Delayed onset muscle soreness peaked 24 h post-match and rate perceived exertion peaked post-match (p < 0.05) and returned to baseline 96 h post-match (p > 0.05). Inflammatory biomarkers peaked at 24 h (p < 0.05) and remained significantly elevated for 72 h after the match (p < 0.05). Muscle damage biomarkers peaked at 24 h (p < 0.05) and remained significantly (p < 0.05) elevated for at least 72 h after the match. Oxidative stress markers peaked at 24 h–48 h (p < 0.05) and returned to baseline 120 h post-match (p > 0.05). In respect to antioxidant responses, these peaked at 24 h–48 h post-match (p < 0.05) and returned to baseline 120 h after the match (p > 0.05). Conclusions: A single futsal match induces short/mid-term changes in performance, inflammation, oxidative stress, and muscle damage markers for about 72 h–96 h post-match.Background: Our aims were to investigate the time-course effects of a futsal match on performance, oxidative stress, and muscle damage markers, as well as inflammatory and antioxidant responses during a 6-day post-match period. Methods: Thirty-four female high-level futsal players were assessed on several oxidative stress, inflammation, subjective muscle soreness, subjective rate perceived exertion, and performance tests before a futsal match, immediately after, and 24 h to 144 h after. Results: Counter movement jump, 20 m, and 10 m sprints performance significantly decreased immediately after the match ( It requmax) , blood lmax) [−1 , and dur−1·min−1 . As it i−1·min−1 ,4,6. How−1·min−1  and espeFutsal players execute a great number of repeated high-intensity actions , which may expose them to high neuromuscular fatigue . FatigueFutsal, on one hand, is characterized by high-intensity efforts of ~2–3 s  with bri−1) [Written consent was obtained from the potential participants after they were thoroughly informed of the aims, procedures, benefits, and potential risks of participating in the study. Prior to the study , all pot−1)  that had−1)  and perc−1) . All pro−1)  and wereThe participants were recruited from women’s futsal top national leagues, first (12 clubs) and second division (12 clubs), and all registered players who competed in these two categories were informed about the study. The 167 players from the first division who expressed their desire to participate were considered as potential participants of the experimental group. However, 92 (55.09%) were not on day 2–8 of the follicular phase of their menstrual cycle during the upcoming intervention of futsal match and served only as co-players or goalkeepers or substitutes in case of need and were excluded from the experimental group. From the remaining players, 40 (23.95%) also did not meet the criteria for inclusion  and were thus excluded. Next, using a computer-generated list of random numbers, the remaining 35 potential participants were randomly assigned as defenders, midfielders, or attackers in 10 different teams where the goalkeepers and the remaining positions were randomly held by healthy previously excluded players. Consequently, five matches were held in parallel in a multi-facility sports center. Potential participants had to play the entire duration of a match , while at the researcher’s signal they would regularly change their field position in a circular manner to ensure that each participant played approximately the same amount of time in all positions . Thus, excluding the players who were used to complete the residual positions of the teams and one potential participant who was injured during the intervention of the futsal match, who was also excluded, the remaining players (20.36%) form the experimental group of the study . The 148 players from the second division who expressed their desire to participate were considered as potential participants of the control group. However, 118 (79.73%) did not meet the criteria for inclusion  and were excluded. The remaining players (20.27%) were selected for the control group .Based on the biomarkers empirical evidence derived from the EROS study  and thei® . The participants were instructed to begin using the test daily starting from the first day of their menstrual cycle. The test detects the LH surge, which typically occurs 24–36 h before ovulation. The participants were advised to interpret the test results according to the manufacturer’s instructions and classify their menstrual cycle phases based on the observed LH surge. They were provided with a standardized guide that outlined the criteria for identifying each phase, including the follicular, ovulatory, and luteal phases. The self-identification of the menstrual cycle phases was conducted in a private and confidential manner to ensure accurate reporting. The test strips and instructions were provided to participants at the beginning of the study, and they were asked to record the date of the LH surge and the corresponding cycle phase in a provided diary. This approach allowed for the collection of menstrual cycle data while minimizing interference with the participants’ daily routines. Additionally, the participants were informed about the potential limitations of self-identification and were encouraged to consult with healthcare professionals for confirmation or further guidance if needed.The participants’ phases of the menstrual cycle were self-identified after the detection of the luteinizing hormone (LH) surge using the Clearblue Digital Ovulation Test®, Seca, Birmingham, UK) and nude body mass to the nearest 0.1 kg  were measured. Harpenden Skinfold Calipers®  were used to measure the thickness of seven skinfolds [An anthropometric assessment was conducted during a preliminary visit to the laboratory one week pre-match. Standing height to the nearest 0.5 cm (Stadiometerkinfolds  and bodykinfolds .2max) was measured in a laboratory one week pre-match and after a preliminary familiarization visit for ⩒O2max assessments and lower limb muscle strength tests, as well as sprint performance tests. For the measurement of ⩒O2max, a portable gas exchange analyzer  that had been calibrated with known oxygen and carbon dioxide gas mixtures and an incremental running test to exhaustion on a treadmill  were used. The ⩒O2max protocol consisted of running at 7 km·h−1 for 1 min and 8 km·h−1 for 30 s. Thereafter, the speed of the treadmill was increased by 0.5 km·h−1 every 30 s until exhaustion; the inclination of the treadmill was 1%. The ⩒O2max and HRmax were determined as the highest values reached in a 15 and 5 s period, respectively, during the last part of the incremental test. The criteria for the attainment of ⩒O2max included two of the following: a respiratory exchange ratio >1.1, HRmax within 10 bpm of the estimated HRmax based on age, a rating of perceived exertion equal to, or higher than 18, or a leveling off in ⩒O2 (<2 mL·kg−1·min−1) with a concomitant increase in treadmill speed [®, Polar Vantage NV monitor; Polar Electro Oy, Kempele, Finland). Throughout the futsal match HR was recorded at 5 s intervals.Maximal oxygen uptake  after a standardized 5 min warm up at 60–70% intensity on a Monark leg cycle ergometer  [®, Newtest, Oulu, Finland) placed at a 100 cm height. The tests were conducted from a standing position, with the front foot placed 30 cm behind the start line.The participants’ lower limb power was measured with counter movement jump (CMJ) with arm swing on a EuroJump Sweden) . The sprFor leg dominance determination, the Waterloo Footedness Questionnaire-Revised was used . Next, e®, F. Hoffmann-La Roche Ltd., Rotkreuz, Switzerland). Moreover, eight blood samples in total per participant were drawn from the basilic or mesobasilic vein with the player in a sitting position in the morning after a 12 h fasting before the match day, immediately after the match, and at 24, 48, 72, 96, 120, and 144 h post-match. Approximately 10 mL from each sample was treated with sodium citrate, centrifuged for 10 min at 2250× g, and the supernatant plasma was stored at −70 °C for later analyses. Approximately 5 mL of the sample was placed in vacutainers without anticoagulant, left to clot, and then centrifuged at 2250× g for 15 min at 4 °C. The serum samples obtained were separated from the packed red cells, placed in Eppendorf tubes, and stored at −70 °C. A small quantity of blood (0.2 mL), which was immediately added to trichloroacetic acid (0.4 mL of 5%) and centrifuged , was analyzed spectrophotometrically at the 412 nm wavelength for reduced glutathione (GSH) and oxidized glutathione (GSSG) determination, as described in previous studies [®, Akishima, Tokyo, Japan) after a reaction with ninhydrin, as described in detail by Suen et al. [®, Elabscience, Houston, TX, USA) according to the manufacturer’s instructions on a standard ELISA reader . The levels of CRP were determined immunonephelometrically on a biochemistry analyzer  according to the manufacturer’s directions. On an MDA180® automated analyzer , fibrinogen (FIB) was measured using a functional assay based on Clauss’ method and the kit Fibriquik , as described elsewhere [®, Roche, Basel, Switzerland) and a reflectance photometer  were used to measure CK. Thiobarbituric acid reactive substances (TBARSs), protein carbonyls (PCs), activity of serum catalase (CAT), and total antioxidant capacity (TAC) were analyzed spectrophotometrically at 530 nm, 395 nm, 240 nm, 540 nm wavelengths  according to Keles et al., Patsoukis et al., Aebi, and Janaszewska and Bartosz, respectively [®, Sigma Diagnostics, St. Louis, MO, USA). Uric acid (UA) and complete white blood cell count (WBC) was determined within 24 h via matching duplicate counts using an automated hematology analyzer . All samples were measured in duplicate and their average was used for statistical analysis .Blood drop samples were collected within 1 min before the match, immediately after the 1st half, before the 2nd half, during half-time, and within 1 min after the completion of the match to measure lactate concentration  mixed analysis of variance (ANOVA) with repeated measures on the time factor. A 2 by 3 and a 2 by 5 (groups × time) mixed ANOVA with repeated measures on the time factor was conducted for the heart rate and lactate measurements. The degrees of freedom (df) for the main effects, interactions, and error terms were adjusted according to Greenhouse–Geisser correction when the assumption of sphericity was violated. For statistically significant interactions, post hoc analyses of simple effects and analytical pairwise comparisons with Bonferroni correction were used to identify significant differences [p2). The participants’ pooled values  of blood lactate concentration after the match for the five matches were analyzed with a one-way ANOVA design (1st match × 2nd match × 3rd match × 4th match × 5th match). The Pearson’s correlation coefficient, r, was used to evaluate the intercorrelations among the biochemical and performance variables. All data were analyzed with Statistical Package for the Social Sciences for Windows  and according to the procedures outlined by Meyers et al. [p < 0.05 for all analyses. Partial η2 and Pearson product moment correlations (r) were calculated and interpreted according to Cohen (1988) [p2), a sample size of 64, two groups in two conditions, and eight repeated measures. The observed power (1 − β) was >0.9. When other power variables  were used, a similar power was calculated.The homogeneity and normality of the obtained data were assessed through the Levene’s and Shapiro–Wilk tests, respectively. Independent ferences  between s et al. . Statistn (1988) . GraphPap > 0.05). Additionally, time measurements did not show statistical significance for the control group in all variables (p > 0.05). Therefore, for significant interactions, statistical findings are presented based on simple effects  and Bonferroni pairwise comparisons between time measurements for the experimental group in all dependent variables. Moreover, as reflected by the after-match blood lactate concentration measurements, participants’ match intensity of the five played matches showed no statistically significant difference between matches as determined by one-way ANOVA  = 0.084, p = 0.987). The descriptive characteristics see  and pre-p < 0.001, ηp2 = 0.81). Bonferroni pairwise comparisons indicated that the lowest CMJ value occurred immediately after the game (p < 0.05), it remained significantly lower than the PRE measurement, 24 h and 48 h after the match (p < 0.05), and returned to baseline levels 72 h after the match. Interclass correlations showed that CMJ 48 h after the match was negatively correlated with TNF-α, cortisol, and WBC , whereas it was positively correlated with HCY . Interclass correlations of the significant differences between the deficit of pre- and post-match performance measurements showed that CMJ was positively correlated with pre-match HCY , negatively with pre-match CK , and positively with the 24 h GSH/GSSG ratio .The results of simple effects analysis on the CMJ measurements for the experimental group showed statistically significant differences  = 137.28, p < 0.001, ηp2 = 0.97) and for the 20 m sprint  = 2069.01, p < 0.001, ηp2 = 0.98) measurements. Bonferroni pairwise comparisons showed that the 10 m sprint had the highest time scores immediately after the match (p < 0.05) and these remained significantly higher than the PRE measurement, 24 h, 48 h, and 72 h after the match (p < 0.05). Similarly, the 20 m sprint had the highest time scores immediately after the match (p < 0.05) and it remained significantly higher than the PRE measurement, 24 h, 48 h, and 72 h after the match (p < 0.05). Furthermore, the 10 m and 20 m