Мета. Аналіз технологій мікроклонального розмноження рослин для створення життєздатних міжвидових гібридів і сортів Actinidia Lindl.Методи. Загальнонаукові – гіпотеза, експеримент, спостереження, аналіз, метод синтезу для формування висновків.Результати. Упровадження технологій in vitro натепер стає панівним комерційним методом масштабного й швидкого отримання саджанців зі стабільним успадкуванням ознак сорту, високим коефіцієнтом розмноження, збереженням господарсько-цінних ознак за відсутності сезонності виробництва та обмежень у часі. Крім розмноження, пришвидшується й селекційний процес, зокрема мутагенез і гібридизація. Важливо одержати не лише стерильний експлант, а й морфогенно активний, тобто такий, що приживеться, і згодом регенерує рослину in vitro. Найліпшим за ефективністю деконтамінації є спосіб оброблення гіпохлоритом та додавання у живильне середовище біоциду РРМ, але за цих умов відзначено найменше виживання експлантів у всіх зразків. На ефективність введення в асептичну культуру на першому етапі мікроклонального розмноження впливають також і біологічні особливості первинних експлантів. У дослідженнях із поживними середовищами для A. arguta встановлено, що з елементів мінерального живлення лише 11 іонів є необхідними для життєдіяльності: п’ять макро- (N, K, P, Mg, S) і шість мікроелементів (Cl, Fe, B, Mo, Na, I). Рослини in vitro мають нижчий уміст сухих речовин та більшу кількість вологи, зокрема й вільної, яка за умови порушення водного балансу швидко втрачається.Висновки. Здатність до регенерації більшою мірою виражена у видів A. chinensis та A. deliciosa, меншою – в А. arguta. Для A. chinensis ефективним є застосування гідропонної технології адаптації регенерантів на етапі ex vitro. | Purpose. Analysis of plant micropropagation technologies for the creation of viable interspecific hybrids and varieties of Actinidia Lindl. Methods. General scientific – hypothesis, experiment, observation, analysis, synthesis method for drawing conclusions. Results. The introduction of in vitro technologies is now becoming the dominant commercial method of large-scale and rapid production of seedlings with stable inheritance of variety traits, high multiplication rate, preservation of economically valuable traits in the absence of production seasonality and time constraints. In addition to reproduction, the breeding process is also accelerated, including mutagenesis and hybridization. It is important to obtain not only a sterile explant, but also a morphogenically active one, that is, a plant that takes roots and subsequently regenerates in vitro. The best in terms of decontamination efficiency is the method of treatment with hypochlorite and the addition of PPM biocide to the nutrient medium, but under these conditions, the lowest survival of explants in all samples was noted. The efficiency of introduction into aseptic culture at the first stage of micropropagation is also affected by the biological characteristics of the primary explants. In studies with nutrient media for A. arguta, it was found that of the elements of mineral nutrition, only 11 ions are necessary for life: five macro- (N, K, P, Mg, S) and six microelements (Cl, Fe, B, Mo, Na, I). Plants in vitro have a lower dry matter content and a greater amount of moisture, including free moisture, which is quickly lost when the water balance is disturbed. Conclusions. The abi­lity to regenerate is more pronounced in the species A. chinensis and A. deliciosa, and to a lesser extent in A. arguta. For A. chinensis, the use of hydroponic technology for the adaptation of regenerants at the ex vitro stage is effective.

Designing an optimal mineral-salt composition medium for plant micropropagation is one of the biggest challenges for the plant biotechnologist. Several micropropagation protocols for Actinidia spp. have been proposed since Harada (1975), although the MS (Murashige and Skoog, 1962) medium is the most commonly used for kiwifruit in vitro culture. In this research, six different culture media currently used for Actinidia micropropagation, MS (Murashige and Skoog), Ch (Cheng), H (Harada), St (Standardi), B5 (Gamborg) and Kh medium (modified Cheng), were tested to assess the multiplication response of in-vitro-cultured Actinidia arguta ‘Issai’. All media were supplemented with 1 mg L-1 6-benzylaminopurine (BAP) and 1 mg L-1 gibberellic acid (GA3). The vegetative growth of explants varied significantly depending on the media used, with the St medium being the most effective, generating healthy (4.1 up to 5) and long (3.4 cm) shoots and the highest number of leaves (27 per explant) with the largest leaf area (43.1 cm2). In contrast, B5 medium promoted low (3.5) and short (1.9 cm) shoots, fewest leaves (18) and smallest leaf area (21.4 cm2), but more importantly, B5 also produced physiological disorders such as undesirable callus and abnormal leaves. The rest of the media produced intermediate and/or not significantly different results compared with St and B5. Standardi medium seems to be the most appropriate medium for good proliferation of Actinidia arguta ‘Issai’, although it clearly needs improving since it also produced physiological side-effects such as callus formation.

