First Report of stem rot in Cymbidium sinense Caused by Fusarium oxysporum (FOSC) in China.

Cymbidium sinense (Jackson ex Andr.) Willd is a perennial terrestrial plant in the orchid family mainly distributed in China, Japan, India and Southeast Asia that occupies a strong position in the flower market due to its bright green leaves and fragrant flowers (Zhang et al. 2013). Cymbidium sinense is not only valued by people for its ornamental and economic value, but its roots have antiasthmatic medicinal properties (Ke et al. 2004). In August 2020, about 15% stem rot on two-year old C. sinense with varying severity was observed in five nursery gardens located in Enshi city (N 30° 16', E 109° 29'), Hubei province, China. Typical symptoms of C. sinense included roots and inner part of the pseudobulbs changing from white to brown and rotting. Leaves became brown and withered from bottom to top, and there was an obvious blight yellow halo at the junction of diseased and healthy tissue, which eventually caused the whole plant to wilt and die (Fig. 1d). To isolate the pathogen, a total of 15 leaf tissues from the disease-health junction (3 × 3 mm) from 5 individual plants (3 leaves/plant) with symptoms were surface sterilized with 75% ethanol for 30 s and 2% sodium hypochlorite (NaOCl) for 3 min. The sterilized tissue was rinsed three times with sterilized water, and then placed on potato dextrose agar (PDA) for incubation at 28°C in the dark for 5 days. Isolated colonies were subcultured by a hyphal tip protocol. Thirteen fungal isolates were obtained. Through preliminary pathogenicity tests, we found that ten isolates induced leaf blight. These ten isolates with pathogenicity showed similar morphological characteristics, with initial white-flocculent aerial mycelium that secreted a lavender pigment and produced colonies with an irregular edge after 3 days on PDA. The ten strains were cultured on PDA plates at 28℃ for 5 and 15 days to observe colony and conidial characteristics. The ten strains were identified as Fusarium based on morphological characteristics (Leslie and Summerell 2006). Strain ML0303 was selected for further identification. Macroconidia were falciform, hyaline, slightly pointed at both ends with two to four septa, 24.0 ± 5.6 µm × 4.7 ± 0.8 µm (n = 50). Microconidia were hyaline, oval, globose, with zero to one septum, 5.5 ± 1.3 µm × 2.2 ± 0.5 µm (n = 50) (Fig. 1c). Total genomic DNA of strain ML0303 was extracted with a CTAB protocol (Stenglein and Balatti 2006). The translation elongation factor (EF-1α), RNA polymerase II second largest subunit (RPB2) and ß-tubulin (Tub2) genes were amplified respectively using primer pairs EF1/EF2, RPB2-5F2/RPB2-7cR and T1/T22 respectively (O'Donnell. et al. 2010, O'Donnell. et al. 1997). The EF-1α, RPB2 and Tub2 (accession numbers-MW719874, OL614838, OL689398, respectively) gene sequences were submitted to GenBank. EF-1α, RPB2 and Tub2 sequences of ML0303 showed 99.5% - 100% identity respectively with Fusarium oxysporum in the Genbank and FUSARIUM-ID databases. The multilocus sequence data was used to infer a phylogenetic tree via a Neighbor-joining (NJ), Maximum-likelihood (ML) and Maximum-Parsimony(MP) together with reference sequences from GenBank. The topology of the three trees was similar; only the NJ tree is presented here. Strain ML0303 and F. oxysporum formed a clade supported with high values (NJ/ML/MP: 96,95,97). The results indicated that the fungus was F. oxysporum based on the phylogenetic analysis and BLASTn queries. For pathogenicity tests, conidia of strain ML0303 were collected by rinsing PDA plates. Two-year-old C. sinense grown in plastic pots filled with sterilized autoclaved sandy loam soil were used for the tests. Three pots (two plants/pot) were included in each treatment. Spore suspensions (106spores/ml) of strain ML0303 were used to irrigate the stem-zone of the plants, and sterile water was used as control. The two treatments were placed in a greenhouse and incubated at 28±2℃ with a 14-hour light/10-hour dark cycle. The experiment was repeated twice. After three weeks, stem rot symptoms were observed on C. sinense inoculated with ML0303, that were the as same as observed in the nursery (Fig. 1e-h). No symptoms were observed on the negative control. Fusarium oxysporum was re-isolated from the infected plants to fulfill Koch's postulates. Partial EF-1α and RPB2 gene sequences were used for molecular identification. Members of the FOSC are notorious for causing many diseases, which includes stem rot of Sulcorebutia heliosa and root rot of Torreya grandis (Garibaldi et al. 2020; Zhang et al. 2016). To our knowledge, this is the first report of stem rot by F. oxysporum on C. sinense in China. The finding of this pathogen provides a clear target for stem rot control.

