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by Maria Magdalena Rivera Salas ,José Basilio Heredia ,Juan Manuel Tovar Pedraza ,Cesar San Martín Hernández ,José Benigno Valdez Torres ,Isabel Cruz Lachica ,Raymundo Saúl García Estrada*
Accepted: 22/July/2024 – Published: 23/August/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2405-11
Abstract Background/Objective. Species of Bacillus are currently gaining interest because its ability to produce secondary metabolites with antifungal properties against various plant pathogenic fungi. The objective of this study was to optimize fermentation temperature and time for antifungal extract production by Bacillus amyloliquefaciens B17 and to verify its activity against plant pathogenic fungi Gilbertella persicaria, Choanephora cucurbitarum, Colletotrichum asianum, and Botrytis cinerea.
Materials and Methods. A central composite design (CCD) with two factors and five levels (fermentation temperature: 23.7, 25, 28, 31, and 32.2 °C and fermentation time: 25, 46, 95, 144, and 164.3 h) was used. Thirteen combinations of temperature and fermentation time were randomly performed. The thirteen crude extracts of B. amyloliquefaciens B17 were obtained from the cell-free fermentation broth by acid precipitation followed by alkaline solubilization. The response variable was the diameter of the inhibition halos generated by placing drops of the different crude extracts onto the medium already inoculated with a suspension of Gilbertella persicaria spores.
Results. The optimal conditions for the production of the extract with the greatest antifungal activity in B. amyloliquefaciens B17 were 26.8 °C and 158.6 h.
Conclusion. The optimized crude extract from B. amyloliquefaciens B17 exhibited a strong ability to inhibit mycelial growth and spore germination of Gilbertella persicaria, Choanephora cucurbitarum, Colletotrichum asianum, and Botrytis cinerea.
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Figure 1. Response surface plot for the inhibition halo diameter as a function of fermentation time and temperature
Table 1. Experimental results of the inhibition halos generated by the crude extracts of B. amyloliquefaciens obtained under different fermentation conditions.
Table 2. ANOVA and regression analysis for the response variable (diameter of the inhibition halo) generated by the crude extracts of B. amyloliquefaciens obtained under different fermentation conditions.
by Erick Ortega Piña ,Daniel Leobardo Ochoa Martínez* ,Reyna Isabel Rojas Martínez ,Alfredo Díaz Lara
Accepted: 26/July/2024 – Published: 12/August/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2402-8
Abstract Background/Objective. In broad bean (Vicia faba) crops established in Montecillo, Texcoco, State of Mexico, 100% of plants were observed with viral symptoms consisting of mosaic, mottling, leaf curling, wilting and a notable decrease in plant development. These symptoms led to a significant reduction in seed yield and quality. In this context, the present study was carried out with the objective of identifying the viral species associated with these symptoms.
Materials and Methods. The experimental strategy consisted of extraction of total RNA from leaves showing the aforementioned symptoms, followed by nextgeneration sequencing.
Results. In the results, the complete genomes of Orthotospovirus impatiensnecromaculae (before impatiens necrotic spot virus) and two isolates of the Potyvirus phaseoluteum (before bean yellow mosaic virus) were obtained. The phylogenetic analysis revealed that the isolate of O. impatiensnecromaculae (INSV CPMEX) shows significant divergences from those previously reported in other plant species. On the other hand, the two P. phaseoluteum isolates (BYMV CP1MEX and BYMV CP2MEX) proved to be different from each other, being related to isolates reported in Sudan and Iran.
Conclusion. This result suggests considerable genetic diversity among the viruses associated with viral symptoms in faba bean crops in the region, which underlines the importance of accurate identification for the management and control of these viral infections.
