RESUMO
RT-qPCR remains a key diagnostic methodology for COVID-19/SARS-CoV-2. Typically, nasal or saliva swabs from patients are placed in virus transport media (VTM), RNA is extracted at the pathology laboratory, and viral RNA is measured using RT-qPCR. In this study, we describe the use of TNA-Cifer Reagent E in a pre-clinical evaluation study to inactivate SARS-CoV-2 as well as prepare samples for RT-qPCR. Adding 1 part TNA-Cifer Reagent E to 5 parts medium containing SARS-CoV-2 for 10 min at room temperature inactivated the virus and permitted RT-qPCR detection. TNA-Cifer Reagent E was compared with established column-based RNA extraction and purification methodology using a panel of human clinical nasal swab samples (n = 61), with TNA-Cifer Reagent E showing high specificity (100%) and sensitivity (97.37%). Mixtures of SARS-CoV-2 virus and TNA-Cifer Reagent E could be stored for 3 days at room temperature or for 2 weeks at 4°C without the loss of RT-qPCR detection sensitivity. The detection sensitivity was preserved when TNA-Cifer Reagent E was used in conjunction with a range of VTM for saliva samples but only PBS (Gibco) and Amies Orange for nasal samples. Thus, TNA-Cifer Reagent E improves safety by rapidly inactivating the virus during sample processing, potentially providing a safe means for molecular SARS-CoV-2 testing outside traditional laboratory settings. The reagent also eliminates the need for column-based and/or automated viral RNA extraction/purification processes, thereby providing cost savings for equipment and reagents, as well as reducing processing and handling times.
RESUMO
BACKGROUND: Bioclimatic variables play an integral part in the life-cycle of Ctenocephalides felis, the most common flea found on companion animals. It is essential that we understand the effects of climate on C. felis distribution as fleas are a major veterinary and public health concern. This study investigated the current distribution of C. felis in Australia and future projections based on climate modelling. RESULTS: Typing of C. felis was undertaken using the cytochrome c oxidase subunit 1 (cox1) mitochondrial DNA (mtDNA) region and current distribution of haplotypes was mapped by Maximum Entropy (Maxent) niche modelling. All C. felis haplotypes have been predicted to persist in environments along the eastern and southern coastlines of Australia and distinct ecological niches were observed for two C. felis haplogroups. Clade 'Cairns' haplogroup thrives under the northern coastal tropical conditions whilst Clade 'Sydney' haplogroup persists in temperate climates along the eastern and southern coasts. The model was then used to predict areas that are projected to have suitable climatic conditions for these haplogroups in 2050 and 2070 under the Intergovernmental Panel on Climate Change (IPCC) climate change scenarios. Under all IPCC Representative Concentration Pathways (RCP) climate change scenarios, the geographical range of all haplotypes was reduced by 5.59-42.21% in 2050 and 27.08-58.82% by 2070. The ranges of all clades were predicted to shift south along the eastern coastline. CONCLUSIONS: As future temperatures exceed critical threshold temperatures for C. felis development in the northern tropical areas, Clade 'Cairns' haplogroup is predicted to shift south along the coastline and possibly outcompete the temperate haplogroup in these areas. If C. felis haplogroups possess distinct climatic niches it suggests a potential for these to be biologically distinct and have differing developmental rates and vector capabilities.