHRs exhibit different TGF-mediated oscillatory hemodynamics, measured by our parameters. The blood flow oscillation driven by the TGF mechanism in Sprague-Dawleys, and the irregular oscillations in SHRs were first observed decades ago in single nephrons with the micropuncture technique45, enabling vascular communication via TGF-mediated signals46, which potentially contributes to efficient autoregulation47. Marsh et al.48 demonstrated such interactions in a 16-nephron vascular network model and predicted synchronous patterns in paired nephrons. Along with technical advances, the modern paradigm has been nudging for ways to assess interactions among ensembles of nephron-vascular network49. Fortunately, the development of laser speckle flowmetry imaging has enabled observation of TGF-conducted nephron synchronization over the whole renal surface in identified microvessels15. With the technique, it was found that synchronized nephron ensembles exist13, and they could be altered with systemic and intra-renal drug administrations16. In hypertensive animal models, the TGF-initiated nephron-nephron interaction is altered50. Specifically, Sosnovtseva et al. found that the synchronous dynamic between the TGF and myogenic (MR) mechanisms among neighboring nephrons  is reduced in hypertensive rats14. 11 pairs and nine pairs of nephrons were measured with a servo-nulling technique in SHRs and normotensive rats, respectively, revealing important differences in nephron synchronization between kidneys of normotensive and hypertensive conditions. However, the small sample size of nephrons is a significant limitation of these previous studies. To validate the difference in synchronization patterns between the two rat strains in a larger set of nephrons, we analyzed about 30 microvessels  per animal, resulting in the assessment of 5725 (SD group) and 2947 (SHR group) non-repeating pairwise combinations of vessels. Our results found that 20 % of the possible pairs were synchronized in the SD group, while only 10 % of the possible pairs were synchronized in SHRs for 10 % of the observation period. For context, a pair of vessels that are synchronized for a mere 32% of the observation period (30 min) still exhibits a high degree of signal synchronization , activating the TGF system55, which can be renoprotective. In hypertensive rats, GLP-1RA did not have an effect on the TGF magnitude, even though there was a substantial increase in the BFI and a decrease in blood pressure. The unchanged AUC in SHRs could indicate reduced autoregulatory capacity when the kidney is faced with acute systemic pressure changes. Possibly, the TGF-mediated hemodynamic change did not occur in the SHR group due to reduced GLP-1R expression in the smooth muscle cells of preglomerular vasculature, as supported by Jensen et al.28. However, other pathways that GLP-1RAs could affect renal hemodynamics should be considered in interpreting our results. For example, clinical evidence suggests that GLP-1RA might counteract angiotensin II activity (RAAS)57, which is a sign of renoprotection against diabetic nephropathy58. Plus, certain neural and metabolic pathways are also thought to be involved in GLP-1RA-induced renal hemodynamic changes59. Further investigation is needed to discern their independent contribution to renoprotection. The efficacy of GLP-1RA on the kidney may also differ based on the mode of delivery60, the duration of treatment, and the severity of renal function impairment23. A future study with longer treatment durations and measurements at various stages of chronic kidney disease progression could address some of the variability mentioned above. In a mouse kidney model of diabetic nephropathy, GLP-1Rs were found in the glomerular capillaries and vascular walls but not in the tubules61. It would be interesting to investigate how acute GLP-1R activation affects the renal hemodynamics in these mice, furthering our understanding of how GLP-1RA may aid in slowing down the functional degradation in diabetic nephropathy. Finally, we acknowledge that there is an inherent limitation of our imaging system. Since we only access the cortical surface of the kidney, the results presented here lack insights into the deeper cortical and medullary hemodynamics.Although GLP-1RA is primarily used as a treatment for type 2 diabetes, clinical studies have shown that it can also be renoprotectiveTo summarize, the microvascular network of the kidney is sophisticated in function and structure. This study demonstrates that systemic hypertension adds another layer of complexity to the hemodynamics mediated by the TGF, not just in terms of individual nephrons but also as synchronized nephron ensembles. Furthermore, the present study unveils some in vivo functional relevance of GLP-1R in the preglomerular vasculature: the enhancement of TGF-mediated hemodynamic oscillation in normotensive rats but not in hypertensive rats. Finally, our data suggest that GLP-1-induced renoprotection does not arise from TGF-mediated microcirculatory changes in hypertensive rats. Nevertheless, many questions remain as to the efficacy of GLP-1RA in renoprotection, and this study is just one piece of the puzzle.The rats were anesthetized with a 5 % sevoflurane gas mixture in an induction box. After the toe pinch test, the animals were laid ventral side up and maintained under anesthesia at 1.8 % through a gas mask. The body temperature on a surgical heating table was maintained at 3762.The drug was obtained from Santa Cruz Biotechnology in a powder form with a 97 % purity. A 50 Experiments alternated between SDs and SHRs each day to minimize surgical bias. Each experiment began with a control imaging period of 30 min. Then, the exenatide infusion was initiated at 50 35. One video zoom lens was set to 4.5 X magnification  and the in-depth data analysis can be found inK as the ratio of the standard deviation to the mean pixel intensity for a temporal kernel of 25. Finally, we calculated the blood flow index (BFI) using All data were analyzed offline using MATLAB (MathWorks). Before the speckle contrast analysis, the high-zoom raw data underwent an image registration step for motion correction, using the myograph wire as a reference mark. The low zoom data did not require a motion correction because the movements were sub-pixel, i.e., the range of translational movement in the x-y plane was within 5.5 37. SLT was selected over continuous wavelet transform due to its superior performance in localizing transient oscillations in both time and frequency of biological data. The frequency of interest was set from 0.01 to 0.05 Hz at 80 bins, with wavelets spanning 15 cycles and the minimum and maximum order set to 1 and 50, respectively. To produce a spatial frequency map in Fig. Low zoom data analysis: first, a binary mask was created with an intensity threshold to exclude pixels covering non-tissue areas such as the kidney frame, fat tissue, and the myograph wire in the blood flow data. Then a Gaussian filter was applied to every frame and spatially averaged to produce a single blood flow time series representing the global renal surface blood flow. To produce the scalogram and the power spectra shown in Fig. BFI was calculated by temporally averaging the blood flow time series over the observation period;The AUC was computed by approximating the integral of the power spectrum of the filtered signal via the trapezoidal method between the frequency of interest (0.015–0.04 Hz);The signal-to-noise ratio (SNR) was calculated by taking the ratio between the signal’s power and the noise’s power.Frequency of the most prominent (dominant) peak in the range of 0.015–0.04 Hz;Prominence measures how much the peak stands out due to its intrinsic height and its location relative to other peaks detected within 0.015–0.04 Hz;High zoom data analysis: individual microvessels (star vessels) were segmented, and blood flow time series were extracted from each vessel. 322 and 227 vessels were segmented for the hemodynamic analysis in SDs and SHRs, respectively. To produce time-averaged power spectra for each vessel, the SLT analysis was applied to individual vessel time series with the same settings as the low zoom analysis. After, several TGF signal parameters were extracted to analyze the TGF-driven hemodynamic changes:All possible pairs were compared to analyze the synchronization among observed vessels in each animal. 5725 and 2947 vessel pairs were analyzed in SDs and SHR groups, respectively. For vessel pair A and B, instantaneous frequencies were computed and parsed for consecutive durations of matching dominant frequencies, referred to as “synchronized duration” henceforth. Two criteria were implemented for calculating the synchronized duration: (1) two synchronized durations interrupted by less than the shortest possible period (25 seconds) were concatenated into a single duration, and (2) any synchronized duration of fewer than two periods was disregarded. To compare the synchronization between SDs and SHRs, the synchronized duration was plotted as a percentage of the observation time of each experiment, and the number of synchronized pairs was plotted as a percentage of all possible pairs. Figure All possible pairs were identified to measure the similarity among observed vessel pairs in each animal, and cross-correlations of respective pairs of filtered signals were computed. Their Euclidean distances, in pixels, were also calculated. Finally, the signal similarity was plotted as a function of distance in Fig. Physiological measurements  were compared using paired t-test (two-tailed). Distributions of TGF parameters were tested for normality using the Anderson-Darling test, and all rejected the null hypothesis of normality at the default 5% significance level. Non-parametric tests were used for subsequent comparisons of non-normally distributed data. The parameters between SD n = 322) and SHR (n = 227) groups were compared using the two-sided Mann–Whitney U test . All methods were performed in accordance with the relevant EU guidelines and regulations. The study is reported in accordance with ARRIVE guidelines. The animals received a 7-day acclimatization period and were caged in a 12-hour light cycle facility with ad libitum access to food and water. Male Sprague Dawleys  and spontaneously hypertensive rats  were used for this study .Supplementary Information.


