Scedosporium and Lomentospora species.This study was aimed to determine the potency of Minocycline (MIN) and azoles, including itraconazole (ITR), voriconazole (VOR) and posaconazole (POS) against G. mellonella infection model demonstrated the in vivo synergistic antifungal effect of MIN and azoles.This study revealed that MIN exhibited no significant antifungal activity against any of the tested strains, whereas in vitro combination of MIN with ITR, VOR or POS showed satisfactory synergistic effects against 8 (80%), 1 (10%), and 9 (90%) strains, respectively. Moreover, combined use of MIN with azoles decreased the minimum inhibitory concentration (MIC) range from 5.33–16 μg/ml to 1–16 μg/ml for ITR, from 0.42–16 μg/ml to 0.21–16 μg/ml for VOR, and from 1.33–16 μg/ml to 0.33–16 μg/ml for POS. Meanwhile, no antagonistic interactions were observed between the above combinations. The Scedosporium and Lomentospora species, while showing a potential for overcoming and preventing azole resistance.The present study demonstrated that combinations between MIN and azoles lead to synergistic antimicrobial effects on  Scedosporium and Lomentospora species are an aggressive and conditional pathogen, causing a rare type of fungal infection, with only 370 cases reported worldwide up to 2007 [Fusarium, and other relatively common hyalohyphomycosis [The incidence of fungal infections is increasing year by year. In particular, invasive fungal infections are increasingly becoming an important factor endangering human health and life . Invasiv to 2007 . The infomycosis .Scedosporium and Lomentospora species is increasing, posing a serious threat to the effectiveness of antifungal drugs [Scedosporium and Lomentospora species are highly resistant to antifungal drugs, highlighting the need for alternative treatment modalities [Lomentospora prolificans infections [Azoles, including itraconazole (ITR), voriconazole (VOR) and posaconazole (POS), are an earlier class of drugs used to treat fungal infections . Accordial drugs . Liu et dalities . A recendalities . Moreovefections . CollectCandida albicans- Staphylococcus aureus dual-species biofilms [Scedosporium and Lomentospora species.Minocycline (MIN) is a semisynthetic broad-spectrum tetracycline antibiotic that displays a similar antibacterial spectrum with tetracycline. MIN can combine with tRNA to achieve antibacterial effect, while being the most potent antibacterial agent among the existing tetracycline antibiotics. Moreover, previous studies have demonstrated that Azoles (fluconazole) in combination with MIN is a potential approach for counteracting biofilms . Based oLomentospora strains.As presented in Table Scedosporium and Lomentospora species (Table Scedosporium minutisporum (8). Meanwhile, no antagonistic interactions were detected in each of the tested combinations.Combination of MIN with ITR, VOR, or POS exhibited synergistic activities against 12 (75.00%), 1 (6.25%), or 15 (93.75%) strains of G. mellonella were infected with Scedosporium apiospermum, and the infected larvae were then treated with MIN and azoles. As shown in Fig. P < 0.05). Together, these findings demonstrated the in vivo synergistic antifungal activity of MIN and azoles against G. mellonella, as evidenced by the increased larval survival.To assess the synergistic effects of MIN and azoles in vivo, Scedosporium apiospermum infection; Uninfected Control-Saline Treatment, wild type larvae injected with saline; Infected Control-No Treatment Group, larvae infected with Scedosporium apiospermum receiving no treatment; POS Group, Scedosporium infected larvae treated with POS only; POS + MIN Group, Scedosporium infected larvae treated with POS in combination with MIN; ITR Group, Scedosporium infected larvae treated with ITR only; ITR + MIN Group, Scedosporium infected larvae treated with ITR in combination with MIN; VOR Group, Scedosporium infected larvae treated with VOR only; VOR + MIN Group, Scedosporium infected larvae treated with VOR in combination with MIN; POS: Chlorhexidine; ITR: Itraconazole; VOR: Voriconazole. The experiment was repeated thrice on different days. *P < 0.05; **P < 0.01; ****P < 0.0001.Uninfected Control Group, wild type larvae without Scedosporium apiospermum spores and hyphae clusters in the infected tissues was detected in all the experimental groups. We observed that treatment with azoles alone resulted in a slight reduction in the number of visible fungal clusters relative to the Scedosporium group, while the number of visible fungal clusters was significantly decreased in the combination treatment groups as compared to the three azoles alone groups.We further performed a histopathological examination of the larvae. As depicted in Fig. Scedosporium and Lomentospora species are important and very recalcitrant conditional pathogens. While the risk factors for infection of Scedosporium and Lomentospora species include a history of immunodeficiency, trauma, and drowning, the infection mainly involves skin, eyes, lungs, brain, even the whole body [Scedosporium and Lomentospora species in vitro [Scedosporium and Lomentospora species. Given the low antimicrobial activity of single drugs, combined use of multiple drugs may enhance their antimicrobial activities. Azole antifungal agents are commonly used in combination with other types of antifungal drugs in clinical practice, such as amphotericin B [ole body , 12. Ovein vitro , 14. Theericin B .Candida albicans [Scedosporium and Lomentospora species. In the present study, we examined the inhibitory effects of ITR, VOR, POS, and MIN alone as well as those of MIN in combination with azoles on Scedosporium and Lomentospora species.MIN has been used clinically as an antibiotic for many years since it was marketed. In recent years, MIN has gained an increasing attention with the research on its non-antimicrobial effect. Previous studies have shown that MIN reduces cell surface hydrophobicity and extracellular levels of 1,3-beta-D-glucan  in biofilms, revealing a mechanism underlying the inhibitory effect of MIN on albicans . Given talbicans , 18. HowScedosporium and Lomentospora species were studied in vitro. While MIN alone showed no in vitro antifungal activity, MIN acted synergistically with ITR, VOR or POS against 8 (80%), 1 (10%), or 9 (90%) isolates tested, showing promising synergistic effects between MIN and azoles against Scedosporium and Lomentospora species. In addition, we found that Lomentospora prolificans displays low susceptibility to current antifungal drugs, being consistent with the previous reports [Scedosporium and Lomentospora species. The in vivo experiments provided more evidence that MIN acts synergistically with azoles against Scedosporium and Lomentospora species  are characterized by the serious clinical manifestations as well as the difficulty in diagnosis and treatment, while y common , 23. As y common , 25. OveScedosporium and Lomentospora species, achieving a synergistic effect between MIN and azoles. Further studies need to be conducted for gaining insights into clinical impacts of the combination of MIN and azoles, as well as understanding clinical relevance of the in vitro data obtained in this study.In conclusion, MIN in combination with azoles may help to enhance the antifungal activities of azoles against Scedosporium aurantiacum (CBS 116910), Scedosporium minutispora (CBS 116911), Scedosporium boydii (CBS 101.22), Lomentospora prolificans (CBS 467.74), Scedosporium dehoogii (CBS 117406), and Scedosporium apiospermum (CBS 116899) were generously provided by Prof. Sybren de Hoog of Fungal Biodiversity Centre (CBS) in Netherlands. The other strains were purchased from Nanjing Institute of Dermatology, China, including Scedosporium boydii (1), Scedosporium boydii (2), Scedosporium boydii (3), Scedosporium apiospermum (4), Scedosporium apiospermum (5), Scedosporium apiospermum (6), Lomentospora prolificans (7), Scedosporium minutisporum (8), Scedosporium apiospermum (9), and Scedosporium boydii (10). The species identity of each isolate was determined based on combined morphological/phenotypic characteristics and molecular sequencing of the internal transcribed spacer (ITS) ribosomal DNA (rDNA). Isolates were sub-cultured from frozen stocks on Sabouraud dextrose agar (SDA) prior to in vitro testing [A total of 16 strains were included in the study. Among them,  testing . The autAll tested agents, including ITR, VOR, POS, and MIN, were obtained from Selleck Chemicals  in powder form and dissolved according to the manufacturer’s protocol. The concentrations of tested agents were in the range from 0.5 to 64 mg/L for ITR, VOR and POS, or from 1 to 64 mg/L for MIN.Scedosporium and Lomentospora species was analysed using a microdilution checkerboard technique [6 CFU/ml with a hemocytometer and then diluted 100 times with RPMI1640 broth containing 0.165 M MOPS (pH 7.0) according to the M38-A2 reference standard. Serial dilutions of tested agents were prepared by dilution with RPMI 1640. For preparation of test microplates, 50 μl of each concentration of azoles  was added to columns 2 to 8, while 50 μl MIN was applied to rows 2 to 8. In each microplate, while row 1 and column 1 contained the azoles and MIN alone, respectively, column 9 was a drug-free well serving as the growth control. One hundred microlitre of fungal inoculum was applied to 96-well plates for a 72 h of incubation in ambient atmosphere at 35 °C, and the data were read visually. MIC values were defined as the lowest concentration of drugs at which the growth was completely inhibited. To assess the interaction outcomes, FICI value was calculated as follows:  + . The interaction of combined drugs was interpreted as FICI ≤0.5 for synergy, 0.5 < FICI< 4.0 for no interaction, or FICI≥4 for antagonism. All the tests were performed in triplicate.In vitro interaction between MIN and azoles against echnique . ConidiaG. mellonella larvae were divided into nine groups (N = 20 per group): control group, Uninfected Control-Saline Treatment group, Infected Control-No Treatment group, MIN group, ITR group, VOR group, POS group, combined ITR and MIN group, combined VOR and MIN group, and combined POS and MIN group. We used several inoculation concentrations for infection. Since lower or higher concentrations resulted in low or high mortality in the control group, we used the optimal concentration for the observation. All nine groups except the control group and Uninfected Control-Saline Treatment group were injected with S. apiospermum (CBS 116899). G. mellonella was pricked into the needle only, but not injected. The experimental group was injected with 10ul of S. apiospermum (CBS 116899) solution at a concentration of 1 × 107. S. apiospermum was injected into the last right proleg. Antifungal agents or a control solution  was introduced after the area was cleaned with an alcohol swab. The control group, MIN group, ITR group, VOR group, POS group, combined ITR and MIN group, combined VOR and MIN group, and combined POS and MIN group were injected with 10ul of saline, 10ul MIN (12μg/ml), 10ul ITR (3μg/ml), 10ul VOR (0.75μg/ml), 10ul POS (1.5μg/ml), 10ul of mixed solution of MIN (12μg/ml) and ITR (3μg/ml), 10ul of a mixture of MIN (12μg/ml) and VOR (0.75μg/ml), and 10ul of mixed solution of MIN (12μg/ml) and POS (1.5μg/ml), respectively. The larvae survival was recorded daily for 6 days. The survival curve of larvae was analysed by Kaplan-Meier method, and the difference was determined by Mantel Cox test. P < 0.05 was considered statistically significant.Larval tissues were collected, fixed in 10% neutral formalin and then dehydrated using an ethanol gradient. Thereafter, tissue samples were embedded in paraffin and xylene, sectioned at a thickness of 8 μm and stained with hematoxylin and eosin (HE). Finally, stained sections were evaluated with FSX100 fluorescence microscopy at magnifications of 10× and 40 × .


