Development and characterization of microsatellite markers for the French endemic Angelica heterocarpa (Apiaceae) and congeneric sympatric species

Objective Angelica heterocarpa (Apiaceae) is a wild endemic French species with special conservation interest in the European Union. It belongs to Angelica complex genus which is widespread throughout the north temperate zone, and is sympatric with other congeneric species. The objective of this work is to develop and characterize microsatellite markers as a new tool for understanding the ecology and evolution of Angelica species complex. Results We identified simple sequence repeat (SSR) regions in a microsatellite‐enriched library from A. heterocarpa and A. sylvestris. All 16 selected SSR regions were found to amplify in these species and were highly polymorphic. Marker transferability was validated in A. razulii and A. archangelica. These markers will help us to better understand the evolutionary dynamic between rare endemics and widespread sister species, and be useful for conservation of the endemic species. Moreover, they can provide new tools for studying the numerous traditional medicinal herbs of the Angelica genus.

Nitric oxide as an indicator for assessing the resistance and susceptibility of cattle to leukemia

The role of allelic variability of inducible nitric oxide synthase (iNOS) is significant in the study of the resistance and susceptibility of animals to leukemia infection. After analyzing the literature data, it can be stated that in the iNOS gene, allele A (with genotype AA) is responsible for resistance to the leukemia virus, and allele B (with genotype BB) is responsible for susceptibility. This is due to the frequency of occurrence of alleles and their genotypes of the polymorphic marker AN13-1 of the inducibeal nitric oxide synthase gene. The iNOS gene is capable of producing a large amount of nitric oxide, compared to other isoforms. In turn, nitric oxide causes death or can stop the growth of pathogenic microorganisms, including viruses. The purpose of this work is to further study nitric oxide as an indicator for determining the resistance and susceptibility of animals to leukemia, as well as the selection of specific primers for PCR-PDRF used in genotyping. Methods. The iNOS gene sequence was analyzed and a pair of specific primers were selected and synthesized using the Vector NTI program. Scientific novelty of this work lies in the fact that we have selected specific primers that are important for the analysis of cattle genotyping by allelic variants of the polymorphic marker AH13-1 of the iNOS gene. Results. Based on this work, a pair of primers iNOSF_new and iNOSR_new, with a calculated annealing temperature of 52 °C, were selected and synthesized, giving an amplicon with a length of 186 bp. The amplicon contains a polymorphic site that distinguishes the A and B alleles. During PCR-RFLP, the following genotype-specific fragments are formed: AA-47/139 bp; AB -186/139/47 bp; BB-186 bp.

Genetic Diversity of Polymorphic Marker Merozoite Surface Protein 1 (Msp-1) and 2 (Msp-2) Genes of Plasmodium falciparum Isolates From Malaria Endemic Region of Pakistan

Background: Understanding the genetic diversity of Plasmodium species through polymorphic studies can assist in designing more effective control strategies of malaria like new drug formulation and development of a vaccine. Pakistan is moderate endemic for Plasmodium falciparum, but little is known about the genetic diversity of this parasite. This study aimed to investigate the molecular diversity of P. falciparum based on msp-1 and msp-2 genes in the malaria-endemic regions of Khyber Pakhtunkhwa, Pakistan.Methods: A total of 199/723 blood samples, tested positive by microscopy for falciparum malaria, were collected from four districts (Dera Ismail Khan, Karak, Mardan, and Peshawar) of Khyber Pakhtunkhwa. Nested PCR amplification technique was employed to target block 2 of msp-1 and the central domain of msp-2 genes, including their respective allelic families K1, MAD20, RO33, FC27, and 3D7/IC, and to detect the extent of genetic diversity of P. falciparum clinical isolates.Results: Among the 199 microscopy-positive P. falciparum samples, a total of 192 were confirmed using PCR. Ninety-seven amplicons were observed for msp-1 and 95 for msp-2. A total of 33 genotypes, 17 for msp-1 (eight K1, six MAD20, and three RO33) and 16 for msp-2 (nine FC27 and seven 3D7/IC), were identified. The specific allelic frequency of the K1 family was higher (44.3%) than that of MAD20 (33.0%) and RO33 (23.0%) for msp-1, while the FC27 allelic family was dominant (60.0%) compared with 3D7/IC (40.0%) for msp-2. No polyclonal infection was observed in msp-1 and msp-2. The expected heterozygosity was 0.98 and 0.97 for msp-1 and msp-2, respectively.Conclusion: It was concluded that the P. falciparum populations are highly polymorphic, and diverse allelic variants of msp-1 and msp-2 are present in Khyber Pakhtunkhwa, Pakistan.