[Follow-up studies for long-term postoperative complications of Warshaw operation].

OBJECTIVE: To explore the long-term postoperative complications of Warshaw operation for distal pancreatectomy with preservation of spleen. METHODS: Retrospectively analyzed the clinical data of distal pancreatectomy on 84 cases between January 2006 and January 2012. Among them, 35 patients underwent Warshaw operation for distal pancreatectomy with preservation of spleen and another 23 had distal pancreatectomy with Kimura operation. All of them were followed up. RESULTS: All procedures were successful. There was no perioperative mortality. During the follow-up period, 2 patients undergoing Warshaw operation were lost. The median follow-up period was 2.6 years (range: 0.75-6.75). Twenty-six patients with the recent neuroimaging data showed: 5 patients were diagnosed of splenic infarction and the incidence rate was 19.2% (5/26) and 6 with gastric varices and the incidence rate was 23.1% (6/26). But none of them developed gastrointestinal hemorrhage or splenic abscess. CONCLUSION: The Warshaw operation for distal pancreatectomy with preservation of spleen may increase the risk of long-term postoperative complications so that its operative indication must be strictly mastered.

Serum Isthmin-1 is negatively correlated with HDL-C in type 2 diabetes mellitus.

BACKGROUND: Isthmin-1 (Ism-1) is a newly identified insulin-like adipokine that increases glucose uptake by adipocytes and inhibits hepatic lipid synthesis. Recent studies have shown that Ism-1 can improve the metabolic disorders associated with type 2 diabetes mellitus (T2DM) and improve lipid metabolism. The classic function of high-density lipoprotein cholesterol (HDL-C) is to transport cholesterol from extra-hepatic tissues to the liver for metabolism. In contrast, disorders of lipid metabolism and inflammation are the leading causes of atherosclerosis (As). Atherosclerosis often manifests as loss of elasticity, lipid accumulation, fibrous tissue proliferation and calcium deposits in the affected arteries, eventually forming plaques. AIM: To illustrate the correlation between HDL-C and Ism-1 in T2DM, and the relationship between lipoprotein cholesterol and carotid plaque. METHODS: A total of 128 patients with T2DM were enrolled in the study and basic information was collected. HDL-C levels were measured chemically. Serum Ism-1 levels were measured using an enzyme-linked immunosorbent assay (ELISA). Linear regression analysis was used to assess the correlation between serum Ism-1 levels and HDL-C in patients with T2DM. Basic information was again collected from 226 patients with T2DM. Independent sample t-tests were performed to explore the relationship between carotid plaque formation and lipids. RESULTS: HDL-C was divided into four groups according to quartiles and there was a between-group difference in Ism-1 (p = 0.040). Multivariable linear regression showed a negative association between Ism-1 and HDL-C in T2DM (ß = -0.235, p < 0.001), even after adjusting for related factors (ß = -0.165, p = 0.009). Low-density lipoprotein cholesterol (LDL-C) and HDL-C showed significant differences between the carotid plaque group and the non-carotid plaque group (pLDL-C = 0.007, pHDL-C = 0.003). CONCLUSION: Serum Ism-1 and HDL-C are negatively correlated in T2DM. LDL-C is significantly higher in carotid plaque group than non-carotid plaque group, while HDL-C is significantly lower than in the non-carotid plaque group.

Induction of detoxification enzymes by quercetin in the silkworm.