Objective: To evaluate the neutrophil-to-lymphocyte ratio (NLR) values’ possible predictive role in fatal and severe cases of COVID-19 disease in pregnant women. Design and data collection: A case-control study was conducted with the inclusion of 45 pregnant COVID-19 patients. All the data were obtained from the hospital information system of Semmelweis University by two of the authors. Results: Statistical analyses showed that NLR values were significantly higher in patients with fatal COVID-19 compared to those who survived the disease, with or without mechanical ventilation. The study also assessed whether NLR values measured on the first day of hospitalization or at their peak provided better markers of disease severity. While both the first-day and peak NLR values were evaluated in patients who did not survive the disease, only the peak NLR values had predictive value regarding patient death. Conclusion: Based on our results, the peak NLR values appear to be useful markers of COVID-19 severity, with a cut-off value of 18.05. However, the authors suggest and hope that larger sample size studies will be conducted to further validate the findings of their research. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or COVID-19) is a ribonucleic acid (RNA) virus that belongs to the Betacoronavirus genus, group 2 ,4. The mTo predict the severity of SARS-CoV-2 virus infection, various methods have been used since the occurrence of the disease ,8,9. OneIn this study, we present the cases of 10 COVID-19 patients who were critically ill during their pregnancies and lost their lives due to the infection, thus evaluating whether the NLR has a prognostic value in the screening of potentially fatal COVID-19 in pregnant patients. We compared the data of the above-mentioned 10 patients with two control groups: one, which consisted of pregnant patients who also needed invasive or non-invasive ventilation but survived the COVID-19 disease, and another control group, in which the pregnant patients also survived but had moderate COVID-19 disease. During the clinical work with COVID-19 patients, our researchers had the hypothesis that those patients who face more severe outcomes due to their infections have higher NLR values on the day of the hospital admission and on the day when the NLR is peaking. Our objective was to evaluate the predictive value of the NLR in assessing the severity of illness in pregnant COVID-19 patients.In this case-control study, 45 patients who received treatment at the clinics of Semmelweis University in Budapest, Hungary, were identified. Semmelweis University is one of the Hungarian COVID-19 centers specializing in the treatment of severe COVID-19 cases in pregnant patients. As a result, the university encountered a higher number of such cases, and nearly all fatal cases involving pregnant COVID-19 patients in Hungary were treated at our institution. The SARS-CoV-2 virus positivity was confirmed using real-time quantitative polymerase chain reaction (RT-q-PCR) technology. Sampling was completed with nasopharyngeal COVID-19 swabs from the upper respiratory tract.n = 15); (2) COVID-19 patients who were hospitalized but did not require respiratory support, only closer medical surveillance (n = 20). From the patients enrolled in the study, all available NLR values and comorbidities were collected from documentations.Two authors, DS and BM, independently collected the data from the hospital information system  of Semmelweis University. A total of 10 pregnant patients with fatal COVID-19 infections were identified and enrolled, and an additional 15 and 20 patients, who survived the disease, were enrolled into two control groups. The control groups were defined as follows: (1) pregnant patients who experienced severe symptoms and required respiratory support (either invasive or non-invasive) but survived COVID-19 disease  of the SARS-CoV-2 virus, as the beta variant was the most dominant during the 3rd wave, while the remaining cases were possibly linked to the delta (B.1.617.2) variant.p < 0.05 both in the Kolmogorov–Smirnov test at the determination of probability and later in other statistical methods. All p-values were adjusted using the Holm method [After obtaining the neutrophil and lymphocyte values, their quotient was taken and measured between groups with the help of IBM SPSS Statistics 29. The measured variables were presented in the tables in the format of mean ± standard deviation (SD). The Kolmogorov–Smirnov test was used to determine if the numerical data matched the normality distribution. Variables were compared using Wilcoxon rank-sum test and Fisher’s exact test. Differences were considered statistically significant at m method .Receiver operating characteristic (ROC) analyses were also carried out to test the sensitivity and specificity of NLR. The ROC analyses were performed in the R for Windows environment  using the pROC R-library (v1.18.0). The coordinates for the best threshold were identified using Youden’s J statistic. Moreover, the effect of various clinical characteristics on NLR values was investigated. The latter analysis was performed using ANCOVA and mixed-effect  models.The 45 patients were divided into the three groups, as described above. Age, gestational age when the infection started measured in weeks, first-day NLR values, and peak NLR values of the study participants are presented in p = 0.3926) nor when comparing to moderate COVID-19 cases . In the case of the peak NLR values, the NLR values of the fatal cases were significantly higher, compared to both control groups. Fatal COVID-19 cases had a peak NLR value of 33.77 ± 14.03, while the ventilated but surviving group had an NLR value of 13.35 ± 11.43 (p = 0.0006). Furthermore, the same could be observed during the comparisons of the fatal and non-ventilated groups , no significant difference could be justified neither in the case when comparing to those patients who needed ventilation support and recovered  and those who needed ventilation support therapy (Group 2) were included. For the first-day NLR values, a cut-off value of 15.22, a sensitivity of 50%, a specificity of 93.33%, and an AUC value of 66.0% were found . The comIt was investigated whether any of the clinical parameters had an effect on NLR values. Two types of models were constructed for both first-day and peak NLRs. First, an ANCOVA model was used to explore the effects of age, gestational week when COVID-19 occurred, and the number of comorbidities. The analysis revealed that neither age, gestational week, nor the number of comorbidities had a significant impact on NLR values . The expSecond, mixed-effect models were constructed to assess whether the number of comorbidities introduced additional variance to the NLR values. The findings indicated that no additional variance was introduced, neither for the first-day NLR nor for the peak NLR values .Neutrophil-to-lymphocyte ratio and its elevation is a widely studied predictive marker in COVID-19: Damar Çakırca et al. collected retrospective data from COVID-19 patients in 2021 and found that NLR is an effective marker for predicting pneumonia in SARS-CoV-2-infected patients . Fois etNLR values are not only elevated in COVID-19 but also in physiological, normal pregnancies , and furOur research group previously reported the case of a 33-year-old pregnant kidney transplant recipient patient. During her COVID-19 illness, her NLR values were elevated, she required non-invasive respiratory support, and she exhibited pre-eclamptic symptoms . After rOur hypothesis is that monitoring NLR values could help physicians initiate potential treatment methods earlier ,36,37. WEven though the NLR value is a useful marker for detecting fatal outcomes, differentiating between surviving patients with and without ventilation is not feasible. In the ventilated group, we observed higher values on the first day of hospitalization, and the mean of the maximums was also higher. However, these results are not significant, likely due to the wide range (high variance) of measured NLR values. These data suggest that NLR values are suitable for differentiating critical cases from others, but they are not useful markers for predicting which patients will require ventilation support and which patients will not.We have reported comparisons of NLR data from three groups on two occasions. Despite the low number of cases, this study underscores the potential prognostic significance of NLR value measurements in pregnant COVID-19 patients. Semmelweis University, as a central hospital in Hungary’s COVID-19 healthcare system, allowed us to present NLR values from 10 pregnant mothers who lost their lives due to the infection. While 10 cases may be relatively high compared to other studies, we encourage fellow researchers who have encountered these unfortunate situations to publish NLR values or other potential markers. This collective effort can aid in reducing the number of such fatalities and contribute to the fight against COVID-19 in pregnant patients.It is important to note that among the patients who died due to COVID-19, there were more comorbidities present, which could have also contributed to the further elevated NLR levels in this group. It is also important to note that, due to the limited number of patients included, ROC statistics may be less reliable in this context.Since the data presented in this study were collected retrospectively, it was not feasible to perform analyses on specific COVID-19 variants. The typification of COVID-19-infected patients was conducted by the Hungarian Public Health Center. The authors do not have access to these data due to Hungarian law; thus, our conclusions regarding which variant infected each patient were based on national statistics, which identified the dominant variant at the time of infection.Our research has revealed that NLR values are valuable for detecting potentially critical and fatal cases of COVID-19, even on the first day of hospital admission. Early identification of elevated NLR values may assist physicians in more closely monitoring these patients and initiating appropriate treatment sooner. It is also essential to emphasize, based on our findings, that NLR values are not effective markers for predicting which patients will require ventilatory support. We encourage other researchers to publish their findings on the same subject, which could potentially lead to the inclusion of a larger number of patients in future studies.


