Genome-informed loop-mediated isothermal amplification assay for specific detection of Pectobacterium parmentieri in infected potato tissues and soil.

Pectobacterium parmentieri (formerly Pectobacterium wasabiae), which causes soft rot disease in potatoes, is a newly established species of pectinolytic bacteria within the family Pectobacteriaceae. Despite serious damage caused to the potato industry worldwide, no field-deployable diagnostic tests are available to detect the pathogen in plant samples. In this study, we aimed to develop a reliable, rapid, field-deployable loop-mediated isothermal amplification (LAMP) assay for the specific detection of P. parmentieri. Specific LAMP primers targeting the petF1 gene region, found in P. parmentieri but no other Pectobacterium spp., were designed and validated in silico and in vitro using extensive inclusivity (15 strains of P. parmentieri) and exclusivity (94 strains including all other species in the genus Pectobacterium and host DNA) panels. No false positives or negatives were detected when the assay was tested directly with bacterial colonies, and with infected plant and soil samples. Sensitivity (analytical) assays using serially diluted bacterial cell lysate and purified genomic DNA established the detection limit at 10 CFU/mL and 100 fg (18-20 genome copies), respectively, even in the presence of host crude DNA. Consistent results obtained by multiple users/operators and field tests suggest the assay's applicability to routine diagnostics, seed certification programs, biosecurity, and epidemiological studies.

Anti-inflammatory activities of Waltheria indica extracts by modulating expression of IL-1B, TNF-α, TNFRII and NF-κB in human macrophages.

In Hawaiian traditional medicinal practices, the indigenous 'uhaloa, Waltheria indica var. Americana is one of the most recognized plants. Waltheria is also known in various cultures as a medicinal plant for the treatment of inflammatory conditions. Results in human subjects and cell and animal models supported anti-inflammatory activity for the Waltheria flavonoid quercetin, and for crude plant extracts, limited animal studies also confirmed anti-inflammatory effects. Yet no systematic studies have examined immune or inflammatory responses affected by these extracts. In order to gain insight into inflammatory cascades modulated by Waltheria extracts, and to uncover the mechanistic bases for the effective use of this medicinal plant as a natural anti-inflammatory agent, we have undertaken analyses of LPS and TNF-α/IF-γ-stimulated human macrophages treated with Waltheria extracts using targeted qRT-PCR and Inflammation Panels to test differential mRNA expression of two hundred immune-related genes, furthermore, ELISA assays and Inflammatory Protein arrays to determine extracts-modulated intracellular and secreted levels of prominent cytokines. Results demonstrate that Waltheria extracts inhibit key inflammatory cytokines and cytokine receptors including protein levels of IL-1B, IL-1ra, IL-8 and IL-6, reduce both mRNA and protein levels of TNF-α and protein levels of its receptor, TNF RII, predicting diminished TNF-α-associated inflammatory signaling that, together with significant reduction of NF-κB mRNA and protein, can effectively diminish activities of multiple pro-inflammatory signaling pathways and mitigate key processes in diseases with inflammatory components.