sprint times returned almost to baseline 120 h and 96 h after the match, respectively. Interclass correlations of the significant differences between the deficit of pre- and post-match performance measurements showed that the 10 m sprint was negatively correlated with pre-match, 24 h, and 96 h after the match IL-6 , positively with pre-match and post-match LDH , positively with 24 h after the match HCY , and positively with post-match and 48 h, 72 h, and 96 h after the match FIB . Similarly, the deficit of the pre- and post-match 20 m sprint was negatively correlated with post-match IL-6 .The results of the simple effects analyses indicated significant differences for the 10 m sprint  = 1008.95, p < 0.001, ηp2 = 0.96), non-dominant leg extensors (NDKE)  = 802.06, p < 0.001, ηp2 = 0.96), dominant leg flexors (DKF)  = 939.53, p < 0.001, ηp2 = 0.97), and non-dominant leg flexors (NDKF)  = 887.33, p < 0.001, ηp2 = 0.96). Bonferroni pairwise comparisons showed that (i) both DKE and NDKE DOMS values peaked 24 h after the match, were significantly higher than the PRE measurements post-match, and 48 h and 72 h after the match (p < 0.05), and returned to baseline 96 h after the match, and (ii) both DKF and NDKF DOMS peaked 24 h after the match, were significantly higher than the PRE measurement post-match, and 48 h, 72 h, and 96 h after the match (p < 0.05), and returned to baseline 120 h after the match. Interclass correlations of the significant differences between the deficit of the pre- and post-match DOMS measurements showed that DKE was positively correlated with pre-match and post-match HCY , as well as with pre-match WBC . On the other hand, NDKE was negatively correlated with pre-match HCY , whereas it was positively correlated with post-match LDH . Regarding the DKF pre- and post-match deficit, a negative correlation was found with post-match UA and TBARSs 72 h after the match . Finally, the NDKF pre- and post-match deficit was negatively correlated with the pre-match and post-match UA , as well as with CK 24 h and 48 h after the match  and positively with WBC 72 h after the match .Statistical analyses indicated significant differences for DOMS among measurements of the experimental group for the dominant leg extensors (DKE)  = 792.10, p < 0.001, ηp2 = 0.95) were found among the RPE measurements of the experimental group. Bonferroni pairwise comparisons showed that RPE peaks occurred immediately after the match (p < 0.05) and were significantly higher after the 1st half, before the 2nd half, and 24 h, 48 h, and 72 h after the match (p < 0.05). The RPEs returned to baseline 120 h after the match. Interclass correlations of the significant differences between the deficit of the pre- and post-match measurements showed that the RPE was positively correlated with CK pre-match and 72 h after the match . On the other hand, RPE was negatively correlated with FIB post-match and 24 h after the match , and with CRP 48 h and 72 h after the match .Statistically significant changes  = 688.58, p < 0.001, ηp2 = 1.00) for the HR measurements of the experimental group and Bonferroni pairwise comparisons indicated that the HR values were similarly high during the 1st and the 2nd half and significantly different to pre-match values (p < 0.05). There was a significant difference  = 143,172.82, p < 0.001, ηp2 = 0.99) and Bonferroni pairwise comparisons showed that the lactate peak occurred immediately after the 1st and the 2nd half (p < 0.05), and remained significantly higher than the pre-match values before the 2nd half as well as 15 min after the match. The simple effects analysis for blood lactate concentration indicated significant difference among the measurements  = 3749.49, p < 0.001, ηp2 = 0.98) and Bonferroni post hoc analysis showed that IL-6 peaked immediately after the match and returned to baseline in the next morning (p < 0.05). The simple effects analysis showed a significant difference among IL-6 responses  = 1538.31, p < 0.001, ηp2 = 0.87) was found for the TNF-α measurements and post hoc analysis showed that TNF-α peaked immediately after the match (p < 0.05) and returned almost to baseline by the next morning. A significant difference  = 217.43, p < 0.001, ηp2 = 0.92) was found for CRP and post hoc analysis indicated that the CRP peak occurred 24 h after the match (p < 0.05), remained significantly higher than the PRE measurement post-match and 48 h and 72 h after the match, and returned almost to baseline 96 h after the match. Statistical significance  = 358.23, p < 0.001, ηp2 = 0.72) for the FIB measurements. Post hoc analysis showed that the FIB peak occurred 24 h after the match, remained significantly higher than the PRE measurement post-match and 48 h and 72 h after the match (p < 0.05), and returned almost to baseline 96 h after the match. Statistical analysis showed a significant difference  = 85.42, p < 0.001, ηp2 = 0.89) for HCY and post hoc analysis showed that the HCY peak occurred 24 h after the match, was significantly higher than the PRE measurement post-match and 48 h and 72 h after the match (p < 0.05), and reached baseline values 96 h after the match.There was a significant difference  = 280.15, p < 0.001, ηp2 = 0.96) was found for the WBC measurements. A post hoc analysis showed that WBC peaked immediately after the match and was higher than the PRE measurement 24 h and 48 h after the match (p < 0.05), and returned almost to baseline 72 h after the match (p < 0.05).A significant difference  = 775.96, p < 0.001, ηp2 = 0.88) of LDH. Pairwise comparisons revealed that the LDH peak occurred 24 h after the match, was significantly higher than the PRE measurement post-match and 48 h and 72 h after the match (p < 0.05), and returned almost to baseline 96 h after the match.The simple effects analysis showed a significant difference among measurements  = 909.19, p < 0.001, ηp2 = 0.88) for the CK measurements. A post hoc analysis showed that CK peaked 24 h after the match and remained significantly higher than the PRE measurement post-match and 48 h, 72 h, 96 h, 120 h, and 144 h after the match (p < 0.05).Statistical analysis showed a significant difference  = 244.69, p < 0.001, ηp2 = 0.69) and post hoc analysis showed that PCs peaked 48 h after the match, remained significantly higher than the PRE measurement post-match, and 24 h, 72 h, and 96 h after the match (p < 0.05), and returned to baseline levels 120 h after the match.The simple effects analysis for PC concentration revealed significant difference among the measurements  = 72.94, p < 0.001, ηp2 = 0.93) was found for the TBARSs measurements and post hoc analysis showed that TBARSs peaked 24 h after the match, remained significantly higher than the PRE measurement post-match and 48 h, 72 h, and 96 h after the match (p < 0.05), and returned to baseline 120 h after the match.A significant difference  = 431.05, p < 0.001, ηp2 = 0.79). A post hoc analysis showed that the lowest GSH value occurred 24 h after the match, was significantly lower than the PRE measurement post-match and 48 h after the match (p < 0.05), and returned to baseline 72 h after the match.The simple effects analysis for GSH concentration showed a significant difference among the measurements  = 125.56, p < 0.001, ηp2 = 0.72) was found for the GSSG measurements and post hoc analysis showed that GSSG peaked 24 h after the match and remained significantly higher than the PRE measurement post-match and 48 h, 72 h, 96 h, 120 h, and 144 h after the match (p < 0.05).A significant difference  = 82.85, p < 0.001, ηp2 = 0.83) was found for the GSH/GSSG ratio and post hoc analysis indicated that the lowest GSH/GSSG ratio value occurred 24 h after the match, was significantly lower than the PRE measurement post-match and 48 h, 72 h, and 96 h after the match (p < 0.05), and returned to baseline 120 h after the match.Statistical significance  = 165.21, p < 0.001, ηp2 = 0.96) for the cortisol measurements. A post hoc analysis revealed that cortisol peaked immediately after the match, was significantly higher than the PRE measurement 24 h, 48 h, and 72 h after the match (p < 0.05), and returned to baseline 96 h after the match.Statistical analysis showed a significant difference  = 747.03, p < 0.001, ηp2 = 0.96) for TAC measurements and post hoc analysis showed that the TAC peak occurred 48 h after the match, remained significantly higher than the PRE measurement post-match and 24 h, 72 h, and 96 h after the match (p < 0.05), and returned almost to baseline 120 h after the match.There was a significant difference  = 762.60, p < 0.001, ηp2 = 0.67) was found for the CAT measurements and post hoc analysis indicated that CAT peaked immediately after the match (p < 0.05) and returned to baseline 24 h after the match.Statistical significance  = 66.74, p < 0.001, ηp2 = 0.91) was found for the UA measurements and post hoc analysis revealed that UA peaked 48 h after the match, was significantly higher than the PRE measurement post-match and 24 h, 72 h, 96 h, and 120 h after the match measurements (p < 0.05), and returned to baseline 144 h after the match.Statistical significance  = 348.62, max and 5.5 mmol·L−1 blood lactate values) induced significant changes in several biochemical markers and performance parameters, i.e., decreased CMJ, 20 m, and 10 m sprints performance, and increased inflammatory, muscle damage, and oxidative stress markers for about 72 h–96 h post-match , 72 h for a 20 m sprint (2.8–8.2%), and 48 h for CMJ (6.4–15.6%) ,25. Our The values of RPE, which is a good indicator of soccer players exertion and fatigue , remaineHowever, intense eccentric muscle activities  result in muscle damage, which is followed by inflammatory responses ,72. In aIn this study, the PC concentration peaked 48 h after the match while TBARSs peaked 24 h after the match. They both remained above their normal range for a 96 h period after the match. These increases might be explained by the increased levels of catecholamines, which assist in the energy metabolism of the muscles, or the increased catabolism of purines that results in elevated levels of UA that also were observed post-match and for several days afterward . OxidatiThe generalizability of the findings of the present study is limited to adult high-level female futsal athletes playing full time (2 × 20 min) on a wooden surface. It was reasonable to assume that all participants in the experimental group had similar weekly schedules and training loads, since all of them are high-level futsal players and compete in the same top national league. On the other hand, they may have different weekly training loads and perhaps some of the participants were more fatigued at the end of the season, which may have affected the measured variables and, therefore, this constitutes another limitation of the study. However, we believe that the eleven days between the end of competitive futsal duties and the intervention match were sufficient for the players to recover from the competitive season. Τhe actual activity pattern and the distances covered in respect to the teams’ and athletes’ positions  were not recorded. However, the participants in the experimental group played almost an equal amount of time in all positions  and match intensity, as determined by the blood lactate concentration after the match , which wFuture studies are needed to evaluate the possible interaction of sex, athletes’ field position, athletes’ covered distance, and physical fitness level with performance indexes, oxidative stress, inflammation markers, and DOMS after a futsal match. Gender hormone fluctuations during the menstrual cycle may have an effect on exercise-induced muscle damage in terms of DOMS and strength loss but not on CK . Muscle At this point, it is worth noting that, to the authors’ best knowledge, this study is the first that has explored the time-course effects of a futsal match on performance indexes, oxidative stress, inflammation, and muscle damage markers, in female futsal athletes, and in six consecutive days post-match using a sound experimental design. Moreover, our outcomes give sufficient evidence to futsal coaches for the usefulness of monitoring post-futsal match residual fatigue, muscle damage, and oxidative stress markers and highlights this procedure as a valuable tool for a better weekly training workload (intensity and volume) adjustment  in favorIn this study, short/mid-term changes in performance indexes, inflammation, oxidative stress, and muscle damage markers for about 72 h to 96 h after a futsal match in female players are evident as in male players. In addition, the association between time course changes in several oxidative stress, inflammation, and muscle damage markers with the subjective muscle soreness, fatigue measurements and the performance indexes after a futsal match is highlighted. Although the findings of the present study derive from players who played the entire match, following a competitive match, female futsal players may also face challenges in their recovery until the next match. For that reason, and due to nature of futsal , coaches should make a large number of substitutions to mitigate the undesirable effects of the match’s high intensity, which are believed to negatively impact players’ health, physical performance, and post-match training capacity. Moreover, trainers should monitor as possible post-match residual fatigue, muscle damage, and oxidative stress, using various markers such as CRP, CK, PCs, TAC, UA, RPE, and DOMS depending on their interest, so as, based on these measurements, to adjust accordingly the weekly workload (intensity and volume) to optimize the athletes’ recovery.


