Molecular Screening of Some Apple Progenies for Detection Vf Gene Using Marker Assisted Selection

Apple scab, caused by Venturia inaequalis is one of the most damaging pathogens that affects apple species. Cross combinations were made between Auriu de Bistrița cv. (female genitor) - a valuable local variety in terms of fruit quality but only tolerant to scab, and Florina cv. (male genitor) used as a donor of Vf resistance gene. It was first detected in Malus floribunda Clone 821, which was later transferred to commercial varieties by different breeding programs. To confirm the presence of Vf gene, progenies resulting from the mentioned combination were tested with MAS (Marker Assited Selection), using two dominant primers pairs (AM19, U1400), and another one codominant (AL07) used to distinguish homozygous and heterozygous genotypes. After the crossing combination, a number of twenty-six hybrids were obtained, of which 50% (13 hybrids) were clasified as resistant (heterozygous), respectively 50% (13 hybrids) as susceptible (recessive homozygotes), so the Mendelian ratio was confirmed.

Pyramiding resistance genes and widening the genetic base of the apple (Malus × domestica Borkh.) crop

Apple breeding is active worldwide and yet the apple crop is in a precarious state as it relies on few dominant cultivars and only the Rvi6 (formerly Vf) gene, that confers resistance to scab, has been extensively exploited in the cultivars entered the market in recent years. However, there are some 20 disease resistance genes described in apple and the apple germplasm includes thousands of accessions in the repositories. In this paper, a breeding programme is described, whereby 36 genotypes, including ancient and contemporary apple cultivars, were crossed to produce a new set of selections that combine extensive genetic resources with pyramided resistance genes to several apple diseases, such as scab and powdery mildew. The 110 cross combinations carried out successfully, of the 260 initially planned, produced 7,876 offsprings, reduced to 2,969 after screening with molecular markers associated with five resistance genes. Selections with three or two resistance genes and good agronomic characteristics were kept for further field observations with the aims of creating new cultivars for the market and new parents for future breeding projects

Allelic Diversity in Apple Germplasm for Fruit Quality and Scab Resistance Using SSR Markers

The present investigation entitled “Allelic diversity in apple germplasm for fruit quality and scab resistance using SSR markers” was conducted in division of plant biotechnology, SKUAST-K, Shalimar during the year 2018. In the present study, four SSR markers namely Md-ACS-1, Md-ACO-1, ALO-7 and AM-19 were used for screening of 40 apple cultivars presently under cultivation in SKUAST-K for quality and scab resistance. The results revealed that out of 40 cultivars evaluated, 3 were found homozygous (ACS-1-2/2), 17 were heterozygous (ACS-1-1/2) and 10 were homozygous (ACS-1-1/1), while as no amplification of ACS-1 was found for 10 cultivars. Similarly, for Md-ACO-1, 3 cultivars were found homozygous, 35 as heterozygous and 2 did not show any amplification. As far as presence of Vf gene conferring resistance against scab disease, ALO-7 amplified at 820bp and 570bp alleles, 570 bp fragment was observed in resistant genotypes and was absent in susceptible genotypes, while the 820 amplicon was observed in both resistant and susceptible genotypes. The markers AM19 led to the amplification of 520 bp fragment in few cultivars.

Comprehensive Virulence Gene Profiling of Bovine Non-aureusStaphylococci Based on Whole-Genome Sequencing Data

ABSTRACTNon-aureusstaphylococci (NAS) are the most frequently isolated pathogens from intramammary infection (IMI) in dairy cattle. Virulence factors (VFs) and mechanisms by which NAS cause IMI are not fully known. Herein, we analyzed the distribution of 191 VFs in 441 genomes of 25 NAS species, after classifying VFs into functional categories: adherence (n = 28), exoenzymes (n = 21), immune evasion (n = 20), iron metabolism (n = 29), and toxins (n = 93). In addition to establishing VF gene profiles, associations of VF genes between and among functional categories were computed, revealing distinctive patterns of association among VFs for various NAS species. Associations were also computed for low, medium, and high somatic cell count (SCC) and clinical mastitis (CM) isolates, demonstrating distinctive patterns of associations for low SCC and CM isolates, but no differences between high SCC and CM isolates. To determine whether VF distributions had any association with SCC or CM, various clustering approaches, including complete linkages, Ward clustering, and t-distributed stochastic neighbor embedding, were applied. However, no clustering of isolates representing low SCC, medium SCC, or high SCC or CM was identified. Regression analysis to test for associations with individual VF functional categories demonstrated that each additional toxin and host immune evasion gene increased the odds of having high SCC or CM, although an overall increase in the number of VFs was not associated with increased SCC or occurrence of CM. In conclusion, we established comprehensive VF gene profiling, determined VF gene distributions and associations, calculated pathogenic potentials of all NAS species, and detected no clear link between VF genes and mastitis.IMPORTANCENon-aureusstaphylococci (NAS) are the most frequently isolated pathogens from milk in dairy cattle worldwide. The virulence factors (VFs) and mechanisms by which these bacteria cause udder infection are not fully known. We determined the distribution and associations of 191 VFs in 25 NAS species and investigated the relationship between VFs and disease. Although the overall number of VFs was not associated with disease severity, increasing numbers of toxin and host immune evasion genes specifically were associated with more severe disease outcomes. These findings suggest that the development of disease and the interactions of VFs with the host are complex and determined by the interplay of genes rather than just the presence of virulence genes. Together, our results provide foundational genetic knowledge to other researchers to design and conduct further experiments, focusing on understanding the synergy between VFs and roles of individual NAS species in IMI and characterizing species-specific effects on udder health.

