Seasonal Stability Assessment of Reference Genes for Quantitative Real-Time Polymerase Chain Reaction Normalization in Bombus terrestris.

Bumblebees (B. terrestris) play a crucial role as highly efficient biological agents in commercial pollination. Understanding the molecular mechanisms governing their adaptation to diverse seasonal environments may pave the way for effective management strategies in the future. With the burgeoning advancement in post-genetic studies focusing on B. terrestris, there is a critical need to normalize quantitative real-time PCR (qRT-PCR) data using suitable reference genes. To address this necessity, we employed RefFinder, a software-based tool, to assess the suitability of several candidate endogenous control genes, including actin (ACT), arginine kinase (AK), elongation factor 1 alpha (EF1), glyceraldehyde-3-phosphate (GAPDH), phospholipase (PLA2), and ribosomal proteins (S18, S28). These genes were evaluated for their efficacy as biological endogenous controls by examining their expression patterns across various environmental conditions corresponding to different seasons (Spring, Summer, Autumn, Winter) and tissues (ovary, fat body, thorax, head) in bumblebees. Moreover, the study investigated the significance of selecting appropriate reference genes for three key genes involved in the juvenile hormone (JH) signaling pathways: Krüppel homolog 1 (Kr-h1), methyl farnesoate epoxidase (MFE), and Vitellogenin (Vg). Our research identifies specific genes suitable for normalization in B. terrestris, thereby offering valuable insights into gene expression and functional metabolic genetics under varying seasonal conditions. This catalog of reference genes will serve as a valuable resource for future research endeavors.

Evaluation of Reference Genes for Real-Time Quantitative PCR Analysis in Tissues from Bumble Bees (Bombus Terrestris) of Different Lines.

Bumble bees are important alternative pollinators and model insects due to their highly developed sociality and colony management. In order to better understand their molecular mechanisms, studies focusing on the genetic and molecular aspects of their development and behavior are needed. Although quantitative real-time polymerase chain reaction (qRT-PCR) can be used to quantify the relative expression of target genes, internal reference genes (which are stably expressed across different lines and tissues) must first be identified to ensure the accurate normalization of target genes. In order to contribute to molecular studies on bumble bees, we used Bombus terrestris to determine the expression stability of eight reference genes (ß-actin (ACT), Arginine Kinase (AK), Phospholipase A2 (PLA2), Elongation factor 1 alpha (EF-1), Ribosomal proteins (S5, S18, S28) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH)) in five different lines and several tissues (ovary, thorax, fat body, and head) using RT-qPCR procedures and four analysis programs (RefFinder, NormFinder, BestKeeper, and geNorm). In general, the S28, S5, and S18 ribosomal protein genes and the PLA2 and EF-1 genes showed the highest stability and were therefore identified as suitable reference genes for the bumble bee species and their defined lines and tissues. Our results also emphasized the need to evaluate the stability of candidate reference genes for any differently designed lines and tissue conditions in bumble bee species.

Estimation of cytogenetic risk among coke oven workers exposed to polycyclic aromatic hydrocarbons.

Polycyclic aromatic hydrocarbons (PAHs) result from the incomplete combustion of natural or synthetic organic materials. The working environment at a coke plant can negatively affect the employed workers who were exposed to coke oven emissions containing PAHs, which formed and released into the environment by the process of pyrolysis of coke. This study aims to analyze the relationship between the exposure of PAHs and the risk of genetic damages such as chromosomal alteration (CA), micronucleus (MN), and DNA damage (PCR-RFLP) in peripheral blood lymphocytes of 27 coke oven workers and equal number of control subjects. The exposed subjects and controls were divided into two groups based on their age (group I<35 years and group II ≥35 years). The exposed subjects were further classified into two groups based on the exposure period (<12 years and ≥12 years). The frequencies of CA and MN in exposed subjects are relatively high with respect to controls. The XRCC1 399 Arg/gln polymorphism showed a substantial smaller difference in allele frequencies between exposed and control subjects. Based on present data, it was concluded that coke oven workers under risk should be monitored for adverse effects of the any long-term exposure.

A comprehensive analysis of plausible genotoxic covariates among workers of a polyvinyl chloride plant exposed to vinyl chloride monomer.

The aim of this study was to assess the frequency of chromosomal aberrations-including chromatid type aberrations (CTAs), chromosomal type aberrations, micronucleus (MN) comet assay, and XRCC1 399 Arg/Gln polymorphism-in peripheral blood lymphocytes of workers occupationally exposed to vinyl chloride monomer (VCM). A total of 52 workers and an equal number of controls were recruited into the study to explore the potential cytogenetic risk of occupational exposure to VCM. Questionnaires were administered to obtain details of habitual cigarette-smoking, alcohol-consumption pattern, and occupation, etc. The exposed subjects and controls were classified into two groups based on age (group I <40 years; group II ≥40 years), and exposed subjects were further classified based on exposure duration (>8 and ≥8 years). CTA, MN, and comet assay frequency were significantly greater in polyvinyl chloride (PVC) factory workers (p < 0.05) with long-duration work. CTA, MN, and comet assay values were found to be increased with age in exposed subjects as well as in controls, with exposed subjects showing a statistically greater degree. An extensively greater MN frequency was observed in smokers exposed to VCM than in the control group (P < 0.05). The mean tail length of exposed subjects was greater compared with controls. The study on XRCC1 399 Arg/gln polymorphism in PVC factory workers showed less significant difference in allele frequency compared with controls. In conclusion, this results of work provides evidence for an apparent genotoxic effect associated with VCM exposure. Our results reinforce the greater sensitivity of cytogenetic assays for biomonitoring of occupationally exposed populations. Statistics indicate that workers exposed to VCM are at carcinogenic risk and should be monitored for long-term adverse effects from their exposure.