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Plant Disease ◽  
2022 ◽  
Author(s):  
Susu Fan ◽  
Fangyuan Zhou ◽  
Xueying Xie ◽  
Xinjian Zhang ◽  
Tielin Wang

Chinese yam (Dioscorea opposita Thunb.), which belongs to the family of Dioscorea, is widely naturalized throughout China, due to its high economic and medicinal value. Since 2019, water-soaked lesions were frequently observed in the underground tubers of Chinese yam located in Xinyang City, Henan Province. To identify the causal agent, ten pieces of tissue from the underground tubers with disease symptoms were collected. Those infected tissues (5×5 mm) were crushed in 500 μL sterilized water after surface sterilization and streaked onto Luria-Bertani agar plates. Pale-yellowish, rod-shaped, slimy single bacterial colonies with smooth margin were observed after 24 hours of incubation, and three bacterial colonies (named CY-1, CY-2 and CY-3) were randomly selected for further biochemical and molecular characterization. These bacteria were gram-negative with the cell length of 1.0 to 3.0 μm, width of 0.5 to 1.0 μm, and with peritrichous flagella. Subsequently, the bacteria were biochemically analyzed through BIOLOG (Hayward, CA) and identified as Pantoea agglomerans with 99% probability. Furthermore, the phylogenetic analysis results based on 16S rDNA, DNA gyrase subunit B (gyrB), and RNA polymerase sigma factor (rpoD) showed these three isolates were most closely related to P. agglomerans. The sequence of 16S rDNA, gyrB and rpoD of each strain was submitted to GenBank with the accession numbers MZ541065 MZ541066 and MZ541067 for 16S rDNA; MZ669846, MZ669847 and MZ669848 for gyrB; MZ669849, MZ669850 and MZ669851 for ropD. Pathogenicity test was performed to complete Koch’s postulates. Tubers of Chinese yam were wounded by sterile needle and inoculated with 500 μL 108 CFU/mL bacterial suspension. Sterilized water was used as a control. Five pots were inoculated for each isolate. Water-soaked lesions appeared after five days incubation at 25°C in a biochemical incubator and no lesions were observed on the control. Bacteria re-isolated from the lesions were similar in phenotypic and molecular characteristics to the original isolates. In brief, based on colony morphology, biochemical tests, characteristic sequence analysis, and pathogenicity verification, the pathogen responsible for the soft rot of Chinese yam in Henan Province was identified as P. agglomerans. In China, P. agglomerans has been reported to associate with bacterial soft rot on Chinese cabbage (Guo et al., 2020). To our knowledge, this work is the first report of bacterial rot caused by P. agglomerans on Chinese yam.


Plant Disease ◽  
2022 ◽  
Author(s):  
Jinhui Wang ◽  
Yuxiang Lu ◽  
Wanxin Han ◽  
Lijun Fu ◽  
Xiaoqing Han ◽  
...  