Quercetin is one of the most abundant flavonoids and the defense secondary metabolites in plants. In this study, the effect of quercetin on the growth of the silkworm larvae was investigated. Cytochrome P450 monooxygenases (P450s), glutathione S-transferases (GSTs), and carboxylesterases (COE) were assayed after exposure to different concentrations of quercetin for 3 d (short-term) and 7 d (long-term), respectively. The results showed that the weight gain of the silkworm larvae significantly decreased after the larvae were treated by different concentrations of quercetin except for the treatment with 0.5% quercetin. Activities of P450, GST, and COE were induced by 0.5 or 1% concentration of quercetin. In the midgut, the induction activity of P450s was reached to the highest level (2.3-fold) by 1% quercetin for 7 d, the highest induction activities of GSTs toward CHP and CDNB were 4.1-fold and 2.6-fold of controls by 1% quercetin after 7 d exposure, respectively. For COEs, the highest activity (2.3-fold) was induced by 0.5% quercetin for 7 d. However, P450s in whole body were higher inducible activities in short-term treatment than those in long-term treatment. The responses of eight cytochrome P450 (CYP) genes belonged to CYP6 and CYP9 families and seven GST genes were detected with real-time polymerase chain reaction. In addition, the genes induced by quercetin significantly were confirmed by qRT-PCR. CYP6AB5, CYP6B29, and GSTe8 were identified as inducible genes, of which the highest induction levels were 10.9-fold (0.5% quercetin for 7 d), 6.2-fold (1% quercetin for 7 d), and 7.1-fold (1% quercetin for 7 d), respectively.

[Observation of laparoscopic distal pancreatectomy in hepatobiliary surgery: a report of 25 cases].

OBJECTIVE: To explore the clinical application of laparoscopic distal pancreatectomy (LDP) in hepatobiliary operations. METHODS: The clinical data of LDP from 25 cases from June 2008 to December 2011 were retrospectively analyzed. Among them, 9 patients underwent LDP with excision of spleen while another 16 patients had LDP with preservation of spleen. And during the same period, 42 patients undergoing open distal pancreatectomy were selected into the control group. And the patient data of two groups were compared. RESULTS: All of them underwent successfully LDP. Among 16 patients with preservation of spleen, 11 patients undergone the Kimura procedure while another 5 undergone the Warshaw operation. The operative duration was (4.5 ± 1.2) hours, volume of blood loss (256 ± 188) ml, length of incision (4.6 ± 0.9) cm, mean time of oral food taking (1.5 ± 0.9) days and mean postoperative hospital stay (7.1 ± 1.9) days. Pathological examinations showed benign tumor (n = 20), malignant tumor (n = 4) and borderline tumor (n = 1). The mean data of LDP group was significantly less than that of open distal pancreatectomy group in terms of anal exsufflation time, length of incision, postoperative hospital stay time and complication rate, etc. (P < 0.05). Four patients were diagnosed of post-LDP pancreatic fistula (3 A level vs 1 B level) and all of them became cured after conservative treatment. There was no mortality. CONCLUSION: As a safe and feasible procedure, LDP is worthy of wider applications.

Yap is essential for uterine decidualization through Rrm2/GSH/ROS pathway in response to Bmp2.

Yap is required for ovarian follicle and early embryo development, but little information is available regarding its physiological significance in decidualization. Here we determine the effects of YAP on decidualization, mitochondrial function, cell apoptosis and DNA damage, and explore its interplay with Bmp2, Rrm2, GSH and ROS. The results exhibited that Yap was abundant in decidual cells and its inactivation impaired the proliferation and differentiation of stromal cells along with the deferral of G1/S phase transition, indicating Yap importance in decidualization. Bmp2 via Alk2 receptor promoted nuclear translocation of Yap where it might interact with Tead and then bind to the promoter of Rrm2 whose activation rescued the faultiness of differentiation program and attenuated oxidative DNA damage caused by Yap impediment. Meanwhile, Yap had an important part in the crosstalk between Bmp2 and Rrm2. Furthermore, inactivation of Yap resulted in an obvious accumulation of intracellular ROS followed by the abnormal GR activity and GSH content dependent on Rrm2. Replenishment of GSH counteracted the regulation of Yap inactivation on stromal differentiation and DNA damage with distinct reduction for intracellular ROS. Additionally, blockage of Yap caused the enhancement of stromal cell apoptosis and brought about mitochondrial dysfunction as indicated by the aberration for ATP level, mtDNA copy number and mitochondrial membrane potential concomitant with the opening of mitochondrial permeability transition pore, but these abnormalities were neutralized by GSH. Administration of mitochondrial antioxidant Mito-TEMPO rescued the fault of stromal differentiation conferred by Yap inactivation. Collectively, Yap was essential for uterine decidualization through Rrm2/GSH/ROS pathway in response to Bmp2.