Late-onset oligomeganephronia (OMN) is a rare chronic kidney disease and has no quantitative criteria for diagnosis yet. The current study aimed to explore its clinicopathological features by histomorphometric analysis.We retrospectively re-reviewed all patients with enlarged and sparse glomeruli by light microscopy at Peking University First Hospital from 2012 to 2021, excluding those with any factor known to contribute to similar changes. Age- and sex-matched patients with thin basement membrane nephropathy were selected as control to establish the cut-off values for glomerulomegaly and rarity. Late-onset OMN cases were then confirmed and the clinicopathological characteristics were summarized.2, giving a lower limit of 211.53 μm for glomerulomegaly and an upper of 2.81 /mm2 for rarity. Seven adults of three females and four males were finally diagnosed as late-onset OMN with a mean age of 26.57 years. They showed mild to moderate proteinuria and/or renal dysfunction at biopsy with the mean proteinuria, serum creatinine (Scr) level, and estimated glomerular filtration rate of 0.50 g/d (0.10–0.95 g/d), 140.9 µmol/L (95.1–227.1 µmol/L), and 58.7 mL/min/1.73m2 (21.3–98.0 mL/min/1.73m2), respectively. Four patients (57.1%) had normal Scr at diagnosis. Six patients with available data showed renal tubular injury with increased urinary microalbumin in all, elevated N-acetyl-β-glucosaminidase in two, and elevated α1 microglobulin in five. Kidney size was normal or slightly reduced. The mean density and glomerular diameter of the seven cases was 0.86 mm2 (0.55–1.41 /mm2) and 229.73 μm (211.88–260.66 μm). Segmental glomerular sclerosis was observed in six (85.7%) with four (66.7%) of perihilar type. Proximal tubule dilation was observed in all, focal to diffuse, lining with enlarged epithelial cells. The mean foot process width was 634.02 nm, wider than 472.54 nm of the control (P = 0.0002).Mean diameter and density of cortical glomeruli in control was 156.53 ± 27.50 μm and 4.07 ± 0.63 /mmLate-onset OMN should be considered a special entity with relatively slow clinical progress characterized by hypertrophy of the sparsely distributed nephron. Oligomeganephronia (OMN), which is also called oligomeganephronic renal hypoplasia, is conditionally defined as a rare type of renal hypoplasia characterized by small kidneys with reduced number of renal lobes mainly occurs in infants or children . HistoloRenal biopsy samples in Peking University First Hospital from 2012 to 2021 were re-reviewed. Those samples that presented with hypertrophic and scattered glomeruli, mild interstitial fibrosis and tubular atrophy (IFTA) of less than 25%, and with no obvious ischemic glomeruli were collected. All patients showed normal bilateral renal pelvis, calyces, ureters, and bladder, with no evidence of postrenal obstruction. Rule out criteria included the conditions that may lead to glomerulomegaly, such as smoking, obesity, diabetes, family history of end-stage renal disease (ESRD), or elevated serum uric acid level. In addition, any other type of histologically confirmed glomerular disease was also excluded. Age- and sex-matched control group with thin basement membrane nephropathy (TBMN) was obtained from percutaneous renal biopsies. Rule in criteria of the control group included: (i) The main clinical manifestation was asymptomatic hematuria; (ii) Proteinuria was absent or trace (< 0.15 g/24 h); (iii) For all patients, there were no abnormal renal function and no conditions listed above that may lead to glomerulomegaly; (iv) Except for thin glomerular basement membrane (GBM), the specimen had no evidence of other glomerular diseases by immunofluorescence (IF), light microscopy (LM), or electron microscopy (EM); (v) The proportion of globally sclerotic glomeruli was within the allowed range, less than the percentage calculated by subtracting 10 from half the patient’s age, namely < (age/2–10)%.Informed consent was obtained from each patient for blood sampling and renal biopsy. The research was in compliance with the Declaration of Helsinki and was approved by the local ethics committee of Peking University First Hospital [IRB number 2017 − 1280].Clinical and laboratory data at the time of renal biopsy including gender, age, body mass index (BMI), blood pressure, albumin, urinary protein per 24 h, hematuria, serum creatinine (Scr), creatinine clearance rate (Ccr), C3 level, urinary microalbumin, transferrin, N-acetyl-β-glucosaminidase (NAG), α1 microglobulin (α1M), immunoglobulin, the range of proteinuria and Scr during the course of the disease, and the past and family history were collected from the patients’ medical records. Glomerular filtration rate (GFR) was evaluated by estimated GFR (eGFR) using the Epidemiology Collaboration Equation . The datThe renal biopsy specimens were processed using standard techniques for direct IF, LM, and EM. The samples were fixed in 4.5% buffered formaldehyde for LM and in 2.5% glutaraldehyde for EM, respectively. Consecutive serial sections were used for histological staining with hematoxylin and eosin, periodic acid-Schiff, silver methenamine, and Masson’s trichrome individually. EM was performed according to standard procedures. After embedded in Epon, ultrathin sections were mounted on copper grids and stained with uranyl acetate and lead citrate prior to being viewed by a transmission EM . Immunofluorescence staining was performed using antibodies against IgG, IgA, IgM, kappa light chain, lambda light chain, C3, and C1q. Two pathologists evaluated the biopsies separately and reach a consensus on the diagnosis.The images of the biopsy tissues were collected by LM (magnification of 100×). AnalySIS Image Processing software  was used to obtain the concerned indicators in each tissue. The following indicators including the density of glomeruli in the renal cortex, the diameter of glomeruli and proximal tubules (PT), the thickness of GBM and the height of epithelial cells lining PT were assessed.Glomerular density was obtained by dividing the number of all glomeruli by the area of renal cortex in each sample. The index of cortical area was got automatically from the software by drawing the outline of the obtained cortex. All glomeruli in the cortex including incomplete or obsolete ones were counted.The mean glomerular diameter was used to represent the size of each glomerulus. Outline of the outer glomerular capillary loops of the tuft and the parameters would be calculated automatically by the software. All glomeruli were measured except those incomplete, those with sclerosis more than 25%, those with significant shrinkage of capillary loops even accompanied by periglomerular fibrosis, and those only with a few capillary loops and with no mesangium observed.We randomly selected PT for measurement through the uniformly spaced test lines produced by the software over each photograph. Only those with non-atrophic intact hit by the test lines could be measured by delineating the outline profile along tubular basement membrane (TBM). In order to minimize the influence of different sections on the size of tubules, the shortest diameter which was given by the software was chosen as the index to represent the size of PT.Intact profile of PT was unnecessary for the height of epithelial cells. The height was indicated by the shortest distance between the TBM and the corresponding luminal surface of the cells, a vertical line perpendicular to the TBM initiating from the intersection point of the test line and TBM to the luminal surface of the cell. It should not be measured when the luminal boundary of the cells was not clear due to shedding of the brush border, and confluence of adjacent or contralateral cells.By EM, the thickness of GBM was evaluated by the distance between the basal aspect of the endothelial membrane and the basal aspect of the podocyte membrane that touches the GBM. Measurements were made along uniform cross-sections of the glomerular capillary wall. FPW of podocytes was obtained to evaluate the degree of foot process effacement as described previously . In brieP < 0.05.All analyses and calculations were performed with the SPSS 14.0 software . The measurement data were expressed as mean ± standard deviation (SD) and/or range for continuous variables and as proportions for categorical variables. The significance between OMN group and the control were determined by Student’s t-test. Differences were considered significant when 2 (19.71–23.12 kg/m2). A total of 96 