The patterns of the distribution of nutrients in kiwiberry (Actinidia arguta (Siebold & Zucc.) Planch. ex Miq.), family Actinidiaceae (Gilg & Werderm), leaves growing under different soil and climatic conditions (Ukraine and China) were studied. Using scanning electron microscopy, significant differences were shown in the distribution of assimilates and mineral nutrients in the leaves of kiwiberry cultivated under different climate and soil conditions (Kyiv city, Ukraine and Jiamusi, China). The leaves of plants grown in China have higher concentration of all of the studied nutrients exception for silicon. The differences found in the content of macro- and microelements in plant tissues are consistent with their total content in the soil, and depend on the synthesis of low molecular weight organic compounds, namely, hydroxybenzoic, benzoic and triterpene acids. An increase in the silicon content in the leaves of kiwiberry plants grown in Ukraine indicates the moisture deficit in the soil. This conclusion is confirmed by the anatomical differences viz. the presence of additional integumentary formations and fewer stomata number per 1 mm2 of leaf surface. The specific feature of ‘Perlyna sadu’ cultivar was high concentrations of sodium and aluminum in the foliar tissues, regardless of the place of growth. The analysis of the distribution of nutrients in the leaves located along the stem showed remobilization of the former within the three layers: the lower one nourishes the roots, the upper one nourishes the leaves in the active growth phase and the middle one allocates the assimilates in both directions. A significant positive relationship was found between the biosynthesis of photosynthetic pigments and electrophysiological activity, especially for the leaves of the lower zone. The revealed differentiation into layers differing in polarity of bioelectric potentials and the distribution of assimilates suggests functional differentiation of the kiwiberry leaves. In particular, the leaves of the lower layer perform a storage function. The middle part is less conservative and characterized by higher sensitivity to environmental factors performs a mainly synthetic function. The upper layer performs an active growth function. The results of the comparative analysis of the indicators of the number of chloroplasts in the mesophyll cells proved that the obtained dependence can be used as a diagnostic criterion in assessing the predicted plant productivity at the early stages of their development.

Actinidia deliciosa is a commercially important plant receiving recognition because of its high nutritive value. This study presents an efficient protocol for the hardening of in vitro raised Actinidia deliciosa plants using a hydroponic method. Leaf explants inoculated on Murashige and Skoog (MS) medium supplemented with 5.00 μM 6-benzylaminopurine (BAP) and 1.00 μM α-naphthaleneacetic acid (NAA) resulted in 5.28 ± 0.14 shoots per explants and 8.33 ± 1.80 cm average shoot length. Rooting was achieved through half-strength MS medium supplemented with 1.5 μM indolebutyric acid (IBA) and 0.6 μM BAP. Maximum root number (12.76 ± 1.08) and 6.78 ± 0.25 cm average root length were recorded from plantlets using half-strength MS medium supplemented with 1.5 μM indolebutyric acid (IBA) and 0.6 μM BAP. In hydroponic system, an average root length of 22.40 ± 0.59 cm, average root number of 21.50 ± 1.24, average leaf number of 4.50 ± 0.40, and average shoot length of 9.71 ± 0.29 cm were observed in Hoagland & Arnon solution. The early development of shoots, roots, and leaves through hydroponics was advantageous in establishment of micropropagated plants in a greenhouse. Complete 100% plant survival was found by following proper acclimatization using hydroponic method. The study underlines the efficient hardening of micropropagated plants of A. deliciosa through hydroponic technique in Himalayan region.

In traditional in vitro culture, the low CO₂ concentration inside the vessels restricts photosynthesis and necessitates the addition of sucrose to the culture medium as the main energy source, thus bringing about changes in the absorption of mineral elements from the culture medium. In this study, we investigated macronutrient absorption and sugar consumption in Actinidia deliciosa Chevalier Liang and Ferguson cv. Hayward (kiwi), cultured on medium supplemented with varying amounts of sucrose (0, 10, and 20 g l⁻¹) under both heterotrophy and autotrophy, flushed with different concentrations of CO₂ (non-ventilation, 300, 600, and 2,000 μl l⁻¹). In ventilated systems with 20 g l⁻¹ of sucrose, sucrose absorption was less than under non-ventilation. The lowest rate of sucrose absorption was recorded when the explants were cultured on medium supplemented with 20 g l⁻¹ of sucrose and flushed with 600 μl l⁻¹ CO₂. Absorption of NO₃ ⁻, PO₄ ³⁻, and Mg²⁺ were high (maximum) at the end of the culture period (40 d) in explants flushed with 600 μl l⁻¹ CO₂ that have been cultured 20 d in the presence of sucrose and then transferred to a sucrose-free medium. These autotrophic conditions promoted maximum plant growth in terms of both fresh and dry mass as well as the length and number of shoots and leaves. The study shows that to maintain an optimum regime of mineral nutrition for prolonged culture of kiwi in vitro, an increased amount of these three ions should be supplemented in Murashige and Skoog's medium.