Effect of Angiotensin-I Converting Enzyme Gene Insertion/Deletion Polymorphism on genome instability in children living in Russian Arctic

Main risks of arterial hypertension manifest in childhood. Children living in the Far North are especially susceptible to this. There is a need for an inexpensive, non-invasive and simple diagnosis of the risk of childhood pathologies. It was previously found that the genotype DD of the in/del polymorphic marker of the ACE gene is found in people at risk of developing cardiovascular pathologies. Buccal micronucleus cytome assay and genetic analysis were used in the work. In total, 77 schoolchildren from the city of Apatity, aged 15-17 years old, were examined. We have shown that carriers of the D allele have a tendency to an increase in the frequency of cells with micronuclei. In the case of homozygous I/I variant, the frequency of occurrence of cells with karyopycnosis is significantly higher than in carriers of allele D. Polymorphic marker in/del of the ACE gene is associated with apoptotic changes in the cells of the studied children. The in/del polymorphic marker of the ACE gene can be used as a prognostic marker of the processes of genome destabilization at the early stages of development of the human body.

Contribution of adipoq genetic polymorphism to the formation and course of comorbidity of non-alcoholic fatty liver disease and renoparinchemal arterial hypertension

Introduction. Nowadays nonalcoholic fatty liver disease (NAFLD) consider as multisystem disease that is primarily associated with components of the metabolic syndrome and is associated with cardiovascular and renal impairment. The comorbidity of NAFLD with renoparenchymal arterial hypertension (RPAH) has not been sufficiently studied. The purpose of the study was to investigate the influence of G276T genetic polymorphism of ADIPOQ on the severity of metabolic disorders, inflammation, liver, artery and kidney damage in the comorbidity of NAFLD and RPAH. Materials and methods. The study included 87 patients with comorbidity of NAFLD and RPAH, grade 2. The mean age of patients was 50.78 ± 9.35 years. The vast majority of patients were overweight or obese. The control group was composed of 20 healthy volunteers. Parameters of carbohydrate and lipid metabolism, liver and kidney’s function, adiponectin, fetuin-A, cytokeratin-18, pro-inflammatory cytokines levels were investigated. For diagnostic of non-alcoholic steatosis and indication parameters of arteries, an ultrasound method was used. Results. It was found that the T allele was detected in 62 (35.6%) patients of the main group, which was significantly (p<0,05) different from the control group (22.5%). In the presence of patients with comorbidity NAFLD and RPAH G/T and T/T genotypes, carbohydrate metabolism disorders are more pronounced than in the G/G genotype. Thus, index HOMA in this group was 4.52 ± 0.87, which was significantly higher than patients with G/G genotype - 3.77 ± 0.53 (p <0.01).The G276T polymorphism of the ADIPOQ is not associated with markers of liver and kidney damage in patients with NAFLD + RPAH. The presence of G/T and T/T genotypes in patients with comorbidity of NAFLD and RPAH is associated with an increase in interleukin-6 and fetuin-A compared to the G/G genotype. Patients with comorbidity of NAFLD + RPAH and with the T allele of the polymorphic marker G276T of the ADIPOQ gene are more likely to have impaired endothelium-dependent vasodilation, indicating more significant vascular reactivity disorders - 7.72 ± 1.25% for the genotype G/G, and 7.00 ± 1.10% for the genotype G/T (p <0,01). Conclusions. The presence of the T allele of the polymorphic marker G276T of the ADIPOQ is associated with the development of comorbidity of NAFLD and RPAH.

