Genetic Complexity and Synteny Analysis of Castanea Genomes: Unveiling the Significance of Chestnut Species in Ecological and Genomic Perspectives

Castanea, a prominent genus within the Fagaceae family, thrives across the expansive woodlands of eastern North America, Europe, and Asia, holding considerable ecological and economic significance. Among the invaluable forest resources, chestnuts play a pivotal role by providing both nourishment and wood products. Furthermore, they assume the status of keystone species due to their indispensable ecological functions in afforestation and the provision of crucial ecosystem services. The genomes of C. mollissima and C. sativa, estimated to be around 800 Mb in size, add to the remarkable genetic complexity of these species. We utilized the powerful Python module jcvi to conduct a thorough synteny analysis, focusing on the Chinese and European chestnut genomes. To detect structural variants between these genomes, we employed SyRI (Synteny and Rearrangement Identifier). Additionally, TBtools was utilized to visually illustrate the syntenic genes across different genomes. Our investigation involved a comprehensive synteny analysis of the Chinese chestnut genome and the Sariaslama cultivar of the European chestnut. Encouragingly, we observed a strong overall synteny between these genomes, indicating significant conservation. To enhance the accuracy and completeness of the genome assemblies, we employed Pacbio sequencing technology, which contributed to the highquality results obtained for both the European and Chinese chestnut genomes.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202

De novo Genome Assembly of Sweet Chestnut (Castanea sativa Mill.) Insights into the Molecular Basis of its Nutritional Properties

The Sweet Chestnut (Castanea sativa Mill.) is a tree species that holds significant economic importance and naturally spreads throughout central-southern Europe and Asia Minor. Its highly nutritious nuts have a unique composition that sets them apart from other nuts, being rich in vitamins, including vitamin C, and B vitamins such as thiamine, niacin, and folate. Over the last few decades, breeding efforts have prioritized the development of sweet chestnut cultivars that are resistant to blight and produce better nuts. However, despite these efforts, molecular genetic studies of the sweet chestnut have been insufficient. To bridge this knowledge gap, we set out to create the first reference genome of the sweet chestnut using whole-genome shotgun paired-end sequencing. Our study involved genome-wide analyses to identify and functionally annotate genes in sweet chestnut, and develop and confirm SSR-SNP markers. Additionally, we have identified and characterized specific genomic loci that enhance the nutritional value of sweet chestnuts. To the best of our knowledge, this is the first study to investigate the genetic loci responsible for determining the nutritional value of chestnuts. We anticipate that our findings will significantly contribute to the development of sweet chestnut cultivars with higher levels of bioactive compounds, minerals, and digestibility, ultimately enhancing the nutritional value of chestnuts.Book of abstract: 4th Belgrade Bioinformatics Conference, June 19-23, 202

Genetic and biochemical differences in populations bred for extremes in maize grain methionine concentration

Background: Methionine is an important nutrient in animal feed and several approaches have been developed to increase methionine concentration in maize (Zea mays L.) grain. One approach is through traditional breeding using recurrent selection. Using divergent selection, genetically related populations with extreme differences in grain methionine content were produced. In order to better understand the molecular mechanisms controlling grain methionine content, we examined seed proteins, transcript levels of candidate genes, and genotypes of these populations. Results: Two populations were selected for high or low methionine concentration for eight generations and 40 and 56% differences between the high and low populations in grain methionine concentration were observed. Mean values between the high and low methionine populations differed by greater than 1.5 standard deviations in some cycles of selection. Other amino acids and total protein concentration exhibited much smaller changes. In an effort to understand the molecular mechanisms that contribute to these differences, we compared transcript levels of candidate genes encoding high methionine seed storage proteins involved in sulfur assimilation or methionine biosynthesis. In combination, we also explored the genetic mechanisms at the SNP level through implementation of an association analysis. Significant differences in methionine-rich seed storage protein genes were observed in comparisons of high and low methionine populations, while transcripts of seed storage proteins lacking high levels of methionine were unchanged. Seed storage protein levels were consistent with transcript levels. Two genes involved in sulfur assimilation, Cys2 and CgS1 showed substantial differences in allele frequencies when two selected populations were compared to the starting populations. Major genes identified across cycles of selection by a high-stringency association analysis included dzs18, wx, dzs10, and zp27. Conclusions: We hypothesize that transcriptional changes alter sink strength by altering the levels of methionine-rich seed storage proteins. To meet the altered need for sulfur, a cysteine-rich seed storage protein is altered while sulfur assimilation and methionine biosynthesis throughput is changed by selection for certain alleles of Cys2 and CgS1

