Patterns of Diversity of Fusarium Fungi Contaminating Soybean Grains.

Soybean is an important, high protein source of food and feed. However, like other agricultural grains, soybean may pose a risk to human and animal health due to contamination of the grains with toxigenic Fusaria and associated mycotoxins. In this study, we investigated the diversity of Fusaria on a panel of 104 field isolates obtained from soybean grains during the growing seasons in 2017-2020. The results of species-specific PCR analyses showed that Fusarium avenaceum was the most common (n = 40) species associated with soybean grains in Poland, followed by F. equiseti (n = 22) and F. sporotrichioides (11 isolates). A set of isolates, which was not determined based on PCR analyses, was whole genome sequenced. Multiple sequence analyses using tef-1α, top1, rpb1, rpb2, tub2, pgk, cam and lsu genes showed that most of them belonged to Equiseti clade. Three cryptic species from this clade: F. clavum, F. flagelliforme and FIESC 31 (lacking Latin binomial) were found on soybean for the first time. This is the first report demonstrating the prevalence of Fusaria on soybean grains in Poland.

Monitoring survey of nitrate content in beetroot, radish and cabbage in Poland.

BACKGROUND: Nitrates, compounds commonly occurring in nature, are present for example in vegetables, where they accumulate and become their contaminants. It is estimated that approximately 70-90% of nitrates intake comes from vegetables, which are therefore the main source of human exposure to these compounds through dietary intake. The European Union legislation limits nitrates as contaminants to three leafy vegetables, i.e. lettuce, spinach and rucola. The EU Member States are obliged to monitor nitrate levels in vegetables which may contain significant levels of these compounds. OBJECTIVE: Objective. To present the results of monitoring surveys on nitrate levels in radish and beetroot as well as in cabbage carried out in Poland between 2012 and 2019. MATERIAL AND METHODS: A total of 966 vegetable samples were tested. Chemical analyses were carried out in accredited laboratories of the State Sanitary Inspection. Analyses were performed by spectrophotometric methods using nitrate reduction on cadmium columns or by HPLC. RESULTS: The median nitrate content in beetroot was 1,260.0 mg NO- 3 /kg, whilst at the 95th percentile level - 3,222.2 mg NO-3 /kg. The levels of nitrates in beetroot preserves were lower: 1,030.3 mg NO-3 /kg (median) and 2337,2 mg NO-3 /kg (95th percentile). The median content of nitrates in radish and cabbage was 1,337.0 mg NO-3 /kg and 369,0 mg NO-3 /kg respectively, while at the 95th percentile the content of these compounds was found to be 3,381.5 mg NO-3 /kg and 1545,8 mg NO-3 /kg, respectively. CONCLUSIONS: The nitrate content in radish and cabbage does not pose a risk to the health of consumers, whilst the consumption of beetroot containing significant amounts of the above mentioned compounds may result in exceeding the ADI especially for young children.

Uncovering Diagnostic Value of Mitogenome for Identification of Cryptic Species Fusarium graminearum Sensu Stricto.

Fungal complexes are often composed of morphologically nearly indistinguishable species with high genetic similarity. However, despite their close relationship, they can exhibit distinct phenotypic differences in pathogenicity and production of mycotoxins. Many plant pathogenic and toxigenic fungi have been shown to consist of such cryptic species. Identification of cryptic species in economically important pathogens has added value in epidemiologic studies and provides opportunities for better control. Analysis of mitochondrial genomes or mitogenomics opens up dimensions for improved diagnostics of fungi, especially when efficient recovery of DNA is problematic. In comparison to nuclear DNA, mitochondrial DNA (mtDNA) can be amplified with improved efficacy due to its multi-copy nature. However, to date, only a few studies have demonstrated the usefulness of mtDNA for identification of cryptic species within fungal complexes. In this study, we explored the value of mtDNA for identification of one of the most important cereal pathogens Fusarium graminearum sensu stricto (F.g.). We found that homing endonucleases (HEGs), which are widely distributed in mitogenomes of fungi, display small indel polymorphism, proven to be potentially species specific. The resulting small differences in their lengths may facilitate further differentiation of F.g. from the other cryptic species belonging to F. graminearum species complex. We also explored the value of SNP analysis of the mitogenome for typing F.g. The success in identifying F.g. strains was estimated at 96%, making this tool an attractive complement to other techniques for identification of F.g.

The in vitro effect of orexin a on the porcine myometrial transcriptomic profile during the early-implantation period.

