Advances and outlook of horticultural bioengineering

A Branch of modern biotechnology for creating unique relevant genotypes is bioengineering that harnesses a spectrum of plant genome modification technologies. The study aimed to analyse the current state of the art in genome modification of fruit and berry crops for more significant (vs. premium pure breeding varieties) deviations of norm in the traits and properties of biotic and abiotic resistance, productivity, fruit quality, etc. First horticultural crop transformation studies aimed at developing protocols based on selectable enzyme marker genes of phosphorylationmediated aminoglycoside antibiotics detoxification. Neomycin phosphotransferase nptII constitutes the most common system of transgenic fruit and berry crop selection. In pome crops, the transgenic selection priorities were resistance to scab (Venturia inaequalis (Wint.) Cke), rust (Gymnosporangium juniper-virginianae Schwein.) and bacterial blight (Erwinia amylovora Burrill, Winslow et al.), higher fruit quality, including bright colouring, and reduced enzymatic browning. In stone crops, it was tolerance to plum pox (PPV), papaya ringspot (PRSV) and Prunus necrotic ringspot (PNRSV) viruses. In berry crops — resistance to Sphaerotheca humuli (DC.) Burrill fungus, grey mould (Botrytis cinerea Pers.), root rot (Phytophthora cactorum (Lebert & Cohn) J.Schrot.) and powdery mildew (Oidium tuckeri Berkeley), as well as higher fruit quality. In citruses — resistance to bacterial canker (Xanthomonas citri sub sp.), citrus ulcer (Xanthomonas axonopodis pv citri), greening disease (Huanglongbing (HLB)) and fungi (Trichoderma harzianum Rifai). In tropical crops — resistance to papaya ringspot (PRSV) and banana streak (eBSV) viruses. Unique FT-phenotype transgenic fruit lines are leveraged in the new FasTrack breeding strategy. Nine fruit and berry transgenic crop lines have now been registered worldwide. Transgenic Arctic apples (Golden, Granny, Fuji), plums (Honey Sweet) and papaya (Rainbow, SunUp, Laie Gold) are industry-approved in fresh and processed form. The transgenic list regulated in the Russian Federation does not include fruit or berry crops.

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.

Successful intergeneric transfer of a major apple scab resistance gene (Rvi6) from apple to pear and precise comparison of the downstream molecular mechanisms of this resistance in both species

Background Scab is the most important fungal disease of apple and pear. Apple (Malus x domestica Borkh.) and European pear (Pyrus communis L.) are genetically related but they are hosts of two different fungal species: Venturia inaequalis for apple and V. pyrina for European pear. The apple/V. inaequalis pathosystem is quite well known, whereas knowledge about the pear/V. pyrina pathosystem is still limited. The aim of our study was to analyse the mode of action of a major resistance gene of apple (Rvi6) in transgenic apple and pear plants interacting with the two scab species (V. inaequalis and V. pyrina), in order to determine the degree of functional transferability between the two pathosystems. Results Transgenic pear clones constitutively expressing the Rvi6 gene from apple were compared to a scab transgenic apple clone carrying the same construct. After inoculation in greenhouse with V. pyrina, strong defense reactions and very limited sporulation were observed on all transgenic pear clones tested. Microscopic observations revealed frequent aborted conidiophores in the Rvi6 transgenic pear / V. pyrina interaction. The macro- and microscopic observations were very comparable to the Rvi6 apple / V. inaequalis interaction. However, this resistance in pear proved variable according to the strain of V. pyrina, and one of the strains tested overcame the resistance of most of the transgenic pear clones. Comparative transcriptomic analyses of apple and pear resistant interactions with V. inaequalis and V. pyrina, respectively, revealed different cascades of molecular mechanisms downstream of the pathogen recognition by Rvi6 in the two species. Signal transduction was triggered in both species with calcium (and G-proteins in pear) and interconnected hormonal signaling (jasmonic acid in pear, auxins in apple and brassinosteroids in both species), without involvement of salicylic acid. This led to the induction of defense responses such as a remodeling of primary and secondary cell wall, lipids biosynthesis (galactolipids in apple and cutin and cuticular waxes in pear), systemic acquired resistance signal generation (in apple) or perception in distal tissues (in pear), and the biosynthesis of phenylpropanoids (flavonoids in apple but also lignin in pear). Conclusion This study is the first example of a successful intergeneric transfer of a resistance gene among Rosaceae, with a resistance gene functioning towards another species of pathogen.

