Subsequent Cultivation Research Articles

Optimization of the marine microalgae Nannochloropsis oculata harvesting by flocculation: Effects on biomass quality and residual medium reuse for a new cycle

ABSTRACT Flocculation using natural plant-based biopolymers from Moringa oleifera seeds and Acacia mearnsii flocculant as Tanfloc® (TSG) has emerged as an efficient strategy for microalgae biomass harvesting. This study optimized the recovery process of Nannochloropsis oculata by varying flocculant concentration, sedimentation time, and culture pH. Results revealed that 1.2 mg.L−1 of TSG at pH 8.9 achieved optimal conditions, although TSG proved effective across different pH ranges. Alkaline flocculation (ALK) also enhanced microalgae harvesting, yielding a high harvesting efficiency (>96%) within 30 minutes of sedimentation. Both methods achieved a concentration factor exceeding 20. However, the use of ALK resulted in reduced lipid and carbohydrate content compared to the control condition. Carotenoid levels showed no significant difference. TSG influenced lipid extraction but not fatty acid quality or carbohydrate extraction. Importantly, the residual medium from TSG flocculation could sustain subsequent cultivation cycles without nutrient supplementation, presenting a cost-effective solution for microalgae cultivation and harvesting.

Two Sporadic Cases of Legionellosis Associated with the Use of Domestic Ultrasonic Humidifiers

Two sporadic cases of legionellosis occurring in consecutive years were confirmed by positive antigenuria to Legionella pneumophila serogroup 1 in individuals with limited mobility who were confined to their homes. Both cases had a history of using ultrasonic humidifiers and of low exposure to other possible sources of infection. This study was conducted through an expanded epidemiological survey and home inspection. Samples were collected from domestic hot water and humidifiers. Environmental isolates were characterised by immunoagglutination and immunofluorescence. The Dresden panel is employed for the determination of groups and subgroups of serogroup 1. The amoebae were isolated by water filtration and subsequent cultivation of the filters. Identification of the isolates was conducted through the sequencing of PCR products. In both cases, epidemiological studies identified the ultrasonic humidifiers as the probable source of infection. The presence of Legionella pneumophila was confirmed in the sanitary water installation and in the humidifiers, where high values of Legionella pneumophila serogroup 1 counts were also obtained. The isolates were characterised as Olda Oxford and Olda OLDA. Furthermore, in addition to the Legionella isolates, the presence of Vermamoeba vermiformis was also identified in samples obtained from domestic hot water and humidifiers in one of the residential addresses under investigation. It has been demonstrated that humidifiers can act as an amplification mechanism for pathogenic microorganisms when they are not cleaned and maintained correctly. This can pose a health danger, especially to people with previous respiratory pathologies, the immunosuppressed, and the elderly. Therefore, it is of the utmost importance that professionals who recommend the therapeutic use of this equipment should issue warnings regarding the importance of its correct use, cleaning, and disinfection. Finally, humidifiers should be considered in epidemiological surveys and in the study of sporadic cases.

Assessment of the Degree Of DNA Methylation in Lymphocytes аfter а Single Blood Irradiation in vitro

DNA methylation is one of the processes of epigenetic regulation of the genome, which is sensitive to the influence of endogenous and exogenous factors. The effect of ionizing radiation on the genome is accompanied by a change in the degree of DNA methylation, which can be dose-dependent and persist for a long time after radiation exposure. The objective of the study was to assess the degree of DNA methylation of blood lymphocytes after a single exposure to gamma radiation at a dose of 1.5 Gy using wide-genome bisulfite sequencing. The study included 10 conditionally healthy male employees of the ionizing radiation facility who were not exposed to radiation and did not suffer from chronic diseases. The material was whole blood: 0 Gy (control samples) and 1.5 Gy (experimental samples irradiated with gamma radiation). After irradiation with subsequent cultivation of whole blood, DNA isolation and bisulfite sequencing of limited sets of genomic loci (Reduced representation bisulfite sequencing) was performed using XmaI restriction enzyme (XmaI-RRBS). 41 genes were identified, including 26 genes (HOXD4, PADI2, FOXK1, FTCD, PRDM16, TOM1, PPP1R14A, FLNB, OR1F1, RARA, CRTAC1, AP5B1, ARL5C, NOC2L, MAMDC4, FGFRL1, PPFIA3, CUX2, ANKRD20A19P, FAM83H-AS1, CBFA2T3, POLN, MIR4458HG, FNBP1, SPIRE2, and ZSCAN10) have a tendency to hypomethylation DNA, and another 15 genes (CHRNA4, SEPTIN9, ZNF174, ELK3, NFAM1, ALG10, SOX8, KLHL30, URI1, HBZ, KLF14, MYO16, MYEOV, DMKN, and PAX7) tend to have hypermethylated state detected in at least 50٪ of the experimental samples. Thus, the genes identified in this study can be promising markers of radiation exposure and, in the future, be used to develop a new type of biological dosimetry – epigenomic dosimetry of personnel in contact with ionizing radiation sources in the course of their professional activities.

