BackgroundExtracellular vesicles (EVs), including exosomes, microvesicles, and apoptotic bodies, can be secreted by most cell types and released in perhaps all biological fluids. EVs contain multiple proteins, specific lipids and several kinds of nucleic acids such as RNAs and DNAs. Studies have found that EVs contain double-stranded DNA and that genetic information has a certain degree of consistency with tumor DNA. Therefore, if genes that exist in exosomes are stable, we may be able to use EVs genetic testing as a new means to monitor gene mutation.MethodsIn this study, EVs were extracted from serum under various storage conditions (4 °C, room temperature and repeated freeze-thaw). We used western blotting to examine the stability of serum EVs. Then, we extracted DNA from EVs and tested the concentration changing under different conditions. We further assessed the stability of EVs DNA s using polymerase chain reaction (PCR) and Sanger sequencing.ResultsEVs is stable under the conditions of 4 °C (for 24 h, 72 h, 168 h), room temperature (for 6 h, 12 h, 24 h, 48 h) and repeated freeze-thaw (after one time, three times, five times). Also, serum DNA is mainly present in EVs, especially in exosomes, and that the content and function of DNA in EVs is stable whether in a changing environment or not. We showed that EVs DNA stayed stable for 1 week at 4 °C, 1 day at room temperature and after repeated freeze-thaw cycles (less than three times). However, DNA from serum EVs after 2 days at room temperature or after five repeated freeze-thaw cycles could be used for PCR and sequencing.ConclusionsSerum EVs and EVs DNA can remain stable under different environments, which is the premise that EVs could serve as a novel means for genetic tumor detection and potential biomarkers for cancer diagnostics and prognostics.

Accurate and quantitative analysis of mycotoxin (such as zearalenone) is particularly imperative in the field of food safety and animal husbandry. Here, we develop a sensitive and specific method for zearalenone detection using competitive surface-enhanced Raman scattering (SERS) immunoassay. In this assay, a functional gold nanoparticle was labeled with the Raman reporter and the zearalenone antibody, and a modified substrate was assembled with the zearalenone-bovine serum albumin. With the addition of free zearalenone, the competitive immune reaction between free zearalenone and zearalenone-bovine serum albumin was initiated for binding with zearalenone antibody labeled on gold nanoparticle, resulting in the change of SERS signal intensity. The proposed method exhibits high sensitivity with a detection limit of 1 pg/mL and a wide dynamic range from 1 to 1000 pg/mL. Furthermore, this method can be further applied to analyze the multiple natural feed samples contaminated with zearalenone, holding great potential for real sample detection.

BackgroundRumen flukes parasitize the rumen and reticulum of ruminants, causing paramphistomiasis. Over the years, there has been considerable debate as to whether Paramphistomum leydeni and Paramphistomum cervi are the same or distant species.MethodsIn the present study, the complete mitochondrial (mt) genome of P. leydeni was amplified using PCR-based sequencing and compared with that of P. cervi. The second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA (rDNA) of P. leydeni specimens (n = 6) and P. cervi specimens (n = 8) was amplified and then sequenced. Phylogenetic relationship of the concatenated amino acid sequence data for 12 protein-coding genes of the two rumen flukes and selected members of Trematoda was evaluated using Bayesian inference (BI).ResultsThe complete mt genome of P. leydeni was 14,050 bp in size. Significant nucleotide difference between the P. leydeni mt genome and that of P. cervi (14.7%) was observed. For genetic divergence in ITS-2, sequence difference between P. leydeni and P. cervi was 3.1%, while no sequence variation was detected within each of them. Phylogenetic analysis indicated that P. leydeni and P. cervi are closely-related but distinct rumen flukes.ConclusionsResults of the present study support the proposal that P. leydeni and P. cervi represent two distinct valid species. The mt genome sequences of P. leydeni provide plentiful resources of mitochondrial markers, which can be combined with nuclear markers, for further comparative studies of the biology of P. leydeni and its congeners from China and other countries.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-0823-4) contains supplementary material, which is available to authorized users.

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BackgroundCystic echinococcosis (CE) in humans and livestock is caused by Echinococcus granulosus (sensu lato). In China where CE is endemic, a number of studies have shown that Echinococcus granulosus (sensu stricto) is majorly responsible for CE. However, E. canadensis (G6) which is the second leading cause of CE is now being detected in most parts of the country. In this study, the species diversity and genetic variation of Echinococcus granulosus (s.l.) in four counties in Tibet Autonomous Region of China were investigated.MethodsInfection with Echinococcus granulosus (s.s.) in yaks and sheep was identified using NADH dehydrogenase subunit 1 and 5 (nad1 and nad5) mitochondrial genes while the genotype G6 of E. canadensis initially diagnosed with NADH dehydrogenase subunit 1 (nad1) was further confirmed by analysis of the complete mitochondrial genome and a phylogenetic network constructed based on the nad2 and nad5 genes.ResultsOut of 85 hydatid cyst samples collected from slaughtered sheep (n = 54) and yaks (n = 31), 83 were identified as E. granulosus (s.s.) G1 (n = 77), G3 (n = 6) and 2 were identified as E. canadensis G6. Analysis of the nad1/nad5 genes revealed 16/17 mutations with 9/14 parsimony informative sites resulting in 15/14 haplotypes, respectively. Haplotype diversity (Hd) and nucleotide diversity (π) of E. granulosus (s.s.) population were 0.650 and 0.00127 for nad1 and 0.782 and 0.00306 for nad5, respectively, with an overall negative Tajima’s D and Fu’s Fs. A low FST indicated no genetic difference between isolates from sheep and yaks.ConclusionPockets of infection with E. canadensis (G6, G7, G8 and G10) have been previously reported in sheep, goats, yaks and/or humans in different parts of China. While the G6 genotype has been previously reported in sheep in the Tibet Autonomous Region, the detection in a yak in the present study represents the first to the best of our knowledge. Therefore, we recommend future surveys and control efforts to comprehensively investigate other potential intermediate hosts for the prevalence and genetic diversity of the E. canadensis group (G6, G7, G8 and G10) across the country and their inclusion into the existing CE control programme.