Although AC patients, when treated with IVT, had a significantly increased risk of 3-month mortality  than WC patients, stroke-related deaths did not differ between AC and WC patients . There were no significant differences between AC and WC patients, when treated with MT ± IVT, regarding 3-month dependency, 3-month mortality and SICH. Functional independence, mortality, and SICH were similar between RC and WC patients. In conclusion, recanalisation therapy might be used in AIS patients with nonmetastatic AC and with RC. Further studies are needed to explore the outcome of AIS patients with metastatic cancer undergoing recanalisation therapy.To date, very few studies focused their attention on efficacy and safety of recanalisation therapy in acute ischemic stroke (AIS) patients with cancer, reporting conflicting results. We retrospectively analysed data from our database of consecutive patients admitted to the Udine University Hospital with AIS that were treated with recanalisation therapy, i.e. intravenous thrombolysis (IVT), mechanical thrombectomy (MT), and bridging therapy, from January 2015 to December 2019. We compared 3-month dependency, 3-month mortality, and symptomatic intracranial haemorrhage (SICH) occurrence of patients with active cancer (AC) and remote cancer (RC) with that of patients without cancer (WC) undergoing recanalisation therapy for AIS. Patients were followed up for 3 months. Among the 613 AIS patients included in the study, 79 patients (12.9%) had either AC ( Atherosclerosis, nonbacterial thrombotic endocarditis, disseminated intravascular coagulation, tumor embolism, and cerebral venous thrombosis represent the most frequent causes of AIS in cancer patients6. In addition, concurrent cancer is able to impair short-term outcome in AIS patients7. On the basis of these considerations, it is fundamental to improve knowledge regarding recanalisation therapy of AIS patients affected by cancer. Recently, a retrospective analysis of a large nationwide sample of inpatient hospitalizations from 1998 to 2015 showed that cancer patients with AIS received intravenous thrombolysis (IVT) about two thirds as often as AIS patients without cancer. Differently, use of mechanical thrombectomy (MT) was similar between patients with and without cancer during the study period8.Patients affected by cancer have an increased risk of acute ischemic stroke (AIS) due to comorbid vascular disease and hypercoagulability16. In addition, it is unknown whether the outcome of AIS patients with remote cancer (RC) is more similar to that of patients with active cancer (AC) or without cancer (WC). Hypothesizing that AIS patients undergoing recanalisation therapy might have a different outcome on the basis of their cancer status, we decided to perform this study comparing AC and RC patients with those without malignancy.To date, very few studies focused their attention on efficacy and safety of recanalisation therapy in AIS patients with cancer, reporting conflicting results18. IVT was performed right after the native CT-scan if intracerebral haemorrhage (ICH) had been ruled out. Patients were considered eligible for MT if the following inclusion criteria were fulfilled: (1) presence of large vessel occlusion (LVO) in the anterior  or posterior  circulation as revealed by CT angiography (CTA); (2) symptoms onset within 6 h; (3) Alberta stroke program early CT score (ASPECTS) > 6 on direct CT scan19. Alteplase administration was continued until the angiographic suite would be ready and during the endovascular procedure. Patients presenting beyond the time window for IVT or with major contraindications to IVT underwent direct MT18. All patients treated with recanalisation therapy were re-scanned approximately 24 h after treatment, or sooner if there was clinical deterioration.We retrospectively analysed data from our database of consecutive patients admitted to the Udine University Hospital with AIS that were treated with recanalisation therapy, i.e. IVT, and MT ± IVT, from January 2015 to December 2019. Patients were followed up for 3 months. Patients showing symptoms onset within 4.5 h received IVT (0.9 mg/kg over 1 h) in accordance with international guidelines9, our patients were classified according to their cancer history into those with AC, RC, and WC. AC patients were those with any current or previous metastatic disease (except brain metastases), those undergoing current treatment for a malignancy or those who were offered treatment but refused it. Patients were also considered as affected by AC when initial diagnosis of malignancy was made during hospitalization after the onset of AIS. RC patients were those whose records indicated an inactive past history of malignancy with no history or evidence of metastatic disease, and who had completed any planned treatments. After the end of treatment, the period of 1 year was used for defining the absence of evidence of active cancer. Patients with end-stage malignancy, i.e. subjects with a life expectancy of less than 6-months or who received palliative care, and those with metastatic brain lesions are not treated with recanalisation therapy at our institution.Based on a previous study by Masrur et al.Comitato Etico Unico Regionale (Ref. No. CEUR-2020-Os-173). All experimental protocols were approved by the local ethics committee, Comitato Etico Unico Regionale. All research was performed in accordance with relevant guidelines/regulations.Written informed consent was obtained from all patients or their representatives. The study was approved by the local ethics committee, 19. Regarding vascular risk factors, we adopted the following definitions: (1) previous transient ischemic attack/stroke was defined if the patient had a history of ischemic (transient attack or stroke) or haemorrhagic cerebrovascular disease; (2) the presence of cardiovascular disease was based on the history of previous ischemic heart disease and/or revascularization treatment using percutaneous coronary intervention/coronary artery bypass grafting; (3) atrial fibrillation was defined if the patient had past medical history of atrial fibrillation that had been confirmed in medical records; (4) high blood pressure was defined as the history of hypertension and/or use of antihypertensive medication; (5) a history of diabetes mellitus that had been confirmed in medical records and/or use of insulin/oral hypoglycaemic agents were considered for defining diabetes; (6) a presence of hypercholesterolemia was based on the use of lipid-lowering medications; (7) information on active tobacco use was used for defining patient as a current smoker. For laboratory tests, venous blood samples were drawn at our Neurology Unit, within 24 h after hospitalization, during the morning hours (range 06.00–08:00) after an overnight fast (at least 12 h).The following variables were collected: age, sex, vascular risk factors, laboratory findings, admission systolic and diastolic blood pressure, admission heart rate and previous pharmacological treatment. In addition, early ischemic changes on native CT-scan within the middle cerebral artery were graded according to the ASPECTS20. Based on previous studies22, we classified our AC patients into those with ECOG score 0–1 versus those with score ≥ 2. Information on underlying causes of death was collected.Regarding malignancy, we collected information on: (1) type of cancer; (2) metastasis status; (3) modality of treatment; (4) impact of cancer on patient’s daily living abilities by means of the Eastern Cooperative Oncology Group (ECOG) performance score. The ECOG score ranges from zero (“fully active”) through three (”capable of only limited self-care”) to five (“dead”)23. The National Institute of Health Stroke Scale (NIHSS) score was adopted for determining stroke severity at admission and at discharge. Patients showing an improvement of ≥ 8 points on the NIHSS from baseline or a NIHSS score of 0 or 1 at discharge were considered as affected by major neurological improvement. Functional outcome was assessed by means of the modified Rankin Scale (mRS) at admission, based on pre-stroke disability, and 3 months after stroke. The mRS score after discharge was recorded at the patients’ routine clinical visit or through telephone interview with patients or their immediate caregivers. Patients with a mRS score comprised between 3 and 5 were considered functionally dependent. The presence of intracranial haemorrhage (ICH), as a consequence of the recanalisation therapy, was defined as any parenchymal hematoma (PH) based on the European Cooperative Acute Stroke Study (ECASS) morphologic definitions (ECASS PH-1 or PH-2)24, whereas presence of symptomatic intracranial haemorrhage (SICH) was based on the ECASS-III protocol25. Regarding recanalisation therapy, we collected information on: (1) time from symptom onset to IVT or MT; (2) time from hospital arrival to alteplase administration (door-to-needle time); (3) time from hospital arrival to groin puncture (door-to-groin time); (3) MT procedure duration; (4) recanalisation rate, assessed at the end of MT using the thrombolysis in cerebral infarction (TICI) classification and defined as successful recanalisation when a TICI 2b-3 was achieved.The Trial of ORG 10172 in Acute Stroke Treatment (TOAST) classification was used to determine AIS subtypes based on their etiologyThe following primary endpoints were analysed: (1) 3-month dependency; (2) 3-month all-cause mortality; (3) presence of SICH. Secondary endpoints were: (1) no major neurological improvement at discharge; (2) in-hospital all-cause mortality; (3) presence of ICH. All the outcome measures were collected as part of our routine clinical practice in patients affected by cerebrovascular events.Data