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Figure 1. Detection of Orthotospovirus impatiensnecromaculae and Potyvirus phaseoluteum in broad bean plants using RT-PCR. A) Product of RT-PCR of a 438 bp fragment of the L segment that codifies the viral replicase of O. impatiensnecromaculae: MPM= Molecular weight marker 100 bp PROMEGA, negative control (-). Lanes 1, 3, 4, 9, 10, 13, 14 and 15 broad bean plant samples with symptoms of mosaic, mottling, leaf curling and wilting. B) Product of RT-PCR of a 319 bp fragment of the ORF of the P. phaseoluteum that codifies for the nucleocapsid MPM= Molecular weight marker 100 bp PROMEGA, negative control (-). Lanes 1, 2, 3, 4, 9, 10, 13, 14 and 15 broad bean plant samples with symptoms of mosaic, mottling, leaf curling and wilting. The sequences of the amplicons obtained had a 99% coverage and 99% identity with P. phaseoluteum and O. impatiensnecromaculae isolates deposited in the GenBank, respectively
Figure 2. Phylogenetic tree of the Potyvirus phaseoluteum CP1MEX and BYMV CP2MEX isolates using the General-Time- Reversible model and the maximum likelihood method with 500 bootstrap replicates.
Table 1. Primers designed to verify the presence of Potyvirus phaseoluteum (Pp) and Orthotospovirus impatiensnecromaculae (Oi) in broad bean foliar tissue with symptoms of mosaic, mottling, leaf curling, wilting and growth reduction.
by Yeison David López Galé* ,Mauricio Fernando Martínez ,Lizeth Paola Palacios Joya ,Nubia Murcia Riaño ,Mario Augusto García Dávila
Accepted: 16/July/2024 – Published: 08/August/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2311-1
Abstract Background/Objective. The level of resistance to Phytophthora cinnamomi in avocado germplasm can be evaluated indirectly through inoculation of the pathogen by wounding the stem. The objective of this work was to compare the conventional graft development method and the etiolated graft method to determine levels of indirect resistance to P. cinnamomi through the stem wound inoculation technique.
Materials and Methods. In the study, three isolates of P. cinnamomi and two avocado genotypes with different levels of resistance to the pathogen were used, Duke-7 (medium resistant) and Hass (susceptible). Clonal multiplication of the genotypes was carried out with buds grafted on rootstocks propagated by Antillean avocado seeds. Inoculation was performed on the shoot at a height of 8 cm and the growth of the lesions was measured for 24 days. With the data, the Area Under the Disease Progress Curve (AUDPC) and the Coefficients Variation (CV) are calculated. The information was analyzed with a completely randomized experimental design with a 2*2*3 factorial arrangement (Method*Genotype*Isolation).
Results. The analysis of variance for the AUDPC showed no differences between methods (p=0.1881); However, there were differences between genotypes, isolates and between the genotype*method and genotype*isolation interactions (p≤0.05). With the conventional method, the development of the outbreaks was late (141-159 days) and the size of the lesions was highly variable (CV=38.9-64.4%), being able to discriminate partial degrees of aggressiveness between isolates, but not levels of resistance between genotypes. The etiolated and greened shoots in the nursery, on the contrary, presented rapid growth (101-107 days) and greater uniformity in the lesions generated by the pathogen (CV=11.1-24.2%).
Conclusion. The development of etiolated shoots in avocado grafts is proposed as a rapid alternative method that can guarantee greater uniformity in the development of lesions within the experimental units of a treatment, thus achieving greater reliability when evaluating and selecting preliminarily. avocado genotypes with indirect resistance attributes to P. cinnamomi.
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Figure 1. Propagation and shoot development process in avocado grafts. A) Conventional graft development method, B) etiolated graft development method. Yellow dots and lines indicate the location where the wound was made and P. cinnamomi was inoculated.
Figure 2. Micrographs of histological sections of Hass and Duke-7 avocado stems developed by the conventional method and the etiolation method (40X). (MP) Medullary parenchyma, (X1) Primary xylem, (X2) Secondary xylem, (CP) Cortical parenchyma, (P) Periderm, (E) Epidermis.
Figure 3. Macroscopic and microscopic characteristics of P. cinnamomi isolate A-Ag- 041. (A) Colony growth on PDA culture medium, (B) mycelium (hyphae), (C) chlamydospores, (D) sporangium.
