Polymerase Chain Reaction-restriction Fragment Length Polymorphism Research Articles

Promoter polymorphisms of neuropeptide Y, interleukin-1B and increased IL-1β levels are associated with rheumatoid arthritis susceptibility in South Gujarat population

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by joint destruction. Interleukin-1B(IL-1β) has been implicated in RA, and neuropeptide Y (NPY) induces the release of pro-inflammatory cytokines including IL1-β, TNF-α and IL-6. Single nucleotide polymorphisms (SNPs) in NPY and IL1B gene may lead to their altered levels leading to RA pathogenesis. Therefore, we aimed to investigate association of NPY rs16147(T > C) SNP, and genotype–phenotype correlation of the IL1B rs16944(C > T) SNP in 302 RA patients and 337 controls by Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Both the genotype and allele frequencies for NPY rs16147(T > C)(p = 0.0019 &p = 0.012) and IL1B rs16944(C > T)(p = 0.0024 &p = 0.0014) SNPs differed significantly between RA patients and controls. The odds ratios of 1.23 and 1.47 for NPY rs16147(T > C) and IL1B rs16944(C > T) SNPs respectively, suggest that these SNPs may be genetic risk factor for RA in South Gujarat population. The plasma IL-1β levels(p = 0.0332) and genotype-phenotype correlation of IL1B rs16944(C > T) SNP suggested increased IL-1β levels, particularly in patients with susceptible ‘TT' genotypes(p < 0.0001). Additionally, plasma IL-1β level was significantly increased in severe RA patients (p = 0.0025) and positively correlated with Rheumatoid factor(RF) (r = 0.9240, p < 0.0001), suggesting the role of IL-1β in disease severity. Overall, the study for the first time suggests that NPY rs16147(T > C) and IL1B rs16944(C > T) SNPs may be associated with RA susceptibility in South Gujarat population. Additionally, the IL1B rs16944 susceptible ‘TT' genotype may lead to increased IL-1β levels, thereby contributing to RA pathogenesis and severity.

Interaction between CETP Taq1B polymorphism and dietary patterns on lipid profile and severity of coronary arteries stenosis in patients under coronary angiography: a cross-sectional study

AimEvidence indicates there are still conflicts regarding CETP Taq1B polymorphism and coronary artery disease risk factors. Current findings about whether dietary patterns can change the relationship of the Taq1B on lipid profile and the severity of coronary arteries stenosis appears to be limited. The present research made an attempt to investigate this possible relationship.MethodsThis cross-sectional study involved 453 male and female participants with a mean age of 57 years. A validated 178-item food frequency questionnaire (FFQ) was used to assess dietary usual intake. Dietary patterns were extracted through principal component analysis (PCA). Taq1B variant was genotyped by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Two-way ANOVA was used to test the interaction between Taq1B polymorphism and dietary patterns.ResultsTwo dietary patterns were detected: the western dietary pattern (WDP) and the traditional dietary pattern (TDP). The frequency of Taq1B genotypes turned out to be 10.4, 72.4, and 17.2% for B1B1, B1B2, and B2B2, respectively. A significant difference was observed in TG and TG/HDL-C levels among TaqIB genotypes in higher adherence to TDP (P = 0.01 and P = 0.03, respectively). Taq1B showed a significant interaction with TDP for modulating TG levels and TG/HDL-C ratio (P = 0.02 and P = 0.04, respectively). Greater compliance to WDP demonstrated a significant difference in TG and TG/HDL-C levels across rs708272 genotypes (P = 0.03) after adjusting for confounding factors. Other lipid components and coronary arteries stenosis scores failed to show any relationship or significant difference across Taq1B genotypes or dietary patterns.ConclusionAdherence to TDP may adjust the association between the Taq1B variant and TG and TG/HDL-C levels in patients undergoing coronary angiography. To better understand the relationships, we suggest prospective studies in different race groups with multivariate approaches.

