ABSTRACT
Growing evidence suggests that metavirome changes could be associated increased risk for malignant cell transformation. Considering Viruses have been proposed as factors for prostate cancer induction. The objective of this study was to examine the composition of the plasma metavirome of patients with prostate cancer. Blood samples were obtained from 49 male patients with primary prostate adenocarcinoma. Thirty blood donors were included as a control group. The obtained next-generation sequencing data were analyzed using a bioinformatic pipeline for virus metagenomics. Viral reads with higher abundance were assembled in contigs and analyzed taxonomically. Viral agents of interest were also confirmed by qPCR. Anelloviruses and the Human Pegivirus-1 (HPgV-1) were the most abundant component of plasma metavirome. Clinically important viruses like hepatitis C virus (HCV), cytomegalovirus and human adenovirus type C were also identified. In comparison, the blood donor virome was exclusively composed of torque teno virus types (TTV) types. The performed HPgV-1 and HCV phylogeny revealed that these viruses belong to commonly detected in Brazil genotypes. Our study sheds light on the plasma viral abundance in patients with prostatic cancer. The obtained viral diversity allowed us to separate the patients and controls, probably suggesting that malignant processes may influence virome composition. More complex and multiple approach investigations are necessary to examine the likely causal relationship between metavirome and its nvolvement in prostate cancer.
ABSTRACT
This work describes the development of a Point-of-Care (POC) Lab-on-a-Chip (LOC) instrument for diagnosis of SARS-CoV-2 by Reverse-Transcription Loop-mediated isothermal amplification (RT-LAMP). The hardware is based on a Raspberry Pi computer ($35), a video camera, an Arduino Nano microcontroller, a printed circuit board as a heater and a 3D printed housing. The chips were manufactured in polymethyl methacrylate (PMMA) using a CO2 laser cutting machine and sealed with a PCR optic plastic film. The chip temperature is precisely controlled by a proportional-integral-derivative (PID) algorithm. During the RT-LAMP amplifications the chip was maintained at â¼ (65.0 ± 0.1) °C for 25 minutes and 5 minutes cooling down, totaling a 30 minutes of reaction .The software interpretation occurs in less than a second. The chip design has four 25 µL chambers, two for clinical samples and two for positive and negative control-samples. The RT-LAMP master mix solution added in the chip chambers contains the pH indicator Phenol Red, that is pink (for pH â¼ 8.0) before amplification and becomes yellow (pH â¼ 6.0) if the genetic material is amplified. The RT-LAMP SARS-CoV-2 diagnostic was made by color image recognition using the OpenCV machine vision software library. The software was programmed to automatically distinguish the HSV color parameter distribution in each one of the four chip chambers. The instrument was successfully tested for SARS-CoV-2 diagnosis, in 22 clinic samples, 11 positives and 11 negatives, achieving an assertiveness of 86% when compared to the results obtained by RT-LAMP standard reactions performed in conventional PCR equipment.
ABSTRACT
To better understand the early events regulating lineage-specific hematopoietic differentiation, we analyzed the transcriptional profiles of CD34+ human hematopoietic stem and progenitor cells (HSPCs) subjected to differentiation stimulus. CD34+ cells were cultured for 12 and 40h in liquid cultures with supplemented media favoring myeloid or erythroid commitment. Serial analysis of gene expression (SAGE) was employed to generate four independent libraries. By analyzing the differentially expressed regulated transcripts between the un-stimulated and the stimulated CD34+ cells, we observed a set of genes that was initially up-regulated at 12h but were then down-regulated at 40h, exclusively after myeloid stimulus. Among those we found transcripts for NFKB2, RELB, IL1B, LTB, LTBR, TNFRSF4, TGFB1, and IKBKA. Also, the inhibitor NFKBIA (IKBA) was more expressed at 12h. All those transcripts code for signaling proteins of the nuclear factor kappa B pathway. NFKB2 is a subunit of the NF-κB transcription factor that with RELB mediates the non-canonical NF-κB pathway. Interference RNA (RNAi) against NFKB1, NFKB2 and control RNAi were transfected into bone marrow CD34+HSPC. The percentage and the size of the myeloid colonies derived from the CD34+ cells decreased after inhibition of NFKB2. Altogether, our results indicate that NFKB2 gene has a role in the early commitment of CD34+HSPC towards the myeloid lineage.