Differences in the cargos and functions of exosomes derived from six cardiac cell types: a systematic review.

Exosomes are bilayer membrane vesicles with cargos that contain a variety of surface proteins, markers, lipids, nucleic acids, and noncoding RNAs. Exosomes from different cardiac cells participate in the processes of cell migration, proliferation, apoptosis, hypertrophy, and regeneration, as well as angiogenesis and enhanced cardiac function, which accelerate cardiac repair. In this article, we mainly focused on the exosomes from six main types of cardiac cells, i.e., fibroblasts, cardiomyocytes, endothelial cells, cardiac progenitor cells, adipocytes, and cardiac telocytes. This may be the first article to describe the commonalities and differences in regard to the function and underlying mechanisms of exosomes among six cardiac cell types in cardiovascular disease.

Hypoxic preconditioning improves long-term functional outcomes after neonatal hypoxia-ischemic injury by restoring white matter integrity and brain development.

AIMS: Neonatal hypoxia-ischemia (H/I) results in gray and white matter injury, characterized by neuronal loss, failure of neural network formation, retarded myelin formation, and abnormal accumulation of oligodendrocyte progenitor cells (OPCs). These changes lead to severe neurological deficits and mortality. Sublethal hypoxic preconditioning (HPC) can protect the developing brain against H/I. However, limited evidence is available concerning its effect on white matter injury. METHODS: In this study, P6 neonatal Sprague-Dawley rats were subjected to normoxic (21% O2 ) or HPC (7.8% O2 ) for 3 hours followed 24 hours later by H/I brain injury. Neurological deficits were assessed by gait, righting reflex, foot fault, and Morris water maze tests. Compound action potential of the corpus callosum was recorded 35 days after surgery, and the correlation between axon myelination and neurological function was determined. RESULTS: Hypoxic preconditioning significantly attenuated H/I brain injury at 7 days and remarkably improved both sensorimotor and cognitive functional performances up to 35 days after H/I. HPC-afforded improvement in long-term neurological outcomes was attributable, at least in part, to restoration of the differentiation and maturation capacity in oligodendrocyte progenitor cells, amelioration of microglia/macrophage activation and neuroinflammation, and continuation of brain development after H/I. CONCLUSIONS: Hypoxic preconditioning restores white matter repair, development, and functional integrity in developing brain after H/I brain injury.

Pharmacokinetics of Eight Flavonoids in Rats Assayed by UPLC-MS/MS after Oral Administration of Drynariae rhizoma Extract.

As a traditional Chinese medicine, Drynariae rhizoma (Kunze ex Mett.) J. Sm. has been used to treat osteoporosis and bone resorption for 2500 years. Based on the previous study and literature references, flavonoids were proved to be the most abundant and main active compounds of Drynariae rhizoma for osteoporosis treatment. In order to make good and rational use of Drynariae rhizoma in future, a rapid, sensitive, and selective ultraperformance liquid chromatography-mass spectrometry (UPLC-MS/MS) method was developed to investigate the pharmacokinetics of eight main flavonoids in rat plasma after oral administration of the Drynariae rhizoma extract, including neoeriocitrin, luteolin-7-O-ß-D-glucoside, astragalin, naringin, eriodictyol, luteolin, naringenin, and kaempferol. Plasma samples' pretreatment involved a solid-phase extraction column. The separation was performed on an ACQUITY UPLCTM BEH C18 column with a gradient mobile-phase system of acetonitrile and 1% acetic acid in water. The detection was performed using a triple quadrupole tandem mass spectrometer equipped with an electrospray ionization interface (ESI) by multiple reaction monitoring (MRM) in the positive ion mode. All calibration curves exhibited good linearity (r 2 > 0.9990) over the measured ranges. The intraday and interday precisions (RSD) were within 13.87%, and the accuracy (RE) ranged from -14.57% to -0.25% at three quality control levels. Extraction recovery, matrix effect, and stability were satisfactory. The pharmacokinetic characteristics of the eight flavonoids of interest were clearly elucidated.

Detection of Zika virus by SYBR green one-step real-time RT-PCR.