Traditional wood-based composites are bonded with synthetic formaldehyde-based adhesives ,2. These2) and micro-silica (mSiO2) [Identifying additives to reduce the total amount of urea-formaldehyde (UF) resin needed without adversely affecting the panel properties is one way to reduce the negative environmental footprint of UF resins caused by the release of formaldehyde. The results provided by Taghiyari et al.  showed t (mSiO2) .One of the possible directions to achieve this goal is the creation of wood composites based on environmentally friendly products, where thermoplastics and their copolymers  are used as adhesives ,37,38,39This Special Issue, entitled “Recent Developments in Eco-Friendly Wood-Based Composites II”, comprises 12 high-quality original research and reviews papers by 62 authors from 10 countries on three continents: Asia , Europe , and Africa (South Africa). The papers provide examples of the most recent developments in eco-friendly wood-based composites.In their paper, Xiao et al.  applied A very interesting study by Bekhta et al.  aimed to3; TS decreased from 3.4 to 1.8%; porosity decreased from 7.0 to 3.0%; MOR increased from 8.19 to 12.26 MPa; MOE increased from 1.67 to 2.87 GPa; and compressive strength increased from 16.00 to 27.20 MPa. The thermal properties of the biocomposite also improved as the particle size reduced. The authors also found that the performance of the biocomposite improved significantly with a lower percentage matrix and filler nanoparticle rather than when increasing the percentage of the matrix. The finding of this research also indicates that the properties of the biocomposite can be improved by reducing the particle size of the filler to nanometers without having to increase the adhesive composition.In the study by Ismail et al. , a new aThe findings of another study  demonstrIn another paper, low-cost wood–plastic composites (WPCs) without any additives were developed from invasive trees without prior processing and low-grade recycled low-density polyethylene . The aut3 were self-bonded. The authors found that widely available bark residues could be successfully utilized as an innovative raw material for efficient eco-friendly thermal insulation products. The thermal conductivity values of the boards were comparable to established insulation boards based on cork or wood fibers. Based on the measured thermal conductivity and zero formaldehyde content, bark fiber insulation panels might be able to compete with conventional insulations if the density can be further reduced, and applications regarding acoustic insulation are also a possibility.In another interesting study, the first attempt to investigate low-density insulation boards made of spruce bark fibers in a wet process was conducted . The insOil palm trunks (OPT) are considered significant waste products. Usually, the trunks remain on the plantation site for nutrient recycling or burning, which increases insect and fungi populations. This causes environmental problems for the new palm generation or air pollution due to fire. Therefore, the comprehensive review conducted by Nuryawan et al.  summarizNanocellulose aerogels are a new category of high-efficiency adsorbents for treating oil spills and water pollution. The review provided by Iskandar et al.  presentsIn their comprehensive review paper, Ramesh et al.  focused The processing technology, bonding mechanism and performance of thermoplastic-bonded wood-based panels are comprehensively summarized and reviewed in another interesting paper . MeanwhiIn recent years, different types of thermoplastic films such as polyethylene, polypropylene, polyvinyl chloride, co-polyamide and co-polyester have been widely used for wood veneer bonding owing to their excellent water resistance, flexibility, easy processing, and secondary melting characteristics. The findings of another study  demonstrA very promising direction is the use of lignin and its derivatives as an ecological alternative to petroleum-based adhesives. However, being the most common renewable source of phenolic compounds of natural origin, only 1–2% of the huge annual production volume (50–70 million tons) is actually used for the production of value-added products. Lignosulphonates (LS) account for 90% of the total market of commercial lignin. In their review paper, Gonçalves et al.  carried The papers from this Special Issue represent only some of the recent developments in eco-friendly wood-based composites. The utilization of recycled plastics, lignin and their derivatives, wood (bark) and agricultural wastes to manufacture wood composites as well as traditional WPCs is highly viable concerning eco-friendliness, and contributes to the improvement of the circular economy. It also saves the usage of virgin materials, thus enhancing sustainability in the production of composite materials. However, most of the proposed methods to manufacture high-performance, eco-friendly wood composites with a lower environmental impact have been studied only in laboratory conditions, can only find use in some nonstructural applications, and have not been introduced in large-scale industrial production as yet. Hence, further research is still needed in order to develop methods for improving reactivity and the selection of suitable crosslinkers for lignin-based adhesives, and modification methods to improve the interfacial adhesion between hydrophilic wood and hydrophobic thermoplastics in order to expand their use in some exterior and structural applications.


The rapid development of coronavirus disease 2019 vaccines during the pandemic has left their long-term effects largely unknown. Instances of autoimmune and subacute thyroiditis showing features of autoimmune/inflammatory syndrome induced by adjuvants have been reported post-vaccination. This case report aims to highlight the autoimmune/inflammatory syndrome induced by adjuvants syndrome after coronavirus disease 2019 vaccination, drawing attention to a possible connection with thyroid dysfunction and urging for further thorough research.We present a case of thyroiditis induced by the COVISHIELD vaccine in a 37-year-old Indian woman. An apparently normal and healthy adult woman developed neck pain and easy fatigability 2 weeks after the second dose of COVISHIELD, which gradually increased and was associated with irritability, decreased sleep, excessive sweating, tremor, palpitation, and weight loss. She presented to the outpatient department after 1 week of symptoms and was evaluated with laboratory tests and imaging. She was diagnosed with thyroiditis due to the coronavirus disease 2019 vaccine and was treated with propranolol.This case report adds to the growing evidence of coronavirus disease 2019 vaccine-related thyroid issues. The development of thyroiditis is rare and underreported post-coronavirus disease 2019 vaccination; hence, research to evaluate the association of coronavirus disease 2019 vaccines with thyroid dysfunction needs to be done in the future.The online version contains supplementary material available at 10.1186/s13256-023-04279-0. The novel coronavirus disease 2019 (COVID-19), which is caused by the virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has resulted in pandemic worldwide. It created a devastating effect on the demographics of the world, causing 3.8 million deaths worldwide [Several adjuvants are employed to enhance the vaccine responsiveness, and they can induce autoimmune and inflammatory responses by promoting immunogenic cross-reactivity in genetically predisposed individuals , 4. SeveA 37-year-old Indian woman who was apparently normal before the COVID-19 vaccination presented to the outpatient department (OPD) with complaints of pain in the neck for 2 weeks, which were present throughout the day. This was accompanied by easy fatigability, irritability, decreased sleep, increased sweating, tremor, palpitation, and weight loss of approximately 3 kg in the previous 3 weeks. One week before the onset of these symptoms, she had been administered the second dose of the COVISHIELD vaccine. There were no records of any viral or respiratory sickness prior to the emergence of the symptoms. The patient received the first dose of COVISHIELD 3 months before the second dose. The patient had no history of adverse events to the first dose of COVISHIELD, except for 2 days of mild fever that resolved spontaneously. The patient had no history of any comorbidities such as diabetes mellitus, hypertension, or cardiac conditions. There was no personal or family history of autoimmune disorders. She was a non-smoker, non-alcoholic, and a teacher by profession. On thyroid examination, there was no visible swelling; however, upon palpation, tenderness was present. Primary hyperthyroidism was found in the thyroid function test. On ultrasound of the neck, multiple colloid cysts on the right lobe of the thyroid along with ill-defined hypoechoic areas in both the lobes were identified. Thyroid scan report was suggestive of thyroiditis with decreased uptake of technetium  guidelines (Additional file This case adds to the growing evidence of COVID-19-vaccine-related thyroid issues. While this report highlights the need for further investigation, it also serves as a reminder of the importance of continuous monitoring and prompt reporting of potential vaccine-related adverse effects.Additional file 1. CARE Checklist.