The efficacy of diuretics in complex treatment of patients with hypertension according to the Gly460Trp polymorphism of the α-adducin gene

The aim of the study was to investigate the effect of thiazide diuretics on blood pressure (BP) depending on Gly460Trp ADD1 gene polymorphism in arterial hypertension (AH) patients of the Ukrainian population in order to predict their individual treatment efficacy. Material and methods. The study included 232 persons: 120 patients with verified stage II AH and 112 healthy individuals. Restriction fragment length polymerase chain reaction (PCR-RFLP) was used to detect genotype (the Gly460Trp-polymorphic locus of the ADD1 gene). The patients received standard therapy, which included ACE inhibitor – ramipril 5 mg, calcium channel antagonist – amlodipine 5 mg, statin – atorvastatin 20 mg, acetylsalicylic acid 75 mg. The patients were randomized into two groups: group I (60 persons) additionally taking treatment with 1.5 mg of indapamide retard and group II (60 persons) – with 25 mg of hydrochlorothiazide. The dynamic reduction of blood pressure has been assessed every 4 weeks for 2 months. Results. Among 120 patients with AH, 91 persons (75.8 %) were homozygous for the G allele (GG), 26 persons (21.7 %) – heterozygous (GT) and 3 persons (2.5 %) – homozygous for the T allele (TT), while the G allele frequency in patients with hypertension was 0.87, and the T allele – 0.13. 98 healthy individuals (87.5 %) were homozygous for the G allele, 13 individuals (11.6 %) were heterozygous, and 1 person (0.9 %) was homozygous for the T allele. The carrier frequency of the G and T alleles was 0.93 and 0.07, respectively. Allelic distribution indicated the predominance of the G allele carriers by Gly460Trp polymorphism of the ADD1 gene among the Ukrainian population, regardless of whether AH symptoms were present. It is noteworthy that the number of the T allele carriers was 2 times large among symptomatic patients than that among healthy individuals. In patients with the T allele, the hypotensive efficacy of indapamide was almost 3 times higher than that in patients with the G allele. The antihypertensive effect of hydrochlorothiazide in patients with the GT and TT genotypes was 2 times greater than that in the GG genotype carriers depending on the presence of the T allele G460T polymorphism of ADD1 gene in the genotype. Conclusions. Allelic distribution indicates the predominance of the G allele carriers by Gly460Trp ADD1 gene polymorphism among the Ukrainian population, regardless of whether AH symptoms are present. Among patients with AH, the accumulation of the T allele G460T polymorphic marker of the α-adducin gene is 2 times more than that in healthy individuals. Patients carrying the T allele demonstrate 2 times higher hypotensive efficacy of indapamide compared with hydrochlorothiazide.

Laboratory predictors of pregnancy in vitro fertilization

Identification of factors determining both of favorable and unfavorable outcome of IVF will increase the effectiveness of this method and optimize infertility treatment. The aim of the research is to analyze the correlation between serum IL-1α concentration, its gene rs1800587 (C/T) genotype carrier and thyroid-stimulating hormone (TSH), thyroid hormones (triiodothyronine (T3) and tetraiodothyronine (T4)), and evaluate the prognostic significance of their combinations in women with tube-peritoneal infertility under the IVF program. 120 patients with tube-peritoneal infertility who applied for an IVF program were examined. Depending on the outcome of the procedure, 2 groups of patients were allocated: 1 group - 40 women who had a pregnancy after IVF, 2 group - 80 patients who did not have a pregnancy. The content of IL-1α, TSH, T3, T4 was determined in blood by ELISA. Genotyping was performed on the rs1800587 (C/T) polymorphic marker of the IL-1α gene. TSH, T3, T4 were within the norm for both groups. In our study, women with a TSH concentration of 0.23 to 1.7 nmol/L had a chance of a favorable IVF outcome 1.4 times higher than with other TSH levels (p = 0.042901); with a T3 level of 1.0 to 1.8 nmol/L had a chance of becoming pregnant 5.7 times higher than with other levels of T3 (p = 0.00002). For T4 concentration, the confidence test was not achieved (p = 0.068505). The individual indicators of IL-1α, TSH, T3 and carrier of the genotype of the gene IL-1α at the preconceptive stage have lower diagnostic value than their combined combination. Three combinations have maximum predictive value: a combination of the T/T genotype of the IL-1α gene and the TSH level of 0.23 to 1.7 nmol/l - OR = 8.1 (p = 0.000048); combination of IL-1α of 28.7 to 85.1 pg/ml, T/T gene genotype IL-1α and TSH level of 0.23 to 1.7 nmol/l - OR = 8.1 (p = 0.000048); combination of IL-1α of 28.7 to 85.1 pg/ml, T/T gene genotype IL-1α, TSH level of 0.23 to 1.7 nmol/l and T3 level of 1.0 to 1.8 nmol/l - OR = 8.1 (p = 0.000146). Thus, proposed new prognostic markers of IVF program effectiveness.