Puccinia suaveolens Causing Leaf Rust on Cirsium setosum in China.

Thistle, Cirsium setosum (Willd.) M. Bieb., is widely distributed in China as a common weed in fields. It is also used as a traditional Chinese medicine for cooling blood, stopping bleeding, dispelling stasis, detoxifying, and resolving carbuncle. In 2023, we found a rust disease on plants of Cirsium setosum in the experimental field of Hebei Agricultural University, Baoding, Hebei Province, China, with incidence of 15% - 25% (Fig. S1 A, B). The diseased leaves turned yellow, and the leaf edges were slightly rolled. The yellow, oil-like pycnia and pycniospores covered the baxial surface of leaves, and brown pustules were produced after 2-3 weeks. On the adaxial surface of the leaves, the brown rust pustules were mainly along the leaf veins. Stems were also be infected later, and dark pustules were scattered. The diseased plants were relatively short and small, and produced relatively small or no flowers compared to healthy plants. A total of 100 plants with typical leaf rust symptoms and signs were collected. To confirm the pathogenicity, healthy plants of thistle were sprayed with 5 ml of urediospores suspension (2.6×105/ml), and plants sprayed with sterile distilled water were treated as control. The sprayed plants were incubated under high moist conditions at 18°C for 24 h, and the inoculated plants were grown at 20°C in a greenhouse. Ten days after inoculation, the plants inoculated with urediniospores showed rust symptoms with uredinia and urediniospores on the leaves (Fig. S1 C), while the control plants were healthy. For morphological characterization, urediospores were picked from the naturally infected plants and placed in a drop of sterile water on a glass slide using a sterile needle, and observed and measured under a microscope. Urediospores were nearly spherical, brown-yellow, and measured 15 - 25 µm in diameter (n=100) (Fig. S1 D). Telia were scattered on the baxial surface of the naturally infected leaves, and teliospores were oval, yellow-brown, double-celled, with very short hyaline pedicels, and measured 15-20 × 15-30 µm (n=100) (Fig. S1 E). For molecular characterization, about 200 µg of urediniospores was collected and placed in a 1.5 ml sterile centrifuge tube, and genomic DNA was extracted using the cetyl-trimethylammonium bromide method (Gawel et al. 1991). The internal transcribed spacer (ITS) region of the rDNA and the D1/D2 domain were amplified using primer pairs ITS1/ITS4 (White et al. 1990) and NL1/NL4 (Borhani et al. 2013) in polymerase chain reaction (PCR), respectively. The PCR products were sequenced, and their sequences were aligned and compared with those deposited in GenBank. The obtained sequences were deposited in GenBank (OR600240 for ITS and OR598614 for D1/D2), which were 100% identical with 100% coverage to the ITS sequence (ON063373.1) and the D1/D2 sequence (ON063379.1) of Puccinia suaveolens (Menzies 1953). Based on the morphological characteristics and DNA sequences, the isolates were identified as P. suaveolens (Fig. S1 and Fig. S2). Thistle rust caused by Puccinia obtegens has been reported in some other parts of China (Zhang 2012). To the best of our knowledge, this is the first report of P. suaveolens causing leaf rust on C. setosum in China. This discovery is helpful for control of leaf rust on thistle grown for Chines medicine and other purposes, and the rust species could be used for biological control of thistle as a weed in crop fields.

circSSPO boosts growth of esophageal squamous cell carcinoma through upregulation of micrRNA-6820-5p-mediated KLK8 and PKD1 expression.

Investigation on a competitive endogenous RNA (ceRNA) network attracted lots of attention due its function in cancer regulation. Here, we probed into the possible molecular mechanism of circSSPO/microRNA-6820-5p (miR-6820-5p)/kallikrein-related peptidase 8 (KLK8)/PKD1 network in the esophageal squamous cell carcinoma (ESCC). Following whole-transcriptome sequencing and differential analysis in collected ESCC tissue samples, circRNA-miRNA-mRNA regulatory network affecting ESCC was investigated. After interaction measurement among circSSPO/miR-6820-5p/KLK8/PKD1, their regulatory roles in ESCC cell functions in vitro and xenograft tumor growth and lung metastasis in vivo were analyzed. The bioinformatics prediction and sequencing results screened that circSSPO, miR-6820-5p, KLK8, and PKD1 were associated with ESCC development. In ESCC, miR-6820-5p was expressed at very low levels, while circSSPO, KLK8, and PKD1 were highly expressed. In vitro cell experiments further proved that circSSPO competitively inhibited miR-6820-5p to induce ESCC cell malignant properties. Moreover, knockdown of KLK8 or PKD1 inhibited ESCC cell malignant properties. circSSPO also promoted the tumorigenic and metastasis of ESCC through the upregulation of KLK8 and PKD1 expression in vivo. We found that circSSPO was an oncogenic circRNA that was significantly abundant in ESCC tissues and circSSPO exhibited an oncogenic activity in ESCC by elevating expression of KLK8 and PKD1 through suppressing miR-6820-5p expression.