are displayed in tables as mean and standard deviation, if not otherwise specified.Differences across the different groups were assessed by means of the Chi square test for categorial variables. One-way analysis of variance for normally distributed continuous variables, and the Kruskal–Wallis test for non-normally distributed continuous variables and for ordinal variables were used. Post-hoc analysis was performed by means of the Bonferroni test or the Dunn test, when appropriate. The Kolmogorov–Smirnov test with Lilliefors significant correction was used to assess normal distribution of data.Rates of primary and secondary endpoints were separately calculated for patients receiving IVT and for those undergoing MT ± IVT.The impact of cancer status on primary outcome measures was evaluated by multiple logistic regression analysis with WC patients as reference category both in patients receiving IVT and in those undergoing MT ± IVT. Age, baseline NIHSS score, and pre-stroke mRS were included in the model as potential confounding variables. Systolic blood pressure was added to other confounders in the analysis that evaluated the association between cancer status and SICH occurrence.All probability values are two-tailed. A p value < 0.05 was considered statistically significant. Statistical analysis was carried out using the SPSS Statistics, Version 22.0 .n = 46; 7.5%) or RC . These data are summarised in the flow diagram of the study , thus the remaining 613 AIS patients were included in the study. Of them, 440 (71.8%) were treated with IVT, and 173 with MT, i.e. 114 (18.6%) received MT combined with alteplase, and 59 (9.6%) undergoing direct MT. Seventy-nine patients (12.9%) had either AC . Causes of death, other than stroke, in AC patients were the following: sepsis (n = 3), and cancer progression (n = 3).Although 3-month mortality was significantly increased in AC patients, we observed that stroke-related deaths were similar between these patients and WC ones , functional dependency , and SICH  did not differ between patients with metastatic and non-metastatic AC.Whereas metastatic AC patients showed higher rates of 3-month mortality . Causes of death, other than stroke, in AC patients were the following: sepsis (n = 4), cancer progression (n = 1), and epigastric artery rupture (n = 1).. 33.3%, p = 0.177), 3-month mortality , and SICH  did not differ between patients with metastatic and non-metastatic AC.Prevalence of 3-month dependency  information on stroke severity, stroke subtype, 90-day functional outcome, cancer staging, and modality of cancer treatment was lacking; (2) the use of administrative data for research depends largely on its accuracy and reliability, thus coding errors were a possibility.To date, only few studies investigated the efficacy and safety of recanalisation therapy in AIS patients affected by cancer. In particular, data are very limited both for AC subjects and for RC ones. The largest investigations were performed by Murthy et al. and Weeda et al. who used the International Classification of Disease (ICD) codes for detecting cancer patients affected by AIS14. Except for two researches conducted in Asian patients undergoing MT15, the remaining literature agreed that, in AIS patients undergoing recanalisation therapy, functional independence was not affected by the presence of concurrent malignancy16.Prevalence of 3-month dependency did not differ between AC and WC patients, regardless of treatment modalities for AIS. A previous Italian study by Sallustio et al. reported similar results9. More recently, prevalence of 3-month mortality has been reported more than double in patients with active malignancy with respect to controls in the case–control study of Sallustio et al. (29.1% vs. 12.5%). However, the most common causes of death were cancer-related and not stroke-related14. The above reported data are perfectly in line with our results. In fact, rates of 3-month mortality were higher in AC patients than in WC ones. However, stroke-related deaths were quite uncommon in patients with active malignancy, occurring in 30% of patients receiving IVT and in 14.6% of patients undergoing MT ± IVT. This lower stroke-related mortality was certainly not attributable to an earlier recognition and treatment of stroke in AC because of pre-existing hospitalization, since all AC patients were not hospitalized at the time of their stroke. Confirming this statement, rates of in-hospital mortality, that is in strong relationship with stroke consequences, were not different between AC and WC patients, regardless of treatment modalities for AIS. Why AC patients treated with MT ± IVT for AIS had lower rates of stroke-related deaths than WC ones remains unknown. Further studies should investigate this specific topic.In 2011 Masrur et al. reviewed their database of AIS patients treated with urgent reperfusion treatment. Of 308 subjects included into the study, 18 and 26 were affected by active and remote malignancy, respectively. Patients with active malignancy had higher in-hospital mortality, but were more likely to die due to medical comorbidities instead of stroke-related causes27. Coagulopathy and consequences of radiotherapy or chemotherapy may increase the odds of haemorrhage transformation after recanalisation therapy in AIS patients with active malignancy28. Data on SICH prevalence in AC patients show an extremely wide range from 0 to 16.2%16. This is because SICH definitions were different among studies. Only Sallustio et al. used our SICH definition based on the ECASS-III protocol. Whereas these authors did not report any SICH in both AC patients and controls, we observed a prevalence of 12% in patients with concurrent malignancy who received IVT and 9.5% in those who underwent MT ± IVT14. Methodological differences between the two studies might explain the discrepancy between our data and those of Sallustio et al.14. In our experience SICH occurrence did not differ between the three groups of AIS patients undergoing MT ± IVT, differently SICH was significantly more common in our AC patients, when treated with IVT, than in the other two groups. This univariate association was not confirmed when confounders, i.e. age, baseline NIHSS score, pre-stroke mRS, and admission systolic blood pressure, were included in the multivariate model. NIHSS at admission represented the only independent predictor of SICH in both IVT and MT ± IVT patients. Thus, it seems that AC patients treated with IVT have more haemorrhages because they have more severe stroke. These findings agreed with previous studies that did not report any significant association between the presence of active malignancy and the occurrence of haemorrhagic transformation after recanalisation therapy16.The risk of SICH after IVT and MT varies from 2 to 7%. 3 months)29. However, our results on metastatic patients should be considered with caution; in fact, these subgroup analyses were conducted on a small sample size having low statistical power.A recent study by Yoo et al. compared 223 patients with nonactive cancer, 105 patients with active nonmetastatic cancer, and 140 patients with metastases. Since presence of metastases increased more than four times the risk of mortality, the authors suggested of using a careful approach when considering recanalisation therapy for AIS in patients with metastases. Similarly, we reported higher rates of 3-month mortality in patients with metastatic AC, but only when they were treated with IVT. In fact, we did not observe any significant difference regarding 3-month dependency, 3-month mortality, and SICH between patients with metastatic and non-metastatic AC when subjects underwent MT. These partial discrepancies might be due to the fact that, differently from our study, Yoo et al. enrolled patients not only treated, but also not treated with recanalisation therapy and used a longer follow-up period  This study has a significant selection bias since only patients undergoing recanalisation therapy for AIS were retrospectively included. We cannot exclude that some cancer patients were not assigned for recanalisation due to their advanced malignant disease and consequently they were not included in the study; (2) this is an observational study performed in a single centre with a limited sample size, which could have resulted in insufficient statistical power to detect true differences between groups; in fact, a power analysis was not conducted before data collection; (3) even after controlling for known confounders, residual confounding from unobserved factors might have affected our results; (4) due to the retrospective nature of the study, new or recurrent unrecognized malignancies might have occurred in RC patients causing a misdiagnosis (RC instead of AC); (5) since some of the 3-month mRS scores were indirectly recorded through telephone interview with patients or their caregivers, it might be possible that post-discharge functional status was not captured as well as in-hospital one; (6) finally, differently from patients treated with MT, for whom recanalisation outcomes are given (measured as TICI result), we were unable to report data on recanalisation rate among patients receiving IVT alone.In conclusion, recanalisation therapy might be used in AIS patients with cancer. In particular, IVT, and MT ± IVT, seem to be safe and effective methods of treatment in subjects with nonmetastatic, active cancer and with nonactive, remote cancer. Regarding metastatic patients, further studies with larger sample size are needed to explore the real outcome of AIS patients with metastatic cancer undergoing recanalisation therapy. Until then, an individualized approach considering life-expectancy is warranted in this subset of patients.