Genetic diversity of Toxoplasma gondii in goats and sheep from the Northeast Region of Brazil destined for human consumption

This study aimed to genotype isolates of Toxoplasma gondii obtained from samples of brain, diaphragm and heart of goats and sheep intended for human consumption in the State of Paraíba, Brazil. Tissue samples from 14 animals, goats (n = 5) and lambs (n = 9), were sourced from public slaughterhouses in seven cities and bio-assayed in mice. The brains of the mice were utilized for DNA extraction. Genotyping was carried out by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) using 10 markers (SAG1, SAG2, SAG3, BTUB, c22-8, PK1, GRA6, L358, c-29-2 and Apico). A total of 10 isolates were fully genotyped (i.e. at all loci), three from goats and seven from sheep, revealing five distinct genotypes: #13 (n = 4); #48 (n = 3); #57 (n = 1); #273 (n = 1); and one new genotype that had not been previously described. Genotype #13 is frequently found in the Northeast of Brazil and represents a clonal lineage circulating in this region and was the most prevalent genotype identified (n = 4). Moreover, in the present study genotypes #13, #48, #57, and #273 were documented for the first time in sheep from Brazil, and the novel genotype was isolated from a goat. Our findings align with previous studies on T. gondii from Brazil, where new genotypes are continuously being identified, highlighting a high level of genetic diversity of T. gondii isolates in the country.

Association of CYP1A2 Gene rs17861162 Polymorphism and Breast Cancer Risk in Iranian Women: A Case-control Study

Background: Breast cancer is the most commonly diagnosed malignant neoplasm among women worldwide. The significance of gene polymorphism in cytochrome P450 (CYP) 1A2 (CYP3A4) in cancer development is generally acknowledged. Objectives: The objective of this study was to investigate the possible association between the rs17861162 polymorphism of the CYP1A2 gene and vulnerability to breast cancer among women of Iranian heritage. Methods: In the current study, a cohort comprising 100 women who were diagnosed with breast cancer was identified as the case group in a case-control inquiry. Another cohort of 100 women in good health and residing in East Azerbaijan province, with similar age and ethnicity attributes to the individuals in the case group, were selected as the control group. The genomic DNA was obtained from leukocytes isolated from peripheral blood through a phenol-chloroform extraction method. The genotyping experiment was performed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Results: The findings of this study indicated that genotypic distribution among individuals diagnosed with breast cancer was 54% CC, 37% CG, and 9% GG. On the other hand, the healthy control group exhibited a distribution of 69% CC, 27% CG, and 2% GG. The statistical analysis revealed notable variations in the genotype and allele frequencies of the rs17861162 polymorphism in the CYP1A2 gene between people diagnosed with breast cancer and healthy control subjects (P&gt;0.05). Conclusions: The results of our study indicated a significant association between breast cancer and the rs17861162 polymorphism in the CYP1A2 gene among Iranian women. In order to gain a comprehensive understanding of the specific ramifications of this polymorphism concerning breast cancer prognosis, diagnosis, and risk, it is imperative to perform additional research that spans a wide range of ethnic and geographical populations.

Phylogenetic Variation of Tri1 Gene and Development of PCR-RFLP Analysis for the Identification of NX Genotypes in Fusarium graminearum Species Complex.

NX toxins have been described as a novel group of type A trichothecenes produced by members of the Fusarium graminearum species complex (FGSC). Differences in structure between NX toxins and the common type B trichothecenes arise from functional variation in the trichothecene biosynthetic enzyme Tri1 in the FGSC. The identified highly conserved changes in the Tri1 gene can be used to develop specific PCR-based assays to identify the NX-producing strains. In this study, the sequences of the Tri1 gene from type B trichothecene- and NX-producing strains were analyzed to identify DNA polymorphisms between the two different kinds of trichothecene producers. Four sets of Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods were successfully developed to distinguish the common type B trichothecene producers and NX producers within FGSC. These promising diagnostic methods can be used for high-throughput genotype detection of Fusarium strains as a step forward for crop disease management and mycotoxin control in agriculture. Additionally, it was found that the Tri1 gene phylogeny differs from the species phylogeny, which is consistent with the previous studies.

Detection of genetic variations in the GDF9 and BMP15 genes in Kazakh meat-wool sheep.