The ongoing Zika virus (ZIKV) outbreak has rapidly spread to new areas of Americas, which were the first transmissions outside its traditional endemic areas in Africa and Asia. Due to the link with newborn defects and neurological disorder, numerous infected cases throughout the world and various mosquito vectors, the virus has been considered to be an international public health emergency. In the present study, we developed a SYBR Green based one-step real-time RT-PCR assay for rapid detection of ZIKV. Our results revealed that the real-time assay is highly specific and sensitive in detection of ZIKV in cell samples. Importantly, the replication of ZIKV at different time points in infected cells could be rapidly monitored by the real-time RT-PCR assay. Specifically, the real-time RT-PCR showed acceptable performance in measurement of infectious ZIKV RNA. This assay could detect ZIKV at a titer as low as 1PFU/mL. The real-time RT-PCR assay could be a useful tool for further virology surveillance and diagnosis of ZIKV.

[Clinicopathological characteristics and prognosis of solid pseudopapillary neoplasm of the pancreas: comparison between tumors ≤5 cm and larger tumors].

OBJECTIVE: To compare the clinicopathological characteristics and prognosis of patients with small (≤5 cm) solid pseudopapillary neoplasm of the pancreas (SPN) and those with large (>5 cm) SPNs. METHODS: We retrospectively analyzed the clinical characteristics, laboratory findings, radiological features, treatment and prognosis of 148 patients with histologically confirmed SPN between August, 2006 and December, 2014 and compared the data between patients with small SPNs (≤5 cm) and those with large SPNs (>5 cm). RESULTS: In the large SPN group, the female-to-male ratio was significantly higher than that in small SPN group (61/8 vs 56/23, P=0.009) and the patients were significantly younger in large SPN group (28.3±12.3 vs 33.0±11.4 years, P=0.016). Small SPNs (≤5 cm) typically presented as inhomogeneous solid or cystic tumors, while large SPNs (>5 cm) often appeared as homogeneous solid and cystic tumors, but they did not show any significant difference in aggressive behaviors (P=0.288). The 5-year disease-free survival of patients with small SPNs was 100%, and the 1-, 3-, and 5-year disease-free survival of patients with large SPNs was 98.6%, 94.9%, and 89.3%, respectively (P=0.030), showing no significant differences in the overall survival between the two groups. CONCLUSION: Small SPNs and large SPNs have different clinical characteristics. Even with complete resection, tumors larger than 5 cm are more likely to have tumor recurrence and metastasis, and close follow-up is recommended for these patients.

Clinicopathological characteristics and prognostic factors of intrahepatic biliary cystadenocarcinoma.

BACKGROUND: Surgical resection is generally considered the main curative treatment for intrahepatic biliary cystadenocarcinoma (IBCA) or suspected IBCAs, but controversy exists regarding the prognosis for IBCAs. This study aimed to describe the clinicopathological characteristics of IBCA and identify prognostic factors that may influence the survival of patients treated with surgical procedures. METHODS: Thirty-four patients with histologically confirmed IBCA treated between January 2000 and June 2014 were included. The clinical characteristics of patients with IBCA were compared with those of 41 patients with intrahepatic biliary cystadenoma (IBC); factors that significant difference were analyzed for prognosis analysis of IBCA using multivariate/univariate Cox proportional hazards regression models. Survival curves were constructed using the Kaplan-Meier method and compared using the log-rank test. RESULTS: IBCAs had a strong female predominance, and the most common presenting symptoms were abdominal pain or discomfort. Compared with IBCs, IBCAs occurred in older patients, in more male patients, and were associated statistically significant abnormal increase in alanine aminotransferase (P = 0.01) and total bilirubin (P = 0.04). Mural nodules were more frequently seen with IBCAs and may associate with malignancy. It was difficult to differentiate between IBC and IBCA based on laboratory examination and imaging findings. Although complete resection is recommended, enucleation with negative margins also achieved good outcomes. Median overall patient survival was 76.2 months; survival at 1, 3, and 5 years was 88.0%, 68.7%, and 45.8%, respectively. Radical resection and noninvasive tumor type were independent prognostic factors for overall survival. CONCLUSIONS: It remains difficult to distinguish between cystadenomas and cystadenocarcinomas based on laboratory examination and image findings. Complete resection is recommended for curative treatment, and patients should be closely followed postoperatively, particularly those with invasive tumors.