Activity of gemmogenesis in regenerated plants of hazelnut cv. ‘Akademyk Yablokov’ and antibiotic therapy for elimination of bacterial contamination in vitro

Background. The problem of obtaining certified planting material with biotechnological methods is important for expanding commercial and homestead plantations of hazelnut cultivars in Belarus. Materials and methods. Regenerated plants of cv. ‘Akademyk Yablokov’, representing the genus Corylus L., were a model object for studying in vitro morphogenesis and effectiveness of antibiotics against bacterial contamination, so that a protocol could be developed to obtain healthy planting material of hazelnut cultivars. The plants produced during this study were included in the duplicate ex situ collection of nut crops preserved in vitro in the active growth state. Results. Single exposure to the antibiotic kanamycin monosulfate at a concentration of 100 mg/L during antibiotic therapy in the stage of in vitro micropropagation eliminated bacterial infection in 83.3% of regenerated plants, and twofold exposure in 100%. Further cultivation revealed its phytotoxic aftereffect manifested in the form of necrosis on most of the regenerated plants and a decrease in the activity of gemmogenesis and growth. Neither single nor twofold exposure to cefotaxime sodium salt at a concentration of 90 mg/L caused elimination of bacterial infection, but gemmogenesis and regenerated plant growth retained their activity during subsequent cultivation on antibiotic-free media. The best development parameters were observed on a modified Murashige–Skoog medium with 6 mg/L 6-BA, 0.01 mg/L IBA, and 0.1 mg/L GA3 (average number of shoots: 2.2; number of microcuttings: 2.3), and a modified DKW medium with 6 mg/L 6-BA, 0.01 mg/L IBA, and 0.1 mg/L GA3 (average number of shoots: 2.05; microcuttings: 2.9). The use of zeatin as a cytokinin to stimulate adventitious morphogenesis or activate the growth of axillary meristems at a concentration of 5 or 6 mg/L was not as effective as 6-BA.

Fungal degradation of phenylacetate focusing on CRISPR/Cas9-assisted characterization of two oxidative enzyme genes of Akanthomyces muscarius AM1091

The degradation of phenylacetate (PA) was investigated as a model to explore aromatic compound breakdown in the fungal system. Fungal strains capable of utilizing PA as their sole carbon source were isolated using a minimal solid medium supplemented with 0.5 % PA. Subsequent cultivation in minimum liquid medium revealed that selected fungal strains, including Trametes versicolor TV0876 and TV3295, Paecilomyces hepiali PH4477, and Akanthomyces muscarius AM1091, efficiently removed PA within 24 h. HPLC analysis of culture supernatants from various fungal strains revealed a time-dependent accumulation of 2-hydroxyphenylacetate (2-HPA) and 4-hydroxyphenylacetate (4-HPA), two key major metabolic products primarily found in ascomycetes and basidiomycetes, respectively. This suggests that the first hydroxylation of PA is catalyzed by two distinct hydroxylases, one for each fungal group. Furthermore, fungal species that make 4-HPA also produce phenylethanol (PE), indicating a distinct catabolic mechanism to remove PA by direct reduction of PA to PE. A. muscarius AM1091, identified as the most efficient PA degrader in this study, was studied further to determine the biochemical pathway of PA degradation. RNA-Seq and RT-PCR analyses of AM1091 revealed two oxidative enzyme genes, CYP1 and DIO4, upregulated in the presence of PA. Targeted disruption utilizing preassembled Cas9-gRNA ribonucleoprotein complexes and homologous DNAs harboring the URA3 gene as an auxotrophic marker resulted in the cyp1 and dio4 mutant strains. The cyp1 mutant was incapable of converting PA to 2-HPA, indicating its involvement in the C2 hydroxylation, whereas the dio4 mutant was unable to degrade 2,5-dihydroxyphenylacetate (2,5-DHPA), resulting in the accumulation of 2,5-DHPA. Our findings indicate that A. muscarius AM1091 degrades PA through the activities of CYP1 and DIO4 for the C2 hydroxylation and subsequent ring-opening reactions, respectively.