Kazakh meat-wool sheep are of great interest because of the intrabreed multifetal type's high productivity of 140 %-160 %. Genes encoding growth differentiation factor-9 (GDF9) and bone morphogenetic protein 15 (BMP15) are promising candidates for studying sheep productivity, as they affect fertility in mammals, including sheep. Thus, the purpose of this study was to assess the fertility of the Kazakh meat-wool sheep breed based on GDF9 and BMP15 candidate genes of fecundity for the selection of animals with valuable genotypes. We selected 300 heads of the Kazakh meat-wool sheep breed from two populations for PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) analysis, 15 of which were subsequently used for sequencing of exon regions of the GDF9 and BMP15 genes. The sheep populations were tested for G1 and G8 mutations of the GDF9 gene and B2 and B4 mutations of the BMP15 gene. The PCR-RFLP analysis revealed that 59 (19.7 %) of the 300 Kazakh meat-wool breed sheep were heterozygous carriers of the G1 mutation (genotype AG) of the GDF9 gene, and sequencing analysis supported these results. The comparative phylogenetic analysis showed a clear separation of Kazakh meat-wool sheep wild types and carriers of the G1 mutation. This mutation was reported to have a relationship with the animals' litter size in other sheep breeds. For this reason, similar relationships should be investigated in Kazakh meat-wool sheep. However, G8, B2, and B4 mutations were not detected among the studied animal populations, showing that these mutations are not characteristic of the Kazakh meat-wool sheep breed.

Epidemiological survey, molecular profiling and phylogenetic analysis of cutaneous leishmaniasis in Khyber Pakhtunkhwa, Pakistan.

Cutaneous leishmaniasis (CL), an emerging vector-borne ailment in Khyber Pakhtunkhwa (KPK), Pakistan, exhibits diverse spread patterns and outbreaks. To comprehend its epidemiology and identify parasite species, we conducted an active survey on suspected CL cases (n=8845) in KPK. Microscopy and internal transcribed spacer-1 PCR-restriction fragment length polymorphism (RFLP) molecular techniques detected Leishmania spp. in blood samples. Phylogenetic analysis gauged genetic affinities with other areas. District Bannu displayed the highest CL impact (14.58%), while Swat had the lowest impact (4.33%) among cases. Annual blood examination rate, parasite incidence and slide positivity rate were 4.96 per 1000 people, 0.0233 and 0.047%, respectively. CL infections were prevalent in 1- to 20-y-olds, with males (57.17%) more vulnerable than females (42.82%). Single lesions occurred in 43.73% of patients, while 31.2% people had two lesions, 17.31% had three lesions and 7.74% had more than three lesions. Most had sand-fly exposure but lacked preventive measures like repellents and bed nets. Leishmania tropica was confirmed via RFLP analysis in amplified samples. Phylogenetic analysis unveiled genetic parallels between L. tropica of KPK and isolates from China, Iran, Afghanistan, India, Syria and Morocco. Urgent comprehensive control measures are imperative. Early detection, targeted interventions and raising awareness of CL and sand-fly vectors are vital for reducing the disease's impact. International collaboration and monitoring are crucial to tackle Leishmania spp.'s genetic diversity and curtail its cross-border spread.

Identification of Genetic Diversity of P21-Activated Kinases (PAK1) Gene in Senduro Goats and Boerawa Goats using PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism)

This study aimed to detect genetic diversity associated with litter size traits of the P21-activated kinases (PAK1) gene in Senduro and Boerawa goats using the PCR-RFLP method. The restriction enzyme used is MSP1 (C*CGG). A pair of primers used in this study, namely Forward ((5'-GCTCAGTGTTGTATTAGCAG-3') and Reverse (5'- CAAGATATAAAGCACAGCCG-3') product length 713 bp (access code ENSCHIG00000003071). The samples used in this study consisted of 80 samples of Senduro goats and 20 samples of Boerawa goats from UPT PT-HMT Singosari, Malang Regency. The results showed that the PAK1 exon 9 gene in Senduro goats and Boerawa goats UPT PT-HMT Singosari was uniform (monomorphic), it was because all samples had GG genotype and G allele frequency of 100%. In conclusion, the PAK1 exon 9 / MSP1 gene is monomorphic and does not associate with litter size in Senduro goats and Boerawa goats, so it cannot be used as a genetic marker.