Simultaneously Targeting the NS3 Protease And Helicase Activities For More Effective Hepatitis C Virus Therapy

This study examines the specificity and mechanism of action of a recently reported hepatitis C virus (HCV) non-structural protein 3 (NS3) helicase-protease inhibitor (HPI), and the interaction of HPI with the NS3 protease inhibitors telaprevir, boceprevir, danoprevir, and grazoprevir. HPI most effectively reduced cellular levels of subgenomic genotype 4a replicons, followed by genotypes 3a and 1b replicons. HPI had no effect on HCV genotype 2a or dengue virus replicon levels. Resistance evolved more slowly to HPI than telaprevir, and HPI inhibited telaprevir-resistant replicons. Molecular modeling and analysis of the ability of HPI to inhibit peptide hydrolysis catalyzed by a variety of wildtype and mutant NS3 proteins suggested that HPI forms a bridge between the NS3 RNA-binding cleft and an allosteric site previously shown to bind other protease inhibitors. In most combinations, the antiviral effect of HPI was additive with telaprevir, boceprevir, minor synergy was observed with danoprevir and modest synergy was observed with grazoprevir

The Molecular Basis of Drug Resistance against Hepatitis C Virus NS3/4A Protease Inhibitors

Hepatitis C virus (HCV) infects over 170 million people worldwide and is the leading cause of chronic liver diseases, including cirrhosis, liver failure, and liver cancer. Available antiviral therapies cause severe side effects and are effective only for a subset of patients, though treatment outcomes have recently been improved by the combination therapy now including boceprevir and telaprevir, which inhibit the viral NS3/4A protease. Despite extensive efforts to develop more potent next-generation protease inhibitors, however, the long-term efficacy of this drug class is challenged by the rapid emergence of resistance. Single-site mutations at protease residues R155, A156 and D168 confer resistance to nearly all inhibitors in clinical development. Thus, developing the next-generation of drugs that retain activity against a broader spectrum of resistant viral variants requires a comprehensive understanding of the molecular basis of drug resistance. In this study, 16 high-resolution crystal structures of four representative protease inhibitors - telaprevir, danoprevir, vaniprevir and MK-5172 - in complex with the wild-type protease and three major drug-resistant variants R155K, A156T and D168A, reveal unique molecular underpinnings of resistance to each drug. The drugs exhibit differential susceptibilities to these protease variants in both enzymatic and antiviral assays. Telaprevir, danoprevir and vaniprevir interact directly with sites that confer resistance upon mutation, while MK-5172 interacts in a unique conformation with the catalytic triad. This novel mode of MK-5172 binding explains its retained potency against two multi-drug-resistant variants, R155K and D168A. These findings define the molecular basis of HCV N3/4A protease inhibitor resistance and provide potential strategies for designing robust therapies against this rapidly evolving virus

Femtosecond laser directed fabrication of optical diffusers

Optical diffusers are widely used in filament lamps, imaging systems, display technologies, lasers, and Light Emitting Diodes (LEDs). Here, a method for the fabrication of optical diffusers through femtosecond laser machining is demonstrated. Float glass surfaces were ablated with femtosecond laser light to form nanoscale ripples with dimensions comparable to the wavelength of visible light. These structures produce highly efficient and wide field of view diffusers. The machined patterns altered the average surface roughness, with the majority of particles in the range of a few hundred nanometers. The optical diffusion characteristic and a maximum diffusion angle of near 172° was achieved with optimum machining parameters. The transmission performance of the diffusers was measured to be ∼30% across the visible spectrum. The demonstrated technique has potential for producing low-cost large area optical devices. The process benefits from the flexibility of the laser writing method and enables the production of custom optical diffusers