Hypoxia-induced ALDH3A1 promotes the proliferation of non-small-cell lung cancer by regulating energy metabolism reprogramming.

Aldehyde dehydrogenase 3A1 (ALDH3A1) is an NAD+-dependent enzyme that is closely related to tumor development. However, its role in non-small-cell lung cancer (NSCLC) has not been elucidated. This study aimed to clarify the mechanism of ALDH3A1 and identify potential therapeutic targets for NSCLC. Here, for the first time, we found that ALDH3A1 expression could be induced by a hypoxic environment in NSCLC. ALDH3A1 was highly expressed in NSCLC tissue, especially in some late-stage patients, and was associated with a poor prognosis. In mechanistic terms, ALDH3A1 enhances glycolysis and suppresses oxidative phosphorylation (OXPHOS) to promote cell proliferation by activating the HIF-1α/LDHA pathway in NSCLC. In addition, the results showed that ALDH3A1 was a target of ß-elemene. ALDH3A1 can be downregulated by ß-elemene to inhibit glycolysis and enhance OXPHOS, thus suppressing NSCLC proliferation in vitro and in vivo. In conclusion, hypoxia-induced ALDH3A1 is related to the energy metabolic status of tumors and the efficacy of ß-elemene, providing a new theoretical basis for better clinical applications in NSCLC.

First Report of Penicillium cellarum Causing Rot Disease on Dioscorea polystachya in China.

Dioscorea polystachya (Chinese yam) is a kind of medicine and food homologous crop, the tubers as its main production organ, with high potassium, low fiber, high protein and rich nutrition characteristics. In 2022, at the Chinese herbal medicine planting experimental site in Anguo, Baoding City, Hebei Province, China, we found the symptoms of Chinese yam decay during the storage, with an incidence of 15%~25%. The diseased part of Chinese yam tuber rots expands from the outside to the inside and sags, with a brown or dark brown discoloration, and the surface covered with a thick grayish green mold. The diseased tissue was first rinsed with clean water to remove dirts from the surface. Thereafter, 3 to 4 mm Chinese yam pieces were picked from rotting edge with a sterilized forceps, sterilized with 75% alcohol for 30 s followed by 0.1% mercuric chloride solution for 1min, and then rinsed three times with sterile water. The sterilized pieces were cultured on potato dextrose agar (PDA). One isolated fungus was obtained, and conidia were observed after incubation for 5 days at 26°C. Pure cultures were isolated by single-spore isolation. Conidia were single spore, round or oval, colorless. Conidiophores produce several rounds of symmetric or asymmetric small stems after multiple branches, which were shaped like brooms. The length and width of 100 conidia were measured, and size ranged from 3 to 4×3 to 4 µm. On the basis of morphological characteristics, the isolate was identified as Penicillium spp. (Uy et al. 2022). To further assess the identity of isolated species, the genomic DNA of the fungal isolate (SYRF1) was extracted by CTAB protocol. The ribosomal DNA internal transcribed spacer (ITS) region and the ribosomal large subunit (LSU) were amplified and sequenced with primers ITS1/4, LR5/LROR respectively (White et al. 1990, Xu et al. 2010). The obtained ITS-rDNA region and LSU sequences (GenBank accession OQ707937 and OQ704185) of the isolate were more than 99% identity to the corresponding sequences of Penicillium cellarum in GenBank (KM249068 and MG714818). Phylogenetic results based on a maximum-likelihood analysis revealed that SYRF1 was grouped with P. cellarum. To determine the pathogenicity of the isolated fungi, tests were carried out by aseptic inoculation of fresh and healthy tubers. Before the experiment, the healthy tubers were washed, surface disinfected and dried. The tubers were then wounded with sterile inoculation needles, and the conidium-bearing hyphal discs (5 mm) were inoculated on the surface of the wounded tubers and covered with wet sterile cotton. Three tubers were inoculated repeatedly each time as the experimental group. Inoculate sterile PDA with three tubers as the control group. Each tuber was inoculated with four mycelium disks, and the pathogenicity test was repeated four times. The inoculated tubers were incubated at 26°C for 14 days with sterile PDA as control. After ten days, the inoculated points showed symptoms similar to those of the initial infection, whereas controls remained symptomless. The reisolated fungus matched SYRF1 based on morphological and sequence analyses, thereby fulfilling Koch's postulates. To the best of our knowledge, this is the first report of Penicillium cellarum as causative agent of postharvest rot of Chinese yam tubers in China. This finding will help inform the prevention and management of postharvest diseases of Chinese yam tubers.

Study on Sustentaculum Tali Fragment Constancy in Intraarticular Calcaneus Fracture.

