Phuntumart, V. | Search bioRxiv

bioRxiv2021

Development and comparison of Loop-Mediated Isothermal Amplification with Quantitative PCR for the specific detection of Saprolegnia spp.

Saprolegniasis is an important disease in freshwater aquaculture, and is associated with oomycete pathogens in the genus Saprolegnia. Early detection of significant levels of Saprolegnia spp. pathogens would allow informed decisions for treatment which could significantly reduce losses. This study is the first to report the development of loop-mediated isothermal amplification (LAMP) for the detection of Saprolegnia spp. and compares it with quantitative PCR (qPCR). The developed protocols targeted the internal transcribed spacer (ITS) region of ribosomal DNA and the cytochrome C oxidase subunit 1 (CoxI) gene and was shown to be specific only to Saprolegnia genus. This LAMP method can detect as low as 10 fg of S. salmonis DNA while the qPCR method has a detection limit of 2 pg of S. salmonis DNA, indicating the superior sensitivity of LAMP compared to qPCR. When applied to detect the pathogen in water samples, both methods could detect the pathogen when only one zoospore of Saprolegnia was present. We propose LAMP as a quick (about 20-60 minutes) and sensitive molecular diagnostic tool for the detection of Saprolegnia spp. suitable for on-site applications.

molecular biology↗

bioRxiv2021

Fecal microbial transplant abates tolerance to methylone-induced hyperthermia

The microbiome-gut-brain axis has been implicated in multiple bodily systems and pathologies, and intentional manipulation of the gut-microbiome has yielded clinically significant results. Here, we examined the effects of bi-directional fecal microbial transplants (FMT) between methylone-induced hyperthermic tolerant (MHT) and methylone-naive (MN) rats. Rats treated with methylone once per week developed tolerance to methylone-induced hyperthermia by the fourth week. Once tolerant, daily bi-directional FMT between the two groups were performed for seven days prior to the next methylone treatment. The FMT abated the developed tolerance in the MHT group. When treated with methylone for the first time following FMT, recipient MN rats displayed significant tolerance to hyperthermia despite it being their initial drug treatment. Post-FMT, MHT rats displayed elevations in norepinephrine and expression of UCP1, UCP3 and TGR5 in brown adipose tissue, with reductions in expression of TGR5 and UCP3 in skeletal muscle. The pre- and post-FMT methylone tolerance phenotypes of transplant recipients are concurrent with changes in the relative abundance of several Classes of Proteobacteria, most evident for Gammaproteobacter and Alphaproteobacter. MHT recipients demonstrated a marked increase in the relative proportion of the Firmicutes Class Erysipelotrichia. These findings suggest that transplantation of gut-microbiomes can confer phenotypic responses to a drug.

pharmacology and toxicology↗