Mixed viral infections in vegetable crops: biochemical and molecular aspects
by Mario Sánchez Sánchez ,Irasema Vargas Arispuro ,Juan Manuel Tovar Pedraza ,Cristóbal González Pérez Pedraza ,Emmanuel Aispuro Hernández ,Eber Addí Quintana Obregón ,Miguel Ángel Martínez Téllez*
Accepted: 10/July/2024 – Published: 06/August/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2404-3
Abstract Mixed viral infections refer to the coinfection of two or more viruses in the plant, which regularly lead to exacerbated symptoms on leaves and fruits. The dynamics of coinfections may follow either a synergistic, antagonistic, or neutral interaction that impacts the severity of the symptoms and infection. Mixed viral infections occur due to the convergence of fundamental characteristics reviewed in this manuscript. The virus‒host plant interrelationship influences the establishment and spread patterns of mixed viral infections. Attention should be drawn to potential changes in the dynamics of transmission and prevalence of plant viral diseases due to the effect of anthropogenic and natural alterations to complex agroecological systems or their components, including hosts, reservoirs, vectors, ecological niches, and the emergence of new virus strains
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![Figure 1. Symptom transition in the lower, middle, and upper leaves of tomato plants agroinoculated with the infectious cloned DNA-A and DNA-B components of isolates of TYLCV from Costa Rica (ToYMoV-[CR:Gre:GR1:90]) and Tomato leaf curl Sinaloa virus (ToLCSiV-[CR:Lib:L1:02]) from Costa Rica and the infectious clone of the genomic DNA of TYLCV from the Dominican Republic (TYLCV-[DO]) individually or in all combinations. (A) TYLCV; (B) non-inoculated tomato plant; (C) ToYMoV and ToLCSiV; (D) ToYMoV and TYLCV; (E) ToLCSiV and TYLCV; (F) ToYMoV, ToLCSiV and TYLCV. Plants were photographed 21 d after agroinoculation (with permission of Maliano <em>et al.,</em> 2022)](img/RMF/Volumenes/NumNormales/V4232024/RMF2404-3/Figure1.jpg)
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Figura 2. Plant components involved in viral mixed infections. Normal communication processes between plant cells involve the interrelation of structures such as plasmodesmata (A) and nucleus (B), the former are conducted in the cell walls that allow the passage of molecules between cytoplasm. In the presence of a viral infection (C), the cell counteracts the action through well-defined biochemical pathways such as inhibition of plasmodesmata function and intracellular trafficking (D), production of specific response metabolites (They can occur before and during the development of the viral infection), (E) or increase in the production of proteins involved in detoxification processes (F). In other hand, after a successful internalization by two viruses occurs synergistic effect of mixed viral infection (G).
Figure 1. Symptom transition in the lower, middle, and upper leaves of tomato plants agroinoculated with the infectious cloned DNA-A and DNA-B components of isolates of TYLCV from Costa Rica (ToYMoV-[CR:Gre:GR1:90]) and Tomato leaf curl Sinaloa virus (ToLCSiV-[CR:Lib:L1:02]) from Costa Rica and the infectious clone of the genomic DNA of TYLCV from the Dominican Republic (TYLCV-[DO]) individually or in all combinations. (A) TYLCV; (B) non-inoculated tomato plant; (C) ToYMoV and ToLCSiV; (D) ToYMoV and TYLCV; (E) ToLCSiV and TYLCV; (F) ToYMoV, ToLCSiV and TYLCV. Plants were photographed 21 d after agroinoculation (with permission of Maliano et al., 2022)
by Rubén Félix Gastélum* ,Gabriel Herrera Rodríguez ,Norma Ávila Alistac ,Elizabeth León
Accepted: 22/July/2024 – Published: 06/August/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2404-6
Abstract White mold (Sclerotinia sclerotiorum) is the main disease of bean and potato in Sinaloa. In the present review, the symptoms and signs of the disease as well as cultural and morphological characteristics of the teleomorph of the pathogen, its ecology and the epidemiology of the disease are addressed. The implementation of a prediction system which includes the carpogenic germination of the sclerotia and the phenology of both bean and potato for the management of the disease is described. This system considers soil temperature ranging from 13 to 19 °C a at depth of 2.5 cm in the soil and the flowering stage in both bean and potato to do the first spray application of synthetic fungicide to prevent the disease. in vitro studies indicated that Trichoderma harzianum, T. viride and T. atroviride reduced mycelial growth rate of S. sclerotiorum. The same antagonistic species exerted control of white mold under field conditions, where an increment of 40% of yield was observed in the treated plots, with respect to those treated with fungicide fluazinam. Future lines of research focusing on the ecology of the pathogen and management of the disease including the antagonistic fungi in the prediction system are suggested.