OBJECTIVES: To establish reproducible measurements of the sustentaculum tali (ST) fragment regarding fracture classification and patient-related factors. DESIGN: Retrospective. SETTING: Trauma center, University Hospital. PATIENTS: A retrospective analysis of the 142 fractured calcanei of 122 patients (101 men and 21 women) treated at our institution between 2012 and 2020 was performed. As control, 62 unaffected calcanei were used. INTERVENTION: Radiographic images were evaluated twice within 2 weeks by 2 orthopaedic surgeons and 1 postgraduate student. Angulation and diastasis were used to distinguish ST fragment constancy based on computed tomography. Using these parameters, the prevalence of inconstant ST fragments was assessed. We also analyzed factors related to ST fragment inconstancy. Patient factors included age, body mass index, smoking, and diabetes. Radiographic factors included the Sanders classification, location of the outermost fracture line of the posterior facet, presence of an intraarticular fracture of the ST, and ST fragment width. MAIN OUTCOME MEASUREMENTS: Angulation and diastasis were used to confirm the ST fragment constancy. Potential risks for inconstant ST fragment subsequently defined. RESULTS: According to the criteria, ST fragment inconstancy was observed in 34.5%. ST fragment width was significantly smaller in the inconstant group ( P < 0.001). Severe comminution of the posterior facet ( P < 0.05), intraarticular fracture of the ST ( P < 0.001), and diabetes ( P < 0.05) were significantly higher in the inconstant group. The cut-off value of the ST fragment width was 20.5 mm. CONCLUSIONS: In intraarticular calcaneus fractures, small ST fragment width, comminuted fracture, intraarticular fracture of the ST, and diabetes were associated with the inconstant group. The ST fragment was expected to be inconstant when the width was less than 20.5 mm.

Functional identification of a novel C7 protein of tomato yellow leaf curl virus.

Tomato yellow leaf curl virus (TYLCV) is a monopartite geminivirus, and one of the most devastating plant viruses in the world. TYLCV is traditionally known to encode six viral proteins in bidirectional and partially overlapping open reading frames (ORFs). However, recent studies have shown that TYLCV encodes additional small proteins with specific subcellular localizations and potential virulence functions. Here, a novel protein named C7, encoded by a newly-described ORF in the complementary strand, was identified as part of the TYLCV proteome using mass spectrometry. The C7 protein localized to the nucleus and cytoplasm, both in the absence and presence of the virus. C7 was found to interact with two other TYLCV-encoded proteins: with C2 in the nucleus, and with V2 in the cytoplasm, forming conspicuous granules. Mutation of C7 start codon ATG to ACG to block the translation of C7 delayed the onset of viral infection, and the mutant virus caused milder virus symptoms and less accumulations of viral DNAs and proteins. Using the potato virus X (PVX)-based recombinant vector, we found that ectopic overexpression of C7 resulted in more severe mosaic symptoms and promoted a higher accumulation of PVX-encoded coat protein in the late virus infection stage. In addition, C7 was also found to inhibit GFP-induced RNA silencing moderately. This study demonstrates that the novel C7 protein encoded by TYLCV is a pathogenicity factor and a weak RNA silencing suppressor, and that it plays a critical role during TYLCV infection.

Virulence and molecular genetic diversity, variation, and evolution of the Puccinia triticina population in Hebei Province of China from 2001 to 2010.

Wheat leaf rust, caused by Puccinia triticina, is one of the most important fungal diseases of wheat in China. However, little is known about the dynamic changes of population structure and genetic diversity of P. triticina during a period of time. In this study, 247 isolates of P. triticina collected from Hebei Province from 2001 to 2010 were tested on 36 Thatcher near-isogenic lines for virulence diversity and detected by 21 pairs of Expressed Sequence Tag derived Simple Sequence Repeat (EST-SSR) primers for genetic diversity. A total of 204 isolates were successfully identified as 164 races, and THTT, THST, PHRT, THTS, and PHTT were the most common races in Hebei Province from 2001 to 2010. The cluster analysis based on virulence showed that P. triticina has a rich virulence polymorphism, which had a certain correlation with the years, while the cluster analysis based on EST-SSR showed that the genetic diversity of the P. triticina population was significantly different between years in Hebei Province from 2001 to 2010. In addition, the population structure of P. triticina may have changed greatly in 2007 and 2009, which was significantly different from that of 2001-2006 on either virulence or genetic characteristics. The variation frequency of the population structure had an increasing trend during this period. From 2001 to 2010, there was a certain degree of gene flow among the P. triticina populations. No significant correlation was found between virulence and molecular polymorphism. The genetic differentiation analysis of the 10 tested populations (each year as a population) showed that the coefficient of genetic differentiation (Gst) was 0.27, indicating that there was a certain genetic differentiation among or within populations of P. triticina in Hebei Province. The genetic variation within populations (73.08%) was higher than that among populations (26.92%), which indicated that the genetic variations were mainly found within populations. Our study provides the foundation for a better understanding of the population structure change and genetic diversity of P. triticina over a period in Hebei Province of China.

Misdiagnosis of Pyoderma Gangrenosum Increases Medical Costs and Prolongs Hospital Stay: A Case Report.