A long-acting κreceptor agonist parenteral analgesic may theoretically improve acute pain and reduce incidence of chronic postsurgical pain (CPSP) after laparoscopic cholecystectomy with minimal drug-related side effects of the traditional μreceptor opioids.n = 40) or placebo (n = 40) after anesthesia induction. Standard multimodal analgesia (MMA) was administered for postoperative pain control. The primary endpoint was pain intensity within 7 days after surgery. The secondary endpoints were incidence CPSP at 3 months and adverse reactions up to 7 days after surgery.Eighty adult patients undergoing elective laparoscopic cholecystectomy were randomly assigned to receive single intramuscular injection of an extended-release sebacoyl dinalbuphine ester . Significantly higher incidence of drug-related adverse events, including dizziness, nausea and injection site reactions, were recorded in the SDE group.The highest visual analogue scale (VAS) and area under the curve of VAS 0 to 48 hours after operation were not different between the two groups and a similar proportion of patients requested rescue parenteral analgesics. Average pain intensities were also not different at 72 hours and 7 days after surgery. Incidence of CPSP was 22.5% and 13.1% in patients who received placebo and SDE treatment, respectively (A single dose of extended-release analgesic SDE given intraoperatively did not provide sufficient add-on effect for acute and chronic pain management after laparoscopic cholecystectomies in patients who received standard postoperative MMA. Intramuscular injection of 150 mg SDE in patients with average body mass causes adverse events that could have been overlooked. More clinical studies are warranted to determine the target populations who may benefit from SDE injections for improvement of acute and chronic postsurgical pain management. Patients may still suffer from acute postoperative surgical-related pain even with the minimally invasive laparoscopic operations. It has been found that up to 80% of patients undergoing laparoscopic cholecystectomies complained of moderate-to-severe pain within the first day after the operation. Most importantly, 20% of the patients may develop persisting chronic postsurgical pain (CPSP) within 1 year postoperatively and they require prolonged use of opioid analgesics to control the CPSP. Several important clinical observational studies have suggested that inadequate control of surgical-related pain at the initial week after cholecystectomy is one of the major risk factors for developing CPSP. Since the length of hospital stays after uncomplicated laparoscopic cholecystectomies are relatively short ,,6 postoperative pain is inevitably managed by repeated doses of oral form analgesics, including opioids. Management of postoperative pain using a multimodal analgesia (MMA) approach is widely recommended by the international scientific societies by administration of drugs with differing mechanisms of actions target pain pathways resulting in additive and/or synergistic effects to opioids. Therefore, a long-acting parental analgesic in addition to the standard MMA could be a viable strategy to achieve optimal control of acute postoperative pain after laparoscopic cholecystectomies in patients who are discharged early (within 24 hours). This strategy could also reduce the incidence of CPSP as plasma therapeutic levels of the parental analgesic can be maintained for several days after injection.Cholecystectomies are a common procedure for the surgical removal of inflammed or otherwise pathological gall bladders and most cholecystectomies are performed laparoscopically in Taiwan (>85%). Sebacoyl dinalbuphine ester  is the prodrug of nalbuphine, and was approved by the Taiwan FDA in 2017. After intramuscular injection, SDE is hydrolyzed by tissue esterase to release the active compound nalbuphine. The systemic bioavailability of nalbuphine is 85.4% with a mean absorption time of up to 145 hours. Therefore, a single parenteral injection of SDE may provide a prolonged analgesic effect for 6 days. Currently, there have been 3 prospective randomized double-blind trials investigating the efficacy and safety of SDE for pain management after hemorrhoidectomy, laparotomy and laparoscopic bariatric surgery. The clinical outcomes of these previous studies show that SDE is superior to the conventional or patient-controlled analgesia for the management of acute postoperative pain with less need for rescue analgesics and more tolerable adverse drug reactions. However, patients who were assigned to control groups in the first 2 studies did not receive the standard MMA treatment, and periods of observation for adverse events were not specified in the study protocols. Furthermore, SDE was administered at least 12 hours before surgical intervention in these studies, which can be a major limitation for the application of SDE in the day-case surgeries, such as laparoscopic cholecystectomy. Therefore, this clinical study was designed to determine the extended analgesic effect of SDE administered after anesthesia induction as a component of MMA for laparoscopic cholecystectomy and evaluate the drug-related adverse events up to 7 days after administration.Nalbuphine is a semi-synthetic opioid that acts as a mixed κreceptor agonist and μreceptor antagonist, but its clinical applications for acute postoperative pain is limited by its relatively short duration of action of 3 to 6 hours.This phase II/III randomized, double-blind, placebo-controlled clinical trial was conducted in E-Da Hospital and E-Da Cancer Hospital  from August 2021 to April 2022. Adult patients aged 20 to 65 years with American Society for Anesthesiologist physical statuses ≤ III who were scheduled for laparoscopic cholecystectomies. This study included patients who were anticipated to stay in the hospital for > 72 hours after operation for feasibility to observe the analgesic effect and potential drug-related adverse events following administration of study drugs. The study was approved by the institutional review board of the E-Da Hospital, Kaohsiung, Taiwan . The trial was registered in March 2021 with Clinicaltrials.gov registration: NCT04808544. All participants provided written informed consent before the commencement of the study.Patients who were scheduled for laparotomies or urgent operations and patients with chronic opioid use, with allergies to Nalbuphine, benzyl benzoate, or sesame oil were excluded from the study. Patients who might require ventilator support after surgery and those who were not able for assessment of pain intensity or refused to participate in questionnaire surveys were also excluded. and other clinical trials.–13 Patient’s treatment allocation was concealed in an opaque envelope and the envelope was opened after the patient was admitted to the holding area of the operating room by the research assistant.Participants were randomly assigned to receive SDE  or vehicle solution (2 mL of mixture of benzyl benzoate and sesame oil) in permuted blocks of 10 using a computer-generated list. Dose of SDE used in this study was derived from the previous pharmacokinetic study,14 Parenteral opioids, including morphine and fentanyl, were individually prescribed by the anesthesiologist according to the patient’s hemodynamic changes and nociceptive responses during operation. At the end of operation, the endotracheal tube was extubated in the operating room. Patients were transferred back to the original wards from the post-anesthetic care unit. Patients were commenced on regular oral analgesics after resumption of normal oral intake, including acetaminophen and non-steroidal anti-inflammatory drugs (NSAID). Rescue parenteral analgesics, including opioids, parecoxib and NSAIDs were prescribed by the anesthesiologists or surgeons when visual analog scale (VAS) remained > 4 after regular oral analgesics were administered. The intraoperative MMA regimen, including SDE, parecoxib and propacetamol were supported by the research funds. The vehicle solutions were generously provided by the Lumosa Therapeutics Co. Ltd. .SDE  and the vehicle solution (solvent containing benzyl benzoate and sesame oil) was prepared in an identical injection syringe within 30 minutes before injection. The research drugs were prepared by a research nurse who did not involve in the clinical care. The surgical procedure was performed under endotracheal intubation general anesthesia and controlled ventilation. Anesthesia was maintained by inhalational anesthetics (desflurane or sevoflurane) at optimal minimal alveolar concentrations and bispectral index levels. After anesthesia induction, SDE or the vehicle solution was injected into the gluteus muscles by the in-charge anesthesiologist under ultrasound guidance. Since study drugs were prepared by the research nurse, the patients, anesthesiologists, surgeons, and research team members who assessed the study outcomes were blinded to the treatment groups. Dexamethasone (5 mg) was given intravenously after induction of anesthesia and intravenous Palonosetron (0.25 mg) was administered in patients who had high risk of developing postoperative nausea and vomiting. Parecoxib (40 mg) and propacetamol (2 g) were administered intravenously at the end of the operation as the standard postoperative MMA. Areas under the curve (AUC) of the VAS were calculated by plotting the individual VAS score as curves in which the x-axis represented measurement time points from baseline  to 48 hours after surgery, and the y-axis represented the VAS score. The secondary endpoints included total doses of rescue parenteral analgesics administered during hospital stay period, length of stay after operation, incidence of CPSP and patient’s quality of life (QoL). CPSP was defined as the newly developed pain at the surgical site or incisional wounds within 3 months after surgery, as other etiologies of pain were excluded. QoL was evaluated by the 12-item short form (SF-12) questionnaire on hospital admission, and at 7 and 90 days after surgery. The scores of SF-12 questionnaire range from 0 to 100, and lower scores indicate more impaired health-related QoL. Systemic and focal drug-related adverse reactions were recorded up to 7 days after surgery.The primary endpoint of this study was the intensity of surgical pain from post-anesthetic care unit to 7 days postoperatively. Pain intensity was assessed by ward nurses using the visual analog scale  in 4-hours intervals from 0 to 48 hours after operation. The numeric rating scale (0 represents “no pain” and 10 represents “worst pain”) was used to evaluate the pain intensity at 72 hours and 7 days postoperatively by the outcome assessors who did not participate in the clinical care. Acute postsurgical pain was defined as unpleasant sensation related to tissue injury during the acute phase after surgical intervention. In comparison to laparotomies, the intensity of postoperative pain is generally reduced in laparoscopic procedures. Therefore, our study proposed that SDE injection might reduce surgical related pain by 20% after laparoscopic cholecystectomies. With a pre-defined 20% standard