Robinia pseudoacacia and Amorpha fruticosa) were selected to study their responses to combined light and defoliation treatments. In a greenhouse experiment, light treatments  and defoliation treatments  were applied at the same time. The seedlings’ physiological and growth traits were determined at 1, 10, 30, and 70 days after the combined treatment. Our results showed that the effects of defoliation on growth and carbon allocation under high light treatments in both species were mainly concentrated in the early stage (days 1–10). R. pseudoacacia can achieve growth recovery within 10 days after defoliation, while A. fruticosa needs 30 days. Seedlings increased SLA and total chlorophyll concentration to improve light capture efficiency under low light treatments in both species, at the expense of reduced leaf thickness and leaf lignin concentration. The negative effects of defoliation treatment on plant growth and non-structural carbohydrates (NSCs) concentration in low light treatment were significantly higher than that in high light treatment after recovery for 70 days in R. pseudoacacia, suggesting sufficient production of carbohydrate would be crucial for seedling growth after defoliation. Plant growth was more sensitive to defoliation and low light stress than photosynthesis, resulting in NSCs accumulating during the early period of treatment. These results illustrated that although seedlings could adjust their resource allocation strategy and carbon dynamics in response to combined defoliation and light treatments, individuals grown in low light conditions will be more suppressed by defoliation. Our results indicate that we should pay more attention to understory seedlings’ regeneration under the pressure of herbivorous insects.Seedlings in regenerating layer are frequently attacked by herbivorous insects, while the combined effects of defoliation and shading are not fully understood. In the present study, two Leguminosae species ( The regeneration of woody plant seedlings is an important component of maintaining the vegetation diversity of forest ecosystem . By affeLight, one of the crucial factors for plant photosynthesis and survival, plays a key role in plant growth and development . As suchPrunus serotina, Cornus sanguinea, and Corylus avellana in low light condition but not in high light condition  and non-structural carbohydrates (NSCs). SCs include lignin, cellulose, semi-fiber, etc., which are involved in plant structure building processes . NSCs arRobinia pseudoacacia and Amorpha fruticosa. R. pseudoacacia is the dominant species in the arbor layer in warm temperate regions of Northern China . The area has a warm temperate monsoon climate, with an annual precipitation of 700 ± 100 mm and an average temperature of 13 ± 1°C. During the experimental period, the microclimate in the greenhouse was monitored with HOBO data loggers . Mean air temperature was 29.6°C (18.7–36.7°C) during daytime and 20.8°C (10.2–27.5°C) during nighttime, and mean relative humidity was 59.3% (28.2–97.8%) during daytime and 93.6% (56.3–100%) during nighttime. Seedlings were maintained at 60–70% of field water capacity throughout the experiment utilizing daily irrigation. The seeds were germinated in deionized water on plates and transferred into plastic pots (26 cm in depth and 24 cm in diameter) containing 7 kg of growth substrate (one seedling per pot). The plant growth substrate was a mixture of air-dried sandy loam and humus soils in proportions of 2:3 by volume.The experiment lasted about 5 months from April to September 2018 at the Fanggan Research Station of Shandong University, Shandong Province, China . The seeds of For each species, 3-month-old seedlings of similar size were selected and randomly assigned to the following treatments. For each species, a factorial experiment of two factors (light and defoliation) was designed. The seedlings were randomly assigned to two light conditions: (1) high light treatment (L+), grown in the greenhouse covered by plastic films ; (2) low light treatment (L−), conducted in greenhouse covered by woven black nylon nets . Under each light treatment, plants were divided into two groups. One group was submitted to top-down 50% defoliation, the upper half leaves were defoliated (DE); and the other was served as a control group without leaf removal (CK). The treatments were conducted from July 12 to September 20, lasting 70 days. There were 16 replicates in each treatment for each species. Four individuals of each species and treatment were randomly selected to measure in each sampling. During the experiment, seedlings were harvested at days 1 (July 13), 10 (July 22), 30 (August 11), and 70 (September 20).Seedling height and basal diameter  were recorded at each harvest. Four seedlings were harvested from each treatment at around noon (12:00–3:00 pm) and separated into roots (include root nodules), stems, and leaves. Then, the samples were oven-dried  and weighed. Total biomass (TB), leaf mass ratio (LMR), stem mass ratio (SMR), root mass ratio (RMR), and root-shoot ratio (R/S) were calculated as follows:where RB is root biomass, SB is stem biomass, and LB is leaf biomass.–1).Non-structural carbohydrates (NSCs) of four replicates per treatment were measured after each harvest. After biomass determination, dried samples were grounded with a ball mill  to analyze NSCs (defined as the sum of starch and soluble sugars) concentration in the leaves, stems, and roots . SolubleA), transpiration rate (E), intercellular carbon dioxide concentration (Ci), and stomatal conductance (Gs) of fully expanded leaves from four replicates (one leaf per seedling) for each measurement were measured using a portable gas exchange measurement system . These measurements were conducted between 9:00 and 12:00 h on sunny days. During the measurements, photosynthetically active radiation, temperature, relative humidity, and CO2 concentration inside the leaf chamber were controlled at 1000 μmol m–2 s–1, 28°C, 50%, and 400 ppm, respectively. The 1000 μmol m–2 s–1 PAR was high enough to obtain the maximum photosynthetic rate according to a pre-experiment. Since all the upper half leaves were defoliated, we measured the gas exchange parameters of mature leaves in the middle position of the individuals in all treatments at days 1 and 10. Due to the reflushing leaves were fully expanded at day 30 after defoliation, we measured the gas exchange parameters on newly mature leaves in all treatments at days 30 and 70. Instantaneous water use efficiency (iWUE) was calculated as follows:Gas-exchange characteristics of seedlings were measured before each harvest. The maximum photosynthetic rate (Specific leaf area (SLA) of four replicates per treatment was also measured after each harvest. Fully expanded and healthy composite leaves  from four seedlings in each treatment were scanned, and images were analyzed with the WinFOLIA Pro 2009a software  to determine leaf area. After scanning, these leaves were oven-dried for 72 h at 75°C and weighed. SLA was calculated as leaf area/leaf dry mass. The collection of leaf samples was the same as the gas exchange measurement.a concentration (Chl a), Chlorophyll b concentration (Chl b), the ratio of chlorophyll a/b (Chl a/b), total Chlorophyll concentration  were calculated as follows:Some other leaf morphological and physiological traits were also measured after 70 days of treatment. Four healthy and fully expanded leaves from the upper reflushing plant part (one leaf per seedling) were selected to determine leaf chlorophyll concentration in each treatment in both species. After extraction with 95% ethanol (v/v), chlorophyll concentration was determined using the spectrophotometric method . The abswhere V is ethanol volume, and FW is fresh weight.Leaf thickness was measured with an electronic digital micrometer for four replicates per treatment and was averaged from 20 fresh leaves per seedling. Leaf lignin concentration was estimated by a spectrophotometric procedure using acetyl bromide, and absorbance (A) was measured at 280 nm . The calculation of the lignin concentration was according to the standard curve .vegan package in R Statistical Software v.4.0.3  was applied to detect the effects of light treatment, defoliation treatment and sampling time for plant traits in each species. One-way ANOVA was used to detect difference among treatments for each species, and Duncan’s multiple comparison tests at α = 0.05 were followed when significant differences were observed. Before ANOVA, the data was checked for normality (Shapiro–Wilk test) and homogeneity of variance (Levene test). ANOVA and Spearman correlation was performed using IBM SPSS Statistics 23.0 , and the figures were illustrated using OriginPro 2016 . To evaluate the relationship between plant traits and experimental treatments, redundancy analysis (RDA) was carried out using the  v.4.0.3 .R. pseudoacacia , but not in the early stage (days 1–10) (For ys 1–10) . Before ys 1–10) . After tys 1–10) . After 7ys 1–10) .A. fruticosa, defoliation and light treatment significantly decreased seedling total NSC concentration at day 1  defoliation did not affect growth in eedlings . Howeverncreased , 3–5. Th0–66% defncreased .2, leaf construction and maintenance costs and the self-shading of the chloroplast , we found that stems soluble sugar concentration decreased significantly after defoliation, while the leaves soluble sugar concentration remained stable in at day 1 . After rat day 1 , indicatat day 1 . Meanwhit day 10 , 3, suggt day 10 . Previout day 10 . The car decline , which aA. fruticosa, but the growth parameters of L−CK treatment was significantly lower than control group. As an adaptation to increasing carbon limitation, plants can prioritize maintaining NSC concentration at the expense of growth, which could help plants survive long periods of minimal C gain (In the late phase of the experiment (days 30–70), growth and carbon allocation recovered rapidly after defoliation under high light conditions but not under low light conditions in both species. With the prolongation of the shading stress, the plant NSC concentration, height, basal diameter, total biomass, and maximum photosynthetic rate in the L− treatments was lower than that of L+ treatments in both species at day 30. Thus, shade stress can inhibit photosynthesis and cause insufficient carbon supply, resulting in carbon limitation, and limited plant growth , which wl C gain .Previous studies showed that herbivore damage could vary significantly between the contrasting light habitats (open vs. shade) on various species . Under hR. pseudoacacia seedlings can recover their total biomass in about 10 days after defoliation under high light condition, but A. fruticosa seedlings could catch up their total biomass with control within 30 days (R. pseudoacacia seedlings recovered faster than A. fruticosa seedlings after defoliation. Low light treatments significantly altered the growth and biomass allocation strategy of A. fruticosa. LMR in L− treatment was significantly higher than the L+ treatment at day 30, while RMR in L− treatment was significantly lower than the L+ treatment in A. fruticosa (It has been shown that  30 days . These rruticosa , which iruticosa .R. pseudoacacia seedlings recovered faster than A. fruticosa seedlings after defoliation under high light condition. Low light treatment could inhibit plant growth and carbon storage in both species. Growth after defoliation would be significantly suppressed under low light conditions, as the production of carbohydrate was not sufficient after 70 days in R. pseudoacacia. Plants can prioritize maintaining NSC concentration at the expense of growth under carbon limitation in A. fruticosa, which could help plants survive long periods of minimal C gain. Future researches should increase field experiments to have a more comprehensive understanding of the adaptation strategies of regenerating layer seedlings to complex habitats.Our results indicated that The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.NW conducted the experiment, wrote the body of the manuscript, and performed sample preparations, and laboratory and data analyses. PZ set up the experimental design and provided funding. TJ and KS performed the experiments. XL, QL, PW, and HS conducted analyses. ND, HW, and RW contributed in editing the manuscript. All authors made a contribution to this work.