Investigation of collections of phytopathogenic bacteria has revealed some strains distinct from known Pectobacterium spp. We report here the draft genome sequences of five such strains, isolated during the period of 1947 to 2012. Based on comparative genomics, we propose a new candidate genomospecies of the genus Pectobacterium , “ Candidatus Pectobacterium maceratum.”

Increasing the accuracy of pathogen identification and reducing the duration of analysis remain relevant for modern molecular diagnostics up to this day. In laboratory and clinical practice, detection of pathogens mostly relies on methods of nucleic acid amplification, among which the polymerase chain reaction (PCR) is considered the “gold standard.” Nevertheless, in some cases, isothermal amplification methods act as an alternative to PCR diagnostics. Upon more than thirty years of the development of isothermal DNA synthesis, the appearance of loop-mediated isothermal amplification (LAMP) has enabled new directions of in-field diagnostics of bacterial and viral infections. This review examines the key characteristics of the LAMP method and corresponding features in practice. We discuss the structure of LAMP amplicons with single-stranded loops, which have the sites for primer annealing under isothermal conditions. The latest achievements in the modification of the LAMP method are analyzed, which allow considering it as a unique platform for creating the next-generation diagnostic assays.

Background: Advantages of loop-mediated isothermal amplification in molecular diagnostics allow to consider the method as a promising technology of nucleic acid detection in agriculture and medicine. A bioinformatics tool that provides rapid screening and selection of target nucleotide sequences with subsequent taxon-specific primer design toward polymorphic orthologous genes, not only unique or conserved common regions of genome, would contribute to the development of more specific and sensitive diagnostic assays. However, considering features of the original software for primer selection, also known as the PrimerExplorer (Eiken Chemical Co. LTD, Tokyo, Japan), the taxon-specific primer design using multiple sequence alignments of orthologs or even viral genomes with conservative architecture is still complicated. Findings: Here, MorphoCatcher is introduced as a fast and simple web plugin for PrimerExplorer with a clear interface. It enables an execution of multiple-alignment based search of taxon-specific mutations, visual screening and selection of target sequences, and easy-to-start specific primer design using the PrimerExplorer software. The combination of MorphoCatcher and PrimerExplorer allows to perform processing of the multiple alignments of orthologs for informative sliding-window plot analysis, which is used to identify the sequence regions with a high density of taxon-specific mutations and cover them by the primer ends for better specificity of amplification. Conclusions: We hope that this new bioinformatics tool developed for target selection and taxon-specific primer design, called the MorphoCatcher, will gain more popularity of the loop-mediated isothermal amplification method for molecular diagnostics community. MorphoCatcher is a simple web plugin tool for the PrimerExplorer software which is freely available only for non-commercial and academic users at http://morphocatcher.ru.

Consumption, white plague, tuberculosis… Only relatively recently, this disease has ceased to be an absolutely death sentence for infected people, but problems of the spread and diagnosis of the disease are still relevant. This paper presents results of the development of a new loop-mediated isothermal amplification (LAMP) assay, named TB-ISATEST, that targets the species-specific gene rv2341 for the differentiation of Mycobacterium tuberculosis from non-tuberculosis mycobacteria. The assay is applicable for quantitative analysis of genomic DNA and allows detecting tenfold difference in concentration. The results of amplification optimization using a unique two-stage protocol, based on the Taguchi orthogonal arrays method, are presented for the first time. A theoretical interpretation of the high amplification efficiency values observed in the LAMP reaction is proposed. Limit of detection of the developed assay is 40 copies of genomic DNA per reaction and amplification requires 15 min. In terms of the combination of characteristics, the TB-ISATEST assay surpasses all the known ways for identifying M. tuberculosis by the LAMP method.