In pigs, early gestation is the most critical period deciding about the reproduction success, and it depends on many processes, involving a significant number of genes and their products. Myometrium was found to be an important source of factors pivotal for a proper course of gestation. The aim of the study was to determine the effect of orexin A (OXA) on the porcine transcriptome, and the determination of relationships among differentially expressed genes (DEG) in the porcine myometrium during implantation using microarray technology. The analyses of gene ontology (GO), DEG assays, biological pathways and networks were performed. OXA affected the expression of 461 genes with fold-change values greater than 1.2 (p < 0.05). The expression of 260 genes were up-regulated and 201 down-regulated in the OXA-treated myometrium. Twelve genes were selected for qPCR validation of differential expression based on their known role in angiogenesis, immune processes, steroid hormone signaling and prostaglandins synthesis. The analysis of relationship between DEG indicated that OXA interacts with genes involved in the inflammatory response, cytokine binding, cytokine activity, interleukin production, leukocyte migration, angiogenesis and embryonic hemopoiesis. The presented results suggest that OXA may play a key role in ensuring optimal conditions for implanting embryos.

The effect of orexin a on the StAR, CYP11A1 and HSD3B1 gene expression, as well as progesterone and androstenedione secretion in the porcine uterus during early pregnancy and the oestrous cycle.

Orexin A (OXA) is primarily known for its involvement in the regulation of feeding behaviour, energy metabolism and sleep/wake cycle. Nevertheless, studies indicate its engagement in the regulation of the porcine reproductive system. Therefore, the aim of this study was to investigate OXA effect (1, 10, 100 nM), in the presence or absence of the selective orexin receptor type 1 antagonist (SB-3348667; 1 µM), on the gene expression of key steroidogenic enzymes: steroidogenic acute regulatory protein (StAR), P450 side-chain cleavage enzyme (CYP11A1) and 3ß-hydroxysteroid dehydrogenase (HSD3B1), as well as on progesterone (P4) and androstenedione (A4) secretion. Endometrial and myometrial tissue explants were collected from gilts on days 10 to 11, 12 to 13, 15 to 16 and 27 to 28 of pregnancy, and on days 10 to 11 of the oestrous cycle (n = 5 per studied period of pregnancy or mid-luteal phase of the oestrous cycle). Gene expression was evaluated by real-time PCR. The level of steroid hormones secreted into the culture medium was examined by radioimmunoassay (RIA). In the present study, in the endometrium, OXA significantly stimulated StAR expression on days 12 to 13, CYP11A1 expression on days 27 to 28 and HSD3B1 expression on days 15 to 16 of pregnancy. Further, in this tissue, OXA decreased StAR mRNA level on days 10 to 11, CYP11A1 mRNA level on days 15 to 16, as well as HSD3B1 mRNA level on days 10 to 11 and 12 to 13 of gestation. Regarding the myometrium, OXA stimulated CYP11A1 gene expression on days 15 to 16 of pregnancy. In this tissue, OXA decreased StAR transcript content on days 15 to 16 and CYP11A1 mRNA level on days 27 to 28. We also demonstrated that OXA alone enhanced P4 secretion in the endometrium on days 10 to 11 and 12 to 13 of gestation. OXA alone has no significant effect on endometrial and myometrial A4 secretion, whereas OXA in combination with OX1R antagonist increased this hormone secretion during all studied stages of pregnancy. Therefore, we can conclude that OXA may affect de novo synthesis and secretion of P4 and A4 in the porcine uterus via participating in the regulation of key steroidogenic enzymes gene expression, as well as modulating steroid hormones secretion during early pregnancy and mid-luteal phase of the oestrous cycle in pigs. However, further research is required to explain the exact role of OXA in the porcine uterus.

Expression of Chemerin and Its Receptors in the Porcine Hypothalamus and Plasma Chemerin Levels during the Oestrous Cycle and Early Pregnancy.

Chemerin (CHEM) may act as an important link integrating energy homeostasis and reproductive functions of females, and its actions are mediated by three receptors: chemokine-like receptor 1 (CMKLR1), G protein-coupled receptor 1 (GPR1), and C-C motif chemokine receptor-like 2 (CCRL2). The aim of the current study was to compare the expression of CHEM and its receptor (CHEM system) mRNAs (quantitative real-time PCR) and proteins (Western blotting and fluorescent immunohistochemistry) in the selected areas of the porcine hypothalamus responsible for gonadotropin-releasing hormone production and secretion: the mediobasal hypothalamus, preoptic area and stalk median eminence during the oestrous cycle and early pregnancy. Moreover, plasma CHEM concentrations were determined using ELISA. The expression of CHEM system has been demonstrated in the porcine hypothalamus throughout the luteal phase and follicular phase of the oestrous cycle, and during early pregnancy from days 10 to 28. Plasma CHEM levels and concentrations of transcripts and proteins of CHEM system components in the hypothalamus fluctuated throughout pregnancy and the oestrous cycle. Our study was the first experiment to demonstrate the presence of CHEM system mRNAs and proteins in the porcine hypothalamus and the correlations between the expression levels and physiological hormonal milieu related to the oestrous cycle and early pregnancy.