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Figure 1. Mycelial growth and sclerotia of Sclerotinia sclerotiorum. A) Fungal growth obtained from bean with sclerotia forming a double circle on PDA in a Petri dish; B) Light brown sclerotia produced on bean plants in the field; C) Fungal growth obtained from potato plant with scattered sclerotia on PDA in a Petri dish; D) Black sclerotia produced on potato plants in the field
by Víctor Manuel Rodríguez Romero ,Ramón Villanueva Arce ,Enrique Durán Páramo*
Accepted: 05/June/2024 – Published: 20/June/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2405-3
Abstract Background / Objetive. Nejayote is an alkaline agroindustrial waste that is generated from the nixtamalization process of corn. The purpose of this work was to demonstrate that nejayote can be used as a culture medium for the growth of Pseudomonas fluorescens NR113647 and to produce metabolites with antifungal activity for the sustainable management of Aspergillus niger, Botrytis cinerea and Fusarium solani.
Materials and Methods. Culture media were formulated with nejayote and nejayote with glycerol, with pH 6 and 12. The bacterial biomass was separated by centrifugation and filtration and the antifungal capacity of the extracts against A. niger, B. cinerea and F. solani was determined. The determination of the metabolites present in the extracts was carried out. P. fluorescens NR113647 was able to grow on all media.
Results. The extracts from nejayote at pH 12 showed inhibition of the growth of all the fungi evaluated; at least five metabolites produced by P. fluorescens NR113647 and involved in the biocontrol of phytopathogens were identified.
Conclusion. Nejayote can be used as a culture medium for P. fluorescens NR113647, to produce biomass and secondary metabolites with antifungal capacity; in addition, nejayote could be used for the cultivation of other microorganisms.
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Figure 1. Identification of the production of hydrogen cyanide in a Petri dish. Negative control of distilled water (A), extracts free of P. fluorescens NR113647 cells in a KB6 medium (B) and extracts free of P. fluorescens NR113647 cells in a N12 medium (C).
Figure 2. Identification of siderophores, Petri dish with distilled water as a control (A) and extracts free of P. fluorescens NR113647 cells (B) (Trejo-Raya et al., 2021) and extracts free of P. fluorescens NR113647 cells in a N12 medium (C).
Table 1. Evaluation of the growth of P. fluorescens NR113647 and pH values of the different treatments after 72 h of incubation
Table 2. Evaluation of the antifungal effect of the extracts obtained from P. fluorescens NR113647, on the mycelial growth and the inhibition of Fusarium solani, Botrytis cinerea and Aspergillus niger.
Table 3. Retention factors of the P. fluorescens NR113647 extracts obtained from treatment N12, comparison against standard compounds
by William Villalobos Muller ,Laura Garita Salazar ,Ana María Conejo Salazar ,Izayana Sandoval Carvajal ,Mauricio Montero Astúa ,Lisela Moreira Carmona*
Accepted: 05/June/2024 – Published: 20/June/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2403-1
Abstract Background/Objective. Bocconia frutescens (Papaveraceae) is a small tree distributed naturally from Mexico to Argentina and the Caribbean Bassin. Bocconia trees showing symptoms resembling phytoplasmas infection, such as little leaves and witches´-broom, were found in Cartago province, Costa Rica. Detection and identification of the potential phytoplasmas associated with B. frutescens little leaf symptoms was the objective out of this study.
Materials and Methods. Evaluation of leaves tissue using transmission electron microscopy (TEM), nested PCR using universal and specific primers to amplify phytoplasmas 16S rRNA and secA genes. Nucleotidic sequences (Sanger method) were obtained from amplicons, and used for BLASTn, phylogenetic analyses, and in silico RFLP’s.
Results. Presence of phytoplasmas into phloem tissue, only in symptomatic trees, was evidenced by TEM. Comparison of partial sequences (16Sr and secA genes) by BLASTn, in silico RFLPs and phylogenetic analyses, showed the occurrence of a Candidatus Phytoplasma pruni related strain in the samples evaluated.