glomeruli were measured and the mean diameter was 156.53 ± 27.50 μm (range 83.21–221.81 μm). Since the measurement data of diameter of glomerular capillary tuft in the control fit the normal distribution, mean ± 2SD was used for expression of the normal value range. Accordingly, the upper limit of diameter for normal glomerular of our controls was 211.53 μm. Similarly, the mean density of cortical glomeruli of the control cases was 4.07 ± 0.63/mm2 (range 3.21–4.92/mm2), giving a lower limit of 2.81/mm2. Therefore, the cut-off values for defining glomerular hypertrophy and rarity were an average glomerular diameter beyond 211.53 μm and a glomerular density below 2.81/mm2. The case that met with both criteria would be confirmed the diagnosis of OMN.There were eight cases in the control, three males and five females with an average age of 26 years (range 21–32 years) and an average BMI of 21.48 kg/mAs shown in Fig. 2 . All seven cases presented with proteinuria and/or mildly impaired kidney function at the time of biopsy. The mean proteinuria, Scr level, and eGFR at biopsy was 0.50 g/d (range 0.1–0.95 g/d), 140.9 µmol/L (range 95.1–227.1 µmol/L), 58.7 mL/min/1.73m2 (range 21.3–98.0 mL/min/1.73m2), respectively. Four patients (57.1%) had normal Scr at diagnosis. Duration the illness of an averaged 24.6 months, proteinuria had ever reached 1.0 g/d in three patients with a maximum of 2.0 g/d, and the maximum of Scr was 240 µmol/L. All patients presented with no to trace hematuria. The minimum of Ccr was 29.1 mL/min/1.73m2. Albumin level was within normal limit in all patients. The data for tubule injury was obtained from six cases with increased urinary microalbumin in all patients ranged from 36 to 1300 mg/L, elevated urinary transferrin in five (range 2.27–83.7 mg/L), NAG in two (21 U/L and 44 U/L), and α1M in five (range 15.3–82.5 mg/L). C3 level was in normal range in all patients except one was slightly depressed. Renal ultrasound demonstrated that the size of the kidney was normal or slightly reduced with the right length ranged from 8.0 to 11.5 cm and the left length from 8.4 to 12.2 cm, all with enhanced echo of renal parenchyma. None of the patients was diagnosed with hypertension except one (Case 2) presented with transient elevated blood pressure, and none of them complained of history of intrauterine growth restriction, maternal diseases, maternal drug intake or premature birth.The patients’ clinical characteristics were shown in Table 2 to 36.6 mL/min/1.73m2. Another three years later, renal function decreased further with a Scr value of 184 µmol/L and eGFR of 32.4 mL/min/1.73m2. Proteinuria was trace during the period of follow-up. No one had progressed to ESRD.The progression of illness was tracked before and after diagnosis for each case. The course of disease before diagnosis ranged from 3 to 27 months. After diagnosis by renal puncture, two cases (Case 2 and Case 6) were followed up for 77 months and 2 months, respectively. Overall, the course of the disease was traced from 3 to 89 months (mean 24.6 months) with Scr fluctuation ranged from 8 to 74 µmol/L. Case 2 had a longest duration of disease for 89 months. She had a slight elevation of Scr from the initial 116.9 µmol/L before biopsy to 169 µmol/L three years later and a declination of eGFR from 60.0 mL/min/1.73m2 which is sufficient for kidney biopsy. LM evaluation revealed much more sparsely distributed and enlarged patent glomeruli in OMN patients than that in the control . The mean glomerular diameter of seven cases was 229.73 μm (range 211.88–260.66 μm). The diameter of the largest glomerulus was 311.27 μm. The ratio of hypertrophic glomeruli in each case ranged from 40.0 to 90.9%. Statistical significance was not demonstrated in glomerular size or density between males and females (P > 0.05).Pathological features of seven cases of OMN were exhibited in Table P = 0.00001 and P = 0.0001), with the value of 87.01 ± 25.30 μm (range 36.44–235.63 μm) vs. 53.91 ± 2.25 μm (range 51.62–57.96 μm) and 13.76 ± 1.98 μm (range 11.82 ± 16.16 μm) vs. 9.23 ± 1.26 μm (range 7.73–11.66 μm) respectively. If the mean diameter of PT greater than upper cut-off value (the mean plus 2 times SD) in the control, namely 58.41 μm, was defined as dilated, the proportion of dilated tubules of OMN cases (range 60.0–94.2%) was much higher than that in the control (range 1.4–10.3%).PT dilation was focal or diffuse and varied in intensity with irregular contour and some were lined by enlarged and focal multilayered epithelial cells . No remarkable change was present in glomeruli, besides mild to moderate foot process effacement of podocytes . However, the cut-off value for the diagnostic criteria of OMN may be influenced by the calculating methods and ethnic differences.Glomerular volume had previously been used to represent the size of glomeruli, which can be evaluated by several methods, such as the Cavalieri method, Weibel-Gomez method, principle dissector method, and the maximal planar area method . AlthougP < 0.001). Hence, dilated PT may be another pathological feature as the diagnostic clue of OMN.Pathologically, the characteristics of late-onset OMN are as follows. Firstly, enlarged and sparsely distributed glomeruli are the main features. When the cut-off values of glomerular diameter and density applied, all nine cases suspicious of OMN met the diagnostic criteria for density while two not for size. However, all cases that met the criteria for hypertrophy also met that for rarity, and vice versa. Therefore, glomerular size may be a more stringent indicator. Based on current data, it is not clear whether late-onset OMN can be determined only by the size of glomeruli. However, this stringent criterion seems enough to screen in all our cases and easy to practice in daily life. Further test and verification of this idea with more cases is necessary. Secondly, segmental sclerosis (SS) was a common phenomenon (6/7) with most of them (4/6) surrounding the vascular pole. The ratio of SS was higher than that reported in the literature (3/7 for the former and 2/3 for the latter). As we know, larger glomeruli are evidence of hyperfiltration because they correlate with both higher single nephron GFR  and incrClinically, by reviewing the sporadic reported cases in the literature and the seven cases in our series, the features of late-onset OMN were different from the classical early-onset type. First, late-onset OMN presented mainly in the third to fourth decade with a gentle male dominance, nine males and five females. Second, its clinical presentation is relatively temperate, with proteinuria and/or elevated creatinine of mild to moderate degree. Third, disease was progressing relatively slowly and no one diagnosed ESRD during the course of the disease. Fourth, the kidney size was either normal or only slightly reduced which is consistent with the fact that reduction in nephron mass in adult-onset OMN is much less than that in pediatric OMN . In addiAs previously mentioned, many factors can lead to enlarged glomeruli and have similar pathological manifestations as late-onset OMN. For example, obesity-related glomerulopathy (ORG) is characterized by glomerulomegaly and focal segmental glomerulosclerosis (FSGS), particularly the perihilar variant, and may show significantly decreased glomerular density too , 38. In PAX2 mutation [RET1 gene mutation [HNF1B gene mutation [PAX2 mutation had been found in one-third of children with OMN and therefore was considered to play a role in the etiology [PAX2 mutation in exon 4 was found [To date, the pathogenesis of OMN, especially the late-onset OMN, has not been elucidated clearly. Several abnormal phenomena had been found in early-onset cases but were controversial because no direct relationship was proved. These findings covered from inadequate embryonic development of the metanephric blastema , low birmutation , chromosmutation , 41, RETmutation , and HNFmutation . Heterozetiology . For latIn conclusion, we investigated the diagnostic criteria of late-onset OMN by histomorphometric analysis for the first time and proposed that late-onset OMN is a special entity different from the early-onset OMN since their disparity in pathogenesis, clinical features and prognosis. Late-onset OMN is an exclusive diagnosis with unknown etiology characterized by typical pathological manifestations of enlarged and sparse glomeruli and expanded tubules with hypertrophic epithelial cells, and relatively slow clinical progress.


