A metagenomic approach to homemade back-slopped yogurts produced in mountainous villages of Turkey with the potential next-generation probiotics

Xylella fastidiosa is a heterogenous gram-negative bacterial plant pathogen with a wide host range covering over 300 plant species. Since 2013, in Europe, the presence of the pathogen is increasing in a part of the Mediterranean area, but it causes in particular severe disease problems in olive orchards in the Southern part of Italy. Various subspecies of the pathogen were also diagnosed in natural outbreaks and intercepted ornamental plants in Europe, among them Olea europaea, Coffea arabica, and Nerium oleander. The host range of the pathogen can vary, depending on the subspecies and even the strain. The availability of fast and reliable diagnostic tools is indispensable in management strategies to control diseases caused by X. fastidiosa. To improve the reliability of the TaqMan assay, currently widely used in surveys, a triplex TaqMan assay was developed in which two specific and sensitive TaqMan assays, previously designed for X. fastidiosa, were combined with an internal control. The triplex assay exhibited the same diagnostic sensitivity as the simplex assays. In addition, the usefulness of a metagenomic approach using next-generation sequencing (NGS) was demonstrated, in which total DNA extracted from plant material was sequenced. DNA extracts from plant material free of X. fastidiosa, from artificially inoculated hosts plants or from naturally infected plants sampled in France, Spain, and Italy, or intercepted in Austria and the Netherlands, were analyzed for the presence of X. fastidiosa using the metagenomic approach. In all samples, even in samples with a low infection level, but not in the pathogen-free samples, DNA reads were detected specific for X. fastidiosa. In most cases, the pathogen could be identified to the subspecies level, and for one sample even the whole genome could be assembled and the sequence type could be determined. All results of NGS-analyzed samples were confirmed with the triplex TaqMan polymerase chain reaction and loop-mediated isothermal amplification.

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Next-generation sequencing-based Actinobacteria community associated with Callyspongia sp. from Kepulauan Seribu Marine National Park, Jakarta, Indonesia

Biodiversitas Journal of Biological Diversity ◽

10.13057/biodiv/d220913 ◽

2021 ◽

Vol 22 (9) ◽

Author(s):

DWI RETNOWATI ◽

DEDY DURYADI SOLIHIN ◽

MUNIF GHULAMAHDI ◽

YULIN LESTARI

Keyword(s):

Next Generation Sequencing ◽

Hot Spots ◽

National Park ◽

Next Generation ◽

Filter Feeding ◽

Next Generation Sequencing Ngs ◽

Metagenomic Approach ◽

Generation Sequencing

Abstract. Retnowati D, Solihin DD, Ghulamahdi M, Lestari. 2021. Next-generation sequencing-based Actinobacteria community associated with Callyspongia sp. from Kepulauan Seribu Marine National Park, Jakarta, Indonesia. Biodiversitas 22: 3702-3708. Sponges are sessile benthic filter-feeding animals, which harbor numerous microbes. The enormous community and abundance of sponge-associated actinobacteria envisage sponges as hot spots of microbes. The metagenomic approach with Next Generation Sequencing (NGS) technology was used to investigate the actinobacteria community of Callyspongia sp. collected from Kepulauan Seribu Marine National Park, Indonesia. Fourteen actinobacteria families were found associated with the sponge. Among them, Microbacteriaceae was the most dominant family. The community of actinobacteria was higher than that of the culturable method from our previous study. The research showed that a metagenomic approach using NGS technology reveals better information regarding the actinobacteria community associated with Callyspongia sp.

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