Molecular characteristics of S-RNase alleles as the determinant of self-incompatibility in the style of Fragaria viridis

Strawberry (Fragaria spp.) is a member of the Rosoideae subfamily in the family Rosaceae. The self-incompatibility (SI) of some diploid species is a key agronomic trait that acts as a basic pollination barrier; however, the genetic mechanism underlying SI control in strawberry remains unclear. Two candidate S-RNases (Sa- and Sb-RNase) identified in the transcriptome of the styles of the self-incompatible Fragaria viridis 42 were confirmed to be SI determinants at the S locus following genotype identification and intraspecific hybridization using selfing progenies. Whole-genome collinearity and RNase T2 family analysis revealed that only an S locus exists in Fragaria; however, none of the compatible species contained S-RNase. Although the results of interspecific hybridization experiments showed that F. viridis (SI) styles could accept pollen from F. mandshurica (self-compatible), the reciprocal cross was incompatible. Sa and Sb-RNase contain large introns, and their noncoding sequences (promotors and introns) can be transcribed into long noncoding RNAs (lncRNAs). Overall, the genus Fragaria exhibits S-RNase-based gametophytic SI, and S-RNase loss occurs at the S locus of compatible germplasms. In addition, a type of SI-independent unilateral incompatibility exists between compatible and incompatible Fragaria species. Furthermore, the large introns and neighboring lncRNAs in S-RNase in Fragaria could offer clues about S-RNase expression strategies.

Toxoplasma gondii in Slaughtered Sheep in High- and Low-Humidity Regions in the South of Iran: Molecular Prevalence and Genotype Identification

Toxoplasma gondii is one of the most common meat-born zoonoses that infect all warm-blooded animals and humans. Sheep (Ovis aries) is one of the main reservoirs of T. gondii worldwide, and the infections induce various sequels, such as abortion and stillbirth. The present study aimed to identify the effects of humidity on the prevalence of T. gondii in sheep in high- and low-humidity regions. Heart samples from 200 slaughtered sheep (140 samples from a high-humidity region and 60 samples from a low-humidity region) were collected from Hormozgan Province (south of Iran). The samples were tested by nested PCR targeting the RE gene. Genotyping was performed by the PCR-RFLP method using the SAG3 and GRA6 genes. Some isolates were sequenced and recorded in the GenBank. T. gondii DNA was detected in 10.71 percent of the samples from the highly humid region, whereas no positive samples were detected in the low-humidity region. Genotyping revealed that all isolates belonged to the T. gondii type III genotype. Our study indicated that humidity is an important factor for the prevalence of T. gondii in sheep. Additionally, our study also showed the dominance of type III strain of T. gondii in sheep in the south of Iran.

Genotype Heterogeneity in Accessions of a Winter Barley Core Collection Assessed on Postulated Specific Powdery Mildew Resistance Genes

Gene bank accessions are necessary for implementing many research and breeding projects. However, a great number of accessions are contaminated or confused. If such accessions are used, the results obtained from these projects are inaccurate and non-reproducible. There are methods that allow almost perfect genotype identification; nevertheless, they are relatively recent and results cannot be compared with the characteristics of the original accessions. Growing resistant cultivars is an environmentally safe and cheap way of disease management and knowledge of diverse resistance genes and their combinations can be used to identify varieties and verify their authenticity and homogeneity. For this purpose, all 172 accessions of the core collection (CC) of the Czech winter barley (Hordeum vulgare) gene bank, originating from 35 countries, were studied. For resistance tests, 51 reference isolates of Blumeria graminis f. sp. Hordei, collected in all nonpolar continents over a period of 63 years and representing the global virulence/avirulence diversity of the pathogen, were used. Only 25 barley accessions were homogeneous (genetically uniform), whereas 147 accessions were heterogeneous due to presence of different genotypes. In total, 17 resistance genes were found singly or in combinations; 76.3% of accessions with identified resistance genes carried alleles at the Mla locus. To purify the CC, progenies of individual plants must be multiplied and authenticity and homogeneity of the seed should be confirmed with resistance tests, and subsequently can be studied with more advanced methods.