Passive Droplet Microfluidic Platform for High-Throughput Screening of Microbial Proteolytic Activity.

Traditional bacterial isolation methods are often costly, have limited throughput, and may not accurately reflect the true microbial community composition. Consequently, identifying rare or slow-growing taxa becomes challenging. Over the past decade, a new approach has been proposed to replace traditional flasks or multiwell plates with ultrahigh-throughput droplet microfluidic screening assays. In this study, we present a novel passive droplet-based method designed for isolating proteolytic microorganisms, which are crucial in various biotechnology industries. Following the encapsulation of single cells in gelatin microgel compartments and their subsequent clonal cultivation, microcultures are passively sorted at high throughput based on the deformability of droplets. Our novel chip design offers a 50-fold improvement in throughput compared to a previously developed deformability-based droplet sorter. This method expands an array of droplet-based microbial enrichment assays and significantly reduces the time and resources required to isolate proteolytic bacteria strains.

Investigation of the effect of macrophage and epithelial cell interaction in the progression of different subtypes of breast cancer.

10 Background: In the realm of tumor progression, the immunosuppressive role of macrophages has been well-established. However, a significant lacuna persists concerning the mechanisms through which epithelial cells regulate macrophages. Furthermore, the precise manner in which macrophages modulate breast cancer cells remains unclear. Hence, the primary objective of the current investigation is to delve into the influence of macrophages on the progression of distinct subtypes of breast cancer. Methods: Human monocyte cell-derived macrophages (M0) were cultivated in the presence of conditioned media (CM) from various subtypes of breast cancer cells. Conversely, human luminal breast cancer cell line T47D, and basal cell line MDA MB 231 were cultured in the presence of CM from macrophages. Results: When macrophages were incubated with CM of T47D, they exhibited a spindle-shaped morphology with pseudopodia. Conversely, exposure to CM of MDA MB 231 induced macrophages to adopt a foamy, round shape with numerous filopodia. Flow cytometry analysis revealed that T47D-CM promoted a higher proportion of CD80+ cells, while MDA MB 231-CM led to an increase in both CD206+ and CD80+ cells. Moreover, both T47D and MDA MB 231-CM escalated the production of various cytokines including IL-1β, IL-2, IL-4, IL-5, IL-7, IL-12(p70), IL-13, IL-17, G-CSF, IFN-γ, and MIP-1β. Intriguingly, MDA MB 231-CM induced a significant increase in IL-6, GM-CSF, and TNF-α. Subsequent cultivation of cancer cells in the presence of macrophage-CM resulted in a marked proliferation of MDA MB 231 cells, while T47D cell proliferation decreased significantly. Correspondingly, there was a notable increase in apoptosis among T47D cells. Scratch wound healing assays revealed heightened migration of MDA MB 231 cells following treatment with macrophage-CM, whereas there was negligible alteration in T47D cell migration. Proteomic analysis unveiled dysregulation of 247 and 596 proteins in MDA MB 231 and T47D cells, respectively. Particularly noteworthy were three proteins—MDHM, H1.5, and LEG8—abundant in MDA MB 231 cells, yet scarce in T47D cells. These proteins have been implicated in promoting tumor growth across various cancer types. Conclusions: This study first time elucidates subtype specific role of macrophage in breast tumour. In triple-negative breast cancer, infiltrating macrophages induce an immunosuppressive effect, altering cytokine expression and fostering the expression of tumor-promoting factors in epithelial cells, thereby facilitating cellular proliferation and metastasis. Conversely, in luminal breast cancer, infiltrating macrophages exhibit a predominantly pro-inflammatory profile, resulting in minimal tumor-promoting effects. Key Words: Breast Cancer, Macrophage polarization, Conditioned Media, Cytokines, Proteomics. Acknowledgements, DBT – Govt of India, NIBMG, Kalyani.