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Aspergillus pulmonary infection in the intensive care unit as shown by respiratory cultures, serum galactomannan and radiological images. Despite advanced intensive care and use of antifungal agents, both patients died eventually. This report illustrates the negative impact of secondary Aspergillosis and calls for the need of increased awareness of COVID19-associated pulmonary aspergillosis.Critically ill patients with coronavirus disease 2019 (COVID-19) are at risk of invasive pulmonary aspergillosis. The clinical manifestations of a superimposed fungal infection in those patients are difficult to underpin. This is compounded by the non-specific radiological findings associated with this infection and the challenges associated with performing bronchoscopy in COVID-19 patients. We would like to present two COVID-19 cases who developed secondary  A novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has led to a worldwide pandemic with devastating consequences in both developed and underdeveloped countries. COVID-19-related respiratory failure, requiring mechanical ventilation, has been associated with poor outcome . In this2) of 85 %, which improved to 92 % on 15 litres/minute (l min−1) oxygen via a non-rebreathing mask. The respiratory rate was 25 breaths/min, temperature of 38.8 °C, blood pressure (BP) of 113/62, and heart rate (HR) of 78/min. Initial chest x-ray revealed bilateral lung infiltrates. He was transferred to the ICU, where he was managed with high-flow nasal oxygen therapy (HFNO) and conscious proning. Lopinavir/ritonavir, hydroxychloroquine and piperacillin/tazobactam were started. Due to severe acute respiratory distress syndrome (ARDS) according to the Berlin definition [A previously healthy patient in their forties, was seen in the emergency room (day 0) with worsening respiratory symptoms, which started 4 days prior to admission. He was recently diagnosed as COVID-19 using real-time PCR (qPCR) . The patient was conscious and alert but hypoxic on room air with oxygen saturation . MIC results in µg ml−1 were as follows: amphotericin B: 0.032, itraconazole: 0.38, caspofungin: 0.004 and voriconazole: 0.023. Subsequently the patient was switched (day 32) to voriconazole 400 mg twice a day followed by 200 mg twice a day via a nasogastric tube due to impaired renal function. The patient continued to be fully dependent on ECMO with no signs of respiratory recovery up to day 34. Few days later (day 37), the patient developed a methicillin-resistant staphylococcus aureus blood-stream infection, which he received vancomycin for. On day 41, there was an improvement in lung compliance as well as CXR appearance, and gradual weaning of sedation and ECMO support was attempted. However, the patient developed septic shock and multiorgan failure due to Pseudomonas aeruginosa and Enterobacter aerogenes bacteremia. The patient's condition continued to deteriorate and he died on day 53 following his hospital admission.Microscopy revealed septate hyphae and accordingly, the patient was started on caspofungin 70 mg followed by 50 mg/day. The chest x-ray (CXR) showed worsening of lung infiltrates . The BAL2 of 86 % on 15 l min−1 oxygen via face mask. The patient was conscious, afebrile and haemodynamically stable. The CXR showed diffuse bilateral lung infiltrates more pronounced in the mid and lower zones. The patient was treated with piperacillin/tazobactam 4.5 g TID and levofloxacin 500 mg OD. . In view of his respiratory distress, the patient was transferred to the ICU, where he was connected to HFNO. The next day (day 1), hydroxychloroquine was added 400 BID, then 200 BID. The patient also received a single dose of tocilizumab 600 mg IV. On day 9, he started to improve from a respiratory point of view and was consequently able to move out of bed for the first time. However, 2 days later (day 14), he developed septic shock and hypoxia (SpO2 84 %) and was eventually intubated and mechanically ventilated. The CXR showed progression in the previously noted bilateral infiltrates. Blood cultures revealed Klebsiella pneumoniae, which was treated with meropenem 500 mg BID. In addition, anidulafungin (200 mg then 100 mg OD) and hydrocortisone (50 mg 4 hourly for 4 days) were added (day 16). In the following days, he was still requiring mechanical ventilation, but showed some improvement in his respiratory parameters. However, on day 23, the patient developed acute kidney injury requiring renal replacement therapy. On day 26, an endotracheal aspirate grew Aspergillus niger. Antifungal susceptibility testing done by E test  revealed the following MICs (µg ml−1): amphotericin B: 0.125, itraconazole: 0.032, caspofungin: 0.002 and voriconazole: 0.094. In addition, his CXR has revealed a large cavitating lesion in the right lung  was positive (6.53). Despite all supportive measures and ongoing antimicrobial therapy, the patient developed multiorgan failure and died eventually (day 31).A patient in their sixties, who suffered from diabetes mellitus, hypertension and ischaemic heart disease, was referred to a secondary hospital (day 0) with COVID-19 pneumonia. On physical examination, the patient was tachypnic with a respiratory rate of 28 breaths/min and SpOght lung . AccordiThe ongoing pandemic of SARS-CoV-2 continues to cause considerable morbidity and mortality. In Kuwait, at the time of writing, the total number of cases has reached 131 743 cases, with subsequent 811 deaths . This reUnlike influenza-associated aspergillosis, where the onset of pulmonary aspergillosis occurs early in the course of the disease (median 3 days from ICU admission), our patients developed CAPA much later (~day 25 post-ICU) . Other sAspergillus infection was confirmed by a positive culture from a BAL specimen in patient A, while patient B had a positive endotracheal aspirate culture accompanied by positive serum galactomannan test. Bronchoscopy can aid in the diagnosis of CAPA by assessment of the tracheobronchial tree for evidence of tracheobronchitis and for obtaining BAL samples for mycological analysis [Aspergillus PCR can also be used. In a recent review of 38 published CAPA cases, Aspergillus DNA was detected in serum or BAL specimens from 13 out of 19 patients [Aspergillus fumigatus, our patients were both infected with Aspergillus niger. The same species has also been reported in Pakistan [The microbiologic diagnosis of analysis . Howeveranalysis . On the analysis . Glucan analysis . Besidespatients . In contPakistan .In terms of radiologic findings, chest computerized tomography (CT) scan was not performed at the time of positive fungal culture, but serial CXR films for patient A have revealed worsening pulmonary infiltrates, while the CXR image of patient B has shown a cavitating lesion. Of note, the radiological findings of invasive pulmonary aspergillosis are usually non-specific among non-neutropenic patients .While both patients were not immune-compromised, they have received steroids before the onset of CAPA. Steroids have recently been shown to improve survival among COVID19 patients , but sucWith regard to anti-fungal treatment, both patients had received caspofungin initially, then shifted to voriconazole and liposomal amphotericin B in patients A and B, respectively. Ideally, the first-line therapy for invasive pulmonary aspergillosis should be intravenous voriconazole or isavuconazole. To ensure optimal therapeutic concentration, voriconazole serum trough level should be carefully monitored .In conclusion, this report attempts to increase the awareness of invasive pulmonary aspergillosis in COVID19 patients with ARDS. Appropriate and timely use of mycological tests, and better communication between different specialties such as intensivists, radiologists and microbiologists are strongly encouraged.


Neuroblastoma is a common extracranial malignant tumor in children. Its main treatment modality is a combination of chemotherapy, radiotherapy, and surgery. Given the advances in chemotherapy regimens and the widespread use of bone marrow transplantation over the decades, there has been improvement in treatment efficacy, which has led to prolonged patient survival. Accordingly, long-term complications have become a growing concern among physicians and patients. This study aimed to analyze the survival rate of patients with neuroblastoma and the risk factors for developing second malignant neoplasms (SMNs).The SEER 18 Regs (1973–2015) and SEER 9 Regs (1973–2015) data of the surveillance, epidemiology, and end results (SEER) database of the US National Cancer Institute were adopted for survival and SMN analysis.p < 0.001) was the only independent risk factor for developing SMNs.The 5-, 10-, and 20-year overall survival rates of patients with neuroblastoma were 67%, 65%, and 62%, respectively. Among 38 patients with neuroblastoma who presented with SMNs, those with abdomen as the primary site accounted for the majority (63.2%), followed by those with thorax (26.3%) and other sites (10.5%). SMNs occurred more commonly in non-specific neuroblastoma (incidence: 0.87%) than ganglioneuroblastoma (incidence: 0.3%). Compared with the general population, the risk of SMN is significantly higher (SIR = 4.36). The risk of developing SMNs was significantly higher in the digestive system (SIR = 7.29), bones and joints (SIR = 12.91), urinary system (SIR = 23.48), brain and other nervous systems (SIR = 5.70), and endocrine system (SIR = 5.84). Multivariate analysis revealed that the year of diagnosis (OR = 2.138, 95% CI = 1.634–2.797, This study identifies the risk factor for developing SMNs in patients with neuroblastoma, which could facilitate individualized screening for high-risk patients, to allow early diagnosis and treatment of SMNs. Neuroblastoma originates from the adrenal medulla or the paravertebral sympathetic nervous system. It is the most common extracranial solid tumor in children. According to the National Cancer Institute surveys in the United States, the incidence of neuroblastoma was 10.54/1000,000 (between 1975 and 2009). Neuroblastoma is a strongly heterogeneous tumor in children aged < 15 years. Some tumors can spontaneously disappear without treatment; however, most are insidious and have metastasized to the whole body at the time of diagnosis, with rapid progression ultimately leading to death. International multi-center collaboration of more than 30 years has facilitated an increase in the 5-year survival rate from 46% in 1974–1989 to 71% in 1999–2004. This is mainly attributed to the widespread adoption of risk-based stratification of therapy, including decreasing and increasing the treatment intensity for low-to intermediate-risk and high-risk patients [Information of patients with neuroblastoma (including other treatment information) from 1973 to 2015 were retrieved from the SEER database using the software SEER*Stat 8.3.6.1 The inclusion criteria were as follows: (1) patients with ICD-O-3 Hist/behave = neuroblastoma (ICD-O-3 = 9500) or ganglioneuroblastoma (ICD-O-3 = 9490); (2) patients with clear pathological classification and staging; (3) age of diagnosis less than 20 years, and (4) patients with complete follow-up information. The exclusion criteria were as follows: (1) patients diagnosed by autopsy; (2) primary site of the tumor is the central nervous system (CNS), and (3) patients with incomplete follow-up information. The survival time investigated in this study was the overall survival (OS) time. SMNs are defined as other malignant tumors appearing at least 2 months after neuroblastoma diagnosis. The standardized incidence ratios (SIRs) of SMNs were computed and analyzed using SEER*Stat 8.3.6.1; further, the corresponding 95% confidence level (CI) was reported. We collected information regarding age at diagnosis, gender, race, pathological type, degree of pathological differentiation, year of diagnosis, primary lesion site, whether radiotherapy was received, whether chemotherapy was received, survival time, and survival status. Authorization was obtained through the SEER website and data were retrieved from the SEER database, with no additional ethical approval being required.p ≤ 0.05. Specifically, the Kaplan–Meier method was used to estimate patient survival time. Univariate and multivariate regression analyses were performed using the Log-Rank test and Cox proportional hazards model, respectively. Independent risk factors for developing SMNs were analyzed using logistic regression. Independent risk factors for death were initially identified using univariate analysis; among them, those with p ≤ 0.1 were included in the multivariate analysis. The calculation of SIRs are calculated by SEER-stat software .Statistical analyses were performed using SPSS 22.0 . Statistical significance was set as This study enrolled 4338 patients, with 3667 patients pathologically-diagnosed with neuroblastoma and 671 with ganglioneuroblastoma. The abdomen (n = 3019) was the most common primary