Pyoderma gangrenosum (PG) is a rare, immunological ulcerative, and necrotic inflammatory skin disease that can be easily misdiagnosed as cellulitis, abscess, diabetic foot ulcer, and other infectious diseases. Misdiagnosing PG leads to unnecessary surgical incision and debridement, which further exacerbates the lesion, ultimately leading to longer treatment periods and higher medical costs. Therefore, early and accurate diagnosis of PG is extremely important for its treatment. In particular, PG should be suspected in patients with inflammatory bowel disease.

Symptomatic accessory soleus muscle: A cause for exertional compartment syndrome in a young soldier: A case report.

BACKGROUND: Accessory soleus muscle (ASM) is a rare congenital variation that is almost asymptomatic, but several papers have recently described symptomatic ASM. The clinical features of this condition are similar to tarsal tunnel syndrome (TTS) and include pain and numbness around the medial side of the ankle. ASM commonly originates from the fibula or soleus muscle and inserts into the Achilles tendon or calcaneus. Usually, it is identified as posteromedial swelling and definitely diagnosed by magnetic resonance imaging. In most cases, treatment is observation, but surgical excision can be considered if symptoms are severe. CASE SUMMARY: A 23-year-old male Korean soldier presented with complaints of bilateral foot and ankle pain and a swelling medial to the Achilles tendon that was more pronounced on the right side. Symptoms first occurred after playing soccer 10 mo before this presentation, worsened after physical exertion, and were relieved by rest. He had no medical history, and no one in his family had the condition. Laboratory results were non-specific. Several tests were performed to exclude common diseases such as tumors or TTS. However, MRI revealed a bulky accessory soleus muscle in both feet, though the patient complained of more severe pain on the right side during physical activity. Accordingly, surgical resection was adopted. At surgery, a large accessory soleus muscle was noted anterior to the Achilles tendon with distinctive insertion from a normal soleus muscle. At 12 mo after surgery, there was no pain, numbness, or swelling of the right foot or ankle, no evidence of recurrence, and the patient could do all sports activities. CONCLUSION: Accessory soleus muscle should be added to the list of differential diagnosis if a patient has pain, sole numbness or swelling of the posteromedial ankle.

Evaluation of the resistance to Chinese predominant races of Puccinia triticina and analysis of effective leaf rust resistance genes in wheat accessions from the U.S. National Plant Germplasm System.

Puccinia triticina, which is the causative agent of wheat leaf rust, is widely spread in China and most other wheat-planting countries around the globe. Cultivating resistant wheat cultivars is the most economical, effective, and environmentally friendly method for controlling leaf rust-caused yield damage. Exploring the source of resistance is very important in wheat resistance breeding programs. In order to explore more effective resistance sources for wheat leaf rust, the resistance of 112 wheat accessions introduced from the U.S. National Plant Germplasm System were identified using a mixture of pathogenic isolates of THTT, THTS, PHTT, THJT and THJS which are the most predominant races in China. As a result, all of these accessions showed high resistance at seedling stage, of which, ninety-nine accessions exhibited resistance at adult plant stage. Eleven molecular markers of eight effective leaf rust resistance genes in China were used to screen the 112 accessions. Seven effective leaf rust resistance genes Lr9, Lr19, Lr24, Lr28, Lr29, Lr38 and Lr45 were detected, except Lr47. Twenty-three accessions had only one of those seven effective leaf rust resistance gene. Eleven accessions carried Lr24+Lr38, and 7 accessions carried Lr9+Lr24+Lr38, Lr24+Lr38+Lr45, Lr24+Lr29+Lr38 and Lr19+Lr38+Lr45 respectively. The remaining seventy-one accessions had none of those eight effective leaf rust resistance genes. This study will provide theoretical guidance for rational utilization of these introduted wheat accessions directly or for breeding the resistant wheat cultivars.

Characteristics of Synovial Fistula of the Ankle Joint: A Case Series.

(1) Background: Little is known about the etiology, clinical features, diagnosis methods, treatments, and the prognosis of synovial fistula of the ankle joint. The purpose of this study is to investigate the clinical features of synovial fistula of the ankle joint. (2) Methods: Between March 2003 and December 2018, 40 cysts associated with synovial fistula of the ankle joint were treated consecutively by two surgeons. Case histories, clinical manifestations, intraoperative findings, surgical treatment methods, and treatment outcomes were evaluated to characterize fistula-associated cysts. The clinical results were assessed using the visual analog scale (VAS) and the American Orthopaedic Foot and Ankle Society (AOFAS) ankle-hindfoot functional scores, preoperatively and at the last follow-up. (3) Results: The main complaints were ankle instability and pain (15 patients), pain only (15 patients), instability (seven patients), and cosmetic problems (three patients). Eleven patients had a cyst with an open skin wound, and eight of these patients had undergone surgery under a misdiagnosis of bursitis. Cysts were located anterior to the lateral malleolus in 22 cases, next to the lateral malleolus in 13 cases, posterior to the lateral malleolus in three cases, and across the entire lateral malleolus in two cases. Mean VAS and AOFAS scores improved from 5.2 (range, from 1 to 7) and 72.3 (range, from 65 to 87) preoperatively to 1.1 (range, from 0 to 3) and 93.6 (range, from 85 to 100), respectively, at final follow-up visits. (4) Conclusions: Cyst occurrence due to a synovial fistula should be considered when treating a cyst around the lateral malleolus. Fistula repair and reinforcement with fibular periosteum provides a good treatment option for cysts attributed to synovial fistula of the ankle that fail to respond to conservative treatment.