Advanced Microfluidic Platform for Tumor Spheroid Formation and Cultivation Fabricated from OSTE+ Polymer

In the evolving landscape of cancer research, 3D cell cultures, particularly tumor cell spheroids, are increasingly preferred in drug screening due to their enhanced mimicry of in vivo tumor environments, especially in drug resistance aspects. However, the consistent formation of uniform spheroids and their precise manipulation remain complex challenges. Among various methodologies, droplet microfluidics emerges as a highly effective approach for tumor spheroid formation. This paper introduces a novel, multifaceted microfluidic system that streamlines the entire spheroid cultivation process: (i) generating tumor spheroids from cell suspensions within individual droplets, (ii) merging these droplets into a continuous aqueous phase once spheroid formation is complete, and (iii) transferring the spheroids to a specialized cultivation area within the chip, equipped with trapping elements for extended cultivation in perfusion mode. Remarkably, this process requires no hydrogel encapsulation or external handling, as all operations are conducted within the microfluidic chip. Fabricated from the innovative OSTE+ (off-stoichiometry thiol-ene epoxy) polymer, the chip is designed for repeated use. To show its efficacy, we successfully formed spheroids from MCF-7, GAMG, and U87 cell lines in our system and compared them with spheroids prepared by a traditional agarose microwell method. Additionally, our methodology has successfully enabled the in-chip release of spheroids from droplets, followed by their effective trapping for subsequent cultivation, a process we have exemplified with MCF-7 spheroids. To our knowledge, this research represents the first instance of a fully integrated droplet microfluidic platform achieving scaffoldless tumor spheroid formation and handling. Our method holds promise for improving high-throughput, automated procedures in the formation, transfer, and cultivation of tumor cell spheroids.Graphical abstract

Algal Biodiversity of Nine Megaliths in South-East Bulgaria.

This paper presents the first data on the biodiversity of lithophytic algae from Bulgarian megaliths obtained after the application of the direct sampling method, subsequent cultivation, and processing by light microscopy. A rich algal flora was found: 90 species and 1 variety of 65 genera from Cyanoprokaryota/Cyanobacteria (29 species, 13 genera), Chlorophyta (40 species and 1 variety, 38 genera), Streptophyta (5 species, 1 genus), and Ochrophyta (16 species, 13 genera). Among them were the globally rare Pseudodictyochloris multinucleata (Chlorophyta), found for the first time in such lowland and warm habitats, and Scotiella tuberculata (Chlorophyta), for which this is the first finding in the country. Three of the recorded species are conservationally important. The low floristic similarity between the sites (0-33%) shows the diversity of the algal flora, with no common species found for all the megaliths studied. The most widespread were the strongly adaptive and competitive Stichococcus bacillaris, Apatococcus lobatus, and Chloroidium ellipsoidium (Chlorophyta). The correlations estimated between the species number and substrate temperature (18.1-49.6 °C) suggest the prospect of future research related to the impact of global warming. In addition, the study points to the safety aspects as it revealed species from nine potentially toxin-producing cyanoprokaryotic genera that could be harmful to visitors' health.

Selection and Genetic Analysis of High Polysaccharide-Producing Mutants in Inonotus obliquus.

Inonotus obliquus, a medicinal fungus, has garnered significant attention in scientific research and medical applications. In this study, protoplasts of the I. obliquus HS819 strain were prepared using an enzymatic method and achieved a regeneration rate of 5.83%. To enhance polysaccharide production of I. obliquus HS819, atmospheric and room temperature plasma (ARTP) technology was employed for mutagenesis of the protoplasts. Through liquid fermentation, 32 mutant strains exhibiting diverse characteristics in morphology, color of the fermentation broth, mycelial pellet size, and biomass were screened. Secondary screening identified mutant strain A27, which showed a significant increase in polysaccharide production up to 1.67 g/L and a mycelial dry weight of 17.6 g/L, representing 137.67% and 15% increases compared to the HS819 strain, respectively. Furthermore, the fermentation period was reduced by 2 days, and subsequent subculture cultivation demonstrated stable polysaccharide production and mycelial dry weight. The genome resequencing analysis of the HS819 strain and mutant strain A27 revealed 3790 InDel sites and mutations affecting 612 functional genes associated with polysaccharide synthesis. We predict that our findings will be helpful for high polysaccharide production through genetic engineering of I. obliquus.