This retrospective study evaluated the characteristics and response of subretinal hyperreflective material (SHRM) to anti-vascular endothelial growth factor (VEGF) treatment in eyes with myopic choroidal neovascularization (CNV). The visual acuity (VA) was assessed at 3, 6, and 12 months after initiating anti-VEGF treatment in 116 patients (119 eyes) with SHRM and myopic CNV. Multimodal imaging, including color fundus photography, fluorescein angiography (FA), and optical coherence tomography angiography (OCT-A), were performed. We compared type 2 neovascularization (NV) (n = 64), subretinal hyperreflective exudation (SHE) (n = 37), NV with hemorrhage (n = 15), and fibrosis (n = 3). The type 2 NV group, and NV with hemorrhage groups showed significant VA improvement after 12 months of treatment (p < 0.05 in both groups); the SHE group failed to show improvement (p = 0.366). All groups showed a significant reduction in central foveal thickness after 12 months of treatment . The SHE group had a significantly higher incidence of interrupted ellipsoid zone than the other groups (p < 0.05). Myopic CNV can present as SHRM on OCT-A. Visual prognoses vary in different SHRM types. OCT-A and FA may help predict the outcomes of different subtypes of myopic CNV. SHE is predictive of outer retinal layer atrophy in patients with various SHRM types. In the east and southeast Asian regions, such as Taiwan and Japan, myopia occurs in 90% of elementary school children4. In Asian countries, the prevalence of high myopia (SE < − 6D) is approximately 7.9–16.6%, and that of very high myopia (SE < − 10D) is 0.08–0.92%6.Myopia, defined as a spherical equivalent refractive error of < − 0.5 D in the least myopic eye, affects 28% of the population worldwide7. On examination, myopic CNV appears as a yellow-greyish patch, which is occasionally accompanied by active hemorrhage. On fluorescein angiography (FA), CNV appears as early hyperfluorescence with late leakage and staining in the late phase of fluorescein angiography, whereas on sturctual optical coherence tomography it can appear as a hyperreflective material located external to the retina and internal to the retinal pigmented epithelium9. This hyperreflective material has been termed “subretinal hyperreflective material (SHRM)” in previous neovascular age-related macular degeneration (AMD) studies10. SHRM can be further classified into neovascular tissue, subretinal hyperreflective exudation (SHE), hemorrhage, and fibrosis according to the fundus photography, FA, and OCT-A findings12.Macular choroidal neovascularization (CNV) is the main cause of visual impairment in patients with high myopia13. The long-term visual prognosis of myopic CNV without treatment is poor15. The RADIANCE and MYRROR studies have demonstrated significant visual improvement in myopic CNV after treatment with ranibizumab and aflibercept21.Anti-vascular endothelial growth factor (VEGF) therapy is the first-line treatment for myopic CNV12. Studies have revealed that, unlike in neovascular AMD or diabetic macular edema, a small amount of intravitreal injection can show sustained and adequate efficacy in the treatment of myopic CNV23. A minimum number of intravitreal ranibizumab injections in eyes with myopic CNV are associated with good visual outcomes and a 5-year visual benefit24. Nevertheless, some patients with myopic CNV who received anti-VEGF treatment experienced poor improvement in visual acuity (VA) and macular atrophy.Previous studies on AMD have attempted to clarify the correlation between the visual outcome and different types of SHRMThe role of SHRM after anti-VEGF therapy, especially in terms of interrupted ellipsoid zone and macular atrophy formation, in patients with myopic CNV has not been studied. Therefore, to clarify the different treatment outcomes of anti-VEGF according to different SHRM patterns in myopic CNV, we conducted a multimodal imaging study to investigate the characteristics and fate of SHRM after anti-VEGF treatment in patients with myopic CNV.We enrolled 119 eyes of 116 patients with SHRM on OCT. These patients were followed up for at least 12 months. The baseline characteristics are shown in Table Among the 119 included eyes, 55 were right eyes, and 64 were left eyes. Among the 116 included patients, 31 were men, and 83 were women. The age of these patients ranged from 33 to 94 years (mean age 60.4 ± 12.1 years). In terms of SHRM type, there were 64 cases of type 2 NV, 37 cases of SHE, 15 cases of hemorrhage, and three cases of fibrosis. The morphological pictures were demonstrated in Fig. 25. The characteristics of the different SHRM groups are summarized in Table The criteria used to classify SHRM were based on those used in previous studiesThe baseline characteristics, including age, sex, axial length, and the number of injections in 1 year, were comparable among the four groups. However, the fibrosis group had a significantly poorer baseline VA, and the hemorrhage group had a higher CFT than the other groups.The mean logMAR VA at baseline was 0.74 ± 0.45, 1.23 ± 0.21, 0.80 ± 0.34, and 0.80 ± 0.32 in the type 2 NV, fibrosis, SHE, and hemorrhage groups, respectively. All patients received anti-VEGF treatment, and further treatment was administered if SRF or IRF persisted. The mean logMAR VA at 3, 6, and 12 months were 0.63 ± 0.57, 0.62 ± 0.55, and 0.60 ± 0.57, respectively, in the type 2 NV group; 0.78 ± 0.44, 0.83 ± 0.55, and 0.82 ± 0.55, respectively, in the SHE group; and 0.52 ± 0.51, 0.53 ± 0.51, and 0.46 ± 0.46, respectively, in the hemorrhage group. At 3, 6, and 12 months, VA in patients with type 2 NV and hemorrhage improved significantly from the baseline VA (p < 0.05)  , between October 2007 and February 2014. The retrospecitve study was approved by the Institutional Review Board of National Taiwan University Hospital(202007059RIND) and the study was conducted in accordance with the tenets of the Declaration of Helsinki. Informed consent was waived by the Institutional Review Board of National Taiwan University Hospital due to retrospective nature of study.All patients underwent slit lamp examination, Snellen best-corrected VA (BCVA) testing, color fundus photography, FA, spectral-domain OCT, and OCT-A. Since the RTVue XR Avanti SD-OCT platform is launched in 2003. Before RTVue XR Avanti was laucnhed, the patient received spectral domain struture OCT and fluorescein angiography. The VA results were converted to logMAR values for further statistical analysis.The inclusion criteria were eyes with myopic CNV, observation of foveal SHRM on FA and OCT-A, initial (pre-treatment) VA of 20/400 or better, and a follow-up period of at least 1 year. Patients with eye diseases that affect VA, including polypoidal choroidal vasculopathy, diabetic retinopathy, retinal detachment, and glaucoma, and those who previously received treatment for myopic CNV were excluded.Myopic CNV was diagnosed using color fundus photography, FA, and OCT. These patients received intravitreal injections of ranibizumab (0.5 mg/0.05 ml) or aflibercept (2.0 mg/0.05 ml) via a 30-gauge needle through the pars plana. All patients underwent monthly follow-up evaluations and received additional injections as needed. Repeat injections were administered to patients with persistent or recurrent intra-retinal fluid (IRF) or subretinal fluid (SRF).BCVA was measured before treatment and at 3, 6, and 12 months after the first injection. The OCT morphological features, including central foveal thickness (CFT), the morphology of the ellipsoid zone, the presence or absence of IRF or SRF, and the presence or absence of subfoveal atrophy at 12 months, were collected from the patients’ medical records. Atrophic changes included the loss of retinal pigment epithelium (RPE) and the outer retinal segment on OCT.25. If multiple components of SHRM were noted on OCT, we classified them according to the SHRM in the foveal center. Myopic SHRM was classified into four groups: type 2 neovascularization (NV), SHE, fibrosis, and hemorrhage. All patients received treatment with anti-VEGF agents.The OCT-A system used in our study  was a spectral domain-OCT device with an A-scan rate of 70,000 scans per second, a light source centered at 840 nm, and a bandwidth of 45 nm to obtain split-spectrum amplitude-decorrelation angiography images. The component and classification were judged according to the criteria described previouslyThe primary outcome measure was the mean change in BCVA after treatment for 12 months in the different groups of SHRM. The OCT morphological features, including CFT, the integrity of the ellipsoid zone, the presence or absence of IRF or SRF, and the presence or absence of atrophy, were analysed in the four groups. The interrupted ellipsodie zone is the continuous signal noted on structured OCT showed discontinuity.t-test to analyze the changes in VA and CFT after treatment in the four groups. The OCT morphological features were compared among the different groups using the chi-square test. P < 0.05 was considered statistically significant.After testing the normality of the data, we used the paired


Cucumis sativus L.) wilt disease caused by Fusarium oxysporum f. sp. cucumerinum (Foc). When using the isolate VJH504 isolated from cucumber rhizosphere soil and identified as Bacillus velezensis, the growth of Foc in the double culture experiment was effectively inhibited. Phenotypic, phylogenetic, and genomic analyses were conducted to enhance understanding of its biocontrol mechanism. According to the result of the phenotype analysis, B. velezensis VJH504 could inhibit cucumber fusarium wilt disease both in vitro and in vivo, and significantly promote cucumber seed germination and seedling growth. Additionally, the tests of growth-promoting and biocontrol characteristics revealed the secretion of proteases, amylases, β-1,3-glucanases, cellulases, as well as siderophores and indole-3-acetic acid by B. velezensis VJH504. Using the PacBio Sequel II system, we applied the complete genome sequencing for B. velezensis VJH504 and obtained a single circular chromosome with a size of 3.79 Mb. A phylogenetic tree was constructed based on the 16S rRNA gene sequences of B. velezensis VJH504 and 13 other Bacillus species, and Average Nucleotide Identity (ANI) analysis was performed using their whole-genome sequences, confirming isolateVJH504 as B. velezensis. Following this, based on the complete genome sequence od B. velezensis VJH504, specific functional analysis, Carbohydrate-Active Enzymes (CAZymes) analysis, and secondary metabolite analysis were carried out, predicting organism’s abilities for biofilm formation, production of antifungal CAZymes, and synthesis of antagonistic secondary metabolites against pathogens. Afterwards, a comparative genomic analysis was performed between B. velezensis VJH504 and three other B. velezensis strains, revealing subtle differences in their genomic sequences and suggesting the potential for the discovery of novel antimicrobial substances in B. velezensis VJH504. In conclusion, the mechanism of B. velezensis VJH504 in controlling cucumber fusarium wilt was predicted to appear that B. velezensis VJH504is a promising biocontrol agent, showcasing excellent application potential in agricultural production.One major issue in reducing cucumber yield is the destructive disease Cucumber ( Due to the ability of Foc to produce chlamydospores and conidia, which have strong resistance to adverse conditions, it is nearly impossible for Foc to be completely eliminated once it colonizes in soil , of which 5.9 and 1.6% are related to antagonistic secondary metabolites and antibiotic resistance, respectively  was taken from the edge of the Foc FJH36 agar plate and placed at the center of a Potato Dextrose Aga (PDA) plate. Bacteria were inoculated 25 mm away from the plug in a straight line. A control plate was prepared by inoculating Foc FJH36 without the bacteria. This process was repeated three times. The plates were incubated at 28°C for 7 days, and the diameter of the pathogenic fungus was measured to calculate the antimicrobial activity  medium. After incubating at 37°C and 160 rpm for 36 h, a biocontrol bacterial solution with OD2.2.2.2.1.Foc FJH36 was cultured on PDA plates at 28°C for 5 days. Then, using a 5 mm punch, 10 plugs containing the pathogenic fungus were transferred to 100 mL of Potato Dextrose Broth (PDB). The culture was incubated on a shaker at 28°C and 180 rpm for 3 days. Afterward, the Foc FJH36 culture was filtered through four layers of sterile gauze, and sterile water was added to dilute the concentration of pathogenic fungal spores to 1 × 108 CFU/mL.The B. velezensis VJH504 was cultured in LB medium at 37°C and 180 rpm for 24 h. After the incubation, the culture was centrifuged at 4,000 xg for 5 min to collect the bacterial cells. The cells were then suspended in LB medium, and further diluted to a concentration of 1 × 108 CFU/mL.The strain 2.2.2.B. velezensis VJH504 for 4 h and rinsed with sterile water for three times. For the control treatment, the seeds were soaked in distilled water for the same duration. The seeds were placed in petri dishes lined with two layers of qualitative filter paper. Each dish contained 40 seeds, and each treatment repeated the process for three times. The filter paper was kept moisture with water and the dishes were placed in a growth chamber at