Recurrence of intratendinous ganglion due to incomplete excision of satellite lesion in the extensor digitorum brevis tendon: A case report.

BACKGROUND: Intratendious ganglions are rare lesions, especially on the foot and ankle. Although several studies have presented the intratendinous ganglion of the foot and ankle, there are only few reported cases, and no cases of recurrence or secondary surgery have been reported. CASE SUMMARY: We present the case of a 32-year-old man with an intratendinous ganglion of the second extensor digitorum brevis (EDB) tendon that recurred after ganglion excision. Magnetic resonance imaging (MRI) performed before the first surgery was reviewed to analyze the causes of the recurrence. We confirmed that there was a lack of satellite detection. After recurrence, MRI revealed an extra-tendinous lesion, tenosynovitis, and intratendinous ganglion of the second EDB tendon. Since the second EDB tendon can compensate for the extrinsic muscle, en bloc resection was performed alone. In addition, meticulous excision and synovectomy were performed for extra-tendinous lesions and tenosynovitis, respectively. The patient returned to daily life without any functional problems or recurrence. CONCLUSION: If removal of the affected tendon is not fatal, en bloc resection should first be considered to prevent incomplete excision and intraoperative leakage. When planning surgical excision, it is necessary to evaluate the presence of satellite lesions along the course of the affected tendon.

Mini-open Repair for Acute Achilles Tendon Rupture: Ring Forceps vs the Achillon Device.

BACKGROUND: Repair of acute Achilles tendon rupture using the Achillon device is a representative mini-open repair technique; however, the limitations of this technique include the need for special instruments and decreased repair strength. A modified mini-open repair using ring forceps might overcome these limitations. PURPOSE: To compare the Achillon device with ring forceps in mini-open repairs of acute Achilles tendon rupture. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: Fifty patients (41 men and 9 women) with acute Achilles tendon rupture on 1 foot were consecutively treated using mini-open repair techniques. The first 20 patients were treated using the Achillon device (Achillon group), and the subsequent 30 were treated using a ring forceps (forceps group). Clinical, functional, and isokinetic results and postoperative complications were compared between the groups at the last follow-up. Clinical evaluations were performed using the AOFAS (American Orthopaedic Foot and Ankle Society) score, Achilles Tendon Total Rupture Score, length of incision, and operation time. Functional evaluations included active range of motion of the ankle joint, maximum calf circumference, hopping test, and single-limb heel rise (SLHR). Isokinetic evaluations were performed using the isokinetic test for ankle plantar flexion. RESULTS: The AOFAS score (P = .669), Achilles Tendon Total Rupture Score (P = .753), and length of incision (P = .305) were not significantly different between the groups (mean ± SD, 90.1 ± 8.7, 88.3 ± 9.9, and 2.7 ± 0.3 cm in the Achillon group vs 92.2 ± 9.4, 89.9 ± 10.9, and 2.5 ± 0.4 cm in the forceps group, respectively). Operation times in the Achillon group were significantly shorter than those in the forceps group (41.4 ± 9.6 vs 62.8 ± 14.1 minutes, P < .001). The maximum height of the SLHR (P = .042) and the number of SLHRs (P = .043) in the forceps group (79.7% ± 7.4% and 72.9% ± 10.2%) were significantly greater than those in the Achillon group (75.3% ± 7.1% and 66.7% ± 11.0%). No significant differences were detected between the groups in mean peak torques for plantar flexion at angular speeds of 30 deg/s (P = .185) and 120 deg/s (P = .271). There was no significant difference in the occurrence of postoperative complications between the groups (P = .093). CONCLUSION: The ring forceps technique is comparable to the Achillon technique with respect to clinical, functional, and isokinetic results and postoperative complications. Given that no special instrument is required, the ring forceps technique could be a better option for acute Achilles tendon rupture repair.

Citrobacter freundii causing Ginger (Zingiber officinale Rosc.) Rot in Tangshan, China.