Composition and diversity of soil microbial communities change by introducing Phallus impudicus into a Gastrodia elata Bl.-based soil

BackgroundThe Gastrodia elata Bl. is an orchid, and its growth demands the presence of Armillaria species. The strong competitiveness of Armillaria species has always been a concern of major threat to other soil organisms, thus disrupting the equilibrium of soil biodiversity. Introducing other species to where G. elata was cultivated, could possibly alleviate the problems associated with the disequilibrium of soil microenvironment; however, their impacts on the soil microbial communities and the underlying mechanisms remain unclear. To reveal the changes of microbial groups associated with soil chemical properties responding to different cultivation species, the chemical property measurements coupled with the next-generation pyrosequencing analyses were applied with soil samples collected from fallow land, cultivation of G. elata and Phallus impudicus, respectively.ResultsThe cultivation of G. elata induced significant increases (p < 0.05) in soil pH and NO3-N content compared with fallow land, whereas subsequent cultivation of P. impudicus reversed these G. elata-induced increases and was also found to significantly increase (p < 0.05) the content of soil NH4+-N and AP. The alpha diversities of soil microbial communities were significantly increased (p < 0.01) by cultivation of G. elata and P. impudicus as indicated with Chao1 estimator and Shannon index. The structure and composition of soil microbial communities differed responding to different cultivation species. In particular, the relative abundances of Bacillus, norank_o_Gaiellales, Mortierella and unclassified_k_Fungi were significantly increased (p < 0.05), while the abundances of potentially beneficial genera such as Acidibacter, Acidothermus, Cryptococcus, and Penicillium etc., were significantly decreased (p < 0.05) by cultivation of G. elata. It’s interesting to find that cultivation of P. impudicus increased the abundances of these genera that G. elata decreased before, which contributed to the difference of composition and structure. The results of CCA and heatmap indicated that the changes of soil microbial communities had strong correlations with soil nutrients. Specifically, among 28 genera presented, 50% and 42.9% demonstrated significant correlations with soil pH and NO3-N in response to cultivation of G. elata and P. impudicus.ConclusionsOur findings suggested that the cultivation of P. impudicus might have potential benefits as result of affecting soil microorganisms coupled with changes in soil nutrient profile.

Screening of cellulose-degrading bacteria and optimization of cellulase production from Bacillus cereus A49 through response surface methodology

Cellulose-degrading microorganisms hold immense significance in utilizing cellulose resources efficiently. The screening of natural cellulase bacteria and the optimization of fermentation conditions are the hot spots of research. This study meticulously screened cellulose-degrading bacteria from mixed soil samples adopting a multi-step approach, encompassing preliminary culture medium screening, Congo red medium-based re-screening, and quantification of cellulase activity across various strains. Particularly, three robust cellulase-producing strains were identified: A24 (MT740356.1 Brevibacillus borstelensis), A49 (MT740358.1 Bacillus cereus), and A61 (MT740357.1 Paenibacillus sp.). For subsequent cultivation experiments, the growth curves of the three obtained isolates were monitored diligently. Additionally, optimal CMCase production conditions were determined, keeping CMCase activity as a key metric, through a series of single-factor experiments: agitation speed, cultivation temperature, unit medium concentration, and inoculum volume. Maximum CMCase production was observed at 150 rpm/37 °C, doubling the unit medium addition, and a 5 mL inoculation volume. Further optimization was conducted using the selected isolate A49 employing response surface methodology. The software model recommended a 2.21fold unit medium addition, 36.11 °C temperature, and 4.91 mL inoculant volume for optimal CMCase production. Consequently, three parallel experiments were conducted based on predicted conditions consistently yielding an average CMCase production activity of 15.63 U/mL, closely aligning with the predicted value of 16.41 U/mL. These findings validated the reliability of the model and demonstrated the effectiveness of optimized CMCase production conditions for isolate A49.