error, a total sample size of 80 patients (including a dropout rate of 10%) was desirable to detect a 20% difference in pain reduction with an α value 0.05 and statistical power 0.8. Categorical variables were compared using the Fisher’s exact test. Comparisons of the continuous variables between the 2 study groups were analyzed using the Student t test or Mann–Whitney U test, as appropriate. Two-way repeated measures ANOVA was used to compare SF-12 scores at baseline and after surgery. All analyses were carried out using the SAS software, version 9.1 .A recent study showed that SDE significantly reduced the VAS by 32% at 48 hours after elective laparotomy surgery.During the study period, a total of 163 patients were evaluated for inclusion and 83 patients were excluded due to ineligibility and patient refusal  (Table P = .893) , and fewer patients required rescue analgesics from 24 to 48 hours postoperatively in both groups  (Table P = .379)  and 26 (68.4%) patients in the placebo and SDE groups, respectively  and nausea (defined as feeling of retching or stomach distress with an urge to vomit) was found in patients who received SDE within 72 hours post-operatively  for acute pain up to 7 days after laparoscopic cholecystectomy. In comparison to the previous controlled studies,–13 there were 2 main differences in our study design. First, the study drug was injected after induction of anesthesia in this study rather than administered > 12 hours preoperatively. Administration of SDE after anesthesia induction may not only reduce the discomfort of deep intramuscular injection in conscious patients, but also allows the treatment for day-case surgeries or when preoperative admission is not mandatory. Although the onset of SDE therapeutic effects could be delayed for 12 hours, this study did not detect clinically significant beneficial responses in pain intensity reduction from 24 hours to 7 days after administration. Secondly, standard parenteral MMA was administered intraoperatively to all participants in this study and the previous trial reported by our research team, whereas the placebo groups in the 2 previous studies received only blank or suboptimal analgesics for postoperative pain control.,12 Nevertheless, our study results are consistent with a previous open-label study, which demonstrated that preoperative administration of SDE did not significantly improve pain intensity and requirement for rescue analgesics after laparoscopic cholecystectomy. The results of these 2 studies suggest that laparoscopic cholecystectomies only induce mild-to-moderate levels of pain in the acute post-operative stage. The postoperative pain could be adequately managed by the standard opioid-sparing MMA regimen, including NSAID, selective cyclooxygenase-2 inhibitors, and acetaminophen.Three previous randomized double-blind placebo-controlled trials have demonstrated that a single preoperative injection of 150 mg SDE provides significant clinical analgesic effects in the management of post-open surgery pain. The presence of CPSP after laparoscopic cholecystectomy is known to increase the risk of chronic opioid use . Therefore, the long-lasting analgesic effect of SDE could theoretically attenuate the development of CPSP. To our knowledge, this is the first study testing the potential beneficial effect of a long-lasting parenteral analgesic in the prevention of CPSP, as acute postoperative pain is more likely to transit into chronic pain if the acute pain is not adequately controlled and it persists beyond the usual tissue healing time of 1 week. Our results showed that the incidence of CPSP at 3 months after surgery in the placebo-controlled group was 22.5%, which is comparable with the findings of previous studies reported by other groups in the range of 3% to 59%.,22,23 The incidence of CPSP was reduced by 41% in the SDE-treated group (22.5% vs 13.1%), but the difference was not statistically significant.After intramuscular injection, SDE is hydrolyzed by tissue esterase and the active compound nalbuphine is constantly released over 145 hours to mediate a longer lasting analgesic effect.,12,20 and retrospective clinical studies. In general, the adverse drug reactions include systemic responses reacting to the active compound nalbuphine  and focal reactions following intramuscular injection (such as soreness and tissue reactions at the injection site). These studies also noted an increase in unexpected pyrexia in patients who received SDE injection.,20 It should also be noted that the plasma concentration of nalbuphine can be maintained at therapeutic levels for up to 6 days after administration, but onset and subsidence time frames of the drug-related adverse responses were not reported in these studies. In our study, the potential drug-related adverse events were recorded up to 7 days post-operatively. We did not record any cases of pyrexia while in hospital. Dizziness was the most common systemic adverse reaction in this study and was significantly higher in the SDE group through postoperative days 1 to 3. 39.5% of the patients in the SDE complained of dizziness on postoperative day 3 compared to only 5% in the placebo group. The incidence of nausea was similar between the placebo and SDE on the first day post-operatively, but significantly more patients in the SDE group remained nauseous on day 2 to day 3 postoperatively. At day 7, most of these systemic adverse events resolved spontaneously, except a small proportion (7.9%) of patients in the SDE group who reported ongoing dizziness. On the contrary, incidence and severity of SDE-related systemic adverse events were significantly reduced in overweight patients (body weight index > 38 kg/m2) who received laparoscopic bariatric surgery, suggesting that these adverse drug reactions might be dose-dependent and the therapeutic doses of SDE should be titrated according to the patient’s total body weight. Although injection site reaction seldom occurs following a single dose of intramuscular nalbuphine, our study and other study groups found that more than 20% of the patients receiving SDE injections experienced local erythema, swelling, and tenderness while only 6% to 12.5% of the patients who only received the vehicle solution injection developed focal reactions. In addition, we observed that injection site reactions  more often developed 48 hours after injections and could last for more than 5 days. However, these focal reactions at injection sites were usually mild and transient, and there were no participants who requested additional medical attention or management in this study. To avoid accidental administration into other subcutaneous tissues, such as the adipose layers, ultrasound-guided intramuscular injection of SDE is highly recommended.The drug-related adverse reactions of SDE were previously reported in the several prospectiveThis study also measured pre- and post-operative QoL using the SF-12 questionnaire, which includes physical and mental health composition domains. The average total SF-12 scores were found to be low both before and at 7 days post laparoscopic cholecystectomy in both groups (75–79). These scores increased significantly to > 90 at postoperative day 90, suggesting an improvement in physical and mental health after surgical removal of the pathological gall bladder. Since there were no differences in the SF-12 scores between the 2 groups at the 3 measurement time points, we suggest that the SDE-related adverse events were most likely tolerable and did not significantly affect the physical and mental recovery after laparoscopic cholecystectomy. Therefore, the preemptive analgesic benefit of SDE might not be available if SDE was administered just before surgical stimulation. Second, all patients were interviewed over telephone at postoperative days 7 and 90 and the development of CPSP at postoperative day 90 was surveyed using a standard questionnaire. Since most of the patients who claimed to develop CPSP were unwilling to return to our pain clinic for further assessment, this study could only confirm that there was newly developed pain at the surgical sites or on the incisional wounds in some of the participants, but the severity and nature of the chronic pain were not assessed by pain specialists. Since postoperative MMA was administered in the 2 treatment groups, the effects of MMA on relieving of injection site reaction were not determined. Nevertheless, QoL was improved similarly in both groups at postoperative day 90, and none of these patients required medical attention for CPSP during the telephone interview. This suggests that the degree of chronic pain was mild, and it did not significantly affect post-operative recovery. Third, although there were no significant differences in the number of patients who requested for rescue parenteral analgesics up to 48 hours after surgery, differences in the total dosage and types of analgesics could exist between the 2 groups. Forth, although the incidence of CPSP was reduced in patients received SDE injection, this study was underpowered to detect a statistically significant effect of SDE to prevent the development of CPSP after laparoscopic cholecystectomy. Lastly, the drug-related adverse reactions could be dose-dependent. Further dose-ranging or divided dosing studies is advised to determine the clinical pharmacodynamics of SDE.Several limitations of this study should be addressed. First, this study tested the postoperative analgesic effects of SDE that was administered after induction of anesthesia rather than > 12 hours before surgical intervention. According to the clinical pharmacokinetics of SDE, plasma therapeutic levels of nalbuphine are achieved at 12 hours after intramuscular injection.In conclusion, a single dose of the extended-release analgesic SDE administered intra-operatively did not provide a sufficient add-on effect to the standard MMA for acute and chronic postsurgical pain after laparoscopic cholecystectomy. On the contrary, intramuscular injection of 150 mg SDE in patients with average body mass can cause certain adverse events that may have been overlooked in previous reports. The adverse reaction profile and appropriate dosing of SDE should be more thoroughly investigated.The authors wish to appreciate Ms. Tzu-Shan Chen  for the advice in statistical analysis of the clinical data.Conceptualization: Ying-En Lee, Chen-Yu Fu, Jen-Lung Chen, Chen-Fuh Lam.Data curation: Ying-En Lee, Chen-Yu Fu.Formal analysis: Ying-En Lee, Chen-Yu Fu, Yow-Ling Shiue, Chu-Yun Lu, Jen-Lung Chen, Chen-Fuh Lam.Funding acquisition: Chen-Fuh Lam.Investigation: Ying-En Lee, Chen-Yu Fu, Chung-Yen Chen, Jian-Han Chen, Jen-Lung Chen, Chen-Fuh Lam.Methodology: Ying-En Lee, Yow-Ling Shiue, Chu-Yun Lu, Jen-Lung Chen, Chen-Fuh Lam.Project administration: Jen-Lung Chen, Chen-Fuh Lam.Resources: Jen-Lung Chen, Chen-Fuh Lam.Software: Yow-Ling Shiue, Chu-Yun Lu, Chen-Fuh Lam.Supervision: Jen-Lung Chen, Chen-Fuh Lam.Visualization: Jen-Lung Chen, Chen-Fuh Lam.Writing – original draft: Ying-En Lee, Chen-Yu Fu, Chung-Yen Chen, Jian-Han Chen.Writing – review & editing: Yow-Ling Shiue, Chu-Yun Lu, Jen-Lung Chen, Chen-Fuh Lam.