Transcriptomic Analysis of Porcine Endometrium during Implantation after In Vitro Stimulation by Adiponectin.

Comprehensive understanding of the regulatory mechanism of the implantation process in pigs is crucial for reproductive success. The endometrium plays an important role in regulating the establishment and maintenance of gestation. The goal of the current study was to determine the effect of adiponectin on the global expression pattern of genes and relationships among differentially expressed genes (DE-genes) in the porcine endometrium during implantation using microarrays. Diverse transcriptome analyses including gene ontology (GO), biological pathway, networks, and DE-gene analyses were performed. Adiponectin altered the expression of 1286 genes with fold-change (FC) values greater than 1.2 (p < 0.05). The expression of 560 genes were upregulated and 726 downregulated in the endometrium treated with adiponectin. Thirteen genes were selected for real-time PCR validation of differential expression based on a known role in metabolism, steroid and prostaglandin synthesis, interleukin and growth factor action, and embryo implantation. Functional analysis of the relationship between DE-genes indicated that adiponectin interacts with genes that are involved in the processes of cell proliferation, programmed cell death, steroid and prostaglandin synthesis/metabolism, cytokine production, and cell adhesion that are critical for reproductive success. The presented results suggest that adiponectin signalling may play a key role in the implantation of pig.

In vitro effect of orexin A on the transcriptomic profile of the endometrium during early pregnancy in pigs.

Orexin A belongs to the group of hypothalamic-derived peptides that are involved in a number of processes, such as regulation of energy metabolism, control of food intake and regulation of the reproductive system, by influencing the hypothalamic-pituitary-ovarian axis. Orexin A is also present in the endometrium, myometrium and placenta, which indicates that it may function as an important local regulator of the reproductive functions. The aim of this study was to explore the effects of orexin A on global gene expression in the endometrium of pigs during early gestation, on days 15 to 16 of pregnancy (implantation period). Orexin A altered the expression of 1,242 genes. In this group, 1,104 genes had a fold change greater than 1.2 (P < 0.05). In the group of genes with a fold change that was greater than 1.2, the expression of 457 genes were increased and 647 decreased because of the effects of orexin A. An analysis of the interactions between differentially expressed genes demonstrated that orexin A interacts with genes that potentially encode intracellular signalling pathways and factors regulating reproductive functions in the endometrium. The data from the present study indicate that orexin A affects a number of genes and processes in the endometrium of pregnant pigs and may be regarded as an important regulator of implantation, depending on maternal nutritional status.

The effect of orexin B on steroidogenic acute regulatory protein, P450 side-chain cleavage enzyme, and 3ß-hydroxysteroid dehydrogenase gene expression, and progesterone and androstenedione secretion by the porcine uterus during early pregnancy and the estrous cycle.

The aim of this study was to investigate the effect of orexin B (OXB) on progesterone (P4) and androstenedione (A4) secretion by porcine endometrial and myometrial tissue explants and on the expression of key steroidogenic proteins and enzymes involved in steroid production. The hormones secretion and the expression of steroidogenic acute regulatory protein (StAR), P450 side-chain cleavage enzyme (CYP11A1), and 3ß-hydroxysteroid dehydrogenase (HSD3B1) were analyzed on days 10 to 11, 12 to 13, 15 to 16, and 27 to 28 of pregnancy and during the luteal phase of the estrous cycle (days 10 to 11). Endometrial and myometrial explants were cultured in vitro in the presence of OXB (1, 10, or 100 nM) and OXB (1, 10, or 100 nM) with 1 µM of JNJ (OX2R antagonist). Gene expression was examined by real-time PCR, and steroid secretion was determined by radioimmunoassay. Orexin B modulated StAR, CYP11A1, HSD3B1 mRNA content depending on the type of uterine tissue, the applied OXB dose, and the stage of pregnancy or the estrous cycle (P < 0.05). Orexin B increased P4 secretion in all stages of early gestation (P < 0.05). Orexin B enhanced the release of A4 on days 12 to 13, 15 to 16, and 27 to 28 of gestation, whereas on days 10 to 11 of early pregnancy, A4 secretion decreased in the endometrium and increased in the myometrium (P < 0.05). These results indicate that OXB affects the expression of key steroidogenic regulators and the secretion of steroid hormones in the porcine uterus during early pregnancy.