a temperature of 30°C and humidity between 60 to 70%. After 24 h, the germination of the seeds in each treatment was examined and the germination rate for each treatment was calculated.The healthy, plump, and uniformly sized cucumber seeds of the variety Bojie 616 were selected and surface sterilized with 75% ethanol (v/v) for 30 s, followed rinsing with sterile water for 5 times. The sterilized seeds were pre-soak in warm water at 55°C for 10 min. After removing excess water from the seed surface using filter paper, the seeds were immersed in a bacterial suspension of 2.2.3.B. velezensis VJH504 suspension every 2 days for a total of three irrigations; (2) After three root irrigations with 20 mL of B. velezensis VJH504 suspension, root irrigation with 20 mL of Foc FJH36 spore suspension; (3) Root irrigation with 20 mL of 0.1% hymexazol once, followed by root irrigation with 20 mL of Foc FJH36 spore suspension after 24 h; (4) Root irrigation with 20 mL of sterile distilled water every 2 days for a total of three irrigations; (5) After three root irrigations with 20 mL of sterile distilled water, root irrigation with Foc FJH36 spore suspension. Measurements were taken at 20 and 40 days after inoculation of the pathogen. The measured parameters included plant height, stem thickness, leaf area, chlorophyll content, aboveground fresh weight, belowground fresh weight, aboveground dry weight, belowground dry weight. The vigor index was calculated as (stem thickness/plant height + root dry weight/aboveground dry weight) × total plant dry weight. The severity and incidence of cucumber wilt disease were recorded at 40 days. Cucumber wilt disease was classified into 5 levels: 0 = no symptoms on leaves; 1 = wilting of leaves less than 1/4 of cucumber seedlings; 2 = wilting of 1/4 to 1/2 of cucumber seedling leaves; 3 = wilting of more than 1/2 of cucumber seedling leaves; 4 = complete wilting and death of the whole cucumber plant. Disease index (DI) was calculated using the formula DI = [(0 × N0) + (1 × N1) + (2 × N2) + (3 × N3) + (4 × N4)]/(T × 4) × 100, where N represents the number of cucumber seedlings with each disease rating, and T is the total number of cucumber seedlings. Disease incidence was calculated as [(N1 + N2 + N3 + N4)/T] × 100%. Control efficacy was determined by the formula (Control DI - Treatment DI)/Control DI × 100%. Leaf chlorophyll content was measured using an SPAD-502 Plus chlorophyll meter. Root length and branch height were measured using a ruler. Root and stem fresh weights were recorded using an analytical balance . Stem thickness at a distance of 2 cm from the crown was measured using a caliper .Bojie 616 cucumber seeds with strong, full and uniform size were selected. The seeds were disinfected with 75% ethanol (v/v) for 30 s and then rinsed with sterile water for 5 times. The seeds were soaked in warm water (55°C) for 10 min. Then soaked in distilled water for 4 h. Wet a sterile cloth with sterile water and rinse the soaked cucumber seeds several times. The washed seeds were neatly arranged on the cloth and placed in a constant temperature incubator at 28°C for germination. Once the seed has germinated and shows a white tip, it is sown in a sterile substrate-filled seedling tray. Six days after sowing, the seedlings with two true leaves were transferred to a pot with a diameter of 10 cm and a height of 10 cm. The pot was filled with 400 g sterile substrate. Five pots of plants were placed in a greenhouse at a temperature of 14 ~ 25°C for 16 h of light and 8 h of darkness, with a light cycle of 4 d. The experiment was conducted using a completely randomized design with five treatments and five replications per treatment. The treatments were as follows: (1) Root irrigation with 20 mL of 2.3.Bacillus velezensis VJH504 was inoculated in skim milk agar  with a single colony, and cultured at 30°C for 48 h (B. velezensis VJH504 was inoculated into carboxymethyl cellulose (CMC) agar  with single colony, and cultured at 30°C for 48 h. Flood the plate with 1% (w/v) Congo Red solution, and then rinse with sterile distilled water. The transparent area around the colony proved that the strain had cellulase activity. Inoculate B. velezensis VJH504 as single colonies in Chrome Azurol S (CAS) blue agar , 50 mL CAS staining solution, and 18 g agar / L, pH 7.2), and culture at 30°C for 48 h. A color change from blue to orange around the colonies indicates the production of siderophores. B. velezensis VJH504 was inoculated into L-tryptophan nutrient broth  with a single colony, and cultured at 30°C for 48 h. After centrifugation at 14,000 xg for 10 min, mix 1 mL of the supernatant with 2 mL of Salkowski reagent, and then allow it to stand at room temperature in the dark for 30 min. A color change from yellow to orange indicates the production of indole-3-acetic acid (IAA)  and cultured at 30°C for 48 h. The transparent area around the colony proved that the strain had β-glucanase activity (for 48 h . The clefor 48 h . B. veleid (IAA) . B. veleactivity .2.4.After 20 dpi of strain inoculation, the leaves were harvested and stored at −80°C. 0.5 g of leaves were ground in liquid nitrogen and 1 mL of extraction buffer was added. After centrifugation at 8000 xg for 10 min, the supernatant was collected for enzyme determination. Enzyme activities were measured using assay kits for PPO, SOD, CAT, PAL, and LOX following the manufacturer’s protocols . Absorbance was measured using a microplate reader .2.5.B. velezensis VJH504 was inoculated in LB medium and cultured at 28°C, 150 rpm for 18 h. The genomic DNA of B. velezensis VJH504 is extracted using the Mini BEST Bacteria Genomic DNA Extraction Kit Ver.3.0 according to the manufacturer’s instructions. A 10 kb insert fragment Pac Bio library was constructed and the whole genome was sequenced using the Pac Bio Sequel II system. The sequencing reads are assembled de novo using HGAP4 and Canu software. The genome coverage depth is analyzed using alignment tools. The assembled complete genome sequence of B. velezensis VJH504 is deposited in NCBI GenBank under accession number CP131928. A circular genome map of B. velezensis VJH504 is constructed using Circos (The single colony of g Circos .2.6.B. velezensis VJH504 was annotated using Glimmer (v3.02) (The genome of  (v3.02) . tRNA an (v3.02)  and RNAm (v3.02) , respect2.7.B. velezensis VJH504 and B. velezensis FZB42, B. velezensis G341, B. velezensis BvL03, B. subtilis YB-04, B. subtilis H1, B. amyloliquefaciens 35 M, B. amyloliquefaciens Ba13, B. licheniformis ATCC 14580, B. licheniformis SRCM 120569, B. pumilus ZB201701, B. pumilus SF-4, B. thuringiensis B13, and B. thuringiensis JW-1  was constructed using the Neighbor Joining method in MEGA7.0  database   . The proon 7.0.0 .2.9.p ≤ 0.05.Statistical analysis was performed using SPSS v21.0. One-way analysis of variance (ANOVA) was conducted to analyze the data. The mean values were compared using Duncan’s multiple range test with a significance level set at 3.3.1.Foc FJH36 in the dual-culture assay, twelve strains were chosen from the numerous colonies with various morphologies, isolated and purified from healthy cucumber rhizosphere soil , the plant height, stem diameter, and aboveground fresh weight of iderably –L. At 20at 20dpi . In compeatments . At the eatments ,K. The seatments .3.5.B. velezensis (14.11%) sequences, followed by B. amyloliquefaciens Y2 (2.11%) and B. amyloliquefaciens CC178 (1.08%)  and B. amyloliquefaciens (0.51%)  . In the  (0.51%) .3.6.B. velezensis VJH504 and 13 other Bacillus strains  values (97% and above) were only observed with B. velezensis strains , Electrochemical Potential-driven Transporters (299), Incompletely Characterized Transport Systems (188), Channel/Pores (91), Group Translocators (69), Accessory Factors Involved in Transport (43), and Transmembrane Electron Carriers (18) (In ers (18) .3.8.B. velezensis VJH504 identified 86 genes as putative CAZymes, including 40 Glycoside Hydrolases (GHs), 18 Glycosyl Transferases (GTs), 17 Carbohydrate Esterases (CEs), 6 Auxiliary Activities (AAs), 3 Polysaccharide Lyases (PLs), and 5 Carbohydrate-Binding Modules (CBMs), which play a crucial role in facilitating the binding of enzymes to their substrates. Among them, three genes, orf01880, orf01885, and orf03428, were classified as both CHs and CBMs  were analyzed for syntenic regions. This was accomplished by using the Mauve software, which effectively identifies conserved genomic regions in multiple genomes, rearrangements, and more. The results show that the three B. velezensis strains and B. velezensis VJH504, share similar genome structures and have no genetic mutations. However, B. velezensis VJH504 and G341 as well as BvL03 have a minor genetic inversion . It is apparent that the ANI values between VJH504 and all B. velezensis strains were above 97%, surpassing the threshold of 96%. Strong suggestion of a higher degree of genomic homogeneity between VJH504 and B. velezensis FZB42 due to the results showed that the highest ANI value of VJH504 with B. velezensis FZB42. Moreover, the comparative genomic analysis revealed that VJH504 strain has the highest number of homologous genes and single-copy genes among the strains examined. The most striking results that VJH504 strain possesses certain specificity, and its biocontrol mechanisms exhibit some differences compared to other B. velezensis strains.Thereafter, the phylogenetic tree analysis using the 16S rRNA gene sequences was carried out, and found that the VJH504 strain clusters were closely with B. subtilis 3,610 formed biofilms on tomato roots, enhancing its colonization ability in the biocontrol process against tomato bacterial wilt. Futhermore it was discovered that the formation ability of biofilms positively correlated with biocontrol efficacy against Ralstonia solanacearum, which is the causal agent of bacterial wilt, by the construction and evaluation of biofilm deficient and enhanced mutants. It has been reported that quorum sensing significantly affects biofilm formation in bacteria   .The exciB. subtilis, B. amyloliquefaciens, and B. velezensis, have been found to inhibit grasses’ spore germination or hyphal growth. In dual cultures of plant pathogens and antagonistic bacteria, the hyphae of the pathogenic fungi exhibit deformities. B. subtilis from grapevine wood, which could inhibit the growth of the grapevine trunk disease pathogen Eutypa lata and induce abnormal hyphal growth. When the concentration of B. subtilis was equal to or greater than 0.8 mg/mL, its antibiotic substances completely inhibited the germination of Eutypa lata spores. B. subtilis ZD01, which exhibited strong antifungal activity, produced volatile organic compounds that caused significant morphological changes in Alternaria solani, inhibiting its conidial germination and reducing the lesion area and pathogen population. The findings of this study mirror those of the previous studies that have examined the morphological changes in fungi. In our case, the microscopic images showed that B. velezensis VJH504 could inhibit the hyphal development of Foc and induce abnormal swelling of spores and hyphae in dual-culture antagonism tests. Additionally, the genome annotation of B. velezensis VJH504 revealed a total of 329 antibiotic resistance genes with a large number of CAZymes genes, indicating its potential for degradation and utilization of fungal polymers as a nutrient source. Its strong capability in combating fungal pathogenswas proved previously  pathway and induce resistance mechanisms in soybeans. Subsequently, research on Bacillus-induced systemic resistance (ISR) in plants has gained significant attention. In this study, analysis of the secondary metabolite gene clusters in B. velezensis VJH504 revealed the presence of gene clusters encoding surfactin and fengycin. Therefore, it can be inferred that B. velezensis VJH504 has the potential to induce similar systemic resistance in plants.B. velezensis VJH504. Based on the experimental and genomic analysis, particularly in the aspects of biofilm formation, antibiotic synthesis, promotion of plant growth, and production of systemic resistance-inducing agents, the biocontrol mechanisms of B. velezensis VJH504 can be inferred as follows: (1) the formation of Biofilm enhance colonization ability in the plant root system, enabling rapid establishment in the rhizosphere and occupying ecological niches where competition with Foc is limited; (2) Secretion of cell wall-degrading enzymes and secondary metabolites such as terpenes, lanthipeptides, bacilysin, fengycin, bacillibactin, bacillaene, and macrolactin H antagonize phytopathogenic fungi; (3) Production of iron carriers and indole-3-acetic acid promote nutrient uptake and plant growth, thereby achieving disease control; (4) Inducing systemic resistance in plants control cucumber wilt disease  can be found at: FY: Conceptualization, Investigation, Methodology, Writing – original draft. HJ: Data curation, Formal analysis, Software, Visualization, Writing – original draft. KM: Funding acquisition, Writing – review & editing. XW: Supervision, Writing – review & editing. SL: Funding acquisition, Supervision, Writing – review & editing. YC: Supervision, Writing – review & editing. YJ: Supervision, Writing – review & editing. BT: Supervision, Writing – review & editing. XS: Supervision, Writing – review & editing.


