Facile Synthesis of Cu-Doped ZnO Nanoparticles for the Enhanced Photocatalytic Disinfection of Bacteria and Fungi.

In this study, Cu-doped ZnO was prepared via the facile one-pot solvothermal approach. The structure and composition of the synthesized samples were characterized by XRD (X-ray diffraction), TEM (transmission electron microscopy), and XPS (X-ray photoelectron spectroscopy) analyses, revealing that the synthesized samples consisted of Cu-doped ZnO nanoparticles. Ultraviolet-visible (UV-vis) spectroscopy analysis showed that Cu-doping significantly improves the visible light absorption properties of ZnO. The photocatalytic capacity of the synthesized samples was tested via the disinfection of Escherichia coli, with the Cu-ZnO presenting enhanced disinfection compared to pure ZnO. Of the synthesized materials, 7% Cu-ZnO exhibited the best photocatalytic performance, for which the size was ~9 nm. The photocurrent density of the 7% Cu-ZnO samples was also significantly higher than that of pure ZnO. The antifungal activity for 7% Cu-ZnO was also tested on the pathogenic fungi of Fusarium graminearum. The macroconidia of F. graminearum was treated with 7% Cu-ZnO photocatalyst for 5 h, resulting in a three order of magnitude reduction at a concentration of 105 CFU/mL. Fluorescence staining tests were used to verify the survival of macroconidia before and after photocatalytic treatment. ICP-MS was used to confirm that Cu-ZnO met national standards for cu ion precipitation, indicating that Cu-ZnO are environmentally friendly materials.

First Report of Leaf Spot on Cherry Tomatoes Caused by Exserohilum rostratum in Hainan Province, China.

Cherry tomatoes (Lycopersicon esculentum var. cerasiforme) is the main tomato variety planted in Hainan Province, China and is prized for its nutritional value and sweet taste (Zheng et al. 2020). During October 2020 to February 2021, a leaf spot disease was observed on cherry tomatoes (cultivar Qianxi) in Chengmai, Hainan Province. The disease incidence was approximately 40% in each of three fields in Yongfa (19°76'-21°08'N, 110°21'-110°51'E). Leaves were initially chlorotic before developing black, irregular-shaped lesions on the leaf margins or tips. After several days, lesions expanded along the mid-vein to encompass the entire leaf. Then, the affected leaves turned gray-brown, leading to defoliation. Severely affected leaves became dry and necrotic. Leaf tissues of 10 diseased plants samples collected from the fields were surface sterilized in 70% ethanol for 30 s, 0.1% HgCl2 for 30 s, rinsed thrice with sterile distilled water for 30 s, placed on a modified potato dextrose agar (PDA) with 30 mg/liter of kanamycin sulfate, and incubated at 28°C in the dark for 3 to 5 days. Three fungal isolates were obtained from the diseased leaves by single-sporing. The mycelia on PDA were white and later became gray or dark gray after 3 to 4 days. Conidia were rostrate, straight to slightly curved, ellipsoidal to narrowly obclavate, dark brown, protuberant with a darker and thicker wall at the basal end. Conidia were 4 to 12 distoseptate and measured 63.92 ± 5.77 × 13.47 ± 1.22 µm (n= 50) Conidiophores were single, cylindrical, dark brown, geniculate, with swollen conidiogenous cells containing a acircular conidial scar. Morphological characteristics of the isolates were similar to those of Exserohilum rostratum (Cardona et al. 2008). A representative isolate (FQY-7) was used for pathogenicity and genomic studies. Genomic DNA was extracted from the mycelium of a representative isolate (FQY-7). The internal transcribed spacer (ITS) region, actin (act), translation elongation factor 1-alpha (tef1-α), glyceraldehydes 3-phos-phate dehydrogenase (gapdh) and ß-tubulin (tub2) genes were amplified with primers ITS1/ITS4 (White et al. 1990), Act1/Act4 (Voigt and Wöstemeyer 2000), EF1-728F/EF1-986R (Carbone and Kohn 1999), Gpd-1/Gpd-2 (Berbee et al. 1999) and T1 (O'Donnell and Cigelnik 1997) + Bt2b (Glass and Donaldson 1995). The consensus sequences (GenBank Accession No. MW036279 for ITS, MW133266 for act, MW133268 for tef1-α, MW133267 for gapdh, and MW133269 for tub2) were aligned using BLAST in GenBank obtaining 100%, 100%, 99%, 100%, and 99% identity to E. rostratum strain CBS706 (LT837842, LT837674, LT896663, LT882546, LT899350). Maximum likelihood analysis based on the combined five gene sequences was conducted under 1,000 bootstrap replicates. The Phylogenetic tree showed that FQY-7 and E. rostratum were located in one clade supported with 99% bootstrap values. Pathogenicity test was performed by depositing 10-µl droplets of a conidial suspension (1 × 106 per ml) into 5 noninoculated leaves (using a sterile needle) of 10 healthy 5-month-old cherry tomato (cv. Qianxi) plants. An equal number of artificially control leaves were received only sterile water to serve as a negative control. The test was conducted three times. Plants were kept at 28°C with 80% humidity and observed for symptoms every day. Two weeks after inoculation, all the inoculated plants showed symptoms of black spots similar to those observed in the field. No symptoms were observed on the controls. FQY-7 was successfully re-isolated from the inoculated leaves and confirmed by morphological characterization and molecular assays as described herein. To the best of our knowledge, this is the first report of leaf spot of cherry tomatoes caused by E. rostratum in China. Confirming the existence of this pathogen in this area will be useful to adopt effective field management measures to control this disease on cherry tomatoes. References: Berbee, M. L., et al. 1999. Mycologia 91:964. Cardona, R. et al. 2008. Bioagro 20:141. Carbone, I. and Kohn, L. M. 1999. Mycologia 91:553. Glass, N. L., and Donaldson, G. C. 1995. Appl. Environl. Microb. 61:1323. White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. O'Donnell K., and Cigelnik, E. 1997. Mol. Phylogenet. Evol. 7:103. Voigt, K., and Wöstemeyer, J. 2000. Microbiol. Res. J. 155:179. Zheng J., et al. 2020. Guangdong Agr. Sci. 47:212. The author(s) declare no conflict of interest.