Bacterial Endophyte Community Dynamics in Apple (Malus domestica Borkh.) Germplasm and Their Evaluation for Scab Management Strategies

The large genetic evolution due to the sexual reproduction-mediated gene assortments and propensities has made Venturia inaequalis (causing apple scab) unique with respect to its management strategies. The resistance in apple germplasm against the scab, being controlled for by more than fifteen genes, has limited gene alteration-based investigations. Therefore, a biological approach of bacterial endophyte community dynamics was envisioned across the apple germplasm in context to the fungistatic behavior against V. inaequalis. A total of 155 colonies of bacterial endophytes were isolated from various plant parts of the apple, comprising 19 varieties, and after screening for antifungal behavior followed by morphological, ARDRA, and sequence analysis, a total of 71 isolates were selected for this study. The alpha diversity indices were seen to fluctuate greatly among the isolation samples in context to microflora with antifungal behavior. As all the isolates were screened for the presence of various metabolites and some relevant genes that directly or indirectly influence the fungistatic behavior of the isolated microflora, a huge variation among the isolated microflora was observed. The outstanding isolates showing highest percentage growth inhibition of V. inaequalis were exploited to raise a bio-formulation, which was tested against the scab prevalence in eight apple varieties under controlled growth conditions. The formulation at all the concentrations caused considerable reductions in both the disease severity and disease incidence in all the tested apple varieties. Red Delicious being most important cultivar of the northwestern Himalayas was further investigated for its biochemical behavior in formulation and the investigation revealed different levels of enzyme production, chlorophyll, and sugars against the non-inoculated control.

Mapping of Quantitative Trait Loci for Scab Resistance in Apple (Malus × Domestica)

Scab caused by Venturia inaequalis (Cke.) Wint. is the most important fungal disease of apple. Fungicide application is a widely practiced method of disease control. The use of chemicals is however, cost intensive, tedious and ecologically unsafe. Development of genetic resistance and breeding of resistant cultivars is most reliable and a safest option. One such source of scab resistance happens to be the variety ‘Shireen’, released from SKUASTKashmir. However, till date the nature of resistance and its genetic control has not been characterized. The aim of this research was to elucidate the genetic basis of scab resistance in Shireen. The present study helped us to identify two quantitative trait loci (QTLs) on chromosome 2 and 8 and six potential CDGs for the polygenic resistance in ‘Shireen’. The genomic region corresponding to the mapped QTLs in LG 2 and LG 8 of ‘Shireen’ was examined for candidate genes possibly related to scab resistance using in silico analysis. The QTLs mapped in the genetic background of Shireen are the novel QTLs and may be transferred to desirable genetic backgrounds and provide opportunities for isolation and cloning of genes apart from their utility in order to achieve durable resistance to scab.

ASSESSMENT OF INTRODUCED APPLE VARIETIES BY SCAB AFFECTION (VENTURIA INAEQUALIS (COOKE) G. WINTER)

The article presents data on the identifying of apple varieties resistant to scab for their further use in breeding. The research was carried out on the basis of the Orenburg branch of the Federal Horticultural Research Center for Breeding, Agrotechnology and Nursery from 2010 to 2020 in the orchard established in 2004. The objects of research were introduced apple varieties and local hybrids. Scab (the causative agent is the marsupial mushroom Venturia inaequalis (Cooke) G. Winter) causes great damage to the apple tree. This disease affects the fruits and leaves, while dramatically reducing the apple productivity and marketability. According to the results of the studies, the greatest development of scab was observed in 2011, 2013 and 2016. In dry years (2010, 2012, 2014, 2015, 2017-2020 years) scab lesions were not observed. We have identified apple introduced varieties, which over the years of observation had a weak scab lesion and few spots on the apple leaves. In the control variant, in summer varieties in 2011, the scab on the leaves of the Serebryanoe kopytse variety was 0.9 points, all other studied varieties had a lower parameter. In autumn varieties, the varieties Sverdlovchanin and Gornist had the lowest parameter — 0.3-0.4 points, respectively, with a control parameter of 0.8 points (Prizemlennoye variety); in winter varieties, all the studied samples were lower than the control variant — 0.9 points. Weather conditions 2013 and 2016 also showed apple varieties less susceptible to scab in the conditions of the Orenburg region. As a result, we identified apple varieties that are weakly affected by scab: summer varieties — Letneye polosatoye, Mechtatel’nitsa, Solntsedar, Chudnoye; autumn varieties — resident of Sverdlovsk, Hornist; winter — Anis Sverdlovsky, Krasa Sverdlovsk, Persian. The obtained data can be used in breeding to create new apple varieties resistant to scab in the conditions of the Orenburg region.