Intraspecifična karakterizacija izolata Vibrio alginolyticus prikupljenih iz sustava akvakulture i morskih biotopa u Tunisu korištenjem PCR-RFLP, ECP i OMPS profiliranja

A total of 54 presumptive Vibrio alginolyticus strains isolated from a variety of Tunisian marine biotopes, such as seawater, sediment samples from some bathing areas, and aquaculture farms, were characterized and identified by several biochemical tests. DNA polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis, extracellular products (ECP) and outer membrane protein (OMP) profiling were used to evaluate their usefulness as a tool to investigate the Vibrio alginolyticus diversity within this complex group. Results showed that there is great heterogeneity in the diversity observed via the PCR-RFLP method related to the number of genotypes generated by the two enzymes SduI and FaqI tested. This heterogeneity was observed not only according to the origin (seawater, sediment, fish and bivalve aquaculture farms) but also within the same type of sample. The two other methods, ECP and OMP resulted in 32 and 26 profiles, respectively. The discriminatory index determined in this study highlighted the good ability of the PCR-RFLP method to discriminate similar V. alginolyticus strains.

Fully automated ApoE4 assay in plasma and the estimation of the APOE polymorphism

AbstractBackgroundApolipoprotein E (ApoE) is a major cholesterol carrier that supports lipid transport and injury repair in the brain. APOE polymorphic alleles are the main genetic determinants of Alzheimer disease (AD) risk: individuals carrying the ε4 allele are at increased risk of AD compared with those carrying the more common ε3 allele, whereas the ε2 allele decreases risk. Moreover, it has been reported that the APOE4 haplotype has a dose‐dependent association with an increased risk of amyloid‐related imaging abnormalities (ARIA) in individuals receiving disease‐modifying therapy (DMT) for AD. Therefore, the importance of APOE genotyping or phenotyping and its expanding clinical role in dementia field has been highlighted. Here, we developed fully automated chemiluminescence enzyme‐immunoassay (CLEIA) kits for ApoE4 and pan‐ApoE, and evaluated its diagnostic concordance with the APOE genotyping.MethodApoE4 and pan‐ApoE assays for fully automated platform, LUMIPULSE, are a two‐step sandwich CLEIA with a specific treatment process to extract the antigen from the lipid surface in plasma samples during the 1st immunoreaction. The turnaround time for each reaction is within 30 minutes. Healthy volunteers’ samples were used for the measurement of ApoE4 and pan‐ApoE with CLEIA or the PCR‐Restriction fragment length polymorphism (PCR‐RFLP) analysis for the APOE polymorphism. The study design was approved by the ethics committee at H.U. Group Holdings, Inc in the 2022 fiscal year (22‐012‐01).ResultThe ApoE4 kit specifically detected the ApoE4 concentration in plasma samples, and could distinguish ApoE polymorphism. Moreover, the polymorphism could be classified clearly by the ratio of ApoE4 and Pan‐ApoE amount in plasma.ConclusionThe new CLEIA ApoE4 and pan‐ApoE assay robustly distinguishes among APOE4 carrier status in plasma. Assessing ApoE isoform by CLEIA may help not only clarify ApoE metabolism, but also ARIA risk.

Probiotics Potential of Lactic Acid Bacteria from Horse Milk in Tambora, West Nusa Tenggara, Indonesia

This study explores the probiotic potential of lactic acid bacteria (LAB) in horse milk from Tambora, West Nusa Tenggara by examining its functional and probiotic attributes, including bile salt tolerance, low pH, simulated gastric juice (SGJ), simulated intestinal juice (SIJ), antibacterial activity, as well as bile salt hydrolase (BSH) genes. Genotyping of LAB was performed using restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR) analysis and then identified based on the 16S rRNA gene. A total of 25 LAB isolates showed the ability to grow at low pH, tolerant to bile, and survived under SIJ and SGJ conditions. The BSH gene was confirmed in three isolates, namely: SK1-28, SK2-30, and SK2-34. Results of RFLP-PCR analysis showed that the LAB isolates were grouped into three groups based on the number and molecular weight of the differences DNA fragments. The 16S rRNA analysis showed that the first two groups were Lacticaseibacillus rhamnosus, whereas the third group was Lactiplantibacillus plantarum. In addition, all Lacticaseibacillus rhamnosusisolates in group I showed the ability to grow at pH 9.0, but not group II. It can be concluded that Lacticaseibacillus rhamnosus and Lactiplantibacillus plantarum can be used as the indigenous probiotic bacteria source from Indonesia.