Vetiver grass cleans up arsenic contaminated field for subsequent safe cultivation of rice with low arsenic in grains: A two year field study

The presence of high concentrations of arsenic (As) in agricultural soils and its subsequent accumulation in rice crop is a serious issue threatening sustainability of agriculture and human health. In the present work, remediation of As contaminated field in Nadia, West Bengal, India was done through the cultivation of Vetiver (Vetiveria zizanoides L. Nash) and the same field was subsequently used for rice (Oryza sativa L.) cultivation. The results showed that V. zizanoides could reduce As concentrations in the field to bring it lower than the maximum permissible limit (20 mg kg−1) in 11 months' time. The rice plants grown in remediated field showed improvement in growth and photosynthesis parameters as compared to that of contaminated field. Importantly, yield related parameters (filled seed, 1000 grain weight, number of panicles etc.) were also significantly higher in remediated field than that in contaminated field. Arsenic concentration in roots, shoot, husk and grains of rice was found to be significantly lower in remediated field than in contaminated field. Grain As decreased from 0.75 to 0.77 μg g−1 dw in contaminated field to 0.15–0.18 μg g−1 dw. In conclusion, replacing rice for single year with V. zizanoides crop can significantly remediate the field and can be a viable option.

Specifics of Cryopreservation of Hydrogel Biopolymer Scaffolds with Encapsulated Mesenchymal Stem Cells.

The demand for regenerative medicine products is growing rapidly in clinical practice. Unfortunately, their use has certain limitations. One of these, which significantly constrains the widespread distribution and commercialization of such materials, is their short life span. For products containing suspensions of cells, this issue can be solved by using cryopreservation. However, this approach is rarely used for multicomponent tissue-engineered products due to the complexity of selecting appropriate cryopreservation protocols and the lack of established criteria for assessing the quality of such products once defrosted. Our research is aimed at developing a cryopreservation protocol for an original hydrogel scaffold with encapsulated MSCs and developing a set of criteria for assessing the quality of their functional activity in vitro. The scaffolds were frozen using two alternative types of cryocontainers and stored at either -40 °C or -80 °C. After cryopreservation, the external state of the scaffolds was evaluated in addition to recording the cell viability, visible changes during subsequent cultivation, and any alterations in proliferative and secretory activity. These observations were compared to those of scaffolds cultivated without cryopreservation. It was shown that cryopreservation at -80 °C in an appropriate type of cryocontainer was optimal for the hydrogels/adipose-derived stem cells (ASCs) tested if it provided a smooth temperature decrease during freezing over a period of at least three hours until the target values of the cryopreservation temperature regimen were reached. It was shown that evaluating a set of indicators, including the viability, the morphology, and the proliferative and secretory activity of the cells, enables the characterization of the quality of a tissue-engineered construct after its withdrawal from cryopreservation, as well as indicating the effectiveness of the cryopreservation protocol.

A New Method for the Production of High-Concentration Collagen Bioinks with Semiautonomic Preparation.

It is believed that 3D bioprinting will greatly help the field of tissue engineering and regenerative medicine, as live patient cells are incorporated into the material, which directly creates a 3D structure. Thus, this method has potential in many types of human body tissues. Collagen provides an advantage, as it is the most common extracellular matrix present in all kinds of tissues and is, therefore, very natural for cells and the organism. Hydrogels with highly concentrated collagen make it possible to create 3D structures without additional additives to crosslink the polymer, which could negatively affect cell proliferation and viability. This study established a new method for preparing highly concentrated collagen bioinks, which does not negatively affect cell proliferation and viability. The method is based on two successive neutralizations of the prepared hydrogel using the bicarbonate buffering mechanisms of the 2× enhanced culture medium and pH adjustment by adding NaOH. Collagen hydrogel was used in concentrations of 20 and 30 mg/mL dissolved in acetic acid with a concentration of 0.05 and 0.1 wt.%. The bioink preparation process is automated, including colorimetric pH detection and adjustment. The new method was validated using bioprinting and subsequent cultivation of collagen hydrogels with incorporated stromal cells. After 96 h of cultivation, cell proliferation and viability were not statistically significantly reduced.