site, followed by the chest (n = 773). Males (52.9%) and females (47.1%) had roughly the same incidence rate. The incidence rates of patients aged < 1 year, 1–5 years, and 6–19 years were 34.4%, 54.1%, and 11.5%, respectively. Table p ≤ 0.1 were included in the multivariate analysis. Univariate analysis identified the following significant factors: age (p < 0.001), year of diagnosis (p < 0.001), histologic type (p < 0.001), primary site (p < 0.001), chemotherapy (p < 0.001), radiotherapy (p < 0.001), degree of pathological differentiation (p < 0.001), gender (p = 0.004), and race (p = 0.007). SMNs (p = 0.175) were not a risk factor for mortality. Factors with p ≤ 0.1 were included in a Cox proportional hazards model, which identified the following independent risk factors for patient OS: age , year of diagnosis , degree of pathological differentiation , primary site , chemotherapy .Radiotherapy (p = 0.08) is not a risk factor for mortality.The 5-, 10-, and 20-year OS rates of the entire neuroblastoma cohort were 67%, 65%, and 62%, respectively. Independent risk factors for death were initially identified using univariate analysis; among them, those with p < 0.05). Univariate analysis showed that the year of diagnosis , radiotherapy (p = 0.001), and chemotherapy (p = 0.059) were significantly correlated with the incidence of SMNs  was the only independent risk factors for developing SMNs. The result showed that patients with earlier year of diagnosis were more likely to develop SMN. The SIR of SMN gradually increased from 3.44 in the 1970s to 8.14 in the 2000s (p < 0.05). Radiotherapy (p = 0.109) and chemotherapy (p = 0.082) were not independent risk factors for developing SMNs.A total of 38 patients were diagnosed with SMNs. The SIR of all the patients was 4.36 , autologous transplantation, and MIBG treatment. Update of treatment concept and advancement of treatment technology have led to a significant increase in survival rates, which has allowed observation of long-term side effects represented by SMNs in long-term survivors.Our findings indicate that the year of diagnosis is an independent risk factor for the incidence of SMNs in patients with neuroblastoma. The result showed that patients with an earlier year of diagnosis are more likely to develop SMN, which may be associated with the extension follow-up time and evolution of treatment during the period. High-risk patients are recommended to receive higher-intensity radiotherapy and chemotherapy. Chemotherapy-related hematological SMN usually appeared within a few years after treatment. This has led to a significant increase in the risk of total SMN. However, due to the limitation in the number of cases in the SMN cohort, this view still needs more long-term follow-up and a larger sample size for verification.Our study shows that the SMN incidence rate of the entire cohort is 0.88%, which is similar to the incidence of several other neuroblastoma SMN cohorts , 4, whicp = 0.001), but multivariate analysis showed that radiotherapy was not a risk factor for SMNs (p = 0.109). 18 of the 38 patients in our SMN cohort had received radiotherapy (47.3%), but we were unable to obtain detailed information such as the field of radiotherapy and the site of the SMNs to assess whether SMN occurred in the radiation field. Another reason that may affect the statistical results is that the SMN cohort has a small sample size, which makes it impossible to calculate statistical differences. Some study showed radiotherapy may not play a role in the occurrence of SMN [Our study reveals that radiotherapy is not the independent risk factor for developing SMNs, although most studies have indicated the carcinogenic role of radiotherapy in neuroblastoma treatment , 7–12. Ce of SMN  mainly be of SMN .Our study indicate that chemotherapy is not an influencing factor of SMNs, which is inconsistent with other reports , 8, and TP53. LFS can cause various cancers, including breast cancer, brain tumors, sarcomas and other cancers. It usually has a clear family history, and most of it occurs at a young age. The probability of aggressive malignant tumors in high-risk family members before the age of 30 is as high as 50%. Osteosarcoma is a common SMN type after childhood tumor treatment, and is related to radiotherapy and chemotherapy but Ewing's sarcoma is not. A patient with Ewing's sarcoma was found in his SMN cohort, which is a rarer type of SMN pathology, meaning that genetic factors, rather than therapeutic factors such as radiotherapy and chemotherapy, may have caused the occurrence of this SMN. Another study found that multiple gene mutations exist in SMN patients with neuroblastoma [Additionally, patients with relapsed and refractory neuroblastoma are more likely to develop SMNs, which could be associated with genetic factors and higher-intensity chemoradiotherapy . Anotherblastoma , but dueThis study has several limitations. First, the treatment information in the SEER database is incomplete; moreover, there is missing key information, including disease stage, radiotherapy dose and genetic information, which impedes deeper analysis, although our study showed that age, year of diagnosis, degree of pathological differentiation, primary site, chemotherapy, and radiotherapy are independent risk factors for the prognosis of patients with neuroblastoma. We assumed that radiotherapy was performed in patients with high-risk disease, which may be a bias in interpreting the results. Neuroblastoma COG (Children's Oncology Group) risk stratification is mainly based on MYCN gene amplification, age, INSS  staging and other factors. Because the SEER database lacks MYCN gene amplification, INSS staging information, the current data cannot be used for risk stratification. Second, the SEER database does not represent the entire US patient population, which may affect the assessment of the actual incidence of SMNs in patients with neuroblastoma. Furthermore, the patient data included in the database cover a large time span with the risk stratification and treatment strategies of neuroblastoma having significantly changed several times since the 1970s. Therefore, the current stratification information and treatment regimens may not match those of past decades, which could introduce bias into the data. Nevertheless, this study revealed some characteristics regarding the incidence of SMNs in patients with neuroblastoma, which provide support for screening of long-term survivors, as well as early detection and intervention of SMNs.This study offers data on the SMNs of patients with neuroblastoma, indicating the risk factors for SMNs, which provide insight for neuroblastoma survivors and their physicians during follow-ups.


Melampsora larici-populina is a macrocyclic rust, and the haploid stage with two nuclei and the diploid of mononuclear sequentially occur annually. During the preservation of dry urediniospores at −80°C, we found that one isolate, ΔTs06, was different from the usual wild-type isolate Ts06 at −20°C because it has mixed polykaryotic urediniospores. However, the other spores, including the 0, I, III, and IV stages of a life cycle, were the same as Ts06. After five generations of successive inoculation and harvest of urediniospores from the compatible host Populus purdomii, the isolate ΔTs06 steadily maintained more than 20% multiple nucleus spores. To test the pathogenesis variation of ΔTs06, an assay of host poplars was applied to evaluate the differences between ΔTs06 and Ts06. After ΔTs06 and Ts06 inoculation, leaves of P. purdomii were used to detect the expression of small secreted proteins (SSPs) and fungal biomasses using quantitative real-time PCR (qRT-PCR) and trypan blue staining. ΔTs06 displayed stronger expression of five SSPs and had a shorter latent period, a higher density of uredinia, and higher DNA mass. A transcriptomic comparison between ΔTs06 and Ts06 revealed that 3,224 were differentially expressed genes (DEGs), 55 of which were related to reactive oxygen species metabolism, the Mitogen-activated protein kinase (MAPK) signaling pathway, and the meiosis pathway. Ten genes in the mitotic and meiotic pathways and another two genes associated with the “response to DNA damage stimulus” all had an upward expression, which were detected by qRT-PCR in ΔTs06 during cryopreservation. Gas chromatography–mass spectrometry (GC-MS) confirmed that the amounts of hexadecanoic acid and octadecadienoic acid were much more in ΔTs06 than in Ts06. In addition, using spectrophotometry, hydrogen peroxide (H2O2) was also present in greater quantities in ΔTs06 compared with those found in Ts06. Increased fatty acids metabolism could prevent damage to urediniospores in super-low temperatures, but oxidant species that involved H2O2 may destroy tube proteins of mitosis and meiosis, which could cause abnormal nuclear division and lead to multinucleation, which has a different genotype. Therefore, the multinuclear isolate is different from the wild-type isolate in terms of phenotype and genotype; this multinucleation phenomenon in urediniospores improves the pathogenesis and environmental fitness of M. larici-populina. Melampsora larici-populina Kleb., an obligate biotrophic parasite, has caused heavy rust infections with disastrous consequences for the poplar industry  Nucleus mutation: factors for nuclear mutation include ion radiation, chemical action, temperature changes, and pH changes  germ tube fusion or hyphal fusion. Since 1989, fusant formation and fusion phenomena of germ tubes, appressoria, substomatal cavities, or hyphae have been observed in a variety of M. larici-populina are homologous to avirulence genes of M. lini. For example, Mlp124272 and Mlp124256 are homologous to AvrP4; Mlp37347 is homologous to AvrL567; Mlp124530 is homologous to AvrP123; and Mlp123932 is homologous to haustorially expressed secreted protein (HESP) HESP-327. Furthermore, specific labeling of Mlp37347 and Mlp123932 was detected at the periphery of haustoria. Mlp124272, Mlp124256, and Mlp124530 showed very strong labeling in the development of uredinia  after purification and separation. The mutant strain, ΔTs06, was significantly different from Ts06 in host selection, pathogenicity, and other biological characteristics. Therefore, to investigate the genetic stability of mutated strains and the mechanism of multinucleation occurrence, we analyzed the stability of the life cycle of ΔTs06 using artificial inoculation on the compatible host P. purdomii for consecutive subcultures. The phenotype responses on different hosts and the expression of SSPs under real-time fluorescence quantitative analysis were used to determine the differences in pathogenicity between ΔTs06 and Ts06. The molecular mechanism of urediniospore multinucleation was explored using transcriptomics combined with real-time fluorescence quantitative PCR on nuclear division-related genes. The research results are of crucial theoretical significance in understanding the life history, pathogenicity variation, mechanism and environmental adaptability of such obligate parasites.Rust fungi are obligate parasites; hence, artificial culture technology is difficult to implement in biological research, including sporulation, mating, and subcellular structure under artificial synthesis media. Therefore, their spore propagulum is usually kept for research in dry conditions at −20°C for 1 year or at −80°C for more than 1 year. However, we found that multinucleation occurred in urediniospores of strain Ts06 (ΔTs06) at ultralow temperature (−80°C) and unmutated dikaryon strain Ts06. P. purdomii is the natural host of M. larici-populina collected in Qinling Mt., China. P. purdomii, the other four hosts (Larix principis-rupprechtii Mayr.) were planted in greenhouse conditions at 25°C, 70%−80% relative humidity (RH), and 16 h photoperiod  for in planta experiments. They are healthy 1-year-old seedlings.Mutant strain Tsur hosts , and the06 were prepared in spore suspensions at a concentration of 0.01 g/ml and were smeared on a 2% water agar medium for germination and on abaxial leaves of P. purdomii for inoculation as described in Zheng et al.  were transferred to a chamber to induce teliospores at 4°C, and basidiospores developed according to the protocol outlined in Yu et al.  and were dated under fluorescence microscope . Urediniospores of ΔTsg et al. . Histolog et al. . Urediniu et al. . Thereafu et al. . The basP. purdomii, P. deltoides, P. szechuanica, P. × euramericana cv. Polska 15A, and P. × euramericana cv. Robusta were selected for pathogenesis tests , uredinia density (number per cm2), uredinia diameter (mm), and uredinia height (mm) were recorded after inoculation. To measure