The oxadiazole ring has long been used for the treatment of several diseases. This study aimed to analyze the antihyperglycemic and antioxidant roles of the 1,3,4-oxadiazole derivative with its toxicity. Diabetes was induced through intraperitoneal administration of alloxan monohydrate at 150 mg/kg in rats. Glimepiride and acarbose were used as standards. Rats were divided into groups of normal control, disease control, standard, and diabetic rats . After 14 days of oral administration of 1,3,4-oxadiazole derivatives  to the diabetic group, the blood glucose level, body weight, glycated hemoglobin (HbA1c), insulin level, antioxidant effect, and histopathology of the pancreas were performed. The toxicity was measured by estimating liver enzyme, renal function, lipid profile, antioxidative effect, and liver and kidney histopathological study. The blood glucose and body weight were measured before and after treatment. Alloxan significantly increased blood glucose levels, HbA1c, alanine transaminase, aspartate aminotransferase, urea, cholesterol, triglycerides, and creatinine. In contrast, body weight, insulin level, and antioxidant factors were reduced compared to the normal control group. Treatment with oxadiazole derivatives showed a significant reduction in blood glucose levels, HbA1c, alanine transaminase, aspartate aminotransferase, urea, cholesterol, triglycerides, and creatinine as compared to the disease control group. The 1,3,4-oxadiazole derivative significantly improved body weight, insulin level, and antioxidant factors compared to the disease control group. In conclusion, the oxadiazole derivative showed potential antidiabetic activity and indicated its potential as a therapeutic. Diabetes mellitus (DM) is often simply referred as a group of metabolic diseases in which a person has high blood sugar as well as altered fat, carbohydrate, and protein metabolism because either the body does not produce enough insulin (Type I) or cells do not respond to the insulin (Type II) . This hiAlthough the future cannot be predicted with certainty, it is surely possible that an extensive epidemic of resistance to antidiabetic drugs could potentially harm millions of people. Given that it takes more than 10 years to establish the efficacy and safety of the new compound, there is an urgent need to restock the antidiabetic agents. Only a few new antidiabetic compounds have received approval from the US Food and Drug Administration . As we lLiterature survey reveals 1,3,4-oxadiazole derivatives , which bα-amylase and α-glucosidase, while preventing oxidative stress is deemed as a practicable strategy for regulating postprandial glucose levels in diabetic patients [α-glucosidase inhibitor in many molecular hybrid designs [1,3,4-oxadiazole derivative exerts its effects on diabetes by inhibiting the carbohydrate hydrolyzing enzymes, i.e., patients . 1,3,4-o designs . Conceiv designs .Oxadiazole occurs in four feasible interpretations of isomers, but 1,3,4-oxadiazole is broadly researched for different functions . OxadiazThe ring of 1,3,4-oxadiazole is attached with the phenol and sulfanyl hydroxyethyl group . In viewAlloxan monohydrate, hydrogen peroxide, sodium chloride, chloroform, sodium hydroxide, pyrogallol, potassium phosphate, sodium carbonate, glimepiride, acarbose, hydrochloric acid, folic acid, dextrose, trichloroacetic acid, and thiobarbituric acid (TBA) were purchased from Merck. Lidocaine was purchased from Pfizer company. DTNB  acid was bought from the local market.Glucometer (Accu check), glucometer strip (Accu check), digital electronics weighing balance , vortex mixture (Vm-300), sonicator (Shakeel and Sons), spectrophotometer (Shimadzu UV-1601), centrifuge (Centurion scientific UK), homogenizer (Wise Tis HG-15A), refrigerator, digital water bath (HH-S4), binocular stereo microscope (YJ-T3c-China), and diagnostic kits  were used in this study.Studies on albino Wister rats (weighing 150–200 g) of both sexes were performed. All rats were kept at room temperature (25 ± 1°C) under normal laboratory conditions for 12 h light-dark period with a humidity of 45–55%. A standard diet of pellets and water was fed before and during the experimental rats. All methods were issued by the Research Ethical Committee (REC/RIPS-LHR/011) of Riphah International University, Lahore, Pakistan.In this study, a single intraperitoneal injection (i.p.) of alloxan (150 mg/kg in 0.9% NaCl solution) was used to induce diabetes . The intrd day of alloxan treatment. Diabetic rats were known to have a blood glucose level greater than 180 mg/dL. All treatments were given orally once a day according to the body weight. The duration of the study was 14 days. Group I: control group with normal saline (5 mL/kg p.o.), Group II: diseased group with alloxan monohydrate (150 mg/kg i.p.), Group III: treated with glimepiride (4 mg/kg p.o.), Group IV: treated with acarbose (40 mg/kg p.o.), Group V: treated with 1,3,4-oxadiazole derivative (5 mg/kg p.o.), Group VI: treated with 1,3,4-oxadiazole derivative (10 mg/kg p.o.), and Group VII: treated with 1,3,4-oxadiazole derivative .Rats were divided into seven groups, and each group contained 6 animals. Treatment with the compound was started on the 3th, 10th, and 14th day from the tail tip using a glass capillary tube. Basal blood glucose is the normal blood glucose content attained before inducing diabetes, while the day on which diabetes is completely induced in rats before starting the treatment is called zero-day [Blood glucose levels were determined using a glucometer before diabetes induction and during treatment time. The blood samples were collected on basal, zero, 5zero-day .The level of HbA1c in blood represents the average blood glucose level over the last 2–3 months. When diabetes is not well regulated, blood glucose levels will also be elevated, which will cause higher levels of HbA1c. Glycosylated hemoglobin is measured by the Nycocard reader by glycosylated hemoglobin kits .th day of treatment after anaesthetizing with isoflurane (diluted with 2% oxygen), and their blood samples were taken from cardiac puncture. The blood was then centrifuged for 15 min at 2500 rpm to extract blood serum. The concentration of insulin in serum was calculated using a commercially available DSL-1600 insulin device . The values for insulin were expressed as μU/mL.All animals were sacrificed on the 15th, 10th, and 15th days of treatment. The effect of 1,3,4-oxidiazole derivative on the body weight of rats was examined on the 5th, 10th, and 15th day of the study.The body weight of rats was examined at basal, zero, 5To prepare tissue homogenate, the pancreas was taken from sacrificed animals. All the tissues (10% w/v) were washed with cold normal saline and then dissolved in 0.1 M of phosphate buffer (pH 7.4).2CO3, 2% sodium potassium tartrate, and 1% CuSO4 was added. The mixture containing 0.5 mL of Folin–Ciocalteau phenol reagent was incubated for 10 min. The sample was incubated for 30 min before reading the absorbance at 620 nm [Tissues were adequately diluted with phosphate buffer for protein estimation, and 5 mL of copper sulfate reagent comprising 1% Nat 620 nm .The protein level was determined using the following equation:2O2) were added into the supernatant (0.05 mL). A change in the absorbance value was spectrophotometrically reported at 240 nm every 5 sec. The CAT activity was expressed as units per milligram of protein as compared to the standard [δ OD is the change in absorbance per min and E is extinction coefficient (0.071 mmol·cm−1) of H2O2 [Phosphate buffer  and 1 mL of 30 mM of hydrogen peroxide (Hstandard .(2)CAT aVT is total mixture volume (4 mL), 1.56 × 105 is the coefficient of molar extinction, WT is the weight of the dissected brain (1 g), and VU is volume of aliquot (1 mL) [The rates of MDA, an indicator of lipid peroxidation, were calculated using the double heating method. The procedure is based on spectrometric measurement of violet colour produced by the TBA reaction. For this purpose, 2.5 mL of TCA solution was added to each centrifuge tube containing 0.5 mL of the supernatant of the tissue preparation. Tubes were incubated in a boiling water bath for 15 min. Upon cooling to ambient temperature, the tubes were centrifuged for 10 min and 2 mL of each supernatant sample was mixed to 1 mL of TBA solution. Each tube was then placed for 15 min in a boiling bath. The absorbance was measured at 532 nm .(3)Concet (1 mL) .Each reaction mixture contained 2.8 mL of potassium phosphate buffer, 0.1 mL of homogeneous tissue, and 0.1 mL of pyrogallol solution. Increased absorbance at 325 nm was spectrophotometrically reported at 30 sec intervals for 5 min .Y is the absorbance at 412 nm of tissue homogenate, DF is the dilution factor (1), BT is brain homogeneous tissue (1 mL), and VU is volume of aliquot (1 mL).For the calculation of reduced glutathione, 1 mL of homogenous tissue with 1 mL of 10% TCA was precipitated. Then, 4 mL of phosphate solution and 0.5 mL of DTNB reagent were applied to the supernatant aliquot, and the absorbance was read at 412 nm.The pancreas was blotted free of mucus from laboratory rats, washed in normal saline, and preserved in 10% of formaldehyde for 24 h, followed by implanting in paraffin. The kidney and liver were collected after dissection and fixed in the blocks of paraffin wax, followed by storing in 10% formalin solution. The pancreas, kidney, and liver were stained with hematoxylin-eosin for histopathological study.The toxicological study of 1,3,4-oxadiazole derivative was observed by contrasting the degree of different parameters of biochemistry between the test group, control group, disease group, and normal group. The method of collecting blood and separating serum from blood has already been described earlier. Preparation of the supernatant from tissue is also described.Liver function tests viz. ALT and AST were analyzed by Crescent Diagnostics CZ 902L and Crescent Diagnostic CZ 904L kit, respectively. The kidney function test, i.e., the urea level was measured by Crescent Diagnostic CS612 kit, and the creatinine level was measured by Crescent Diagnostic CS604 kit. Triglyceride and cholesterol were analyzed by Crescent Diagnostic CS611 and Crescent Diagnostic CS603, respectively.Superoxide dismutase was calculated according to the method of Kim et al. . GSH andThe kidney and liver were collected and fixed in the blocks of paraffin wax, followed by saving in 10% formalin solution. The kidney and liver were stained with hematoxylin-eosin for histopathological observations.P < 0.05 and ∗∗P < 0.01) and as highly significant (∗∗∗P < 0.001).Results were presented as the mean ± SEM (standard error mean). For graphical interpretation, GraphPad prism was applied. The test of two-way ANOVA  was applied. The results were intimated as moderately significant  increased the body weight  attenuation of elevated HbA1c levels and was estimated as 0.56 ± 0.4, 0.44 ± 0.2, and 0.39 ± 0.3%, respectively, in contrast to the normal control group (0.37 ± 0.4%). The 1,3,4-oxadiazole derivative produced a dose-dependent reduction in the HbA1c level . The CAT level was significantly (P < 0.05) increased to 123 ± 1.0 μgmole/min/mg at a dose level of 15 mg/kg as compared to the disease control group which was recorded as 96 ± 0.5 μgmole/min/mg. 1,3,4-oxadiazole derivative at a dose level of 15 mg/kg showed a significant (P < 0.05) increase in the level of the GSH level as compared to the disease control group. Administration of 1,3,4-oxadiazole derivative at 15 mg/kg to the diabetic rats reduced the level of MDA to 0.370 ± 0.005 μmole/mg, which was a nonsignificant reduction as compared to the disease control group (0.390 ± 0.010 μmole/mg). 