COVID-19 pandemic emerged worldwide at the end of 2019, causing a severe global public health threat, and smoking is closely related to COVID-19. Previous studies have reported changes in smoking behavior and influencing factors during the COVID-19 period, but none of them explored the main influencing factor and high-risk populations for smoking behavior during this period.We conducted a nationwide survey and obtained 21,916 valid data. Logistic regression was used to examine the relationships between each potential influencing factor  and smoking outcomes. Then, variables related to smoking behavior were included based on the results of the multiple logistic regression, and the classification and regression tree (CART) method was used to determine the high-risk population for increased smoking behavior during COVID-19 and the most profound influencing factors on smoking increase. Finally, we used accuracy to evaluated the performance of the tree.The strongest predictor of smoking behavior during the COVID-19 period is acceptance degree of passive smoking. The subgroup with a high acceptation degree of passive smoking, have no smokers smoked around, and a length of smoking of ≥ 30 years is identified as the highest smoking risk (34%). The accuracy of classification and regression tree is 87%.The main influencing factor is acceptance degree of passive smoking. More knowledge about the harm of secondhand smoke should be promoted. For high-risk population who smoke, the “mask protection” effect during the COVID-19 pandemic should be fully utilized to encourage smoking cessation.The online version contains supplementary material available at 10.1186/s12889-023-17224-z. At the end of 2019, COVID-19 spread globally. In March 2020, WHO declared it a pandemic . SmokingSome previous studies have reported changes in smoking behavior during the COVID-19 pandemic and identified influencing factors. Some studies suggest that smoking behavior has decreased during the pandemic due to concerns about the perceived harm of smoking during COVID-19 , difficuClassification and Regression Tree Analysis (CART) is a decision tree method developed by Breiman and colleagues. Using CART, it is possible to identify the most significant influencing factors for relative risk and explore the interaction between influencing factors and the most critical influencing factors to form the branches of the classification and regression tree, dividing the population into high-risk subgroups . It is aOverall, smoking is a risk factor for COVID-19 infection and severity. Prior studies have analyzed the influencing factors of smoking behavior during the pandemic, but these studies only explored the relationship between influencing factors and changes in smoking behavior. This study aims to address the limitations of these studies. Specifically, CART analysis was used to explore the factors that most deeply influence smoking behavior in the population and to analyze the interactions between this factor and other influencing factors to identify high-risk populations for increased smoking behavior.The data used in this study is conducted in 23 provinces, 5 autonomous regions, and 4 municipalities directly under the central government from June 20, 2022, to August 31, 2022. In this time, China was still experiencing the peak of the COVID-19 pandemic, with an increase of 442 − 77,402 cases per day , 22.https://www.wjx.cn/) to distribute the electronic questionnaire to each person to collect data. All participants obtained the participating respondent and record the questionnaire number issued to that person. Subjects were included in the study if they were ≥ 12 years old, provided written informed consent, and volunteered to participate in the study. A total of 23,414 questionnaires were collected, and after identifying and removing duplicate values, missing values and outliers with logical problems, a final sample of 21,916 was obtained, with a valid response rate of 93.6%.The survey used Equal-probability sampling and non-equal-probability sampling. Equal-probability sampling (stratified sampling) at the provincial, municipal, district, township/subdistrict, and community/village levels (stratified sampling) and non-equal-probability sampling (quota sampling) at the community/village to individual level At least one surveyor or a panel of surveyors were recruited in each city. Investigators set up questionnaire points at health service centers or relevant health service stations in the sampling communities under their responsibility to conduct face-to-face surveys. And if they cannot conduct them due to the epidemic, the user uses the Online Questionnaire Star platform , family characteristics (family income) and personal health status (chronic), COVID-19 related , Negative events, Smoking status (length of smoking), exposure to secondhand smoke . See Supplementary Table Self-efficacy was measured using the New General Self-Efficacy Scale short form (NGSES-SF3) . The scaDepression was measured using the Patient Health Questionnaire-9 (PHQ-9) . It is aAnxiety was measured using the Generalized Anxiety Disorder Questionnaire (GAD-7) developed by Robert L Spitzer . The scaSocial support was measured using the Perceived Social Support Scale short form (PSSS-SF3) based on the Zimet Perceived Social Support Scale . A 3-iteFirst, we used EmpowerStats for descriptive analysis. Continuous variables were presented as mean ± standard deviation and categorical variables were reported as frequency N (%) and assessed using t-tests for continuous variables and chi-square tests for categorical variables. Next, we conducted univariate and multivariate logistic regression using Stata version 16.0 to examine the relationships between each potential predictor  and smoking outcome. Than, based on the results of the multivariate logistic regression, variables related to smoking behavior were included and a CART analysis was performed using R to identify high-risk populations for increased smoking behavior during COVID-19 and the factors that most deeply influenced the increase in smoking behavior.Finally, we used R to evaluate the performance of CART. We calculated sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of the CART model. In this study, sensitivity refers to the probability of correctly predicting smokers as smokers. Specificity refers to the probability of not predicting non-smokers as smokers. Positive predictive value represents the proportion of true smokers among the sample units predicted as smokers. Negative predictive value represents the proportion of true non-smokers among the sample units predicted as non-smokers. Accuracy refers to the proportion of correctly categorized smokers and non-smokers out of the total.A total of 21,916 valid data were collected. Table Univariate regression analysis showed that the COVID-19 Impact of lockdown on Livelihoods was not associated with smoking behavior during COVID-19 P = 0.246) , high positive predictive value (71%), high negative predictive value (88%), and a high accuracy rate (87%). But sensitivity (20%) is low, it may be due to category imbalance that large sample size gap between smokers and non-smokers in this study Table .Smoking is a closely related factor to COVID-19, and controlling smoking behavior is of great significance for the prevention and treatment of COVID-19. Identifying high-risk subgroups for smoking during the pandemic can enable targeted prevention and effective reduction in smoking behavior. Therefore, this study used CART analysis to identify high-risk subgroups for smoking behavior during COVID-19 and determine the factors that have the deepest influence.Lockdown is an important factor influencing smoking behavior during COVID-19. It refers to the measures taken by various countries to prevent the spread of COVID-19, such as home quarantine, closure of entertainment venues, and isolation and quarantine measures . Due to According to the CART model, currently, the subgroup with a high acceptation degree of passive smoking, have no smoker smoked around them, and a length of smoked of 30 years or more has the highest smoking rate during the COVID-19 pandemic. The acceptation degree of passive smoking is the main determinant of smoking behavior during the pandemic. This may be because people are more attentive to personal health protection during the COVID-19 pandemic and are more sensitive to the perceived harmfulness of tobacco , which mAccording to the CART model, during the COVID-19 pandemic, people are more likely to smoke when they are in the presence of have no smoker smoked around, which is contrary to previous research results. Previous studies have shown that individuals are more likely to start smoking when family and friends around them smoke , 32. ThiHaving a chronic illness is also a significant predictor, as non-chronically ill individuals are more likely to smoke. Smoking is strongly associated with chronic diseases , and ChiRegarding these issues, first, more attention should be paid to long-term smokers, and more specialized smoking cessation help should be provided to them. For example, the smoking cessation clinic actively promoted in China is an effective method . SecondlFinally, this study explored the high-risk groups for smoking, and future studies should also delve deeper into the triggers for smoking cessation to provide a guiding direction for tobacco control policies and to form a continuity study to enrich policy guidelines.Our study conducted a national survey using quota sampling, which can balance differences between regions and reflect the situation nationwide. Secondly, we focused on smoking behavior during the COVID-19 period and comprehensively analyzed the factors that influence smoking behavior in the context of epidemic prevention and control. Finally, our study results