Differing Von Hippel Lindau Genotype in Paired Primary and Metastatic Tumors in Patients with Clear Cell Renal Cell Carcinoma

In sporadic clear cell renal cell carcinoma (CCRCC), the von Hippel Lindau (VHL) gene is inactivated by mutation or methylation in the majority of primary (P) tumors. Due to differing effects of wild-type (WT) and mutant (MT) VHL gene on downstream signaling pathways regulating angiogenesis, VHL gene status could impact clinical outcome. In CCRCC, comparative genomic hybridization analysis studies have reported genetic differences between paired P and metastatic (M) tumors. We thus sequenced the VHL gene in paired tumor specimens from 10 patients to determine a possible clonal relationship between the P tumor and M lesion(s) in patients with CCRCC. Using paraffin-embedded specimens, genomic DNA from microdissected samples (>80% tumor) of paired P tumor and M lesions from all 10 patients, as well as in normal tissue from 6 of these cases, was analyzed. The DNA was used for PCR-based amplification of each of the 3 exons of the VHL gene. Sequences derived from amplified samples were compared to the wild-type VHL gene sequence (GenBank Accession No. AF010238). Methylation status of the VHL gene was determined using VHL methylation-specific PCR primers after DNA bisulfite modification. In 4/10 (40%) patients the VHL gene status differed between the P tumor and the M lesion. As expected, when the VHL gene was mutated in both the P tumor and M lesion, the mutation was identical. Further, while the VHL genotype differed between the primary tumor in different kidneys or multiple metastatic lesions in the same patient, the VHL germline genotype in the normal adjacent tissue was always wild-type irrespective of the VHL gene status in the P tumor. These results demonstrate for the first time that the VHL gene status can be different between paired primary and metastatic tissue in patients with CCRCC

Evaluation of a remote-controlled laparoscopic camera holder for basic laparoscopic skills acquisition : a randomized controlled trial

Unsteady camera movement and poor visualization contribute to a difficult learning curve for laparoscopic surgery. Remote-controlled camera holders (RCHs) aim to mitigate these factors and may be used to overcome barriers to learning. Our aim was to evaluate performance benefits to laparoscopic skill acquisition in novices using a RCH. Novices were randomized into groups using a human camera assistant (HCA) or the FreeHand v1.0 RCH and trained in the (E-BLUS) curriculum. After completing training, a surgical workload questionnaire (SURG-TLX) was issued to participants. Forty volunteers naïve in laparoscopic skill were randomized into control and intervention groups (n = 20) with intention-to-treat analysis. Each participant received up to 10 training sessions using the E-BLUS curriculum. Competency was reached in the peg transfer task in 5.5 and 7.6 sessions for the ACH and HCA groups, respectively (P = 0.015), and 3.6 and 6.8 sessions for the laparoscopic suturing task (P = 0.0004). No significance differences were achieved in the circle cutting (P = 0.18) or needle guidance tasks (P = 0.32). The RCH group experienced significantly lower workload (P = 0.014) due to lower levels of distraction (P = 0.047). Remote-controlled camera holders have demonstrated the potential to significantly benefit intra-operative performance and surgical experience where camera movement is minimal. Future high-quality studies are needed to evaluate RCHs in clinical practice. ISRCTN 8373397

Essential oils from Egyptian aromatic plants as antioxidant and novel anticancer agents in human cancer cell lines