Permafrost degradation is accelerating beneath the bottom of Yanhu Lake in the Hoh Xil, Qinghai-Tibet Plateau.

Lakes on the Qinghai-Tibet Plateau (QTP) have notably expanded over the past 20 years. Due to lake water level rise and lake area expansion, the permafrost surrounding these lakes is increasingly becoming submerged by lake water. However, the change process of submerged permafrost remains unclear, which is not conducive to further analyzing the environmental effects of permafrost change. Yanhu Lake, a tectonic lake on the QTP, has experienced significant expansion and water level rise. Field measurement results indicate that the water level of Yanhu Lake increased by 2.87 m per year on average from 2016 to 2019. Cold permafrost, developed in the lake basin, was partially submerged by lake water at the end of 2017. Based on the water level change and permafrost thermal regime, a numerical heat conduction permafrost model was employed to predict future changes in permafrost beneath the lake bottom. The simulated results indicate that the submerged permafrost would continuously degrade because of the significant thermal impact of lake water. By 2100, the maximum talik thicknesses could reach approximately 7, 12, 16, and 19 m under lake-bottom temperatures of +2.0, +4.0, +6.0, and +8.0 °C, respectively. Approximately 291 years would be required to completely melt 47 m of submerged permafrost under the lake-bottom temperature of +4 °C. Note that the permafrost table begins to melt earlier than does the permafrost base, and the decline in the permafrost table occurs relatively fast at first, but then the process is attenuated, after which the permafrost table again rapidly declines. Compared to climate warming, the degradation of the submerged permafrost beneath the lake bottom occurred more rapidly and notably.

First Report of Diaporthe longicolla Causing Leaf Spot on Kalanchoe pinnata in China.