Exploring the Influence of VDR Genetic Variants TaqI, ApaI, and FokI on COVID-19 Severity and Long-COVID-19 Symptoms.

There is increasing evidence regarding the importance of vitamin D in the prognosis of coronavirus disease 2019 (COVID-19). Genetic variants in the vitamin D receptor (VDR) gene affect the response to vitamin D and have been linked to various diseases. This study investigated the associations of the major VDR genetic variants ApaI, FokI, and TaqI with the severity and long post-infection symptoms of COVID-19. In total, 100 Jordanian patients with confirmed COVID-19 were genotyped for the VDR ApaI, FokI, and TaqI variants using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. COVID-19 severity, the most commonly reported long-COVID-19 symptoms that lasted for >4 weeks from the onset of infection, and other variables were analyzed according to VDR genetic variants. In this study, ApaI and FokI polymorphisms showed no significant associations with COVID-19 severity (p > 0.05). However, a significant association was detected between the TaqI polymorphism and the severity of symptoms after infection with the SARS-CoV-2 virus (p = 0.04). The wild-type TaqI genotype was typically present in patients with mild illness, whereas the heterozygous TaqI genotype was present in asymptomatic patients. With regard to long-COVID-19 symptoms, the VDR heterozygous ApaI and wild-type TaqI genotypes were significantly associated with persistent fatigue and muscle pain after COVID-19 (p ˂ 0.05). Most carriers of the heterozygous ApaI genotype and carriers of the wild-type TaqI genotype reported experiencing fatigue and muscle pain that lasted for more than 1 month after the onset of COVID-19. Furthermore, the TaqI genotype was associated with persistent shortness of breath after COVID-19 (p = 0.003). Shortness of breath was more common among individuals with homozygous TaqI genotype than among individuals with the wild-type or heterozygous TaqI genotype. VDR TaqI is a possible genetic variant related to both COVID-19 severity and long-COVID-19 symptoms among Jordanian individuals. The associations between VDR TaqI polymorphisms and long-COVID-19 symptoms should be investigated in larger and more diverse ethnic populations.

LEP (G2548A-G19A) and ADIPOQ (T45G-G276T) gene polymorphisms are associated with markers for metabolic syndrome

Background and aimsThere is a link between genetics with metabolic balance and adiposity homeostasis on metabolic syndrome (MetS). Polymorphism in adipokine genes such as leptin and adiponectin may play an important role in its development. This study aimed to determine the association of the individual and general components of MetS with genetic alterations in LEP (rs7799039 and rs2167270) and ADIPOQ (rs1501299 and rs2241766) genes in the Mexican population.Methods and resultsThe polymorphisms of the LEP gene rs7799039 and rs2167270, together with rs1501299 and rs2241766 polymorphisms of the ADIPOQ gene were genotyped using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) on 328 individuals (n = 131 MetS). The rs7799039 under the recessive inheritance model was found to be associated with increased risk of MetS (OR = 2.16, 95% CI = 1.06–4.37), dyslipidemia (OR = 7.97, 95% CI = 2.17–29.36), low HDL (OR = 7.01, 95% CI = 1.65–29.71) and hypertension (OR = 13.02, 95% CI = 1.76–96.44); the heterozygote demonstrate a protective effect on MetS (OR = 0.48, 95% CI = 0.28–0.88) and diabetes (OR = 0.09, 95% CI = 0.02–0.53) under the over the dominant model. Haplotype analysis showed linkage disequilibrium between the SNPs of ADIPOQ rs1501299/rs2241766, and their association as risk factors for low HDL and hypertension.ConclusionThe association of rs7799039 with the presence of MetS, suggests a risk factor for the development of dyslipidemia, as well as its heterozygous as a protective factor for DM. There is a linkage disequilibrium between the SNPs of ADIPOQ.