Мета. Аналіз технологій мікроклонального розмноження рослин для створення життєздатних міжвидових гібридів і сортів Actinidia Lindl. Методи. Загальнонаукові – гіпотеза, експеримент, спостереження, аналіз, метод синтезу для формування висновків. Результати. Упровадження технологій in vitro натепер стає панівним комерційним методом масштабного й швидкого отримання саджанців зі стабільним успадкуванням ознак сорту, високим коефіцієнтом розмноження, збереженням господарсько-цінних ознак за відсутності сезонності виробництва та обмежень у часі. Крім розмноження, пришвидшується й селекційний процес, зокрема мутагенез і гібридизація. Важливо одержати не лише стерильний експлант, а й морфогенно активний, тобто такий, що приживеться, і згодом регенерує рослину in vitro. Найліпшим за ефективністю деконтамінації є спосіб оброблення гіпохлоритом та додавання у живильне середовище біоциду РРМ, але за цих умов відзначено найменше виживання експлантів у всіх зразків. На ефективність введення в асептичну культуру на першому етапі мікроклонального розмноження впливають також і біологічні особливості первинних експлантів. У дослідженнях із поживними середовищами для A. arguta встановлено, що з елементів мінерального живлення лише 11 іонів є необхідними для життєдіяльності: п’ять макро- (N, K, P, Mg, S) і шість мікроелементів (Cl, Fe, B, Mo, Na, I). Рослини in vitro мають нижчий уміст сухих речовин та більшу кількість вологи, зокрема й вільної, яка за умови порушення водного балансу швидко втрачається. Висновки. Здатність до регенерації більшою мірою виражена у видів A. chinensis та A. deliciosa, меншою – в А. arguta. Для A. chinensis ефективним є застосування гідропонної технології адаптації регенерантів на етапі ex vitro.

The design of plant tissue culture media remains a complicated task due to the interactions of many factors. The use of computer-based tools is still very scarce, although they have demonstrated great advantages when used in large dataset analysis. In this study, design of experiments (DOE) and three machine learning (ML) algorithms, artificial neural networks (ANNs), fuzzy logic, and genetic algorithms (GA), were combined to decipher the key minerals and predict the optimal combination of salts for hardy kiwi (Actinidia arguta) in vitro micropropagation. A five-factor experimental design of 33 salt treatments was defined using DOE. Later, the effect of the ionic variations generated by these five factors on three morpho-physiological growth responses – shoot number (SN), shoot length (SL), and leaves area (LA) – and on three quality responses - shoots quality (SQ), basal callus (BC), and hyperhydricity (H) – were modeled and analyzed simultaneously. Neurofuzzy logic models demonstrated that just 11 ions (five macronutrients (N, K, P, Mg, and S) and six micronutrients (Cl, Fe, B, Mo, Na, and I)) out of the 18 tested explained the results obtained. The rules “IF – THEN” allow for easy deduction of the concentration range of each ion that causes a positive effect on growth responses and guarantees healthy shoots. Secondly, using a combination of ANNs-GA, a new optimized medium was designed and the desired values for each response parameter were accurately predicted. Finally, the experimental validation of the model showed that the optimized medium significantly promotes SQ and reduces BC and H compared to standard media generally used in plant tissue culture. This study demonstrated the suitability of computer-based tools for improving plant in vitro micropropagation: (i) DOE to design more efficient experiments, saving time and cost; (ii) ANNs combined with fuzzy logic to understand the cause-effect of several factors on the response parameters; and (iii) ANNs-GA to predict new mineral media formulation, which improve growth response, avoiding morpho-physiological abnormalities. The lack of predictability on some response parameters can be due to other key media components, such as vitamins, PGRs, or organic compounds, particularly glycine, which could modulate the effect of the ions and needs further research for confirmation.