The variability of Venturia inaequalis CKE. races in the Czech republic and the accumulation of resistance genes in apple germplasm

The growing of resistant apple varieties against the scab, impacts the Venturia inaequalis CKE. races development that can overcome the resistance. For this reason the main breeders object is to cumulate the different genetic mechanisms of resistance against this disease. Presented in this paper is the first genetic study of monosporic isolates in the Czech Republic. By means of RAPD and UPGMA methods which characterised the variability of 10 monosporic isolates from different localities and apple varieties. The monosporic isolate derived from the resistant genotype (Vf gene) proved a 79% genetic similarity with the isolate derived from sensitive variety Top Red. The genetic similarity of other isolates did not prove the dependence either on a locality or a host variety. The Vf and Vm genes accumulation in apple germplasm by means of specific PCR markers was studied. It was confirmed that Vf gene donors are always heterozygous. Concurrently it was statistically confirmed that the donor of Vm gene (OR-45-T-132) is heterozygous, too. The accumulation of Vf and Vm major genes against the scab was validated in 25% of seedlings of the cross.  

Alleles controlling apple skin colour and incompatibility in new Czech apple varieties with different degrees of resistance against Venturia inaequalis CKE.

The skin colour of 21 varieties was assessed on a molecular level. Four varieties had yellow skin colour with the genotype constitution a¹a¹, a¹a² or a²a². Seventeen varieties had dominant red colour. Homozygous A¹A¹ constitution was present in 7 varieties, and heterozygous constitution A¹a¹ or A¹a² was found in the rest of the assessed varieties. The S-alleles controlled the incompatibility system of the pistil. Their detection is possible by means of the PCR method on the basis of allele specific primers. Six S-alleles (S1, S2, S3, S5, S7 and S9) were studied in the same collection of 21 diploid Czech varieties. This paper brings new findings on S-allele characterization, because the Czech varieties have not yet been studied on a molecular level. Both types of S-allele were found in 12 varieties. Only one type of S-allele was described in 9 varieties. Simultaneously, the presence of the Vf gene was screened in the collection of 21 Czech apple varieties. Ten varieties with a field resistance against the scab had a heterozygous constitution of the Vf gene. All 11 susceptible varieties were recessive homozygous vfvf.

Six promising selections from the Hungarian apple breeding program for multiple resistance

An apple breeding program has been carried out at the Department of Fruit Science for more than a decade. Several apple selections have been released from the progenies of crosses in 1992 and 1993. Six candidates were submitted for national recognition out of the hybrids examined for more than a decade. The six selections are resistant against all the three most important apple diseases (apple scab, powdery mildew and fire blight). Scab resistance is controlled by the Vf gene originating from the species Malus floribunda 821 and transmitted by cultivar Prima. Heterozygote Vfvf genotype of the six cultivar candidates was proved by molecular genetic examinations of Dept. Genetics and Plant Breeding. Characteristics of these selections from 'Prima' progenies are shown on the base of our own observations.

Application of DNA markers for detection of scab resistant apple cultivars and selections

A DNA marker-based study was undertaken to identify the occurrence of major scab resistance genes in some apple cultivars and selections of importance for apple breeding. Unfortunately none of the RAPD-based markers previously reported to detect the Va, Vb, Vr and Vx genes produced unambiguous results. By contrast, the CAPS marker MI8 produced the expected three bands in all cultivars and selections already known or suspected to have the Vf gene, as well as in the Russian cultivar 'Antonovka Polotora Funtovaja' suspected to have Va resistance which however may be allelic to Vf. Vf-carrying selections and newly named cultivars 'Frida' and 'Fredrik' are grown successfully in Sweden without fungicides, suggesting that the Vr resistance breaking scab races 6 and 7 have not yet become a problem. The SCAR marker B12 detected the Vm gene in 'Prairifire', 'Rouville', clones 'OR45T132' and 'OR48T70', and selection '16-36-193'. The SSR locus 0102b10 detected one band at 118 by in 'Reka'. This is presumed to be identical to the Vr gene marker previously reported.