Inhibitors of tumor growth using extracts from aromatic plants are rapidly emerging as important new drug candidates for cancer therapy. The cytotoxicity and <em>in vitro</em> anticancer evaluation of the essential oils from thyme, juniper and clove has been assessed against five different human cancer cell lines (liver HepG2, breast MCF-7, prostate PC3, colon HCT116 and lung A549). A GC/MS analysis revealed that α-pinene, thymol and eugenol are the major components of Egyptian juniper, thyme and clove oils with concentrations of 31.19%, 79.15% and 82.71%, respectively. Strong antioxidant profiles of all the oils are revealed <em>in vitro</em> by DPPH and β-carotene bleaching assays. The results showed that clove oil was similarly potent to the reference drug, doxorubicin in prostate, colon and lung cell lines. Thyme oil was more effective than the doxorubicin in breast and lung cell lines while juniper oil was more effective than the doxorubicin in all the tested cancer cell lines except prostate cancer. In conclusion, the essential oils from Egyptian aromatic plants can be used as good candidates for novel therapeutic strategies for cancer as they possess significant anticancer activity.<br><br>Los inhibidores de crecimiento de tumores usando extractos de plantas aromáticas están emergiendo con rapidez como nuevos e importantes medicamentos para el tratamiento del cáncer. La citotoxicidad y la acción anticancerígena <em>in vitro</em> de aceites esenciales de tomillo, enebro y clavo han sido evaluadas en cinco líneas celulares de cáncer humano (hígado HepG2, mama MCF-7, próstata PC3, colon HCT116 y pulmón A549). Los análisis de GC/MS mostraron que α-pineno, timol y eugenol son los principales componentes de los aceites egipcios de enebro, tomillo y clavo, con concentraciones de 31,19%, 79,15% y 82,71%, respectivamente. Se demuestra, mediante ensayos <em>in vitro</em> de blanqueo de DPPH y β-caroteno, el enérgico perfil antioxidante de todos los aceites. Los resultados mostraron que el aceite de clavo fue similar de potente al fármaco de referencia, doxorrubicina en las líneas celulares de próstata, colon y pulmón. El aceite de tomillo fue más efectivo que la doxorrubicina en las líneas celulares de mama y de pulmón, mientras que el aceite de enebro fue más eficaz que la doxorrubicina en todas las líneas celulares de cáncer ensayados, excepto en la de cáncer de próstata. En conclusión, los aceites esenciales de plantas aromáticas egipcias se pueden utilizar como buenos candidatos para nuevas estrategias terapéuticas para el cáncer al poseer una significativa actividad anticancerígena

Relationship between Helicobacter pylori Virulence Genes and Clinical Outcomes in Saudi Patients

Helicobacter pylori has been strongly associated with gastritis, gastric and duodenal ulcers, and it is a risk factor for gastric cancer. Two major virulence factors of H. pylori have been described: the cytotoxin-associated gene product (cagA) and the vacuolating toxin (vacA). Since considerable geographic diversity in the prevalence of H. pylori virulence factors has been reported, the aim of this work was to determine if there is a significant correlation between different H. pylori virulence genes (cagA and vacA) in 68 patients, from Saudi Arabia, and gastric clinical outcomes. H. pylor was recognized in cultures of gastric biopsies. vacA and cagA genes were detected by polymerase chain reaction (PCR). The cagA gene was obtained with 42 isolates (61.8%). The vacA s- and m- region genotypes were determined in all strains studied. Three genotypes were found: s1/m1 (28%), s1/m2 (40%) and s2/m2 (26%). The s2/m1 genotype was not found in this study. The relation of the presence of cagA and the development of cases to gastritis and ulcer was statistically significant (P < 0.05). The study showed a significant correlation between the vacA s1/m2 genotype and gastritis cases, and a significant correlation between vacA s1/m1 genotype and peptic ulcer cases. The results of this study might be used for the identification of high-risk patients who are infected by vacA s1/m1 genotype of H. pylori strains. In conclusion, H. pylori strains of vacA type s1 and the combination of s1/m1 were associated with peptic ulceration and the presence of cagA gene