The ENPP1 K121Q polymorphism is associated with obesity-related parameters in Iranian normoglycemic male subjects

Objectives: The K121Q polymorphism of the Ectoenzyme Nucleotide Pyrophosphate Phosphodiesterase 1 (ENPP1) gene has been variably associated with obesity in several populations. However, this association has not been studied in Iranian subjects. It was hypothesized that the K121Q variant might be associated with obesity and related risk factors in subjects with normal glucose tolerance and normal fasting glucose.&#x0D; Methods: The K121Q genotypes were determined by PCR-restriction fragment length polymorphism in 377 non-diabetic subjects.&#x0D; Results: In males, the frequency of the Q allele was 20.8% and 15.9% (p=0.269) in obese and non-obese subjects, respectively. The ENPP1 genotype (KQ+QQ) was not associated with systolic and diastolic blood pressure, triglyceride, cholesterol, LDL-C, HDL-C, HOMA-IR, or insulin levels in both genders. Male carriers of the KQ+QQ genotype had significantly higher values for BMI (p=0.079), waist (p=0.01), and WHR (p=0.006) than subjects with the KK genotype. Obesity-related parameters were not significantly different between obese and non-obese female subjects.&#x0D; Conclusion: Our results suggest that the ENPP1 121Q allele might be associated with obesity and related parameters only among Iranian normoglycemic male subjects.

Polymorphism and promoter methylation-regulated mRNA expression of IFI6 gene affect meat quality in pigs

This study examined the association, promoter methylation of Interferon alpha-inducible protein 6 (IFI6) gene, and their effects on mRNA expression and meat quality traits in pig. Muscle samples (n=300) were randomly collected from the Longissimus thoracis et lumborum (LTL) to determine meat quality traits including drip loss, thawing loss, cooking loss, meat pH [at 45 minutes (pH45min) and at 24 hours (pH24h) post-mortem], shear force (for tenderness), and lightness (L*) in pigs. Genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Muscle samples (5 per group) with divergent meat pH (low vs high) were selected for promoter methylation study using bisulfate and pyrosequencing. Muscle samples from animals (5 per group) with divergent phenotypes (low vs high meat pH and low vs high drip loss) were used for mRNA expression study by quantitative real-time polymerase chain reaction (qRT-PCR). Results showed that a polymorphism in IFI6 gene (g.370A>G) was significantly (P < 0.01) associated with meat pH24h. Furthermore, four CpG regions were identified in IFI6 promoter, of which DNA methylation in a CG site was higher in low meat pH24h group compared to high meat pH24h group (P < 0.05). The mRNA expression of IFI6 gene was significantly (P < 0.01) downregulated in low meat pH24h group than high meat pH24h group. This result postulated that high DNA methylation at CG site in the promoter region might downregulate the IFI6 gene expression, hence lower the muscle pH that might lead to higher drip loss and lower tenderness. Therefore, porcine IFI6 gene might be a potential candidate gene for meat quality trait in pig. Nevertheless, further research is needed to confirm the role of IFI6 gene in controlling meat quality traits in other commercial populations.

The gene polymorphisms of eNOS and MTHFR modulates the development of preeclampsia in Han population

We explored the effects of the endothelial nitric oxide synthase (eNOS) gene −786 T > C and 894 G > T locus polymorphisms and the methylenetetrahydrofolate reductase (MTHFR) gene 1298 A > C and 677 C > T locus polymorphisms on preeclampsia (PE) in pregnant women in Quanzhou area and provide reliable and stable predictors PE. This study included 160 normal pregnant women (normal control group) and 160 women with preeclampsia (PE group). Polymorphisms in eNOS gene and MTHFR were analyzed by the polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) technique. eNOS 894 G > T locus and MTHFR 1298 A > C locus had no significant difference between the two groups. In the PE patients, eNOS −786 T allele (OR: 2.07, p = 0.03) and MTHFR 677 C allele (OR: 1.83, p = 0.04) had significantly lower frequency. The nitric oxide (NO) level in patients with eNOS −786 C C was significantly lower than that in those with −786 TT. The homocysteine (Hcy) level in patients with MTHFR 677 TT was significantly higher than that in those with 677 C C. In conclusion, the frequency of the eNOS −786 C C genotype and MTHFR 677 TT genotype are higher in women with PE, which cause lower NO level and higher Hcy level.