Identification of Echinococcus granulosus Genotypes G1 and G3 by SNPs Genotyping Assays

Echinococcus granulosus sensu lato (s.l.) is the causative agent of cystic echinococcosis in animals and humans. Different E. granulosuss.l. genotypes exhibit great diversity in their life cycle, host selectivity and pathogenicity. For this reason, the study of genetic variation within Echinococcus species is of importance for their epidemiological implication. We employed two SNP genotyping technologies to distinguish G1 and G3 E. granulosus sensu stricto (s.s.). genotypes. The genotypes of DNA samples (n = 28) extracted from hydatid cysts of different animal species were identified by amplification and sequencing of a fragment of the mitochondrial nad5 gene. Two SYBR green and three TaqMan real time PCR assays were developed for targeting of three nad5 informative positions (SNP758, 1123, and 1380) known to be able to discriminate G1 from G3. Genotyping by SYBR Green PCR based on cycle threshold (Ct) with melting temperature (Tm) analysis and performed on SNP1123 and SNP1380 failed to identify one DNA sample. TaqMan assays for SNP758, 1123 and 1380 effectively confirmed genotype identification obtained by Sanger sequencing. Our results demonstrated that the combination of the three Taqman assays developed in this study represents a valuable and cost effective tool alternative to DNA sequencing for E. granulosus s.s. genotyping.

Phylogenetic Characterization of the 5′ Untranslated Region of Untypable HCV Genotypes Circulating in Pakistan

<b><i>Introduction:</i></b> Commercial methods for HCV genotyping is challenged by the increased prevalence of untypable genotypes in Pakistan. <b><i>Objective:</i></b> The aim of the current study was to perform nucleotide sequencing of 5′ UTR region for genotyping of viral isolates circulating in Peshawar, Pakistan. <b><i>Methods:</i></b> The total number of commercially untypable samples were 94 in which 18 samples were sequenced for the characterization of 5′ UTR region. Post-sequencing analysis was performed for genotype identification (<i>n</i> = 18) and molecular phylogenetic analysis. <b><i>Results:</i></b> The current study reveals different genotypes, that is, 10/18 viral isolates were found to be genotype 3a (55.55%), 3 isolates (genotype 3b, 16.66%), 2 isolates (genotype 6h/6g, 11.11%), 2 (6g/d, 11.11%), and 1 sample (genotype 1c, 5.55%). In addition, genotype 3a is the dominant representative of HCV circulating in Pakistan and has been increasing across the country. <b><i>Conclusion:</i></b> The current study also reveals that genotype 6 (2 were genotype 6h/6g and 2 were 6g/d) is also circulating in Pakistan and not restricted to South China and Hong Kong.

Genotype identification and diversity analysis in Kiwifruit (Actinidia spp.) using RAPD markers

Kiwifruit (Actinidia spp.) is a significant plantation crop belonging to family Actinidiaceae, having deciduous, dioecious, and scrambling vines with chromosome number 2n=58. Dioecy in kiwifruit forms the basis for several breeding programs. The present study was carried out for diversity analysis in kiwifruit genotypes using RAPD markers. 7 kiwifruit genotypes (2 males viz. Allision & Tomuri and 5 females viz. Hayward, Bruno, Allision, Monty & Abbott) were analysed for molecular polymorphism using 94 RAPD primers, out of which 23 primers amplified the genomic DNA in all the genotypes. RAPD data was analysed using NTSYS-pc software and dendrogram construction was done using UPGMA method. Two separate clusters of male and female genotypes were formed. Similarity matrix indices showed maximum similarity between Tomuri (M) and Allision (M) with a similarity coefficient of 0.719 while Abbott (F) and Allision (M) were found to have least similarity having a similarity coefficient of 0.521. Four RAPD primers amplified unique amplicons in Monty, Hayward, Bruno, Allision (M) and Abbott and two primers amplified unique amplicons in Allision (M) & Tomuri (M) along with the male and female plants of Allision genotype respectively. Therefore, these primers can help in distinguishing the genotypes of kiwifruit and can also be validated as putative markers for the sex identification in kiwifruit.