Ginger (Zingiber officinale Rosc.) is an important economic crop and its rhizome can be used as seasoning agent and traditional medicine in China. During July 2018 and 2019, decay symptoms occurred in the ginger planting area of Tangshan City, Hebei Province, with incidence rates of 15%～20%. The pathogen infected the rhizomes and leaves. The symptoms included leaves chlorosis and gradually wilting, even the whole plant wilted, the rhizome became soft and presented light brown maceration. In serious cases, the interior of rhizome was completely eroded, gray-white juice overflowing the epidermis, and with foul smell. The rhizome surfaces of ginger plants were disinfected with 1% NaOCl, and colonies were isolated and purified on nutrient agar (NA) solid medium by streaking. Eight isolates were obtained from 15 diseased tissue samples. Further morphological, physiological and biochemical identification of the pure cultured bacteria were carried out. Three strains of bacteria were picked for further analysis. All of the three strains were gram-negative, short rod-shaped，nonmotile bacillus. Colonies were round and milky yellow, smooth raised, and moist after incubation at 28°C for 24h on NA. Physiological and biochemical test results showed that strains were facultatively anaerobic, negative for indole, methyl red, the Voges-Prauskauer test (V-P) and urease; positive for glucose, sucrose, sorbitol, inositol, mannitol, citrate utilization and hydrogen sulfide production; gelatin liquefaction. A typical hypersensitive reaction was induced on 12-week-old tobacco (Nicotiana benthamiana) leaves, which were inoculated by injecting suspensions of the isolated strain (108 CFU/mL) at 25 ℃ after 24h. These characteristics were consistent with Citrobacter freundii (Werkman and Gillen 1932). To further assess the identity of the strains, the genomic DNA was extracted from one bacterium（JXJ4）. The partial 16S rRNA region (Lane 1991) and specific rpoB and gyrB genes (Mollet et al. 1997, Brady et al. 2013) were amplified and sequenced with primers 27F/1492R, CM7/CM31b and UP1f/UP2r, respectively. The obtained 16S, rpoB and gyrB sequences (GenBank accession MN148645, MN158728 and MW199734) of the isolate showed 99.93%, 99.51% and 99.82% identity to the corresponding sequences of C. freundii in GenBank (CP024679.1, CP024677.1 and KM509081.1). Maximum likelihood analysis was performed, and the phylogenetic tree clustered with C. freundii (MEGAX, Bootstrap n=1000). The pathogenicity of the isolates was tested on ginger plants and rhizomes tissue. The bacterial suspensions (108 CFU/mL) of three isolates were injected into the basal stem and rhizomes center of 9 healthy ginger seedlings respectively, and Control groups were treated with sterile water. The inoculated plants were kept in a moist chamber (28°C, 16-h light and 8-h dark period) and ginger rhizomes were placed in the incubator (30°C, 16-h light and 8-h dark period). Seven days after inoculation, the ginger tubers showed symptoms of decay, and 20 to 25 days later, the ginger plant leaves browned and died. The pathogenicity test was repeated 4 times and all controls were healthy. Pathogens were reisolated from symptomatic plants and rhizomes and identified as C. freundii based on the morphological, biochemical and molecular methods described previously, fulfilling Koch's hypothesis. To our knowledge, this is the first report of ginger rot caused by C. freundii in China.

Randomized comparative study between extensile lateral and sinus tarsi approaches for the treatment of Sanders type 2 calcaneal fracture.

AIMS: No randomized comparative study has compared the extensile lateral approach (ELA) and sinus tarsi approach (STA) for Sanders type 2 calcaneal fractures. This randomized comparative study was conducted to confirm whether the STA was prone to fewer wound complications than the ELA. METHODS: Between August 2013 and August 2018, 64 patients with Sanders type 2 calcaneus fractures were randomly assigned to receive surgical treatment by the ELA (32 patients) and STA (32 patients). The primary outcome was development of wound complications. The secondary outcomes were postoperative complications, pain scored of a visual analogue scale (VAS), American Orthopaedic Foot and Ankle Society (AOFAS) score, 36-item Short Form health survey, operative duration, subtalar joint range of motion (ROM), Böhler's angle and calcaneal width, and posterior facet reduction. RESULTS: Although four patients (12.5%) in the ELA groups and none in the STA group experienced complications, the difference was not statistically significant (p = 0.113). VAS and AOFAS score were significantly better in the STA group than in the ELA group at six months (p = 0.017 and p = 0.021), but not at 12 months (p = 0.096 and p = 0.200) after surgery. The operation time was significantly shorter in the STA group than in the ELA group (p < 0.001). The subtalar joint ROM was significantly better in the STA group (p = 0.015). Assessment of the amount of postoperative reduction compared with the uninjured limb showed significant restoration of calcaneal width in the ELA group compared with that in the STA group (p < 0.001). CONCLUSION: The ELA group showed higher frequency of wound complications than the STA group for Sanders type 2 calcaneal fractures even though this was not statistically significant. Cite this article: Bone Joint J 2021;103-B(2):286-293.

First Report of Colletotrichum cereale Causing Anthracnose on Avena nuda in China.

