Contamination control conditions for the in vitro sowing of orchid seeds
DOI:
https://doi.org/10.18265/2447-9187a2025id8794Palavras-chave:
Cattleya perrinii, Cattleya bicolor, fungicides, micropropagation, seed germinationResumo
Orchids are highly valued for their ornamental and economic importance, with increasing demand for their cultivation in horticulture and the floral industry. However, the in vitro sowing of orchid seeds presents considerable challenges, particularly regarding contamination control, which is critical for successful germination. This issue becomes even more pronounced in laboratories with limited infrastructure, where the absence of a laminar flow hood necessitates methodological adaptations. This study aimed to evaluate the effectiveness of various contamination control strategies by analyzing the impact of pH, culture medium composition, inoculation techniques, light regimes, and fungicide (Chlorothalonil) application on the contamination rate and germination potential of Cattleya perrinii and Cattleya bicolor seeds. Both species are endemic to Brazil and possess significant conservation and ornamental value. Three experimental conditions were tested: E1 (control), E2 (pH adjustment with a synthetic medium), and E3 (light regime alteration and fungicide application). The results indicated that E2, which involved pH optimization and the use of a synthetic medium, significantly reduced contamination, achieving a contamination index (CI) of 0%. In contrast, E3, which involved changes in light exposure and fungicide application, resulted in a CI ranging from 50% to 70%, with no significant reduction in contamination levels. These findings suggest that effective contamination control is achievable even in the absence of a laminar flow hood, particularly when pH and culture medium are appropriately adjusted. This highlights the feasibility of implementing the protocol in low-cost settings, such as small-scale laboratories or conservation initiatives. While pH and culture medium composition have proven effective without compromising seed germination, other factors, including light exposure and fungicide treatment, exhibited minimal or no influence on contamination control. Further methodological refinements are necessary, particularly regarding these variables' independent and combined effects, to enhance the reproducibility and consistency of the in vitro sowing process.
Downloads
Métricas
Referências
ABDALLA, N.; EL-RAMADY, H.; SELIEM, M. K.; EL-MAHROUK, M. E.; TAHA, N.; BAYOUMI, Y.; SHALABY, T. A.; DOBRÁNSZKI, J. An academic and technical overview on plant micropropagation challenges. Horticulturae, v. 8, n. 8, 677, 2022. DOI: https://doi.org/10.3390/horticulturae8080677.
BRAY, J. R.; CURTIS, J. T. An ordination of the upland forest communities of southern Wisconsin. Ecological Monographs, v. 27, n. 4, p. 325-349, 1957. DOI: https://doi.org/10.2307/1942268.
BROWN, D. M.; GROOM, C. L.; CVITANIK, M.; BROWN, M.; COOPER, J. L.; ARDITTI, J. Effects of fungicides and bactericides on orchid seed germination and shoot tip cultures in vitro. Plant Cell, Tissue and Organ Culture, v. 1, p. 165-180, 1982. DOI: https://doi.org/10.1007/BF02318914.
CAOVILA, L. E.; GIANINI, P. F.; PEDROSO-DE-MORAES, C. Concentração de sacarose e índices de pH no crescimento in vitro de Oncidium flexuosum SIMS. (ORCHIDACEAE). Revista em Agronegócio e Meio Ambiente, v. 9, n. 3, p. 531-545, 2016. DOI: http://dx.doi.org/10.17765/2176-9168.2016v9n3p531-545.
CAMPOS, D. M. Orquídeas: manual prático de cultura. 3. ed. Rio de Janeiro: Expressão e Cultura, 2002. In Portuguese.
COLOMBO, L. A.; FARIA, R. T.; CARVALHO, J. R. F. P.; ASSIS, A. M.; FONSECA, I. C. B. Influência do fungicida clorotalonil no desenvolvimento vegetativo e no enraizamento in vitro de duas espécies de orquídeas brasileiras. Acta Scientiarum Agronomy, v. 26, n. 2, p. 253-258, 2004. DOI: https://doi.org/10.4025/actasciagron.v26i2.1893. In Portuguese.
COUTO, J. M. F.; OTONI, W. C.; PINHEIRO, A. L.; FONSECA, E. P. Desinfestação e germinação in vitro de sementes de mogno (Swietenia macrophyla King). Revista Árvore, v. 28, n. 5, p. 633-642, 2004. DOI: https://doi.org/10.1590/S0100-67622004000500002. In Portuguese.
DEBERGH, P. C.; ZIMMERMAN, R. H. Micropropagation: technology and application. Dordrecht: Kluwer Academic Publishers, p. 30-37, 1991. DOI: https://doi.org/10.1007/978-94-009-2075-0.
DIMMOCK, J. P. R. E.; GOODING, M. J. The influence of foliar diseases, and their control by fungicides, on the protein concentration in wheat grain: a review. The Journal of Agricultural Science, v. 138, n. 4, p. 349-366, 2002. DOI: https://doi.org/10.1017/S0021859602002058.