further revealed the mutual interactions between the most important risk factors and other influencing factors, thus identifying the high-risk group for smoking during the COVID-19 period.However, this study also has some limitations. First, the study is a cross-sectional survey and does not establish causal relationships. Second, there may be other risk factors that affect smoking behavior during the COVID-19 period that were not included in this study. Finally, the sensitivity of CART in this study was relatively low, probably because of the small number of smokers in this study, which was large sample size gap between smokers and non-smokers. But even so, the accuracy of CART was high.In general, this study was based on a national sample and used CART analysis to explore the high-risk population for increased smoking behavior during the COVID-19 period. The results showed that people with a high acceptation degree of passive smoking, have no smokers smoked around, and a length of smoking of ≥ 30 years were the subgroups with the highest smoking behavior during the COVID-19 period. Acceptation degree of passive smoking was the strongest predictor of smoking behavior during the COVID-19 period. It is important to pay more attention to long-term smokers and non-chronic disease patients, raise awareness of the hazards of smoking and secondhand smoke, and take advantage of the “mask effect” during the epidemic period to reduce smoking behavior during the COVID-19 pandemic.Below is the link to the electronic supplementary material.Supplementary Material 1: Variable description, NGSES-SF3, PHQ-9, GAD-7, PSSS-SF3 and assignment criteriaSupplementary Material 2: The R package of classification and regression tree (CART)


Telephone-assisted cardiopulmonary resuscitation (T-CPR) has proven to be a crucial intervention in enhancing the ability of lay responders to perform cardiopulmonary resuscitation (CPR) during telehealth emergency services. While the majority of established T-CPR protocols primarily focus on guiding individual rescuers, there is a lack of emphasis on instructing and coordinating multiple lay responders to perform resuscitation collaboratively.This study aimed to develop an innovative team-based tele-instruction tool to efficiently organize and instruct multiple lay responders on the CPR process and to evaluate the effectiveness and feasibility of the tool.We used a mixed methods design in this study. We conducted a randomized controlled simulation trial to conduct the quantitative analysis. The intervention groups used the team-based tele-instruction tool for team resuscitation, while the control groups did not have access to the tool. Baseline resuscitation was performed during the initial phase (phase I test). Subsequently, all teams watched a team-based CPR education training video and finished a 3-person practice session with teaching followed by a posttraining test (phase II test). In the qualitative analysis, we randomly selected an individual from each team and 4 experts in emergency medical services to conduct semistructured interviews. The purpose of these interviews was to evaluate the effectiveness and feasibility of this tool.–1 vs median 112, IQR 106-120.8 min–1; P=.04 in phase I and median 117.5, IQR 112.3-125 min–1 vs median 111, IQR 105.3-119 min–1; P=.03 in phase II). In the intervention group, there was a delay in the emergency response time compared to that in the control group . However, this delay was partially mitigated after the phase II test. The qualitative results confirmed the compatibility and generalizability of the team-based tele-instruction tool, demonstrating its ability to effectively guide multiple lay responders through teamwork and effective communication with telecommunicators.The team-based tele-instruction tool significantly improved the quality of chest compression in both phase I and phase II tests. The average compression rates were more appropriate in the intervention groups compared to the control groups (median 104.5, IQR 98.8-111.8 minThe use of the team-based tele-instruction tool offers an effective solution to enhance the quality of chest compression among multiple lay responders. This tool facilitated the organization of resuscitation teams by dispatchers and enabled efficient cooperation. Further assessment of the widespread adoption and practical application of the team-based tele-instruction tools in real-life rescue scenarios within the telehealth emergency services system is warranted. Technological advancements, particularly in telehealth services, have opened up new possibilities for improving the quality and accessibility of health care ,2. The lThe existing T-CPR protocols for lay responders primarily focus on single-rescuer CPR and lack guidance on promoting collaboration among multiple lay responders to enhance prearrival care. Studies have shown that 43.1% of OHCA rescue events involve the presence of multiple lay responders at the scene . In our Introducing a team-based tele-instruction plan for telecommunicators could serve as a cost-effective yet highly beneficial strategy to optimize the effectiveness of team cooperation among multiple lay responders. This plan would ensure the consistent delivery of high-quality CPR for patients who experience OHCA. In contrast to periodic mass training initiatives for team-based CPR, implementing the team-based tele-instruction plan through telecommunicators leverages its widespread reach to effectively instruct a large number of lay responders, resulting in enhanced resuscitation quality. Hence, this study aimed to develop an innovative team-based tele-instruction tool with the aim of expanding the use of telehealth emergency services. We evaluated the effectiveness and feasibility of this tool through a randomized controlled simulation trial and semistructured interviews.Study DesignWe used a mixed methods approach in this study. We designed an innovative team-based tele-instruction tool for lay responders based on updated guidelines and an extensive literature study ,4,13. SuIn cooperation with a Chinese company specializing in dispatch systems, we innovatively designed a team-based tele-instruction protocol and developed a test tool for the medical dispatch system; however, it is currently used only for research purposes. If the response to the question “Can you find other people to help you?” was answered as “Yes,” the system would link to the team-based tele-instruction protocol.The protocol was applied to resuscitation teams of three or more lay responders, and the medical emergency response process was divided into 2 parts: pre-CPR instructions and team-based CPR instructions. The pre-CPR instructions included information registration, identification of cardiac arrest, and evaluation of lay responders’ CPR skills. Team-based CPR instructions included chest compressions, mouth-to-mouth ventilation, automated external defibrillator (AED) support, and continuous encouragement . The proFrom June 1 to December 31, 2021, 132 participants aged 18-60 years were recruited. Participants who had undergone clinical practice, were engaged in medical or emergency-related occupations, or were physically unable to perform high-intensity testing and training were excluded. To ensure that all participants possessed the necessary skills in single-rescuer CPR, we initially conducted a basic life support training called WeCan CPR. This training encompassed essential skills, such as contacting the emergency dispatch center (dispatch help number: 1-2-0), performing chest compressions, providing mouth-to-mouth ventilation, and using an AED .Subsequently, participants who passed the single-rescue CPR skill test were randomized into teams of 3. Each team was randomly assigned to 1 of the 2 study groups: the control group and the intervention group. In the control group, participants did not receive any guidance from a telecommunicator. In contrast, in the intervention group, one of the researchers (LZ) instructed the participants to use the team-based tele-instruction tool. Each team was asked to perform resuscitation before the team-based CPR training (phase I test) at baseline. Following the phase I test, each team underwent a 30-minute team training separately, following the same protocol as our previous study . The teaIn the team-based CPR simulated scenario, each team was required to assign and finish tasks, including contacting the emergency telecommunicator, performing chest compressions, mouth-to-mouth ventilation, and retrieving and using an AED. In the simulated scenario, the telecommunicator was blinded to the team’s CPR performance and activated the medical emergency response as soon as the call went through from the team-based T-CPR group. The scenario commenced as each team entered the room and concluded 10 minutes after they initiated the first chest compression. Participant B left the room to retrieve the AED  and was instructed to return to the room after 5 minutes.No feedback was provided to the participants once the scenario began. Quantitative data were collected from manikin feedback , video recordings, and the evaluations of researchers and submitted to a blinded assessor. A total of 3 cameras captured the algorithm time data, CPR technique, and communication with the telecommunicator from different angles . The quality of chest compressions and ventilation was recorded using SimPad PLUS . The AED performance was assessed by researchers using a dichotomous (yes or no) format. Time intervals, including hands-off time and a delay at the beginning, were measured using a stopwatch. The participants’ age, sex, and self-reported body weight were also documented.Semistructured interviews were conducted to evaluate the effectiveness and feasibility of the tool from the perspectives of telecommunicators and lay responders.Semistructured