density, a 1-cm2 piece of paper was placed on the back of the leaves. The number of uredinia in at least 5-cm2 of paper per leaf was randomly counted to calculate the average density value. The uredinium diameter and height were observed and measured using an anatomical lens. All analyses were performed with SPSS 22.0 . The means of the densities were determined using Duncan's multiple range test at p <0.05. Simultaneously, leaves of P. purdomii were collected using a kit  at 0, 12, 24, 48, 96, and 168 h post inoculation (hpi) to obtain qualified RNA. The iScript™ cDNA kit was used for cDNA synthesis . The SsoFast™ EvaGreen® Supermix kit was used for qPCR  with SSP primers, as shown in aTUB) and elongation factor-1-alpha (ELF1a) of M. larici-populina were selected as reference genes, and amplification reaction conditions were the same as in Ye et al. . Clean reads were compared with the genome sequence of M. larici-populina  using hierarchical indexing for spliced alignment of transcripts (HISAT) software , and the relationship between interacting genes was obtained using homology and known proteins.TsaTUB) and elongation factor-1-alpha (ELF1a) of M. larici-populina were used as reference genes . In addition, 0.1 g of urediniospores from Two80 and 0.1 g from CK80 were selected to detect fatty acids using gas chromatography–mass spectrometry   of 0.05 g of urediniospores were used to detect concentrations of H06 and Ts06 urediniospores. Random amplified microsatellites (RAMs) were applied to screen for differences in whole genomic DNA between ΔTs06 and Ts06. The RAM primers were designed   were significantly higher in ΔTs06 than in Ts06, and ΔTs06 produced more intercellular hyphae than Ts06. At the 168th h, the expression levels of three proteins  were significantly higher in ΔTs06 than in Ts06, and ΔTs06 produced more urediniospores than Ts06.Except for the incompatible host ur hosts . No signe <0.05) , and thepurdomii . These rat 7 dpi . During n of SSP . HoweverAn average of 42.5 M of clean reads per sample were collected . A totalThe GO analysis of these 55 genes revealed that the enriched items primarily included “drug metabolism” (GO: 0017144), “reactive oxygen species metabolism” (GO: 0072593), “antibiotic metabolism” (GO: 0016999), and “response to oxidative stress” (GO: 0006979) . AccordiTen genes in the mitotic and meiosis pathways and two genes of “responses to DNA damage stimulus” were selected for qRT-PCR verification . The exp2O2 in Two20 and Two80 were significantly higher than those in One20 and One80, while the concentrations of H2O2 in One20 and One80 were significantly higher than those in CK20 and CK80  were found in greater quantities in ΔTs06 than in Ts06  were upregulated, suggesting that these genes responded to environmental stimuli. Except for these genes, expressions of the protein kinase activity gene MELLADRAFT_102486 (PKA) and the nucleic acid metabolism gene MELLADRAFT_61177  also increased, and the downstream genes related to mitosis and S-phase meiotic division were both activated. For example, MELLADRAFT_36075 (Dmc1), MELLADRAFT_104525 (Spo11), BGI-novel_G000167 (ORC), and MELLADRAFT_115157 (MCM) exhibited upward trends of expression at −80°C as cryopreservation time increased. Abnormal mitosis and abnormal S-phase progression of urediniospore nuclei in ΔTs06 were believed to cause polynuclear phenomena in this study.Nuclear mutation is a main way to form multinucleation phenomena  are important factors in eukaryotic signaling networks  and “trehalose biosynthesis process” (MELLADRAFT_60322) were significantly upregulated . TrehaloM. lini  and C. quercuum are highly similar. Zemodeme II originates from zemodeme I after karyogamy, or the latter is a haploid from the former .The datasets generated for this study can be found in the NCBI Sequence Read Archive (PRJNA673784, ZY and ZC conceived and planned the study. WZ performed the field investigation and characterization experiments and analyzed the data. ZY and WZ contributed key ideas, analyzed the data, and wrote the manuscript. ZP and SP assisted in the data analysis. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Recurrence of pelvic organ prolapse (POP) after transvaginal mesh (TVM) implantation pelvic floor reconstruction surgery remains an unresolved problem in clinical practice. In this retrospective observational study, clinical and pelvic floor ultrasound (PFUS) parameters were analyzed in order to identify high-risk factors of POP recurrence.The clinical and PFUS data from September 2013 to November 2019 of patients who underwent TVM were retrospectively analyzed. The patients with prolapse recurrence on postoperative follow-up diagnosed by PFUS were selected as case group, the clinical and PFUS parameters of them were compared with the control group in which the patients had no sign of prolapse recurrence. Univariate and multivariate regression analyses were performed based on age, BMI, gravidity, parity, surgical history (non-POP hysterectomy and incontinence-or-POP surgery), preoperative POP stage, follow-up in years, levator avulsion and hiatal area (HA) on Valsalva.P < 0.001). The area under the ROC curve was 0.775 .Altogether 102 patients entered the study and the median interval between PFUS and TVM surgery was 2.5 years. Univariate analysis showed that levator avulsion and HA were significantly different between case group and control; multivariate regression analysis showed that only HA was related to prolapse recurrence after TVM (OR = 1.202, 95% CI 1.100–1.313, Hiatal area on Valsalva was related to prolapse recurrence after TVM surgery and it is an important parameter for postoperative follow-up of TVM surgery. Transvaginal mesh (TVM) pelvic floor reconstruction is an effective treatment method for moderate to severe pelvic organ prolapse (POP) in middle-aged and elder women . CompareDiagnostic methods for POP prolapse recurrence include clinical examination, questionnaires, and imaging , 6. The In this study, PFUS was used in Chinese patients who underwent TVM pelvic floor reconstruction surgery to diagnose prolapse recurrence. The mesh position and movement during Valsalva maneuver was recorded. The support failure types of anterior compartment mesh were reported. Clinical and PFUS parameters were compared between case and control group to find high-risk factors for POP recurrence.This study was approved by the Ethics Committee of Peking Union Medical College Hospital of Chinese Academy of Medical Sciences.This was a cross-sectional study. The clinical and PFUS volume data of patients who underwent TVM pelvic floor reconstruction surgery and were followed up with PFUS from September 2013 to November 2019 in our hospital were retrospectively analyzed. Our hospital is one of the largest diagnosis and treatment centers for pelvic floor disorders in China. All TVM surgeries were performed by the same doctors team. Inclusion criteria were the following: patients who underwent TVM surgery because of POP in our hospital; the patient had clinical review data as well as PFUS data; the interval between PFUS and surgery was at least 10 months. All patients were grouped into case and control group according to post operation PFUS. The patients with prolapse recurrence in any compartment diagnosed by PFUS were grouped to case group. The patients without any of the above abnormality constituted the control group. The general data and clinical and ultrasound parameters between the case group and the control group were compared in order to identify the risk factors for POP recurrence after TVM.The patients’ inpatient medical records were collected, including patient age, gravidity, parity, history of gynecologically-related surgery , preoperative prolapse ICS POP-Q stage , and date of surgery. The body mass index (BMI) of the patients at the date of PFUS examination were recorded. Follow-up in years was calculated as the interval between surgery and first ultrasound showing prolapse recurrence (cases) and the interval between surgery and last scan (controls).GE Voluson E8 or E10 diagnostic ultrasound machine (GE Company) with transabdominal 4D probes were used. The PFUS of all patients were finished by one doctor (L. Tan). With bladder lithotomy position, the patients were asked to perform the maximum Valsalva maneuver . The midsagittal view was taken to observe the descent of pelvic structures including the bladder, vault of vagina, bowels, and rectum. A horizontal reference line (H line) that passed the posteroinferior margin of the symphysis pubis was used as the reference line. In addition, three-dimensional volumetric imaging was performed to observe the integrity of the levator ani in the TUI model during maximal pelvic floor contraction; the HA was measured during the maximum Valsalva maneuver. The mesh-anchor failure type of cystocele cases was reviewed by two doctors (Z. Liu and L. Chen), and were classified into anterior failure, apical failure and global failure according to reference .The diagnostic criteria for significant ultrasound prolapse were : cystoceP < 0.10 were entered into Logistic regression analysis. ROC curve was plotted and the area under the curve (AUC) was calculated. P < 0.05 (two-tailed) was considered to be statistically significant.SPSS 26.0 software was used; measurement data conforming to normal distribution were expressed as mean ± standard deviation, and those not conforming to normal distribution were expressed as range, median, 25% and 75% quartiles. Age, BMI, gravidity, parity, surgical history, preoperative prolapse ICS POP-Q stage, follow-up in years, levator avulsion, and HA of case and control groups were compared using t-test , U test  and chi-square test . Afterward, variables with Altogether 102 patients entered our analysis. The patients mean age was 65 years (53–77). Mean parity was 2 (0–5). Mean BMI when they came for review was 24.49 (18.82–30.08). 10 of them had a history of non-POP hysterectomy; 15 had surgery history of POP or incontinence. Concomitant hysterectomy was performed in all of the remaining patients. 10 patients received concomitant incontinence surgery . The median interval between PFUS and TVM surgery was 2.5 years (0.8–10.8 years).On PFUS, cystocele was diagnosed in 37 patients, among whom 6 were in combination with enterocele or rectocele; Enterocele, rectocele, or both but without cystocele were diagnosed in 17 patients. Altogether 54 patients composed the case group. Anterior vaginal wall meshes could in all cases be observed, and mesh movement was recorded on Valsalva Maneuver  and HA  were significantly different between the two groups.The distribution and comparison of general data, clinical characteristics, and ultrasound features of patients in the case group, control group and total patients were shown in Table P = 0.001, < 0.001 and 0.087, respectively). The analysis results showed that only HA was related to recurrence of POP after TVM, OR = 1.202, 95% CI 1.100–1.313 (P < 0.001). The ROC curve for HA in prolapse recurrence diagnosis was shown in Fig. CI 0.684–0.867, P < 0.001).Levator avulsion, HA and POP or incontinence surgery history entered into Logistic regression analysis (CI 1.100–1.313), which means that for every 1 cm2 increase in HA, the probability of recurrence of prolapse after TVM increased by 20%. The prediction of HA for prolapse recurrence showed an AUC of 0.775 (95% CI 0.684–0.867).In this cross-sectional study, we analyzed a number of clinical and ultrasound parameters to find high-risk factor(s) of prolapse recurrence after TVM surgery. The parameters included age, BMI at the time of clinical review, gravidity, parity, previous surgical history, preoperative prolapse stage, follow-up in years, levator avulsion, and HA. The prolapse recurrence in our study was diagnosed by ultrasound as cystocele, enterocele or rectocele (at least one of the three). Altogether 54 patients of case group and 48 patients of control group entered our study. The multivariate regression analysis showed that only HA was related to prolapse recurrence with an OR 1.202  + Pb ” on the ICS POP-Q if also measured on Valsalva may act as a surrogate for HA. Previous studies have shown the correlation between value of (Gh + Pb) in POP-Q scoring and HA in PFUS , 18. AndIn this retrospective study, hiatus area under Valsalva was related to prolapse recurrence after TVM surgery at a median follow-up length of 2.5 years. Ultrasound could help clinicians focus on patients who are at high risk of prolapse recurrence. Clinical physicians should pay more attention to patients with Hiatus area enlargement and actively take interventions in order to achieve better prognosis of these patients.