DUNLEAVY, J. M. An economical laminar-flow microbe-free chamber for culturing small plants. Proceedings of the Iowa Academy of Science, v. 93, n. 2, p. 51-53, 1986. Available at: https://scholarworks.uni.edu/pias/vol93/iss2/7. Accessed on: 17 apr. 2025.
FALLON, M. E.; MATHEWS, R.; HINDS, M. T. In vitro flow chamber design for the study of endothelial cell (patho)physiology. Journal of Biomechanical Engineering, v. 144, n. 2, 020801, 2022. DOI: https://doi.org/10.1115/1.4051765.
GEORGE, P. S.; RAVISHANKAR, G. A. In vitro multiplication of Vanilla planifolia using axillary bud explants. Plant Cell Reports, v. 16, n. 6, p. 490-494, 1997. DOI: https://doi.org/10.1007/BF01092772.
GILADI, I.; ALTMAN, A.; GOREN, R. A method for aseptic culture for bud explants from citrus trees. Scientia Horticulturae, v. 10, n. 4, p. 357-362, 1979. DOI: https://doi.org/10.1016/0304-4238(79)90095-5.
HERNÁNDEZ-MEJÍA, J. A.; ROSA-MANZANO, E.; DELGADO-SÁNCHEZ, P. Ecosystem services provided by orchids: a global analysis. Botanical Sciences, v. 102, n. 3, p. 671-685, 2024. DOI: https://doi.org/10.17129/botsci.3478.
HOSSAIN, M. M.; KANT, R.; VAN, P. T.; WINARTO, B.; ZENG, S.; TEIXEIRA DA SILVA, J. A. The application of biotechnology to orchids. Critical Reviews in Plant Sciences, v. 32, n. 2, p. 69-139, 2013. DOI: https://doi.org/10.1080/07352689.2012.715984.
JOLMAN, D.; BATALLA, M. I.; HUNGERFORD, A.; NORWOOD, P.; TAIT, N.; WALLACE, L. E. The challenges of growing orchids from seeds for conservation: an assessment of asymbiotic techniques. Applications in Plant Sciences, v. 10, n. 5, e11496, 2022. DOI: https://doi.org/10.1002/aps3.11496.
KHEYRODIN, H.; RAJABI L.; KIANIAN M.K. Study of potato dextrose agar (PDA). Journal of Bio Innovation, v. 7, n. 4, p. 511-520, 2018. Available at: https://www.jbino.com/docs/Issue04_03_2018.pdf. Acessed on: 10 jul. 2023.
KIKUCHI, S.; BEVILACQUA, E. Semeadura – Multiplique! Revista Como Cultivar Orquídeas, v. 14, p. 16-19, 2005. São Paulo: Casa Dois Editora. In Portuguese.
KINDLMANN, P.; KULL, T.; MCCORMICK, M. The distribution and diversity of orchids. Diversity, v. 15, n. 7, 810, 2023. DOI: https://doi.org/10.3390/d15070810.
KNUDSON, L. Nonsymbiotic germination of orchid seeds. Botanical Gazette, v. 73, n. 1, p. 1-25, 1922. DOI: https://doi.org/10.1086/332956.
LAEZZA, C.; SALBITANI, G.; CARFAGNA, S. Fungal contamination in microalgal cultivation: Biological and biotechnological aspects of fungi-microalgae interaction. Journal of Fungi, v. 8, n. 10, 1099, 2022. DOI: https://doi.org/10.3390/jof8101099
LEIFERT, C.; RITCHIE, J. Y.; WAITES, W. M. Contaminants of plant-tissue and cell cultures. World Journal of Microbiology and Biotechnology, v. 7, p. 452-469, 1991. DOI: https://doi.org/10.1007/BF00303371.
LEIFERT, C.; MORRIS, C. E.; WAITES, W. M. Ecology of microbial saprophytes and pathogens in tissue culture and field-grown plants: reasons for contamination problems in vitro. Critical Reviews in Plant Sciences, v. 13, n. 2, p. 139-183, 1994. DOI: https://doi.org/10.1080/07352689409701912.
LEMES, C. S. R.; SORGATO, J. C.; SOARES, J. S.; NUNES, D. P.; RIBEIRO, L. M. Initial in vitro establishment of the native Cerrado orchid Miltonia flavescens. Floresta e Ambiente, v. 27, n. 4, e20180221, 2020. DOI: https://doi.org/10.1590/2179-8087.022118.