interviews were conducted after the phase II test to explore the acceptability of the protocol. We selected 1 individual from each T-CPR team using a random number table. All interviews included open-ended questions to elicit detailed accounts of the participants’ experiences with the protocol, with subsequent probing questions based on the interviewees’ responses. The questions were divided into 2 dimensions: participants’ attitudes toward teamwork and their interactions with the telecommunicator. A total of 5 themes were identified, including operational skills, team cooperation, communication with the telecommunicator, team-based CPR instructions, and other opinions.The interviews began with a question about the individual’s role in the simulation, followed by seeking their viewpoints on instructions provided by telecommunicators. Questions were specifically tailored to teamwork and leadership during resuscitation and quality efforts in CPR and other areas. Lastly, we inquired about the overall processes related to the tool, their views about this tool’s best practices, and the areas where they felt improvement was most needed. With the permission of the participants, each interview was audio-recorded and fully transcribed by independent professional transcriptionists within 24 hours of each interview and reviewed by the interviewers for accuracy. Data analysis was performed concurrently with data collection.Expert consultations were conducted to enhance the clarity and efficiency of instructing the caller on the required actions and procedures while also ensuring that the protocol remained scientifically sound, logical, and feasible. Interviews were conducted to address 3 aspects of the protocol: interior logic verification, exterior contrast analysis, and generalizability analysis. The 4 experts involved in the study were experienced specialists in the fields of emergency medical services and T-CPR dispatch protocol. They can play a substantial role in supervising the team-based T-CPR instruction protocol and participating in the development of the tool promotion plan in China.P values <.05 were considered statistically significant.Quantitative data are presented as frequencies with percentages for categorical variables and median (IQR) for continuous variables. Differences in categorical outcomes were assessed using the chi-square test, and continuous variables were compared between the 2 groups using the Wilcoxon rank sum test. All statistical analyses were performed using SPSS version 24.0 (IBM Corp). Two-sided A qualitative analysis was conducted by 2 researchers using Colaizzi’s phenomenological method . ResearcEthical approval was obtained from the Joint Research Ethics Board of the Shanghai Jiao Tong University School of Public Health and Nursing (SJUPN-201913). All participants were verbally informed about the intentions of the study, and they provided written informed consent. To protect the privacy of the participants, the data were anonymized by assigning participant IDs to individuals for representation. Our team provided free basic life support training and a 50 RMB (approximately US $8) present to each participant upon completion of the study to compensate for their time and feedback.A total of 132 participants were randomly divided into 44 teams of 3 members each; of these, 48 (36.4%) were male. The age of the participants ranged from 18 to 60 years, with a median age of 20 (IQR 18-30) years in the team-based T-CPR group and 19 (IQR 18-30) years in the team-based CPR group. The mean weight was 63.6 (SD 12.7) kg and 63.6 (SD 14.9) kg in the team-based T-CPR and team-based CPR groups, respectively. The demographic characteristics of the 2 groups were not significantly different.–1) in both phase I and II tests   . The telctively) C. This pctively) D.P=.02). The percentage of clearing, while the AED delivered shock in the phase I test, was significantly higher in the intervention group .The compression depth of the intervention group was significantly greater in the first minute of the phase I test A, and thP=.03), time to open the airway , and the time to the first shock . Training partially reduced the delay in emergency response time, as indicated in the phase II test, where the difference in the time to first chest compression was not statistically significant compared with the time to open the airway  and the time to the first shock .In the phase I test, the emergency response time of the team-based T-CPR group was significantly longer than that of the team-based CPR group , includiWhen inquired about teamwork, most responses were positive, particularly regarding team cooperation.When I felt tired, my teammates would take over chest compression, which significantly improved the efficacy.Male, 21 years, role AWe had good coordination. When one person was pressing, the others could communicate with the telecommunicator, so that the telecommunicator could adjust the instructions according to the situation timely.Female, 42 years, role BIn addition, team-based instructions were widely recognized. Participants described the protocol as “understandable,” “precise,” and “efficient.” The beneficial effects of instruction on team cooperation were widely acknowledged.The instructions helped us form a well-organized team, which is a process that would take a long time without the telecommunicator involved.Female, 23 years, role CThe experts also spoke highly of the protocol. The advantages of task allocation and closed-loop communication protocols were recognized during the simulation.The protocol is in strict compliance with the AHA guidelines and can play a good instructing role on the event of OHCA. It allows multiple lay responders to have a clear division of labor and help each other as a team, which is important for providing high-quality CPR.However, there was little feedback on the shortcomings of the protocol, which included a high demand for the telecommunicator and the requirement for more detailed instructions during the process of switching to the chest compression implementer.The significant quantitative findings were integrated with qualitative remarks to reveal the participants’ classic behavioral performance that affected the quality of CPR. Performance was divided into 2 themes: procedural issues and team factors. A total of 12 items were extracted .OHCA is a major cause of death worldwide, and high-quality CPR facilitates neurological outcomes and overall prognosis . An orgaThe tele-instruction tool for multiple lay responders has good universality and promotional value. It was reported that 43.1% of OHCA resuscitation incidents involved multiple lay responders, more than half of which involved 3 or more lay responders . In the CPR has long been considered the hallmark of OHCA management, and chest compressions with an appropriate frequency and depth are a significant component of high-quality CPR. The team-based tele-instruction tool demonstrated a remarkable advantage in terms of chest compression quality. By counting loudly with a telecommunicator, the intervention group maintained a more suitable average compression rate. This was particularly obvious when the rescuers had just begun chest compressions or had just changed roles. Team-based instructions could help rescuers achieve a better effect quickly after beginning compression, shorten the time of adaptation, and facilitate efficient intrateam role alternation. In addition, the intervention group achieved a relatively greater compression depth (within 5-6 centimeters). It was assumed that team-based instructions could realize better team cooperation and more timely rotation, avoiding poor-quality resuscitation due to fatigue.Prompt emergency response is crucial for OHCA rescue -25. We fFeedback from the participants and experts provided support for the validity of the protocol and highlighted issues requiring improvement. First, many participants did not actively report to the telecommunicator during resuscitation, which may have wasted their valuable time. Second, telecommunicators should pay more attention to feedback from rescuers, adjust instructions in a timely manner, and offer psychological support and encouragement, which requires abundant work experience and an effective improvisation ability. However, it may be difficult for telecommunicators to achieve this on their own. Some studies used machine learning to assist telecommunicators in OHCA recognition and achieved better results ,30. InteNevertheless, this study had some limitations. First, a simulation trial was conducted with people who had mastered single-rescuer CPR skills to perform a targeted evaluation of the tool. However, in real-life OHCA situations, it is not certain whether lay responders equipped with resuscitation skills would be recruited. Therefore, we will further expand the compatibility by collecting more representative data to demonstrate the efficacy of the protocol. Second, due to some practical barriers, such as the disconnection of the call and the time-consuming task of information registration, our test may not have been completely simulated. Finally, although the study design could reduce the effect of confounding factors between the groups, the practice effect did exist and could have led to motivational bias.In this study, we developed an innovative team-based tele-instruction tool and assessed its effectiveness in ensuring the delivery of high-quality CPR. This tool facilitated the efficient organization of resuscitation teams, promoted effective cooperation, and enhanced compression quality among multiple lay responders. Consequently, we propose that the efficacy and applicability of the team-based tele-instruction tool should be evaluated in the telehealth emergency service systems to enhance resuscitation outcomes in real-life rescue scenarios.