In August 2020, ginger (Zingiber officinale) rhizomes (cv. Mianjiang) showing soft rot symptoms were observed in a field in Tayang Village, Fengrun District, Tangshan, Hebei Province (North China). The disease incidence in that field (15 ha in size) was more than 20%. Symptomatic rhizomes (brown and water-soaked) were surface-sterilized in 75% ethanol for 60 sec and then three successive rinses with sterile distilled water. Rhizomes were cut into pieces ca. 0.5 cm in length, and then were soaked in 500 µl 0.9% saline for 20 min. Aliquots (20 μl) of three tenfold dilutions of the tissue specimen soaking solution were plated onto the lysogeny broth (LB) medium. And LB plates were incubated at 28°C for 24 h. Five single colonies were picked from each LB plate and restreaked three times for purity. Endophytic bacteria were also isolated from asymptomatic rhizomes as control. The bacterial gDNA was extracted using the EasyPure Bacteria Genomic DNA Kit (TransGen Biotech, Beijing, China). The 16S rDNA region was amplified by PCR using the universal primer pair 27F/1492R (Weisburg et al. 1991) and sequenced. The results of BLASTN against NCBI nr of the 16S rDNA amplicons suggested that the most isolates (8/10) obtained from the rotten rhizomes belonged to the genus Pectobacterium, and few isolates (2/10) were Enterobacter spp.. Only Enterobacter spp. were isolated from asymptomatic rhizomes. Since all Pectobacterium isolates showed identical 16S rDNA sequence, thus, only two isolates were selected for further analysis. Pectobacterium isolates TS20HJ1 and TS20HJ2 (MZ853520, MZ853521) represent isolates from two plant individuals. To determine the species of the rhizome rot Pectobacterium isolates, multi-locus sequence analysis (MLSA) was performed with five housekeeping genes acnA, icdA, mdh, proA and rpoS (MZ994717-MZ994726) (Ma et al. 2007; Waleron et al. 2008), and a phylogenetic tree was reconstructed using RAxML v8.2.12 (github.com/stamatak/standard-RAxML). No sequence variation was observed at any MLSA locus between the two isolates. The result of phylogenetic analysis showed that the ginger rhizome isolates clustered with P. brasiliense type strain IBSBF1692T (Duarte et al. 2004; Nabhan et al. 2012). Ginger seedlings (cv. Mianjiang) were inoculated with the isolate TS20HJ1 by injecting 10 µl of bacterial suspensions (108 CFU·mL-1) into the rhizomes, or injected with 10 µl of 0.9% saline solution as control. The seedlings were grown at 28°C and 50% relative humidity. Ten days after inoculation, only the bacteria-inoculated rhizomes showed diseased symptoms resembling to those observed in the field. Bacterial colonies were obtained from the infected rhizomes and were identified with MLSA gene sequencing, fulfilling Koch’s postulates. P. brasiliense causes soft rot of a wide range of economically important crops (Oulghazi et al. 2021). To our knowledge, this is the first report of P. brasiliense causing rhizome rot of ginger in China. The rhizome rot caused 20-25% yield loss on average in Tangshan region in 2020, which poses a significant threat to the local ginger farming. Further research on epidemiology and disease management options is needed.


Plant Disease ◽  
2022 ◽  
Author(s):  
Xinhua Ding ◽  
Chongchong Lu ◽  
Mingxia Hao ◽  
Lingguang Kong ◽  
Lulu Wang ◽  
...  

Rice (Oryza sativa L.) is the largest grain crop, accounting for about 40 % of the total grain production in China. In mid-July 2021, bacterial leaf streak-like disease emerged in rice varieties Chunyou584 and Yongyou2604 in Linyi city, Shandong Province, China. Disease incidences of the disease ranged from 80% to 90% in the surveyed fields. Infected rice leaves displayed dark green to yellowish-brown water-soaked thin streaks, and a large amount of beaded yellow oozes were observed on the lesions. After drying, there were gelatinous granules that were not easy to fall off and spread between leaf veins (Fig.S1A). According to the field symptoms of this disease, it was preliminarily suspected to be rice bacterial leaf streak caused by Xanthomonas oryzae pv. oryzicola (Xoc), which is a guaranteed disease in China. To isolate the causal agent, leaf discs (~1 cm2) of diseased leaves were collected from the margins of the lesions, surface sterilized and ground into pieces in sterile double distilled water. The 10-3, 10-4 and 10-5 dilutions were spread onto peptone sugar agar (PSA) and incubated at 28°C for 36 hours. Yellow mucous bacterial colonies were consistently obtained on PSA medium. To identify the pathogen, fragments of the 16S rDNA, leuS and rpoB were amplified and sequenced using the primers previously reported (Yu et al. 2021). Three strains (LY01, LY02 and LY03) showed identical colony morphology and LY01 was used for further analyses. Sequence analyses showed that the fragments of 16S rDNA (955 bp, GenBank accession number: OK261898), leuS (755 bp, GenBank accession number: OK298387) and rpoB (926 bp, GenBank accession number: OK298388) of strain LY01 shared 99.16%, 99.46% and 100% similarities with those of Pantoea ananatis TZ39 (GenBank accession numbers: CP081342.1 for 16S rDNA, MW981338.1 for leuS and MW981344.1 for rpoB), respectively, which suggest the pathogenic bacterial strain LY01 isolated is P. ananatis. In addition, the single colony of P. ananatis LY01 was shown as Fig. S2B. Furthermore, pathogenicity tests were also performed according to the following steps. Bacterial suspension at OD600=0.1 was inoculated into eight rice leaves of four healthy rice plants (Chunyou 584) at 25-33°C and 60%-80% relative humidity in the field using a clipping method (Yang et al. 2020) or spraying methods, and sterile distilled water was as negative control. The clipped leaves (Fig. S1B) and spray-inoculated leaves (Fig. S1C) showed dark green water-soaked streaks at 14 days after inoculation, respectively, which showed similar symptoms with those samples collected from the fields (Fig. S1A). On contrary, the control rice leaves remained healthy and symptomless (Fig. S2A). The bacterium was re-isolated in the inoculated rice leaves and the re-isolated bacterial isolates, which was confirmed by sequencing 16S rDNA, leuS and rpoB, incited the same symptoms as in fields, which fulfills Koch’s postulates. In the past decade, P. ananatis was reported to result in grain discoloration and leaf blight in China (Yan et al. 2010; Xue et al. 2020, Yu et al. 2021), which could result in 40% - 60% yield losses. To our best knowledge, this is the first report of the bacterial leaf streak-likely disease occurred in Shandong Province caused by P. ananatis, so we named it as Pantoea leaf streak of rice. Although P. ananatis was also reported in Zhejiang province and Jiangxi province, which caused leaf streak lesions on rice, the disease symptoms are completely different from those of Pantoea leaf streak of rice. To the best of our knowledge, this is the first report of Pantoea leaf streak of rice caused by P. ananatis. This study provides sloid evidence that Pantoea leaf streak of rice in Eastern China can be caused by the new pathogen, P. ananatis, rather than Xoc as traditionally assumed. Disease development and quarantine of the new Pantoea leaf streak of rice disease caused by P. ananatis on rice need more attention in the near future.