MONTARROYOS, A. V. V.; COELHO, R. S. B.; FERRAZ, G. M. G.; SANTOS, R.; SANTOS, V. F.; ANDRADE, P. P. Efeitos de meio de cultura, fontes de carbono e nitrogênio, pH e regime luminoso no crescimento de Mycosphaerella musicolla. Summa Phytopathologica, v. 33, n. 1, p. 86-89, 2007. DOI: https://doi.org/10.1590/S0100-54052007000100014. In Portuguese.
MOREIRA, D. M.; BOFF, L.; ARAÚJO, G. A. C.; SILVA, S. M. Ecological inferences in Orchidaceae species from the Brazilian subtropical Atlantic Forest based on morphological and functional anatomical traits. Flora, v. 317, 152558, 2024. DOI: https://doi.org/10.1016/j.flora.2024.152558.
MURASHIGE, T.; SKOOG, F. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologia Plantarum, v. 15, n. 3, p. 473-497, 1962. DOI: https://doi.org/10.1111/j.1399-3054.1962.tb08052.x.
NADAL, M. C.; MACHADO, N. B.; SANTOS, C. S.; FLORES, J. H. N.; DÓRIA, J.; PASQUAL, M. Impacto of monochromatic lights on the in vitro development of Cattleya walkeriana and effects on acclimatization. Ornamental Horticulture, v. 29, n. 2, p. 238-248, 2023. DOI: https://doi.org/10.1590/2447-536X.v29i2.2610.
NIMS, R. W.; PRICE, P. J. Best practices for detecting and mitigating the risk of cell culture contaminants. In Vitro Cellular & Development Biology-Animal, v. 53, p. 872-879, 2017. DOI: https://doi.org/10.1007/s11626-017-0203-9.
ODA, M. L.; FARIA, R. T.; FONSECA, I. C. B.; SILVA, G. L. Avaliação da fitotoxicidade de fungicidas e germicidas na propagação in vitro de Oncidium varicosum Lindl. (Orchidaceae) para o controle de microorganismos. Revista Semina: Ciências Agrárias, v. 24, n. 2, p. 273-276, 2003. DOI: https://doi.org/10.5433/1679-0359.2003v24n2p273. In Portuguese.
OLIVEIRA, R. P.; SILVEIRA, D. G.; SILVA, S. O. Efeito da desinfestação e do uso de meios de indicadores de contaminação na micropropagação da bananeira. Revista Brasileira de Fruticultura, v. 22, n. 1, p. 57-61, 2000. Available at: https://fruticultura.org/admin/files/anexo_revista/file_TxgWJUsRDE2e.pdf. Accessed on: 17 apr. 2025. In Portuguese.
OLIVEIRA, C. F.; OLIVEIRA, D. C.; PARISI, J. J. D.; BARBEDO, C. J. Deterioração de sementes de espécies brasileiras de Eugenia em função da incidência e do controle de fungos. Revista Brasileira de Sementes, v. 33, p. 520-532, 2011. DOI: https://doi.org/10.1590/S0101-31222011000300015. In Portuguese.
PANT, M.; NEGI, A.; SINGH, A.; GAUTAM, A.; RAWAT, M. Cattleya orchids: a mini review. Journal of Critical Reviews, v. 7, n. 12, p. 2394-5125, 2020. Available at: https://jcreview.com/archives/volume-7/issue-12/6390. Accessed on: 17 apr. 2025.
PARVEEN, S.; WANI, A. H.; BHAT, M. Y. Effect of culture filtrates of pathogenic and antagonistic fungi on seed germination of some economically important vegetables. Brazilian Journal of Biological Sciences, v. 6, n. 12, p. 133-139, 2019. DOI: https://doi.org/10.21472/bjbs.061212.
PINHEIRO, C. L.; ZAMPIROLLO, J. B.; MENDES, M. M.; SANTOS, V. F.; MARTINS, J. P. R.; SILVA, D. M.; TOGNELLA, M. M. P.; CASSOL, D.; FALQUETO, A. R. Exposition of three Cattleya species (Orchidaceae) to full sunlight: effect on their physiological plasticity and response to changes in light conditions. Ornamental Horticulture, v. 29, n. 1, p. 57-67, 2023. DOI: https://doi.org/10.1590/2447-536X.v29i1.2527.
RASMUSSEN, H. N.; DIXON, K. W.; JERSÁKOVÁ, J.; TĚŠITELOVÁ, T. Germination and seedling establishment in orchids: a complex of requirements. Annals of Botany, v. 116, n. 3, p. 391-402, 2015. DOI: https://doi.org/10.1093/aob/mcv087.
SETIAJI, A.; ANNISA, R. R.; SANTOSO, A. D.; KINASIH, A.; RIYADI, A. D. Factors affecting mass propagation of Vanda orchid in vitro. Cell Biology and Development, v. 5, n. 2, 2021. DOI: https://doi.org/10.13057/cellbioldev/v050201.