Kiwifruit has high economic value but is susceptible to fungal infection after harvest. Botrytis cinerea (B. cinerea) is one of these fungal pathogens that cause huge economic losses and potential food safety problems. Auxin is a vital plant growth regulator that has been reported to play a role in plant defense. In this study, postharvest kiwifruit was treated with different concentrations of indole-3-acetic acid (IAA), after which the average incidence rate and lesion area were analyzed to determine the appropriate concentration of IAA for disease resistance. Defense enzymes, such as superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were detected. Ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS)-based widely targeted metabolomic analysis was used to investigate the mechanism of kiwifruit active compounds during IAA treatment and B. cinerea infection. Exogenous IAA treatment (especially at 50 μg/ mL) enhanced the resistance of kiwifruit to B. cinerea, resulting in reduced disease incidence and lesion area, and showed higher pathogen resistance-related defense enzyme activity in response to B. cinerea infection. Further metabolomic analysis revealed 776 metabolites in kiwifruit after IAA treatment, among which 51 metabolites in IAA_24h (24 h after IAA treatment) vs. CK (sterilized ddH₂O treatment) fruit, 55 metabolites in IAA_72h (72 h after IAA treatment) vs. CK, and 67 metabolites in IAA_24h vs. IAA_72h were found to be significantly different. KEGG analysis showed that the resistance induced in kiwifruit by IAA treatment was closely related to the activation of phenylpropanoids, including flavonoids and phenols, terpenoids, carbohydrate metabolism, and hormone signaling pathways. These results might reveal the mechanism of IAA-induced resistance in kiwifruit and provide a new theoretical basis for the safe and efficient control of postharvest diseases in kiwifruit.

Kiwifruit (Actinidia spp., Family Actinidiaceae) is a recently domesticated crop with highly endangered genetic resources in field genebanks and in the centre of diversity. Improved genotypes of several species in the genus have been commercialised and several other species are used in breeding programmes. Cryopreservation is considered a safe and cost-effective option for ex-situ conservation of clonal crops. In the current research we tested if the currently available method of droplet vitrification (DV) cryopreservation could be simplified whilst improving plant regeneration. Plant regeneration in the nine accessions belonging to A. chinensis var. chinensis, and A. chinensis var. deliciosa, A. arguta, A. macrosperma and A. polygama after cryopreservation ranged from 59 to 88% with the improved method where cold acclimation and antioxidant treatment steps were eliminated while sucrose pretreatment of shoot tips has been reduced to two steps in two days. Use of smaller (0.5–1 mm compared to 2 mm) in vitro shoot tips from younger (two weeks after subculture), fast-growing shoots and the use of liquid sucrose for shoot tip pretreatment are considered to be the reasons for this improved response. We also introduced the use of sodium dichloroisocyanurate as an alternative to household bleach as a sterilant for kiwifruit tissue culture establishment. The improved DV protocol is being used to cryopreserve valuable kiwifruit genetic resources with greater efficiency.

A highly efficient in vitro micropropagation protocol, using newly emerged sprouts coming from greenhouse-grown plants, was developed to produce Actinidia melanandra Franch. and Actinidia rubricaulis Dunn. This is the first report of in vitro culture of A. rubricaulis Dunn. Axenic culture was possible using one- or two-node explants or terminal buds, surface-sterilised with commercial bleach tablets. Nearly 90% success rate in establishment was observed. The best axillary shoot proliferation and maximum adventitious shoots elongation were achieved on Quoirin-Lepoivre medium supplemented with 1.5 mg L⁻¹ 6-benzyladenine and 30 g L⁻¹ sucrose. Although used as an inert support, agar brand has a significant effect. Kobe agar at 8 g L⁻¹ gave the best response. 100% of in vitro rooting was observed. Roots length and their quality were highly improved by the use of vermiculite in transfer medium. Over 99% of rooted plants survived after acclimatisation.

Angular, necrotic leaf spot, longitudinal cracks along the petiole, oozing and wilting of branches were observed during summer 2008 on Actinidia chinensis (yellow kiwifruit) cultivar Hort16A, cultivated in different orchards located the province of Latina (central Italy). Symptoms closely resembled those incited by Pseudomonas syringae pv. actinidiae on kiwifruit A. deliciosa. Isolates obtained from typical lesions were assessed by means of biochemical, pathogenicity and host range tests and compared with some Pseudomonas syringae pathovars by enterobacterial repetitive intergenic consensus (ERIC-PCR) analysis. The isolates belong to Pseudomonas LOPAT group Ia, incited the death of pot-cultivated A. chinensis cv. Hort 16A and A. deliciosa cv. Hayward plants in few days, but did not cause any symptoms to the other inoculated plant species. Upon ERIC-PCR analysis, all the isolates showed similarity with P. syringae pv. actinidiae NCPPB 3739, type-strain of the pathovar. This is the first report of this pathogen on A. chinensis in Italy and, as far as we currently know, in the world.