Efeito residual de fontes de fósforo na presença e ausência de composto orgânico na produção de beterraba e chicória em cultivo sucessivo de brócolis

ABSTRACT The utilization of organic fertilizer for a short cycle crop may allow planting, in sequence, another short cycle crop, without the need for new fertilizers, reducing the costs of a new fertilization. The objective of this study was to evaluate the residual effect of fertilization applied to broccoli with phosphorus sources in the presence and absence of organic compost, on beet and chicory production in subsequent cultivation. In broccoli cultivation, four sources of phosphorus (thermophosphate, natural phosphate, natural reactive phosphate, and triple superphosphate) (600 kg/ha of P2O5) were evaluated in the presence and absence of organic compost (Visafértil®, 55 t/ha) before planting. After the removal of the cultural remains of broccoli, beet and chicory, seedlings were transplanted in the same plots of the first crop treatments. For each species eight treatments were evaluated, in a 4 x 2 factorial scheme. The first factor were the four sources of phosphorus and the second one was the application or not of organic compost before broccoli (hybrid BRO 68) planting. Treatments with organic compost showed greater production of beet and chicory, compared to the absence of compost. Fertilization with organic compost associated to the phosphorus sources provides necessary residual effect to beet and chicory production. The best P source for beet was triple superphosphate, and for chicory, in the absence of organic compost, triple superphosphate was the best source. In the presence of organic compost, the best sources were triple superphosphate and natural phosphate.

ВИРОЩУВАННЯ СОРГО ЗЕРНОВОГО НА КИСЛИХ ГРУНТАХ В УМОВАХ ЗАКАРПАТСЬКОЇ ОБЛАСТІ

The work aims to conduct production field research to determine the potential yield level of grain sorghum on acidic soils, as well as work out the generally accepted basic elements of cultivation technology with its subsequent cultivation in the Zakarpattia region. Materials and methods. In 2023, the production range for growing grain sorghum at Karpatiagroproduct LLC was located in the lowland zone of Uzhgorod district, Zakarpattia region. The article describes agrotechnical operations related to soil cultivation, application of fertilizers and plant protection products that were used in the farm, the main of which was the application of mineral fertilizer diammonium phosphate (NP 18:46) at the rate of 160 kg during sowing. The sowing rate was 165,000 seeds per hectare with a row width of 37 sm. Results. Sorghum is not picky about soils, so it can grow on both clay and sandy soils. The landfill where sorghum was grown is characterized by a low content of mobile phosphorus (6.33 mg/kg), a very low content of total nitrogen (N-NO3- + N-NH4+ – 8.92 mg/kg). According to the degree of acidity, the soil belongs to a very acidic soil (the hydrogen index of the saline extract of the soil is 4.08). In the economic conditions of 2023, the yield of sorghum grain hybrid Iceberg was 4.4 t/ha, which is 1.0 t/ha more than the average yield of sorghum in Ukraine over the past 10 years. Taking into account the genetic potential of this crop and the sufficient level of moisture supply of the territory, the level of the sorghum harvest should be twice as large. Because sorghum is not recommended to be grown on acidic, waterlogged soils with a close occurrence of groundwater, the underachieved level of the harvest to some extent may be related to this. In addition, the low content of mobile phosphorus and very low content of available nitrogen, which is characteristic of acidic clay soils, to some extent negatively affected the processes of crop yield formation. Conclusions. The obtained sorghum yield at Karpatiagroproduct LLC - 4.4 t/ha is lower than the average yield in Italy, by 2.6 t/ha, and in France - by 0.6 t/ha. However, it was in the territory of Ukraine that the record yield of sorghum in the world was obtained - 14.43 t/ha, which testifies to the high potential of grain sorghum and the unlimited potential of scientific and industrial research to reveal the genetic expression of this crop. Keywords: grain sorghum, yield, soil pH, «Iceberg» hybrid, agricultural technology.

Електросапа – культиватор «Мрія»