Conclusion. Phytoplasmas were found only in the symptomatic B. frutescens trees evaluated. The phytoplasmas were identified as a Ca. Phytoplasma pruni related strain. This is the first report of B. frutescens as a natural host of Ca. Phytoplasma pruni.
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Figure 1. Bocconia frutescens (Papaveraceae) trees showing symptoms reminiscent of phytoplasmas infection, observed in Cartago province, Costa Rica; A) witches’-broom and bunches of little leaves observed in some branches; B) Witches’- broom, defoliation, and die-back; C) Detail of axillar proliferation with leaves reduced in size.
Figure 2. (A) Pleomorphic phytoplasmas bodies (») found inside phloem of Bocconia frutescens trees with little leaves and witches’ broom symptoms observed in Cartago province, Costa Rica. CW = cell wall. (B and C) Profiles of virtual RFLPs obtained at iPhyClassifier to 16Sr F2/R2n fragment of phytoplasmas associated to B. frutescens little leaf (BfLL, GenBank Acc. No. PP353584) using BstUI (C) and HpaII (D). Molecular weight (MW) fragment sizes (bp): 1353, 1078, 872, 603, 310, 281, 271, 234, 194, 118, 72
Figure 3. Dendrogram obtained from phylogenetic analysis using Maximum Parsimony method of partial 16S rRNA gene sequences from Bocconia frutescens little leaf (BfLL) phytoplasma, 29 sequences of 16SrIII group´s strains, seven ‘Ca. Phytoplasma species´ and Acholeplasma laidlawii as outgroup. The bootstrap test selected was 1000 replicates. GenBank accession number of every sequence is shown in parentheses. BfLL phytoplasma is displayed in bold. * = tentatively new subgroups
Figure 4. Dendrogram obtained from phylogenetic analysis with Maximum Parsimony using partial secA gene sequences from Bocconia frutescens little leaf associated phytoplasma, ten sequences of secA from different 16SrIII subgroups, ‘Ca. Phytoplasma trifolii´ and Bacilus subtilis (outgroup). The bootstrap test selected was 1000 replicates. GenBank accession number of every sequence is shown in parentheses. * = tentatively new subgroup.
Table 1. Localities at Cartago province, Costa Rica, where trees of Bocconia frutescens were observed with little leaf symptoms associated with phytoplasmas.
Table 2. Data about oligonucleotides (primers) and thermocycle profiles herein used for the detection of phytoplasmas genes 16S rRNA and secA translocase gene.
Table 3. Unique oligonucleotide sequence regions (UOSR) in the 16S rRNA gene of ‘Candidatus Phytoplasma pruni’ rrnA (JQ044393, Davis et al., 2013) compared to 16Sr sequence of BfLL associated phytoplasma (PP353584). Position in the respective sequence is shown in parenthesis, every nucleotide corresponding to a SNP is displayed in standard DNA nucleotides color, bold font and underlined.
Identification of phytoplasmas associated with Bunchy Top disease of papaya in Colima, Mexico
by Pedro Valadez Ramírez ,Daniel Leobardo Ochoa Martínez* ,Guadalupe Valdovinos Martínez ,Edith Blanco Rodríguez ,Sergio Aranda Ocampo ,Candelario Ortega Acosta ,Marco Tulio Buenrostro Nava ,Jetzajary Ayerim Rodríguez Barajas ,Luis Rafael De la Torre Velázquez ,Carlos Luis Leopardi Verde
Accepted: 19/May/2024 – Published: 18/June/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2403-2
Abstract Background/Objective. Phytoplasmas, rickettsiae and viruses have been detected in papaya plants with Bunchy Top disease (BT). In 2019, papaya plants with BT-like symptoms were observed in agroecosystems of Colima, Mexico. In order to determine the BT-associated phytoplasmas species or subgroups, asymptomatic and symptomatic plants were collected from papaya agroecosystems in four papaya producer municipalities, as well as papaya-associated weeds and insects.
Materials and Methods. Phytoplasma detection and identification was conducted by PCR, sequencing and phylogenetics of translocase subunit SecA (secA) and 16S ribosomal RNA (16Sr) genes, and PCR-RFLPs in vitro and in silico for 16Sr gene.