SILVA, J. L.; TEIXEIRA, R. N. V. Esporulação e crescimento micelial de Fusarium solani em diferentes meios de cultura e regimes de luminosidade. Revista Agro@mbiente On-line, v. 6, n. 1, 47-52, 2012. DOI: https://doi.org/10.18227/1982-8470ragro.v6i1.604. In Portuguese.
SINGH, A.; DUGGAL, S. Medicinal orchids: an overview. Ethnobotanical Leaflets, v. 2009, n. 2, p. 399-412, 2009. Available at: https://opensiuc.lib.siu.edu/ebl/vol2009/iss2/11. Accessed on: 17 apr. 2025.
STANCATO, G. C.; FARIA, R. T. In vitro growth and mineral nutrition of the lithophytic orchid Laelia cinnabarina Batem. (Orchidaceae): effects of macro and microelements. Lindleyana, v. 11, n. 1, p. 41-43, 1996.
STANCATO, G. C.; MAZZAFERA, P.; BUCKERIDGE, M. S. Effects of light stress on the growth of the epiphytic orchid Cattleya forbesii Lindl. x Laelia tenebrosa Rolfe. Revista Brasileira de Botânica, v. 25, n. 2, p. 229-235, 2002. DOI: https://doi.org/10.1590/S0100-84042002000200011.
SU, J.-F.; CHEN, S.-P.; HSIEH, T.-F. Strategies in orchid health maintenance. In: LEE, y. i.; YEUNG, E. T. (ed.). Orchid propagation: from laboratories to greenhouses. Methods and protocols, New Yourk: Springer, p. 447-460, 2018. DOI: https://doi.org/10.1007/978-1-4939-7771-0_24.
SUZUKI, R. M.; MOREIRA, V. C.; NAKABASHI, M.; FERREIRA, W. M. Estudo da germinação e crescimento in vitro de Hadrolaelia tenebrosa (Rolfe) Chiron & V.P. Castro (Orchidaceae), uma espécie da flora brasileira ameaçada de extinção. Revista Hoehnea, v. 36, n. 4, p. 657-666, 2009. DOI: https://doi.org/10.1590/S2236-89062009000400006. In Portuguese.
TISSERAT, B. Factors involved in the production of plantlets from date palm callus cultures. Euphytica, v. 31, p. 201-214, 1982. DOI: https://doi.org/10.1007/BF00028323.
VILCHERREZ-ATOCHE, J. A.; IIYAMA, C. M.; CARDOSO, J. C. Polyploidization in orchids: from cellular changes to breeding applications. Plants, v. 11, n. 4, 469, 2022. DOI: https://doi.org/10.3390/plants11040469.
WAES, J. Effects of activated charcoal on in vitro propagation of Western European orchids. Acta Horticulturae, v. 212, p. 131-138, 1987. DOI: https://doi.org/10.17660/ActaHortic.1987.212.21.
WRIGHT, N.; ALDERSON, P. G.; DULLFORCE, W. M. Micropropagation in horticulture: practice and commercial problems. Institute of Horticulture, 1986.
YUAN, S.-C.; LEKAWATANA, S.; AMORE, T. D.; CHEN, F.-C.; CHIN, S.-W.; VEGA, D. M.; WANG, Y.-T. The global orchid market. In: CHEN, F. C.; CHIN, S. W. (ed.). The orchid genome, p. 1-28, 2021. DOI: https://doi.org/10.1007/978-3-030-66826-6_1.
ZAYNAGABDINOV, R.; GABITOV, I.; BAKIEV, I.; GAFUROV, I.; KOSTAREV, K. Optimum planning use of equipment in agriculture. Journal of Industrial Engineering and Management, v. 13, n. 3, p. 514-528, 2020. DOI: https://doi.org/10.3926/jiem.3185.
Downloads
Publicado
Como Citar
Edição
Seção
Licença
Copyright (c) 2025 Izabela Gomes Schelb, João Sebastião de Paula Araújo, Felipe Zuñe, Rosana Conrado Lopes
Este trabalho está licenciado sob uma licença Creative Commons Attribution 4.0 International License.
Esta revista, seguindo as recomendações do movimento de Acesso Aberto, proporciona seu conteúdo em Full Open Access. Assim os autores conservam todos seus direitos permitindo que a Revista Principia possa publicar seus artigos e disponibilizar pra toda a comunidade.
A Revista Principia adota a licença Creative Commons 4.0 do tipo atribuição (CC-BY). Esta licença permite que outros distribuam, remixem, adaptem e criem a partir do seu trabalho, inclusive para fins comerciais, desde que lhe atribuam o devido crédito pela criação original.
Os autores estão autorizados a enviar a versão do artigo publicado nesta revista em repositório institucionais, com reconhecimento de autoria e publicação inicial na Revista Principia.
Demais informações sobre a Política de Direitos Autorais da Revista Principia encontram-se neste link.
(precisa estar logado)
