Multiplex Real-Time PCR with HRM for Detection of Lactobacillus sakei and Lactobacillus curvatus in Food Samples

SAŽETAK Za optimiranje procesa potrebno je pratiti sastav starter kulture i njezin rast tijekom fermentacije. Većina starter kultura sadržava srodne mikroorganizme. U današnje se vrijeme za identifikaciju većeg broja srodnih vrsta često upotrebljava postupak analize krivulje mekšanja DNK visoke razlučivosti. Stoga je u radu za detekciju i razlikovanje bakterija Lactobacillus sakei i L. curvatus upotrijebljena lančana reakcija polimeraze (PCR) u stvarnom vremenu, te je provedena analiza krivulje mekšanja DNK visoke razlučivosti. Odabran je par početnica za ciljano mjesto vezivanja u genu rpoA, izoliranom iz bakterija roda Lactobacillus spp. S tim je parom početnica uspješno ispitano 1 1 starter kultura i 15 uzoraka fermentiranih kobasica poznatog sastava bakterija.Optimization of fermentation processes requires monitoring the species composition of starter cultures and their growth during fermentation. Most starter cultures contain closely related species. Nowadays, high-resolution melting (HRM) analysis is extensively used for multiplex identification of closely related species. In the present paper, we applied real-time polymerase chain reaction (PCR) with HRM analysis for the detection and differentiation of Lactobacillus sakei and L. curvatus. A primer pair was selected for the site of the rpoA gene of Lactobacillus spp. Eleven starter cultures and fifteen fermented sausages with a known bacterial composition were successfully tested using real-time PCR with HRM analysis with the developed primer pair

Multiplex Real-Time PCR with HRM for Detection of Lactobacillus sakei and Lactobacillus curvatus in Food Samples

Optimization of fermentation processes requires monitoring the species composition of starter cultures and their growth during fermentation. Most starter cultures contain closely related species. Nowadays, high-resolution melting (HRM) analysis is extensively used for multiplex identification of closely related species. In the present paper, we applied real-time polymerase chain reaction (PCR) with HRM analysis for the detection and differentiation of Lactobacillus sakei and L. curvatus. A primer pair was selected for the site of the rpoA gene of Lactobacillus spp. Eleven starter cultures and fifteen fermented sausages with a known bacterial composition were successfully tested using real-time PCR with HRM analysis with the developed primer pair

The effects of genetic drift and genomic selection on differentiation and local adaptation of the introduced populations of Aedes albopictus in southern Russia

Background Asian tiger mosquito Aedes albopictus is an arbovirus vector that has spread from its native habitation areal in Southeast Asia throughout North and South Americas, Europe, and Africa. Ae. albopictus was first detected in the Southern Federal District of the Russian Federation in the subtropical town of Sochi in 2011. In subsequent years, this species has been described in the continental areas with more severe climate and lower winter temperatures. Methods Genomic analysis of pooled Ae. albopictus samples collected in the mosquito populations in the coastal and continental regions of the Krasnodar Krai was conducted to look for the genetic changes associated with the spread and potential cold adaptation in Ae. albopictus. Results The results of the phylogenetic analysis based on mitochondrial genomes corresponded well with the hypothesis that Ae. albopictus haplotype A1a2a1 was introduced into the region from a single source. Population analysis revealed the role of dispersal and genetic drift in the local adaptation of the Asian tiger mosquito. The absence of shared haplotypes between the samples and high fixation indices suggest that gene flow between samples was heavily restricted. Mitochondrial and genomic differentiation together with different distances between dispersal routes, natural and anthropogenic barriers and local effective population size reduction could lead to difficulties in local climatic adaptations due to reduced selection effectiveness. We have found genomic regions with selective sweep patterns which can be considered as having been affected by recent selection events. The genes located in these regions participate in neural protection, lipid conservation, and cuticle formation during diapause. These processes were shown to be important for cold adaptation in the previous transcriptomic and proteomic studies. However, the population history and relatively low coverage obtained in the present article could have negatively affect sweep detection

Performance of Multiplexed XY Resistive Micromegas detectors in a high intensity beam

We present the performance of multiplexed XY resistive Micromegas detectors tested in the CERN SPS 100 GeV/c electron beam at intensities up to 3.3 × 10 5e−/(s-cm2). So far, all studies with multiplexed Micromegas have only been reported for tests with radioactive sources and cosmic rays. The use of multiplexed modules in high intensity environments was not explored due to the effect of ambiguities in the reconstruction of the hit point caused by the multiplexing feature. For the specific mapping and beam intensities analyzed in this work with a multiplexing factor of five, more than 50% level of ambiguity is introduced due to particle pile-up as well as fake clusters due to the mapping feature. Our results prove that by using the additional information of cluster size and integrated charge from the signal clusters induced on the XY strips, the ambiguities can be reduced to a level below 2%. The tested detectors are used in the CERN NA64 experiment for tracking the incoming particles bending in a magnetic field in order to reconstruct their momentum. The average hit detection efficiency of each module was found to be 96% at the highest beam intensities. By using four modules a tracking resolution of 1.1% was obtained with 85% combined tracking efficiency.ISSN:0168-9002ISSN:1872-957