Results. In papaya, phytoplasma groups 16SrI (subgroup AF), 16SrX, and 16SrXIII were identified in 2.08% (4 out of 192) symptomatic samples. The results of RFLPs in silico analysis showing the presence of 16SrX and 16SrXIII (sub)groups. In papaya-associated weeds and insects, phytoplasmas of group 16SrI (subgroups AF and B) were identified in 1.7% (3 out of 174) and 1.1% (2 out of 185) evaluated samples, respectively. Phytoplasma-carrying weeds were Amaranthus palmeri and Echinochloa colona; positive insects were Micrutalis calva and Balclutha mexicana.
Conclusion. It is the first time that phytoplasmas 16SrI-AF, 16SrX y 16SrXIII are associated with Bunchy Top disease of papaya in agroecosystems from Colima, Mexico. Phytoplasmas 16SrX y 16SrXIII are first reported in papaya plants at the world level and in Mexico, respectively. Phytoplasma-carrying weeds and insects are new records as natural reservoirs and potential vectors.
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Figure 1. Detection of phytoplasmas in agroecosystems papaya with Bunchy Top disease in Colima, Mexico. A) Papaya plants showing stems with shortened internodes and yellow, chlorotic leaves, positive to phytoplasmas. B) in vitro RFLPs profiles of gene 16Sr (primers R16F2n/R16R2) of the phytoplasmas under study. Lanes 1-4: samples 1S-4S- papaya, Lanes 5-6: samples 5S-6S-Amaranthus palmeri; Lane 7: sample 7S-Echinochloa colona; Lane 8: sample 8S-Micrutalis calva; and Lane 9: sample 9S-Balclutha mexicana. The DNA was digested with 10 U of every enzyme and the products were separated by electrophoresis in 3% (w/v) agarose gels in 1×TAE buffer. Lane M: 1 kb plus molecular weight marker (Thermo Scientific, EUA).
Figure 2. in silico RFLPs profiles of gene 16Sr of the phytoplasmas under study, generated with iPhyClassifier with 17 restriction enzymes. A) sample 1S-papaya; B) sample 2S-papaya; C) sample 3S-papaya; D) sample 4S-papaya, E) sample 5S-Amaranthus palmeri; F) sample 6S-Amaranthus palmeri; G) sample 7S-Echinochloa colona; H) sample 8S-Micrutalis calva; I) sample 9S-Balclutha mexicana. Lanes MW: φX174 DNA digested with Hae III
Figure 3. Phylogenetic trees of the secA (A) and 16Sr (B) trees obtained using the neighbor-joining method of 16Sr groups of phytoplasmas identified in papaya agroecosystems in Colima, Mexico (black circles) and from phytoplasma groups from other parts of the world. Accession number are shown in parentheses. Samples 1S-4S: papaya, samples 5S-6S: Amaranthus palmeri, sample 7S: Echinochloa colona, sample 8S: Micrutalis calva, sample 9S: Balclutha mexicana. The gene sequence of secA of Bacillus subtilis and the 16Sr gene sequence of Acholeplasma laidlawii were used as outgroups. In each phylogeny, bootstrap values (from 1000 replicates, greater than 70%) are shown on the branches. The scale bar indicates the number of nucleotide substitutions per site
Table 1. Detection of phytoplasmas in papaya (Carica papaya), weeds and insects associated with the crop in Colima, Mexico, in November and December of 2019.
Table 2. Phytoplasmas identified in papaya, weeds and insects associated with the crop in Colima, Mexico, in November-December of 2019.
by María Emilia Belingheri Lagunes ,Rosario Medel Ortiz* ,Alejandro Salinas Castro ,Dora Trejo Aguilar
Accepted: 18/January/2024 – Published: 07/June/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2401-1
Abstract Background/Objective. The objective of this work was to evaluate the in vitro antagonistic capacity of a strain of Clonostachys sp. against five species of fungi associated with diseases in economically important crops.
Materials and Methods. Five fungal species associated with crop diseases were tested: Alternaria alternata, Colletotrichum kahawae, C. musae, Fusarium oxysporum and F. solani. Dual cultures were performed with five replicates plus controls. Growth was recorded every 24 hours, until 360 hours were completed. Interactions were determined, the degree of antagonism and the percentage of colonization was calculated. Statistical analyses were performed with a generalized linear model (GLM).