Kalanchoe pinnata (Lam.) Pers. [syn.: Bryophyllum pinnatum (Lam.) Oken] is an important medicinal agent in southern China. The succulent leaves of this plant are used in the treatment of cholera, bruises, uri-nary diseases and whitlow. In Oct. 2019, leaf spots were detected on K. pinnata plants in Chengmai County, Hainan Province, China. Lesions with brown to black margins were irregularly shaped and associated with leaf margins. Spots coalesced to form larger lesions (Fig. S1-A), with black pycnidia present in more mature lesions. Symptomatic K. pinnata were found with 10-20% incidence during the humid winters of Hainan Province. Leaf tissues of 10 symptomatic plants were collected and surface sterilized in 70% ETOH for 30s, 0.1% HgCl2 for 30 s, rinsed 3x with sterile distilled water for 30s, placed on potato dextrose agar (PDA) amended with 30mg/L of kanamycin sulfate, and incubated at 25°C in the dark for 3-5 days. Four fungal isolates were obtained using a single-spore isolation method. The colonies were floccose, dense, and white with forming on older colonies grown on PDA (Fig. S1-B-1&2). Alpha conidia exuded from ostiole, rostrate, long-beaked pycnidia in creamy-to-yellowish drops. Alpha conidia were hyaline, ellipsoidal, separated and averaged 6.3µm (SD ± 1.13) long × 1.9µm (SD ± 0.33) wide (n=50). Beta conidia were not seen. The morphological characteristics matched the previous description of Diaporthe longicolla (syn. Phomopsis longicolla) (Hobbs et al. 1985). Mycelial genomic DNA of the representative isolate LDSG3-2 was extracted as template. The internal transcribed spacer (ITS) , translation elongation factor 1α gene (TEF) and ß-tubulin (TUB2) regions were amplified. These loci were amplified using primer pairs ITS4/ITS5 (White, et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999) and Bt2a/Bt2b (Glass and Donaldson 1995), respectively. A BLAST search of GenBank showed ITS (MN960195), TEF (MN974483) and TUB2 (MN974482) sequences of the isolate were 99%, 100%, and 99% homologous with D. longicolla strains DL11 (MF125048, 557/563 bp), D55 (MN584792, 347/347 bp) and DPC-HOH-32 (MK161506, 502/504 bp). Maximum likelihood trees based on concatenated nucleotide sequences of the three genes were constructed using MEGA 7.0, and bootstrap values indicated the isolate was D. longicolla (Fig. S1-D). Pathogenicity testing was performed using isolate LDSG3-2 by depositing 5µl droplets of a conidial suspension (1 × 106 ml-1) into 5 artificially wounded leaves (using a sterile needle) of 10 healthy 3-month-old K. pinnata plants. An equal number of artificially wounded control leaves were inoculated with sterile water to serve as a negative control. The test was conducted three times. Plants were kept at 25°C in 80% relative humidity and observed for symptoms. Two weeks after inoculation, no symptoms were observed on control plants (Fig. S1-C-1) and all inoculated plants showed symptoms (Fig. S1-C-2) similar to those observed in the field. The fungus was re-isolated from the infected tissues and showed the same cultural and morphological characteristics of the strain inoculated and could not be isolated from the controls fulfilling Koch's postulates. To our knowledge, this is the first report of leaf spot on K. pinnata caused by D. longicolla in China. This disease is of concern since Phomopsis diseases are common in K. pinnata fields and can cause significant reduction in yield. References: White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. DOI: 10.1016/0167-7799(90)90215-J Carbone, I., and Kohn, L. M. 1999. Mycologia. 91:553. DOI: 10.2307/3761358 Glass, N. L., and Donaldson, G. C. 1995. Appl. Environ. Microbiol. 61:1323. DOI: 10.1002/bit.260460112 Hobbs, T. W. et al. 1985. Mycologia. 77: 535. DOI: 10.2307/3793352.

Risk assessment of potential thaw settlement hazard in the permafrost regions of Qinghai-Tibet Plateau.

Climate warming could exacerbate the occurrence of thaw settlement hazard in the permafrost regions of the Qinghai-Tibet Plateau (QTP), which would threaten the stability of engineering infrastructure in cold regions. The risk associated with permafrost settlement, valuable for the regional sustainable development, remains poorly assessed or understood on the QTP. In this study, three common Geo-hazard indices were used to assess the settlement risks in the permafrost regions of the QTP, including the settlement index, the risk zonation index, and the allowable bearing capacity index. However, large spatial differences existed in simulating the risk maps by using the abovementioned Geo-hazard indices. Hence, we developed a combined index (Ic) by integrating the three indices to reduce the uncertainty of the simulations. The results indicated that the ground ice is a critical factor for assessing the settlement risk in permafrost regions. We also applied the Ic to assess the settlement risk along the Qinghai-Tibet Railway (QTR). The proportion of low-risk area along the QTR would be the highest (45.38%) for the future periods 2061-2080 under Representative Concentration Pathway 4.5. The medium-risk area combined with the high-risk area would be accounted for more than 40%, which were located at the boundary of the present permafrost regions. Therefore, the corresponding adaptation measures should be taken to reduce the potential economic losses caused by the high-risk regions to the infrastructure. Overall, the results would present valuable references for engineering design, construction and maintenance, and provide insights for early warning and prevention of permafrost thaw settlement hazard on the QTP.

First Report of Colletotrichum gloeosporioides Causing Anthracnose on Canna edulis Ker in China.

