First Report of Streptomyces brasiliscabiei Causing Common Scab on Potato in China.

Severe typical deep-pitted lesions of Potato Common Scab (PCS) disease were observed in two locations in China, Dingxi, Gansu Province, and Shuozhou, Shaanxi Province, in 2021. Potato farms in Dingxi growing cultivar Huangxin 226 (26 hectares) exhibited a scab disease incidence of 10%, while cultivar Jinshu 15# (4 hectares) in Shuozhou showed a disease incidence of 30% (Fig. 1). During harvest, tubers displaying PCS symptoms were collected for pathogen isolation. To obtain pathogen isolates, surface-sterilized tuber tissue with scab lesions was ground in sterile water, serially diluted, and plated onto ISP5 agar medium plates (Handique et al. 2022). Five pure colonies of Streptomyces isolate were obtained, designated as ZRIMU1503, ZRIMU1502, ZRIMU1320, ZRIMU1321and ZRIMU791. Genomic DNA was extracted and sequenced using Illumina technology. Sequencing data of the 5 isolates were uploaded to NCBI GenBank and annotated (Accession numbers: JBBAYL010000000, JBBAYM010000000, JBBAYN010000000, JBBAYO010000000 and JBBAYP010000000, respectively) using the PGAP pipeline (Tatusova et al. 2016). Average Nucleotide Identity (ANI) values (97.52 %, 97.53 %,97.54 %,97.57 % and 97.52 %, respectively) indicated the identity of the five isolates to the type strain S. brasiliscabiei IBSBF 2867T. Additionally, pairwise comparisons of Digital DNA Hybridization (DDH) value (76.2%, 76.3%, 76.4%, 76.4% and 76.2% respectively) of all the Streptomyces type strains show the highest identity to S. brasiliscabiei IBSBF 2867T. Twelve housekeeping genes (acnA, atpD, dnaN, gap, gyrA, gyrB , infB, mdh, recA, rplB, rpoB, and trpB) were extracted from the genome sequence of the five isolates to construct a multi-locus sequence analysis (MLSA) tree. The evolutionary distance of the five isolates was constructed using MEGAX software (Kumar et al., 1994), along with other Streptomyces strains that are known to cause PCS. The resulting cladogram demonstrated the isolated strains clustered together with S. brasiliscabiei IBSBF 2867T (Fig.2). Koch's postulates were fulfilled by inoculating a perlite potting mix with spore suspensions of each isolate (104 CFU/ml), planting tubers (cv. Favorita), and reproducing PCS symptoms at harvest after three months. Negative control received water treatment. The plants were kept in greenhouse with 12 h of light per day and irrigated regularly. The experiment was repeated twice, once in April 2022 and again in April 2023. On harvest, all five isolates exhibited development of severe symptoms of PCS (Fig.1), while the negative controls had no lesions. The pathogen was reisolated from the lesions and confirmed to be identical to the original isolate by 16S rRNA gene sequences. To our knowledge, this is the first report of S. brasiliscabiei causing PCS in China. S. brasiliscabiei was identified as a new species to cause PCS in Brazil and was identified based on morphology, pathogenicity, and genetic features (Corrêa et al. 2021). Multiple pathogen-causing PCS has been recognized in China and a surge of disease incidence in potato fields has been reported (Handique et al. 2022; Wu et al. 2023). S. brasiliscabiei causes severe symptoms which makes potatoes unmarketable. The disease epidemiology of this pathogen needs to be investigated.

Isolation, characterization and application of noble bacteriophages targeting potato common scab pathogen Streptomyces stelliscabiei.

Bacteriophages have emerged as promising alternatives to pesticides for controlling bacterial pathogens in crops. Among these pathogens, Streptomyces stelliscabiei (syn. S. stelliscabiei) is a primary causative agent of potato common scab (PCS), resulting in substantial global economic losses. The traditional management methods for PCS face numerous challenges, highlighting the need for effective and environmentally friendly control strategies. In this study, we successfully isolated three novel bacteriophages, namely Psst1, Psst2, and Psst4, which exhibited a broad host range encompassing seven S. stelliscabiei strains. Morphological analysis revealed their distinct features, including an icosahedral head and a non-contractile tail. These phages demonstrated stability across a broad range of temperatures (20-50°C), pH (pH 3-11), and UV exposure time (80 min). Genome sequencing revealed double-stranded DNA phage with open reading frames encoding genes for phage structure, DNA packaging and replication, host lysis and other essential functions. These phages lacked genes for antibiotic resistance, virulence, and toxicity. Average nucleotide identity, phylogenetic, and comparative genomic analyses classified the three phages as members of the Rimavirus genus, with Psst1 and Psst2 representing novel species. All three phages efficiently lysed S. stelliscabiei in the liquid medium and alleviated scab symptom development and reduced pathogen abundance on potato slices. Furthermore, phage treatments of radish seedlings alleviated the growth inhibition caused by S. stelliscabiei with no disease symptoms. In soil potted experiments, phages significantly reduced disease incidence by 40%. This decrease is attributed to a reduction in pathogen density and the selection of S. stelliscabiei strains with reduced virulence and slower growth rates in natural environments. Our study is the first to report the isolation of three novel phages that infect S. stelliscabiei as a host bacterium. These phages exhibit a broad host range, and demonstrate stability under a variety of environmental conditions. Additionally, they demonstrate biocontrol efficacy against bacterial infections in potato slices, radish seedlings, and potted experiments, underscoring their significant potential as biocontrol agents for the effective management of PCS.