1,3,4-oxadiazole derivative at a dose of 5, 10, and 15 mg/kg showed an increase in the protein level which was significantly different  in the disease control group .The level of SOD was increased to 53.5 ± 0.5 ol group .β cells of Langerhans.Histology of the pancreas demonstrated normal acini and cellularity in the normal control group. In diseased animals, extensive damage to islet of Langerhans and hemorrhagic condition can be seen clearly in P < 0.001) reduction in ALT level (48.95 ± 1.75 unit/L) was observed after 14 days administration of 15 mg/kg of 1,3,4-oxadiazole derivative with respect to the disease control (84.41 ± 4.38 unit/L). Similar results were observed in AST level (39.50 ± 0.50 unit/L) with administration of 15 mg/kg of 1,3,4-oxadiazole derivative as compared to the disease control group (74.85 ± 0.25 unit/L)  .P < 0.001) different as compared to the disease control group (199 ± 1.0 mg/dL). The administration of 1,3,4-oxadiazole derivative at the dose level of 15 mg/kg for 14 days significantly (P < 0.001) reduced the triglyceride level (131 ± 1.0 mg/dL) as compared to the disease control group (166 ± 1.0 mg/dL). Similarly, a significant (P < 0.001) reduction in the urea level was observed in the presence of 15 mg/kg of an 1,3,4-oxadiazole derivative (65.9 ± 0.4 mg/dL) as compared to the disease control group (127 ± 2.4 mg/dL) , which was significantly (4 mg/dL) .μmole/mg as compared to the disease control group which was found as 1.300 ± 0.009 μmole/mg in the liver. In addition, SOD and GSH activities at a dose level of 10 and 15 mg/kg of 1,3,4-oxadiazole derivative were significantly (P < 0.05) increased as compared to the disease control group in the liver. Similarly, the CAT activity in hepatic tissue was significantly (P < 0.01) increased by adding 1,3,4-oxadiazole derivatives at 15 mg/kg (0.891 ± 0.005 μgmole/min/mg) as compared to the corresponding disease control group (0.450 ± 0.005 μgmole/min/mg). Similarly, 15 mg/kg of 1,3,4-oxadiazole derivative significantly (P < 0.05) increased the protein level (454 ± 0.5 μg/mg) as compared to the disease control group (341 ± 0.1 μg/mg). However, in the kidney, the treatment with 1,3,4-oxadiazole derivative at a dose level of 15 mg/kg significantly increased (12.12 ± 0.52 μg/mg) the level of reduced GSH on the 15th day as compared to the disease control group (4.57 ± 1.4 μg/mg). Treatment with 10 and 15 mg/kg of 1,3,4-oxadiazole derivative significantly (P < 0.05) elevated the level of CAT as compared to the disease control group in the kidney. Treatment with 1,3,4-oxadiazole derivative at a dose level of 10 and 15 mg/kg showed a significant (P < 0.01) increase in SOD activity (48.75 ± 0.54 μg/mg and 48.90 ± 0.10 μg/mg) as compared to the disease control group (44.05 ± 0.54 μg/mg) in the kidney. In the case of protein level, only 15 mg/kg of 1,3,4-oxadiazole derivative showed a significant (P < 0.05) increase in the protein level in the kidney. Treatment with 1,3,4-oxadiazole derivative at a dose level of 5 and 10 mg/kg showed a significant (P < 0.01) reduction in MDA level as compared to the disease control group.Hepatic microscopic examination of the normal group showed healthy liver cells with a well-preserved cytoplasm, nucleus, core nuclei, and major vein . The disIn normal animals, the histology of the glomeruli and the Bowman capsule is appropriate. When lipid is deposited in the glomeruli of diseased rats, the basement membrane of the arterioles of the glomeruli becomes slightly thicker. This condition is known as glomerular lipidosis, and it includes fibrosis and cellular proliferation in the mesangial as well as congestion in the Bowman's as shown in Diabetes mellitus is the condition of hyperglycemia and derangement in carbohydrates, proteins, and fats due to the lack of insulin . WorldwiInsulin is a hormone made by the pancreas that binds to the glycoprotein receptor on the cell membrane of the cell and stimulates the transportation of glucose across the cell membrane by glucose transporter GLUT4 . In diab2−) to nonreactive H2O2 and molecular oxygen [P < 0.001) rise in the SOD level when compared to the diabetic group. Catalase performs its activity to convert H2O2 into water and O2 and protect the cell from oxidative stress [2O2 can rapidly diffuse across cell membranes, and in the presence of transition metal ions, it can be converted to hydroxyl radicals via Fenton chemistry [2+ and H2O2 according to the Fenton reaction. The imbalance ratio of the antioxidant enzymes and oxidants (ROS) results in the emergence of illness [−) into water. In this mechanism, hydroxyl production is affected [P < 0.05) increase in the GSH level. Similarly, 10 and 15 mg/kg of 1,3,4-oxadiazole derivative showed a significant (P < 0.01) increase in the GSH level in the kidney as compared to the diabetic group.Diluric acid is oxidized to alloxan to generate the superoxide radical . The SODr oxygen . At a doe stress . Howeverhemistry . Highly  illness . Glutathaffected . Our resP < 0.001) and kidney (P < 0.01), whereas nonsignificant reduction was reported in the pancreas as compared to the diabetic group. The protein level was increased in 1,3,4-oxadiazole derivative as compared to the diabetic groups. MDA is the mutagenic in bacteria and mammalian cells [The current data revealed that consistent hyperglycemia via alloxan-generated ROS caused marked oxidizing effects, as evidenced by an increase in diabetic animal MDA rates compared to the nondiabetic animals . High lean cells .β cells.Histology of the pancreas demonstrates normal acini and normal cellularity in the control islets of Langerhans. In diabetic animals, extensive damage to Langerhans islets was treated and reduced islet dimensions were observed too. On the other hand, at a dose level of 15 mg/kg, 1,3,4-oxadiazole derivative showed hypertrophy and vacuolations of Langerhans In the serum of diabetic rats, hepatic enzymes have increased . This maCholesterol and triglyceride play a major role in the pathogenesis of DM-related complications. The unusually high serum cholesterol and triglyceride content in diabetics were primarily due to the increased production of free fatty acids from peripheral fat depots. As lipase is suppressed by insulin, insulin insufficiency or resistance may result in dyslipidemia . The groP < 0.001) decreased the plasma urea level. Similarly, the level of creatinine was decreased (P < 0.001) after the administration of 1,3,4-oxadiazole derivative in comparison to the diseased group.Diabetic hyperglycemia contributes to an increase in urea and creatinine plasma levels, which are regarded as important markers of renal dysfunction . With reHepatic photomicrographs displayed the normal liver cells through a well-preserved cytoplasm, nucleus, core nuclei, and main vein in the normal group. The standard lobular structure was conserved in diabetic rats. The main vein detrimentally clogged up. Focal hemorrhage was also established. The accumulation of fat was noticeable. Kupfer cell activation and congestion of the central vein can be seen in the liver treated with 15 mg/kg of 1,3,4-oxadiazole derivative. In normal animals, the histology of the glomeruli and the Bowman capsule is appropriate. For diabetic rats, the basement membrane of the arterioles of glomeruli is slightly thickened, including a slight change in mesangial mesangium density. Epithelial proteins were deposited in the lumen of the renal tubules after 15 mg/kg of oxadiazole derivative treatment.The potential mechanism for oxadiazole derivative action was found to be a high-affinity ligand of the receptor-activated peroxisome proliferator, a member of the superfamily of nuclear receptors that cause transcription of insulin-responsive genes . The gluNumerous studies have been conducted on Nrf2 as a potential antioxidant signaling mechanism . Nrf2 acVarious groups of heterocyclic and fused heterocyclic molecules have been described in search of antiobesity, antidiabetes, anti-inflammatory, antibiotic, and anticancer agents through scientific analysis and drug design approach . OxadiazIt can be concluded that the imbalance between oxidants and antioxidant enzymes play a pivotal role in development and progression of diabetes. In this study, the 1,3,4-oxadiazole derivative produced pronounced antidiabetic activity at a dose of 15 mg/kg, showing substantial decrease in the blood glucose level, body weight, and Hb1Ac level. Other biochemical metabolic parameters including ALT, AST, urea, creatinine, triglyceride, and cholesterol were also reduced by 1,3,4-oxadiazole derivative. The antioxidant enzyme activity was significantly increased by 1,3,4-oxadiazole derivative. The exact mechanism by which 1,3,4-oxadiazole derivative decreases the level of blood glucose in diabetic rats requires further analysis. Moreover, further studies are needed to explore the pharmacological profile of 1,3,4-oxadiazole derivative.