Naked oats (Avena nuda L.) is rich in protein, fat, vitamin, mineral elements and so on, and is one of the world's recognized cereal crops with the highest nutritional and healthcare value. In July 2019, leaf spot was detected on A. nuda in Zhangbei experimental station of Hebei Agricultural University. The incidence of disease is 10% to 20%. The symptoms were similar to anthracnose disease, the infected leaves had fusiform or nearly fusiform yellowish-brown spots, yellow halo around the spots. Numerous acervuli with black setae diagnostic of fungi in the genus Colletotrichum were present on necrotic lesions. To identify the pathogen, ten symptomatic leaves were collected, and only one disease spot was isolated from each leaf. Small square leaf pieces (3 to 5 mm) were excised from the junction of diseased and healthy tissues with a sterile scalpel and surface disinfested with 75% alcohol for 30s, 0.1% corrosive sublimate for 1 min, rinsed three times in sterile water. Plant tissues were then transferred on potato dextrose agar (PDA), and incubated at 25°C for 7 days. Two fungal isolates were obtained and purified by single-spore isolation method. All fungi have the same morphology and no other fungi were isolated. The aerial mycelium was gray black. The conidia were colorless and transparent, falcate, slightly curved, tapered toward the tips, and produced in acervuli with brown setae. The length and width of 100 conidia were measured and size ranged from 1.86 to 3.84 × 8.62 to 29.81 µm. These morphological characteristics were consistent with the description of Colletotrichum cereale (Crouch et al. 2006). To further assess the identity of the species, the genomic DNA of two fungal isolates (LYM19-4 and LYM19-10) was extracted by a CTAB protocol. The ribosomal DNA internal transcribed spacer (ITS) region as well as, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin (ACT), and the beta-tubulin 2 (Tub2) partial genes were amplified and sequenced with primers ITS4/5, GDF/GDR, ACT-512F/ACT-783R, and T1/Bt2b, respectively (Carbone et al. 1999; Templeton et al. 1992; O'Donnell et al. 1997; Glass et al. 1995). The sequences of the ITS-rDNA region (MW040121, MW040122), the GAPDH sequences (MW052554, MW052555), the ACT sequences (MW052556, MW052551) and the Tub2 sequences (MW052552, MW052553) of the two single-spore isolates were more than 99% identical to C. cereale isolate CGMCC3.15110 (JX625159, KC843517, KC843534 and JX625186). Maximum likelihood tree based on concatenated sequences of the four genes were constructed using MEGA7. The results showed the strains isolated from A. nuda were closely related to C. cereale, as supported by high bootstrap values. A pathogenicity test of the C. cereale isolates was performed on first unfolding leaves of A. nuda. Koch's postulates were carried out with isolates by spraying a conidial suspension of 106 conidia/mL on leaves of healthy A. nuda. Four replicated pots were inoculated at a time, 10 leaves each pot, while sterile distilled water was used as the control. All treated plants were placed in a moist chamber (25°C, 16-h light and 8-h dark period). Anthracnose symptoms developed on the inoculated plants 7 days post inoculation while all control plants remained healthy. Microscopic examination showed the surface of infected leaves had the same acervuli, setae, and conidia as the original isolate. The pathogenicity test was repeated three times. C. cereale was previously reported as the causal agent of anthracnose on feather reed grass in US (Crouch et al. 2009). To our knowledge, this is the first report of C. cereale as the causal agent of A. nuda anthracnose in China.

First Report of Alternaria alternata Causing Leaf Spot on Avena nuda in Zhangbei, China.

Naked oats (Avena nuda L.) is an independent species of Avena, which can be used as both food and forage for rich nutritional value. In August 2019, leaf spot was observed at a naked oats planting base in Zhangbei County, Zhangjiakou City, Hebei Province. The incidence of disease was 40% to 50%. The symptoms of the lesions were chlorosis and gradually developing light brown spots with light yellow halos. The spots were irregular, enlarged and even coalesced to form large areas of necrosis on leaves. To identify the pathogen, twenty symptomatic leaves were collected, and one disease spot was isolated from each samples. Small square leaf pieces (3 to 5 mm) were excised from the junction of diseased and healthy tissues with a sterile scalpel and were sterilized with 75% alcohol for 30s, 0.1% mercuric chloride solution for 1 min, and then rinsed three times with sterile water, then transferred cultured on potato dextrose agar (PDA) at 25°C for 7 days. Four fungal isolates were obtained and purified by single-spore isolation method. All fungi have the same morphology and no other fungi were isolated. Colonies of the isolates had round margins, and thick fluffy aerial mycelia with brown coloration after 7 days on PDA. Conidiophores were brown, straight or flexuous, septate, single or in clusters. Conidia were obclavate or oval, dark brown, and size ranging from 4.61 to 15.68 × 6.61 to 35.49µm (n=100), with longitudinal and transverse septa varying from 1 to 3 and 1 to 7, respectively. The transverse median septum of the central section was especially thick. On the basis of morphological characteristics, the isolates were identified as Alternaria spp. (Simmons 2007). To further assess the identity of the species, the genomic DNA of pathogenic isolate (YM3) was extracted by CTAB protocol. The ribosomal DNA internal transcribed spacer (ITS) region, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH), the RNA polymerase II second largest subunit (RPB2), and the plasma membrane ATPase genes were amplified and sequenced with primers ITS1/4, gpd1/2, RPB2-6F/7cR and ATPDF1/ATPDR1 respectively (Nishikawa and Nakashima 2015; Woudenberg et al. 2015). Sequences of ITS, GAPDH, RPB2 and ATPase (MN646900, MT233043, MT233042, MN640794) of the isolate was 99.82%, 99.68%, 100% and 99.51% similar to the fungus A. alternata (MK461082.1, MK451978, KP124770.1, MK804115). A neighbor-joining phylogenetic tree was constructed by combining all sequenced loci in MEGA7. The isolate YM3 clustered in the A. alternata clade with 100% bootstrap support. Therefore, the pathogen was identified as A. alternata based on the morphological characteristics and molecular identification. A pathogenicity test of the A. alternata isolates was performed by placing mycelial disks (5 mm) with conidia on the surface of the first unfolding leaves of naked oats. Each leaf was inoculated with three disks. The pathogenicity test was repeated four times, and 10 leaves were inoculated in each repetition, while sterile PDA was used as the control. All treated plants were placed in a moist chamber (25°C, 16-h light and 8-h dark period). Leaf spot symptoms developed on the inoculated plants about 10 days post inoculation while all control plants remained healthy. The similar isolates were re-isolated from the inoculated and infected leaves and identified as A. alternata by DNA sequencing, fulfilling Koch's postulates. It has been reported that A. alternata can cause leaf spots on A. Sativa(Chen et al. 2020). However, to our knowledge, this is the first report of A. alternata causing leaf spots on A. nuda in China. It can be concluded that A. alternata can cause leaf spot disease of oats (A. Sativa and A. nuda). The spots disease is worthy of our attention for its harm to the production of oats.