Ectopic pregnancy is one of the leading causes of pregnancy-related mortality; the treatment strategies associated with this condition entail complications, such as recurrence of ectopic pregnancy or infertility. The objective of this study was to evaluate the recurrence and fertility rate after salpingostomy in patients with tubal ectopic pregnancy.This cross-sectional retrospective study was conducted at four referral centers of Obstetrics and Gynecology, under the supervision of Shiraz University of Medical Sciences (Iran). The medical records of 125 patients with tubal pregnancy were reviewed. These patients underwent laparoscopic salpingostomy from April 2009 to March 2016.Data on maternal age, BMI, history of previous EP, genital tract infection, IUD insertion, history of previous surgery, and infertility were further obtained. The patients were followed up for approximately 1 to 7 years. The recurrence of EP and subsequent pregnancy rate were assessed during the follow-up period.P = .425); however, the post-salpingostomy recurrence of EP was correlated with BMI (P = 0.001), previous history of EP (P = 0.001), genital tract infection (P = 0.001), and IUD insertion (P = 003). Among 95 women who had no contraception, pregnancy occurred in 51 cases (53.6%) and recurrence of EP was observed in 16 patients (12.8%).There was no statistically significant relationship between post-salpingostomy recurrence and maternal age, previous abdominopelvic surgery, and history of infertility(Our results suggest that salpingostomy is a safe method with a low risk of recurrence and good fertility outcomes for women who consider future pregnancy. In ectopic pregnancy (EP), a fertilized ovum is implanted outside the uterine cavity . EP occuThe exact etiology of EP is unclear, but it may be caused by impaired tubal transport . Other rAlthough it may be asymptomatic, EP frequently presents with pain and vaginal bleeding in the first trimester of pregnancy , 10; howβ-human chorionic gonadotropin (β-hCG) until its return to the normal level), medical treatment (methotrexate), and surgery [Treatment of EP has different strategies, such as expectant management  and conservative technique  by either laparotomy or laparoscopy are the two most common surgical approaches . LaparosThis cross-sectional retrospective study was conducted at four referral centers of Obstetrics and Gynecology under the supervision of Shiraz University of Medical Sciences (Iran) . The medical records of all patients with tubal pregnancy who underwent laparoscopic salpingostomy from April 2009 to March 2016 were reviewed. Demographic data such as maternal age, BMI, history of the previous EP, genital tract infection, IUD insertion, history of previous surgery, and infertility were extracted from the patient’s available records.Indications for salpingostomy included hemodynamic stability, unruptured tube, contraindications to methotrexate or failed medical treatment. During surgery, the appearance of contralateral tube was checked.The exclusion criteria were 1) patients with non-tubal ectopic pregnancy, 2) those treated with salpingectomy or other conservative surgical management (such as laparoscopic milking) or treated by methotrexate, and 3) subjects who were unwilling to participate, refused to answer the question, or had incomplete data in the hospital chart.After surgery, patients were followed by serial serum B-HCG level until it became undetectable. The patients were followed up for approximately 1 to 7 years, during which period the recurrence of EP and the subsequent pregnancy rate were assessed .Women reporting no desire of subsequent pregnancy after the salpingostomy, and who, therefore, did not actively try to conceive or were using a contraceptive method were excluded. Also, patients who had received medical treatment with methotrexate after surgery, patients with male factor infertility, those who had hospitalization due to PID (pelvic inflammatory disease) after surgery, and patients that their age become more than 45 years in the follow-up period were excluded. These strict exclusion criteria were considered to reduce the likelihood of bias.Patients who actively tried to conceive after the salpingostomy and who accepted to be involved in the study were included.Finally, 125 patients were assessed in this study, the demographic data such as BMI, reassessed in the follow up period, which had no significant changes.Data regarding the past medical history and demographics were extracted from hospital paper records to decrease recall bias. Also, inquired future outcomes (the recurrence of EP and the subsequent pregnancy) were rechecked with hospital records to decrease the chance of bias.P value < 0.05 was considered as statistically significant.The study protocol was approved by the institutional Ethics Committee (code: IR.SUMS.MED.REC.1395.s72). Data were analyzed using SPSS version 19. Pearson correlation was used to determine the correlations and An overall of 3434 patients who were diagnosed with EP in our centers from 2009 to 2016 were enrolled in the present study. Among them, 2244 (65.34%) patients were treated with non-surgical methods and 1190 patients had surgery. Salpingectomy and salpingostomy were carried out for 1025 (29.85%) and 165 (4.8%) cases, respectively.Finally, 165 patients were selected, 40 of whom were excluded because they were not willing to answer the questions, gave the wrong phone numbers, or did not answer their phones.2 (maximum = 37.1 and minimum = 15.87). The mean B-HCG level was 2706 ± 3.39 mIU/ml  and the mean size of the ectopic mass was 8.8 ± 6.8 cm2 (1-35) at the time of admission.Among the 125 patients, the mean age was 26.496 ± 5.3 years (maximum = 40 and minimum = 17) and the mean BMI was 24.46 ± 3.6 kg/mDuring laparoscopy, the appearance of bilateral tubes was checked, and salpingostomy was done as a treatment for unruptured ectopic pregnancies.66 (528%) cases developed ectopic pregnancy in the right adnexa and 59 (47.2%) in the left adnexa Table . In all The complication of salpingostomy in persistent trophoblastic disease was detected in 5-15%of cases in some studies (5). In our study, the patients were followed up based on B-HCG level, none presented with persistent diseases, and there was no need for treatments, such as methotrexate or second surgery.Also, no surgical complications were reported after salpingostomy in our patients.The evaluated risk factors for postoperative recurrence were maternal age, BMI, history of the previous EP, genital tract infection, IUD consumption , history of previous surgery, and infertility. History of infertility was observed in 32% of the patients; furthermore, previous pelvic surgery, genital tract infection, previous EP, and IUD insertion were found in 28, 18.4, 16.8, and 10.4% of the cases, respectively. Those who had a previous history of EP were treated with non-surgical methods (25% with methotrexate and 75% without medication) only with expectant management.P = .425); however, as seen in Table P = 0.001), previous history of EP (P = 0.001), genital tract infection (P = 0.001), and IUD insertion (P = 0.03).There was no statistically significant correlation between maternal age and post-salpingostomy recurrence P = .45; howeveMultiple logistic regression test was done, and the results showed that for every unit of BMI increase, the chance of recurrence rose by 20%.The chances of recurrence in patients with a positive history of EP were 9.75 times higher than those with no such history (odds ratio = 9.75).No statistical correlation was found between recurrence of EP post-salpingostomy and previous abdominopelvic surgery (sig = 0.757) or history of infertility (sig = 0.945). Among the 95 women who had no contraception, pregnancy occurred in 51 cases 53.6%) and EP recurrence was seen in 16 cases (12.8%), 0.33% at the site of salpingostomy and0.67% in contralateral tube. All the patients with recurrence (16 people) had at least one risk factor in their life (Table .6% and EChlamydia trachomatis or Neisseria gonorrhea) and tubal surgery  [Among the several well-known factors that augment the risk of ectopic pregnancy, mention can be made of pelvic inflammatory disease (hesions) . Althoughesions) . In the Bangsgaard observed that a history of the previous EP increased the risk of future EP up to 10–25% . ChandraP value = 0.003) and EP recurrence following salpingostomy.Oron et al. showed that the risk of EP using norgestrel-containing IUD was lower than that with a copper IUD. This might be attributed to the reduced overall pregnancy rate in women using norgestrel-containing IUDs . Based oP value = 0.001); this indicates that more studies should be conducted concerning this topic.The risk of EP increased with the rise in maternal age over 35 years. This association may be ascribed to most other risk factors related to an increase in age . HistoryLanger et al. reported that the EP recurrence rate was 7% if the contralateral tube was normal and 18-25% if the opposite tube was abnormal or absent . In a prAgdi and Tulandi reviewed the literature on different treatments for tubal pregnancy. Among patients who attempted to conceive, the subsequent intrauterine pregnancy (IUP) rate was 61.4% after salpingostomy, 38.1% following partial or total salpingectomy, and 54% after treatment with methotrexate. The recurrent ectopic pregnancy rates after salpingostomy, salpingectomy, and methotrexate treatment were 15.4, 9.8, and 8%, respectively . Ding etOur study showed that among the women who attempted to conceive (95 cases), 53.68% had IUP after salpingostomy, which seems to be a good result in comparison with other studies. Finally, the recurrence of EP after salpangostomy in our study was estimated at about 12.8%, which is similar to previous studies.Our results suggest that salpingostomy is a safe method with a low recurrence risk and good fertility outcomes for women who have a desire for future pregnancies.