Canna edulis Ker has been an important economic plant in southern China. The tuberous stems are the most valued plant part and in the past were routinely used as animal feeds. In June 2019, leaf spot disease were detected on C. edulis plants in Chengmai, Hainan Province. Symptoms of the disease were characterized by oval-shaped, initially pale to yellow lesions that become necrotic (brown) with yellow borders, As the lesions expanded, the disease could encompass the entire leaves. which were seen as concentric rings typical of anthracnose disease (FigS1-A). A survey of C. edulis plants revealed that the disease caused serious damage during the summer in Hainan Province, with 50 to 60% incidence in plants. To isolate the pathogen, ten pieces of diseased leaf samples were plated and seven pieces yielded fungal colonies after 5 to 6 days of incubation at 25 °C. The Colonies were single-spored to obtain pure cultures. Pure cultures on potato dextrose agar (PDA) appear white to gray, with white margins and aerial hyphae, and the reverse of the colonies was gray to brown (FigS1-B). Conidia were single-celled, hyaline, cylindrical to slightly curved with a rounded apex and truncated base that measured 13.3 to 18.1(length) × 3.7 to 5.5 (width) µm (n=50) (FigS1-C). The morphological characteristics and measurements of this fungal pathogen matched the previous descriptions of Colletotrichum gloeosporioides (Prihastuti et al. 2012). Isolate JO-3 was identicated by molecular analysis, sequences of the internal transcribed spacer (ITS), actin (ACT) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) regions were obtained. These loci were amplified from isolates using the primer pairs ITS1/ITS4 (Mills et al. 1992), ACT-512F/ ACT-783R (Weir et al. 2012) and GDF/GDR (Templeton et al. 1992) respectively. A BLAST search of GenBank showed that the ITS (MN913584), ACT(MN919196) and GAPDH (MN919195) sequences of the isolate were 99% , 100% and 100% homologous with C. gloeosporioides (GenBank accession nos. MH930419, JX009931 and KX885158). Maximum likelihood trees based on concatenated sequences of the three genes were constructed using MEGA7.0. The results showed the strains isolated from C. edulis were closely related to C. gloeosporioides, as supported by high bootstrap values (FigS1-D). Pathogenicity test was performed with isolate JO-3 by depositing 10-µl droplets of a suspension (1 × 106 conidia/ml) on the surfaces of five artificially wounded leaves (a small hole made with a pushpin) of ten healthy 1-year-old C. edulis plants. An equal number of control leaves were inoculated with sterile water to serve as a negative control. The test was conducted three times. Plants were kept at 25°C with 80% humidity and observed for symptom every day. One weeks after inoculation, all the inoculated plants showed symptoms of yellow sunken spots similar to those observed in the field. No symptoms were observed on the controls. The fungus re-isolated from the infected tissues showed the same cultural and morphological characteristics of the strain inoculated, fulfilling Koch's postulates. C. gloeosporioides was previously reported as the causal agent of anthracnose on Hymenocallis littoralis (Zhao et al. 2019), Abelmoschus esculentus (L.) Moench (Shi et al. 2019) and Sorbaria sorbifolia (Li et al. 2019) in China. To our knowledge, this is the first report of anthracnose on C. edulis caused by C. gloeosporioides in China. This disease can seriously affect the yield and quality of C. edulis in China. Given its wide host range, C. gloeosporioides has great potential to become an economically important plant pathogen. The project was partially funded by Hainan Provincial Research Institute of technology development projects (Collection, Evaluation and Domestication Cultivation of Wild Vegetable Germplasm Resources in Hainan), Hainan Provincial Key Laboratory for Vegetables and Biology，Hainan Provincial Engineering Research Center for Melon and Vegetable Breeding, Major scientific and technological projects in Hainan Province(ZDKJ2017001)，Key R & D projects in Hainan Province (ZDYF2019066), The third Survey and Collection of Crop Germplasm Resources in China. References: Mills, P. R., et al. 1992. FEMS Microbiol Lett. 98:137-144 Weir, B. S., et al. 2012. Stud. Mycol. 73:115. Templeton, M.D. et al. 1992. Gene. 122:225. Prihastuti, H., et al. 2009. Fungal Divers. 39:89 C. D. Zhao, et al. 2019. Plant Dis.103:3286 Y. X. Shi, et al. 2019. Plant Dis.103:303 X. Y. Li, et al. 2019. Plant Dis.103:242.

First Record of Leaf Spot Disease on Costus speciosus Caused by Nigrospora oryzae in Hainan, China.