Lymecycline reverses acquired EGFR-TKI resistance in non-small-cell lung cancer by targeting GRB2.

Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) were first-line treatments for NSCLC patients with EGFR-mutations. However, about 30 % of responders relapsed within six months because of acquired resistance. In this study, we used Connectivity Map (CMap) to discover a drug capable of reversing acquired EGFR-TKIs resistance. To investigate Lymecycline's ability to reverse acquired EGFR-TKIs resistance, two Icotinib resistant cell lines were constructed. Lymecycline's ability to suppress the proliferation of Icotinib resistant cells in vitro and in vivo was then evaluated. Molecular targets were predicted using network pharmacology and used to identify the molecular mechanism. Growth factor receptor-bound protein 2 (GRB2) is an EGFR-binding adaptor protein essential for EGFR phosphorylation and regulation of AKT/ERK/STAT3 signaling pathways. Lymecycline targeted GRB2 and inhibited the resistance of the cell cycle to EGFR-TKI, arresting disease progression and inducing apoptosis in cancer cells. Combined Lymecycline and Icotinib treatment produced a synergistic effect and induced apoptosis in HCC827R5 and PC9R10 cells. Cell proliferation in resistant cancer cells was significantly inhibited by the combined Lymecycline and Icotinib treatment in mouse models. Lymecycline inhibited the resistance of the cell cycle to EGFR-TKI and induced apoptosis in NSCLC by inhibiting EGFR phosphorylation and GRB2-mediated AKT/ERK/STAT3 signaling pathways. This provided strong support that Lymecycline when combined with EGFR targeting drugs, enhanced the efficacy of treatments for drug-resistant NSCLC.

Race and Virulence Analysis of Puccinia triticina in China During 2011 to 2013.

Wheat leaf rust, caused by Puccinia triticina, is a common fungal disease of wheat in China. In order to identify races and determine the individual virulence of isolates in different wheat-growing regions in China, leaf rust samples collected from 18 provinces in 2011 to 2013 were tested on 37 Thatcher near-isogenic lines each carrying a different single leaf rust resistance gene. A total of 158 races were identified. Races THTT (19.5%), THTS (16.9%), PHTT (7.7%), THJS (5.0%), THJT (4.2%), and PHTS (4.0%) were the most predominant races in 2011 to 2013. All of these races were avirulent to resistance genes Lr9 and Lr24. The two most frequent races, THTT and THTS, were widely distributed. The frequencies of the isolates with virulence to Lr1, Lr2c, Lr3, Lr16, Lr26, Lr17, LrB, Lr10, Lr14a, Lr3bg, Lr14b, Lr33, Lr37, and Lr50 exceeded 90%. Frequencies of virulence to Lr2a, Lr3ka, Lr11, Lr30, Lr2b, and Lr32 exceeded 70% but were less than 90%. Frequencies of virulence to Lr18, Lr21, Lr15, Lr23, Lr33+34, Lr36, Lr39, and Lr44 were below 70%, whereas the frequency of virulence to Lr25 was less than 1%. All isolates were avirulent to Lr9, Lr19, Lr24, Lr28, Lr42, Lr29, Lr38, and Lr47. The identified races and individual virulence frequencies provide a basis for selection of effective leaf rust resistance genes for use in breeding programs and can also provide information for the study of race evolution of P. triticina.