In this article, we propose the design of a universal electric hoe - a cultivator with a balancing drive wheel for inter-row cultivation of row crops. Currently, chemical methods and mechanical loosening are widespread in modern agriculture, but concerns about the use of herbicides, especially in "organic farming," leave no choice. We propose a machine that allows for inter-row cultivation of any row crops, even by a child. The proposed device is to be equipped with an electric motor with an electric battery, which will allow for greater mobility, low noise and vibration levels, as well as a set of tools to perform all technological operations when growing row crops. Ninety percent of manual labor in the cultivation of garden crops is spent on sowing and subsequent cultivation of row spacing. Therefore, the device must have a certain set of tools, such as a "hiller paw" for cutting furrows and hilling; a "one-sided paw" for removing weeds from the row spacing; a "lancet paw" for deep loosening and crust destruction. As can be seen from the above, manual weeding is the most effective method of weed control for working on a personal plot. However, due to the high labor intensity of the work, the task of creating a device for processing both inter-row and intra-row weeding is now being addressed. Field tests have shown that this device performs all the tasks in full, and the battery capacity, subject to continuous processing, is enough for 400 m2. The next step in improving the functional characteristics of the proposed design is to equip the electric chopper-cultivator with a device for inter-row cultivation. Inter-row cultivation is a method of weed control that involves removing weeds from the internal space between plants in one row. This process can be an important step in crop care and is aimed at reducing competition between weeds and crops. It is important to perform inter-row weeding in a timely manner, especially in the early stages of crop growth when weeds are just beginning to gain weight. It is economically feasible for the operator to control the machine manually. After a short practice, the control accuracy can reach several centimeters.

Comparison of the Effectiveness of Different Mechanical Tools for the Termination of Cover Crops Using a Modular Prototype

Highlights Cover crops are crops that cover the bare soil, providing multiple benefits. Cover crops must undergo a termination which can be based on mechanical methods. A modular prototype has been developed to evaluate different types of tools. The synergic use of multiple tools in combination yields better results. Abstract. Cover crops are crops that occupy the soil between the end of a cash crop and the beginning of the next one, providing both agronomic and environmental benefits. To allow for the subsequent cultivation of the cash crop, cover crops must undergo an appropriate termination process based on winter frost, herbicide chemical treatments, or mechanical methods. For this latter approach, previous published research has only examined the effectiveness of termination obtained with individual mechanical tools without considering the possible synergistic effects of multiple tools with complementary actions. To address this gap, a modular multi-tool prototype has been developed in this work. It allows for the evaluation of the effects of different types of tools, both individually and in combination. The working modules of the prototype include: 1) tine cultivator; 2) disk harrow; and 3) crimper roller. Field trials were carried out in 2021 and 2022 on three fields of approximately 6000 m2, which were divided into 60 plots each and sown with vetch and barley. The plots were mechanically terminated with the prototype in different configurations, and the effectiveness of the different modules and their combinations was subsequently evaluated by measuring the NDVI index using ground sensing technology. The results indicate that the disk harrow 15°, used alone and in combination with other tools considered (crimper roller and tine cultivator for barley, crimper roller for vetch), offered the best termination effectiveness for both barley and vetch, with a termination rate of greater than or equal to 85%. Keywords: Conservation agriculture, Cover crops, Crimper roller, Disk harrow, Mechanical termination, Tine cultivator.

The Importance and Challenges of Primary Chicken Embryo Liver Cells in Studies of Poultry Viral Diseases: A Review

Primary chicken embryo liver (CEL) cells are derived from the liver tissue of chicken embryonated eggs (CEE) using an aseptic isolation technique and growth under a controlled atmosphere in an artificial environment for cell attachment and proliferation. Although this primary cultured cell has been established for more than six decades, utilization of primary cells is still the preferable medium nowadays as the “gold standard” due to several advantages over other diagnostic techniques. Cells provide better adaptability of the viruses and easily mimic the natural host environment with high virus titration. The volume of virus suspension could be increased by applying an immortal chicken embryo liver-derived cell line. The current review aimed to highlight the importance and challenges of using primary chicken embryo liver cells in poultry virus studies. Primary CEL cells are widely used as an alternative host for diagnosis of infectious poultry viruses, cultivation and passaging of virus isolates, and vaccine production. Yet, there are some challenges and limitations in handling this primary cell, which requires appropriate facilities and environment to sustain the rapid growth of confluent monolayer cells, as highlighted in this paper. The availability of specific pathogen-free CEE is a major concern due to limited resources globally, thus creating a challenge for vaccine manufacturers to upscale the cultured cells. Future improvement of primary cell culture preparation necessitates new technology by applying cellular microcarrier in the bioreactor machine for efficient cell growth and subsequent routine virus cultivation. This study can help the researchers understand the advantages of primary CEL cells and their applications due to their significant impact on poultry viruses.