Costus speciosus (Koen.) Smith has been an important medicinal agent in the various traditional and folk systems of medicine in southern China. In September 2018, leaf spot disease was detected on C. speciosus plants in Chengmai County, Hainan Province. A survey of C. speciosus plants revealed that the disease caused serious damage during the typhoon season of September to November in Hainan Province, with 80 to85% incidence in plants. Early symptoms were yellow-to-brown, irregular-shaped lesions on the leaf margin or tip. After several days, lesions expanded along the mid-vein until the entire leaf was destroyed. Then, the infected leaves turned gray brown, leading to defoliation. Heavily infected leaves became dry and died. The pathogen was consistently isolated from the lesions and pure single-spore cultures were obtained. Twenty pieces of diseased leaf samples were plated and seven pieces yielded fungal colonies after 5 to 6 days of incubation at 25 °C. Colonies on potato dextrose agar (PDA) were white and later became gray to black. Conidia were unicellular, terminal, black, elliptical that measured 10 to 13 (length) × 12 to 16 (width) µm (n=30), growing aerial mycelium covering the entire petri dish (9 cm in diameter). The morphological characteristics and measurements of this fungal pathogen matched the previous descriptions of Nigrospora oryzae (Wang et al. 2017). To confirm identity the internal transcribed spacer (ITS) region of the ribosomal DNA was amplified using primers ITS1/ITS4 (Mills, P. R., et al. 1992), and the 530-bp product (GenBank Accession No. MK280694) of the ITS showed 99% sequence identity to N. oryzae isolates TLFa21 (GenBank Accession No. MH790146) and xsd08022 (GenBank Accession No. EU918714). Pathogenicity tests were conducted. Three leaves of three C. speciosus plants were wounded and inoculated with mycelial plugs (5×5mm) , and an additional five plants were inoculated with PDA plugs as a control. All plants were placed in the field and temperature ranged from 23 to 29°C. Ten days after inoculation, all the inoculated plants showed typical leaf spot symptoms, a yellow-to-brown mildew appeared at the points of inoculation. No symptoms were observed on the controls. The fungus was re-isolated from the infected tissues, fulfilling Koch's postulates. N. oryzae was previously reported as the causal agent of leaf spot on cotton (Zhang. et al. 2012), dendrobium candidum (Wu. et al. 2014) and Aloe vera (Zhai. et al. 2013) in china. To our knowledge, this is the first report of leaf spot of C. speciosus caused by N. oryzae in China. The project was partially founded by Hainan Provincial Research Institute of technology development projects (Screening and application of endophytic bacteria with high resistance to Fusarium Wilt of Sauropus androgynus), Hainan Provincial Key Laboratory for Vegetables and Biology，Hainan Provincial Engineering Research Center for Melon and Vegetable Breeding, Major scientific and technological projects in Hainan Province(ZDKJ2017001)，Third Survey and Collection of Crop Germplasm Resources in China, Collection, identification and preservation of pathogenic bacteria of inverted season vegetable in Hainan. References: L. F. Zhai., et al.2013. Plant Dis.97:1256 L. X. Zhang., et al.2012. Plant Dis.102:2029 J. B. Wu., et al.2014. Plant Dis.98:996 Mills, P. R., et al. 1992. FEMS Microbiol Lett. 98:137-144 Wang et al. 2017. Persoonia 39: 118-142.

Soil organic carbon and total nitrogen pools in permafrost zones of the Qinghai-Tibetan Plateau.

There are several publications related to the soil organic carbon (SOC) on the Qinghai-Tibetan Plateau (QTP). However, most of these reports were from different parts of the plateau with various sampling depth. Here, we present the results from a systematic sampling and analysis of 200 soil pits. Most of the pits were deeper than 2 m from an east-west transect across the plateau. The SOC and total nitrogen (TN) pools of the 148 × 104 km2, the area of the permafrost zone, for the upper 2 m soils calculated from the vegetation map were estimated to be 17.07 Pg (interquartile range: 11.34-25.33 Pg) and 1.72 Pg (interquartile range: 1.08-2.06 Pg), respectively. We also predicted the distribution of land cover types in 2050 and 2070 using decision tree rules and climate scenarios, and then predicted SOC and TN pools of this region. The results suggested that the SOC and TN pools will decrease in the future. The results not only contribute to the carbon and nitrogen storage and stocks in the permafrost regions as a whole but most importantly, to our knowledge of the possible changes of C and N storage on the QTP in the future.