The effect of TRIM5 variants on the susceptibility to HIV-1 infection and disease progression in the Polish population.

Tripartite motif containing 5α protein is a factor contributing to intracellular defense mechanisms against human immunodeficiency virus-1 (HIV-1) infection. The studies of TRIM5 variants effects on the risk of HIV-1 infection and the clinical course of disease provided inconclusive results in different ethnic groups. The aim of this study was to investigate the influence of TRIM5 variants on susceptibility to HIV-1 infection and clinical parameters among Polish HIV-1-infected patients. In our study, we investigated 301 HIV-1-infected patients and 186 age-matched seronegative controls. Seven variants of the TRIM5 gene (rs7127617, rs3824949, rs3740996, rs11601507, rs10838525, rs11038628, and rs28381981) were genotyped using both sequencing and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) techniques. The frequencies of rs7127617 TT genotype and T allele occurrence were lower in HIV-1-infected subjects compared to controls (0.14 vs. 0.26 for T/T genotype and 0.45 vs. 0.54 for T allele), suggesting their possible protective effect (p=0.005 and p=0.007, respectively). Heterozygosity and presence of the T allele at rs3740996 were enriched in controls compared to HIV-1-infected group (0.19 vs. 0.12 for C/T genotype and 0.11 vs. 0.07 for T allele; p=0.03 and p=0.02, respectively). Moreover, rs3824949 CC genotype carriers had a lower viral load than patients bearing rs3824949 GG/CG genotypes (4.0 vs. 4.6 log copies/mL; p=0.049); however, none of the variants affected CD4+ cell count. In conclusion, our data confirm the role of TRIM5 variants in the HIV-1 transmission and the clinical course of HIV-1 infection. The presence of rs7127617 TT genotype and T allele seems to protect against HIV-1 transmission in examined population.

Correlation of Plasma Adiponectin Levels and Adiponectin Gene Polymorphisms with Idiopathic Atrial Fibrillation

Introduction: Adiponectin is a cellular protein secreted by adipocytes, which is closely related to a variety of diseases, including atrial fibrillation (AF). Idiopathic atrial fibrillation (IAF) is defined as AF without hypertension, diabetes, and other underlying diseases. Genetic polymorphism of adiponectin affects serum adiponectin concentration. However, the association of serum adiponectin concentration and its genetic polymorphism with IAF has not been studied. This study investigated the relationship between serum levels of adiponectin, adiponectin gene polymorphisms, and the risk of developing IAF in a Chinese Han population. Methods: Patients with IAF (n = 172, IAF group) and healthy individuals (n = 150, control group) were consecutively and randomly recruited and fasting peripheral blood samples were collected. All participants were examined for serum adiponectin concentrations and the polymorphisms SNP45T>G (SmaI locus, rs2241766) and SNP276G>T (BsmI locus, rs1501299) of the adiponectin gene were genotyped using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Related clinical data from the two groups were also collected. Results: Plasma adiponectin levels in the IAF group were significantly lower than those in the control group (9.9 ± 2.6 mg/L vs. 16.1 ± 7.0 mg/L, p = 0.006). There were no significant differences among the three genotypes (wild type, mutant heterozygote, and homozygote) of SNP45T>G or SNP276G>T in the prediction value of IAF. The frequency of the T allele of SNP45 T>G was 70.3% in the IAF group and significantly different from that of the control group (71.3%; p = 0.02). In the case of SNP276G>T, the frequency of the G allele was 68.61% in patients with IAF compared to 73.34% in the control group (p = 0.35). Furthermore, a comparison of the clinical data of individuals in the two groups revealed that the left atrial diameter (LAD) in patients in the IAF group was obviously higher than that in the control group (43.3 ± 6.7 mm vs. 37.9 ± 5.1 mm, respectively; p < 0.001). The left ventricular ejection fraction (LVEF) in the IAF group was obviously reduced than that in the control group (54.7 ± 11.9% vs. 60.2 ± 5.6%, respectively; p < 0.001). Conclusions: Low plasma adiponectin levels were significantly associated with IAF. Hypoadiponectinemia can thus serve as an important factor for the incidence of IAF. The genotypes of SNP45T>G and SNP276G>T in the adiponectin gene may not correlate with the occurrence of IAF. However, our results demonstrate that the T allele of SNP45T>G may be responsible for IAF development in the Chinese Han population.