2022 ◽  
Author(s):  
Jing Lin ◽  
Ke Huang ◽  
Jing-Yu Huang ◽  
Yuan-Ru Xiong ◽  
Meng-Meng Wei ◽  
...  

Abstract A Gram-stain-negative, aerobic, chemoheterotrophic bacterium, characterized with rod shape and mobility, designated as LST-1T, was isolated from wild Stevia rebaudiana Bertoni and subjected to polyphasic taxonomic analysis. The LST-1T strain grew optimally at 37 °C and pH 6.0–7.0 in the presence of 0.5 % (w/v) NaCl. Phylogenetic sequence analysis based on 16S rDNA from LST-1 indicated that it is close to Lelliottia jeotgali (99.85%), Lelliottia nimipressuralis (98.82%), and Lelliottia amnigena (98.54%). Multi-locus sequence typing analysis of concatenated partial recA, atpD, and infB was performed to improve resolution, and clear distinctions between the closest related type strains were exhibited. Meanwhile, the results from average nucleotide identify analyses and DNA–DNA hybridization with four species (16S rDNA similarity > 98.65%) were less than 90% and 40% respectively, verifying the distinct characteristics from other species of Lelliottia, The cellular fatty acid profile of the strain consisted of C16:0, Summed Feature3, and Summed Feature8 (may be 16:1 w6c/16:1 w7c and 18:1 w6c) as major components. The major polar lipids included phosphatidylethanolamine, phosphatidylglycerol, aminophospholipid, three non-characteristic phospholipids, and a non-characteristic lipid. The genome of LST-1T is 4,611,055 bp, with a DNA G + C content of 55.02%. Combination of several phenotypic, chemotaxonomic, and genomic characteristics proved that the LST-1T strain does represent a novel genus, for which the name Lelliottia sp. LST-1 was proposed. The type strain is LST-1T (= CGMCC 1.19175T = JCM 34938T).


Animals ◽  
2022 ◽  
Vol 12 (2) ◽  
pp. 149
Author(s):  
Haibo Fu ◽  
Wenjing Li

The division of hard and soft feces is an effective digestion strategy in the order Lagomorpha. Although previous studies have reported that hard and soft feces differ in morphology and component, the discrepancy in the microbiome remains unclear. This study explored the microbiomes of hard and soft feces in plateau pikas by sequencing the V3 and V4 regions of 16S rDNA. We found that hard feces harbored higher Firmicutes, while soft feces harbored higher Akkermansia. Increased rare bacterial taxa were observed in hard feces compared with soft feces. Moreover, hard and soft feces displayed a greater difference in terms of core operational taxonomy units (OTUs) compared to the total OTUs. The soft feces showed enhancements in all predicted Kyoto Encyclopedia of Genes and Genomes (KEGG) functions, indicating an advancing microbial metabolism compared to hard feces. The significantly upregulated pathways in soft feces were mainly enriched in metabolism of energy and carbohydrate, glycan biosynthesis, cofactors and vitamins, and amino acids—all of which are associated with increased contents of microbial proteins, vitamins, and short-chain fatty acids. Our study reports, for the first time, the differential microbiomes between hard and soft feces of pikas and provides direction for the future studies on cecotrophy.


