DNA Damage Induced by T-2 Mycotoxin in Human Skin Fibroblast Cell Line-Hs68.

T-2 mycotoxin is the most potent representative of the trichothecene group A and is produced by various Fusarium species, including F. sporotrichioides, F. poae, and F. acuminatum. T-2 toxin has been reported to have toxic effects on various tissues and organs, and humans and animals alike suffer a variety of pathological conditions after consumption of mycotoxin-contaminated food. The T-2 toxin's unique feature is dermal toxicity, characterized by skin inflammation. In this in vitro study, we investigated the molecular mechanism of T-2 toxin-induced genotoxicity in the human skin fibroblast-Hs68 cell line. For the purpose of investigation, the cells were treated with T-2 toxin in 0.1, 1, and 10 µM concentrations and incubated for 24 h and 48 h. Nuclear DNA (nDNA) is found within the nucleus of eukaryotic cells and has a double-helix structure. nDNA encodes the primary structure of proteins, consisting of the basic amino acid sequence. The alkaline comet assay results showed that T-2 toxin induces DNA alkali-labile sites. The DNA strand breaks in cells, and the DNA damage level is correlated with the increasing concentration and time of exposure to T-2 toxin. The evaluation of nDNA damage revealed that exposure to toxin resulted in an increasing lesion frequency in Hs68 cells with HPRT1 and TP53 genes. Further analyses were focused on mRNA expression changes in two groups of genes involved in the inflammatory and repair processes. The level of mRNA increased for all examined inflammatory genes (TNF, INFG, IL1A, and IL1B). In the second group of genes related to the repair process, changes in expression induced by toxin in genes-LIG3 and APEX were observed. The level of mRNA for LIG3 decreased, while that for APEX increased. In the case of LIG1, FEN, and XRCC1, no changes in mRNA level between the control and T-2 toxin probes were observed. In conclusion, the results of this study indicate that T-2 toxin shows genotoxic effects on Hs68 cells, and the molecular mechanism of this toxic effect is related to nDNA damage.

Immunosensors-The Future of Pathogen Real-Time Detection.

Pathogens and their toxins can cause various diseases of different severity. Some of them may be fatal, and therefore early diagnosis and suitable treatment is essential. There are numerous available methods used for their rapid screening. Conventional laboratory-based techniques such as culturing, enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) are dominant. However, culturing still remains the "gold standard" for their identification. These methods have many advantages, including high sensitivity and selectivity, but also numerous limitations, such as long experiment-time, costly instrumentation, and the need for well-qualified personnel to operate the equipment. All these existing limitations are the reasons for the continuous search for a new solutions in the field of bacteria identification. For years, research has been focusing on the use of immunosensors in various types of toxin- and pathogen-detection. Compared to the conventional methods, immunosensors do not require well-trained personnel. What is more, immunosensors are quick, highly selective and sensitive, and possess the potential to significantly improve the pathogen and toxin diagnostic-processes. There is a very important potential use for them in various transport systems, where the risk of contamination by bioagents is very high. In this paper, the advances in the field of immunosensor usage in pathogenic microorganism- and toxin-detection, are described.

T-2 Toxin-The Most Toxic Trichothecene Mycotoxin: Metabolism, Toxicity, and Decontamination Strategies.

Among trichothecenes, T-2 toxin is the most toxic fungal secondary metabolite produced by different Fusarium species. Moreover, T-2 is the most common cause of poisoning that results from the consumption of contaminated cereal-based food and feed reported among humans and animals. The food and feed most contaminated with T-2 toxin is made from wheat, barley, rye, oats, and maize. After exposition or ingestion, T-2 is immediately absorbed from the alimentary tract or through the respiratory mucosal membranes and transported to the liver as a primary organ responsible for toxin's metabolism. Depending on the age, way of exposure, and dosage, intoxication manifests by vomiting, feed refusal, stomach necrosis, and skin irritation, which is rarely observed in case of mycotoxins intoxication. In order to eliminate T-2 toxin, various decontamination techniques have been found to mitigate the concentration of T-2 toxin in agricultural commodities. However, it is believed that 100% degradation of this toxin could be not possible. In this review, T-2 toxin toxicity, metabolism, and decontamination strategies are presented and discussed.

The Existing Methods and Novel Approaches in Mycotoxins' Detection.

Mycotoxins represent a wide range of secondary, naturally occurring and practically unavoidable fungal metabolites. They contaminate various agricultural commodities like cereals, maize, peanuts, fruits, and feed at any stage in pre- or post-harvest conditions. Consumption of mycotoxin-contaminated food and feed can cause acute or chronic toxicity in human and animals. The risk that is posed to public health have prompted the need to develop methods of analysis and detection of mycotoxins in food products. Mycotoxins wide range of structural diversity, high chemical stability, and low concentrations in tested samples require robust, effective, and comprehensible detection methods. This review summarizes current methods, such as chromatographic and immunochemical techniques, as well as novel, alternative approaches like biosensors, electronic noses, or molecularly imprinted polymers that have been successfully applied in detection and identification of various mycotoxins in food commodities. In order to highlight the significance of sampling and sample treatment in the analytical process, these steps have been comprehensively described.

Molecular Aspects of Mycotoxins-A Serious Problem for Human Health.

Mycotoxins are toxic fungal secondary metabolities formed by a variety of fungi (moulds) species. Hundreds of potentially toxic mycotoxins have been already identified and are considered a serious problem in agriculture, animal husbandry, and public health. A large number of food-related products and beverages are yearly contaminated by mycotoxins, resulting in economic welfare losses. Mycotoxin indoor environment contamination is a global problem especially in less technologically developed countries. There is an ongoing effort in prevention of mould growth in the field and decontamination of contaminated food and feed in order to protect human and animal health. It should be emphasized that the mycotoxins production by fungi (moulds) species is unavoidable and that they are more toxic than pesticides. Human and animals are exposed to mycotoxin via food, inhalation, or contact which can result in many building-related illnesses including kidney and neurological diseases and cancer. In this review, we described in detail the molecular aspects of main representatives of mycotoxins, which are serious problems for global health, such as aflatoxins, ochratoxin A, T-2 toxin, deoxynivalenol, patulin, and zearalenone.

Melittin-A Natural Peptide from Bee Venom Which Induces Apoptosis in Human Leukaemia Cells.

Bee venom is a very complex mixture produced and secreted by the honeybee (Apis mellifera). Melittin is a major component of bee venom that accounts for about 52% of its dry mass. A vast number of studies have been dedicated to the effects of melittin's regulation of apoptosis and to the factors that induce apoptosis in various types of cancer such as breast, ovarian, prostate, lung. The latest evidence indicates its potential as a therapeutic agent in the treatment of leukaemia. The aim of our present study is to evaluate melittin's ability to induce apoptosis in leukaemia cell lines of different origin acute lymphoblastic leukaemia (CCRF-CEM) and chronic myelogenous leukaemia (K-562). We demonstrated that melittin strongly reduced cell viability in both leukaemia cell lines but not in physiological peripheral blood mononuclear cells (PMBCs). Subsequent estimated parameters (mitochondrial membrane potential, Annexin V binding and Caspases 3/7 activity) clearly demonstrated that melittin induced apoptosis in leukaemia cells. This is a very important step for research into the development of new potential anti-leukaemia as well as anticancer therapies. Further analyses on the molecular level have been also planned (analysis of proapoptotic genes expression and DNA damages) for our next research project, which will also focus on melittin.