Molecular diagnosis, phylogenetic analysis, and antifungal susceptibility profiles of Candida species isolated from neutropenic oncological patients

BackgroundNeutropenia is the most important cause of life-threatening invasive fungal infections (IFIs). Here, we studied the frequency and antifungal susceptibility profiles of Candida species that colonized or caused infections among neutropenic patients with solid or hematological malignancies.MethodsA total of 362 clinical samples were collected from 138 patients. After initial isolation using a mix of mycological methods, isolates were screened using chromogenic culture media. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was applied for molecular identification. Positive or suspected cases were confirmed using the reference method of sequencing. Antifungal susceptibility testing for voriconazole and caspofungin was carried out using the microbroth dilution method. An in-silico assay was applied for phylogenetic analysis.ResultsThirty-four Candida strains were isolated. C. albicans (47.06%) and C. glabrata (29.41%) were the most frequent strains. Antifungal treatment reduced the chance of Candida colonization by almost 76% in neutropenic patients (OR: 1.759; 95% CI: 1.349 to 2.390; p value: 0.000). An unusual and non-resistant strain, C. lambica, was reported from the bloodstream of a 56-year-old man with hematologic malignancy (HM). Eight isolates were non-susceptible, and one isolate was resistant to voriconazole. Also, four isolates were non-susceptible to caspofungin.ConclusionWe can conclude that there is a cause-and-effect relationship between neutropenia, HM background, and Candida species separated from neutropenic patients, which can lead to possible infections. Further and repetitive studies are recommended using different molecular methods for better prediction and management of fungal infections in neutropenic patients.

Prevalence of 4a/4b polymorphic variants of the eNOS gene introne in patients with different types of encephalopathies

Questions related to the disruption of vasoregulatory processes as essential factor in neurological pathology require further study. The leading role in the vasoregulatory mechanisms is played by endothelial NO synthase which gene has 15 allelic variants. Recent data indicated a probable association between eNOS gene polymorphism and cerebrovascular diseases. The aim of the present work was to study the prevalence of the 4a/4b introne polymorphism of the eNOS gene in patients with various types of encephalopathies and to evaluate the influence of a particular genotype of the studied gene on the occurrence and/or progression of encephalopathy. A total of 96 patients with encephalopathies of various genesis: chronic traumatic encephalopathy, chronic alcohol-induced encephalopathy, chronic vascular encephalopathy, post-infectious encephalopathy were involved in the study. The patients received inpatient treatment in the neurological department of “Ternopil Regional Clinical Psychoneurological Hospital”. Molecular and genetic differentiation of the studied gene variants was carried out by allele-specific PCR or PCR-restriction fragment length polymorphism analysis by standard operational protocols. Analysis of the frequency distribution of eNOS gene 4a/4b polymorphic variant showed that 4b allele prevailed among patients with all types of encephalopathies carriers­. Relative to practically healthy individuals the difference was found only in patients with chronic vascular encephalopathy (CVE), among whom about 39% were carriers of the 4a allele. The presence of the 4a allele was shown to increased the risk of CVE occurrence and/or progression by 4.5 times. The results obtained suggest the reasonability to include the 4a/b intron polymorphism of the eNOS gene in a genetic panel to monitor patients CVE. Keywords: 4a/4b introne, encephalopathies, endothelial NO-synthase, eNOS gene polymorphism