2022 ◽  
Vol 9 ◽  
Author(s):  
Laura Maretto ◽  
Saptarathi Deb ◽  
Samathmika Ravi ◽  
Claudia Chiodi ◽  
Paolo Manfredi ◽  
...  

The microbial diversity is, among soil key factors, responsible for soil fertility and nutrient biogeochemical cycles, and can be modified upon changes in main soil physicochemical properties and soil pollution. Over the years, many restoration techniques have been applied to restore degraded soils. However, the effect of these approaches on soil microbial diversity is less understood and thus requires more investigation. In this study, we analyzed the impact, on soil microbial diversity of a patented novel technology, used to restore degraded soils. Soil samples were collected from three nearby sites located in Borgotrebbia, Piacenza, Italy, and categorized as reconstituted, degraded, and agricultural soils. After total soil DNA extraction, 16S rDNA multi-amplicon sequencing was carried out using an Ion GeneStudio S5 System to compare soils’ bacterial community profiles. Sequenced reads were processed to assign taxonomy and then key microbial community differences were identified across the sampling sites. Species diversity featured significant abatement at all rank levels in the degraded soil when compared to the agricultural control. The 5 year restoration technique showed full recovery of this index at the genus level but not at the phylum level, displaying a rank-dependent gradient of restored richness. In parallel, the abundance of genes involved in the nitrogen (N) biogeochemical cycle was assessed using quantitative Real-Time PCR (qPCR). Total DNA content was significantly higher (p < 0.05) in degraded (μ = 12.69 ± 2.58 μg g−1) and reconstituted (μ = 11.73 ± 1.65 μg g−1) soil samples when compared to the agricultural soil samples (μ = 2.39 ± 0.50 μg g−1). The taxonomic diversity of each soil site was significantly different, with some instances unique of the agricultural soil even at the phylum level. The analysis of N functional genes showed that the relative abundance of bacterial amoA (p < 0.05) and nosZ (p < 0.01) genes were significantly lower in the agricultural than in the reconstituted and degraded soils. We concluded that the application of the soil reconstitution technique appears to enhance the active microbial community, with distinct diversity and functionality towards genes involved in N biogeochemical cycle, as compared to both the degraded and the agricultural soil.


2022 ◽  
Vol 79 (2) ◽  
Author(s):  
Chairmandurai Aravindraja ◽  
Dharmaprakash Viszwapriya ◽  
Alaguvel Valliammai ◽  
Shunmugiah Karutha Pandian

2022 ◽  
Vol 9 (1) ◽  
Author(s):  
Honghai Kuang ◽  
Xi Ye ◽  
Zhiyi Qing

This study is based on the processing of computed microtomography images of rock samples. In this study, a finite automation is constructed using the grey value, red-green-blue (RGB) value and Euler number of polarized images of carbonate rocks from the Jingfengqiao–Baidiao area. The finite automaton is used to perform black and white binary processing of the polarized images of the carbonate rocks. The porosity of the carbonate rock is calculated based on the black and white binarization processing results of the polarized images of the carbonate rocks. The obtained porosity is compared with the carbonate porosity obtained by use of the traditional carbonate research method. When the two porosities are close, the image processing threshold of the finite automata is considered to be credible. Based on the finite automata established using the image processing threshold, the black and white binary images of the polarized images of the carbonate rocks are used to establish a rock pore image using I mage J2X. The polarized images of the carbonate rocks are classified according to their RGB values using the finite automata for the porosity classification, and the obtained images are used as textures to paste onto a cube to construct a three-dimensional data model of the carbonate rocks. This study also uses 16S rDNA analysis to verify the formation mechanism of the carbonate pores in the Jingfengqiao–Baidiao area. The results of the 16S rDNA analysis show that the pores in the carbonate rocks in the Jingfengqiao–Baidiao area are closely related to microorganisms, represented by denitrifying bacteria.


RSC Advances ◽  
2022 ◽  
Vol 12 (2) ◽  
pp. 985-997
Author(s):  
Harinee Subramanian ◽  
Muthukumar Krishnan ◽  
Ashok Mahalingam

Drug-resistant superbugs (DRS) were isolated from hospital sewage waste and confirmed by a 16S rDNA molecular technique as B. filamentosus, B. flexus, P. stutzeri, and A. baumannii.


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