Results. All species evaluated showed antagonism of the overgrowth type. The degree of antagonism was classified into three classes, with class two being present in three of the species. The percentage of colonization was 100% at 216 h for three of the species and 264 h for the other two. There was no significant difference in the percentage of colonization (p =0.0073), but there was a significant difference in the time of invasion (p< 0.0001).
Conclusion. Dual assays to test the antagonistic effect in vitro form the basis for the selection of candidates for biological control of fungi.
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Figure 1. in vitro antagonism of Clonostachys sp. (Csp.) against Alternaria alternata (Aa). A: After 24 hours of evaluation. B. 120 hours of evaluation. C. 216 hours of evaluation. D. 360 hours of evaluation.
Figure 2. in vitro antagonism of Clonostachys sp. (Csp.) against Colletotrichum kahawae (Ck) and Colletotrichum musae (Cm). A-D) Colletotrichum kahawae (Ck). A: 24 hours after evaluation. B: 192 hours of evaluation. C: 360 hours of evaluation. D: reverse Petri dish at 360 hours of evaluation. E-H) Colletotrichum musae (Cm) against Clonostachys sp. (Csp.). E: 24 hours of evaluation. F: 192 hours of evaluation. G: 360 hours of evaluation. H: Conidiophores of Clonostachys sp. growing on acervuli of C. musae
Figure 2. in vitro antagonism of Clonostachys sp. against Fusarium spp. A-D) Fusarium solani (Fs) against Clonostachys sp. (Csp.) A: after 24 hours of evaluation. B: at 96 hours of evaluation. C: at 360 hours of evaluation. D: masses of aqueous conidiophores at the margin of Clonostachys sp. E-H) Fusarium oxysporum (Fo) against Clonostachys sp. (Csp.) E: 24 hours of evaluation. F: 144 hours of evaluation. G: 192 hours of evaluation. H: 360 hours of evaluation
Table 1. Qualitative scale of the degree of antagonism of Clonostachys sp. and the evaluated fungi associated to diseases.
Table 2. Percentage of growth of Clonostachys sp. against the fungi.
by Alejo Jairo Cristóbal* ,José María Tun Suárez ,Arturo Reyes Ramírez ,Alberto Uc Várguez ,Silvia Edith García Díaz
Accepted: 01/July/2024 – Published: 12/July/2024 – DOI: https://doi.org/10.18781/R.MEX.FIT.2405-5
Abstract Background/Objective. In the state of Yucatan, Mexico, 10 million forest plants were produced in the last five years for various conservation and restoration actions. The main limitations in the production of these plants in nursery are disease induced by the genus Fusarium spp., that cause stem and root rots and plant production losses of up to 50%. The objective of the work was to identify the causal agent associated with stem and root rot and necrosis of cedar (Cedrela odorata) and mahogany (Swietenia macrophylla) and their in vitro sensitivity to conventional fungicides.
Materials and Methods. C. odorata and S. macrophylla plants were collected at three and six weeks of germination, respectively, with symptoms of necrosis and rot indicated; from where five fungal isolates were obtained and morphologically and molecularly identified. The Minimum Inhibitory Concentration (MIC) of spores and the Minimum Lethal Concentration (MLC) of six conventional fungicides of recurrent application in the region (Prochloraz, Carbendazim, Benomyl, Fosetyl Al, Captan and Mancozeb) were determined in vitro by the microdilution method and validate their effectiveness and viability in the management of this problematic.
Results. The morphology and molecular sequences of the isolates were similar to the reported for Fusarium solani. The MIC of F. solani spores for Prochloraz, Carbendazim, Benomyl, Captan and Mancozeb were 2.44. 11.38, 14.06, 7.81 and 7.81 ppm, respectively; Fosetyl Al, did not inhibit spore germination normal mycelial growth of the fungus was observed at the concentration evaluated.
Conclusion. Prochloraz and Mancozeb had the lowest MLC with 2.44 and 7.81 ppm, respectively.
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Figure 1. Plants with symptoms of necrosis and rot in the stem and root. A, B, and C) Cedrela odorata plants. D, E, and F) Swietenia macrophylla plants
Figure 2. A and D) View of 14-day-old Fusarium solani isolated from Cedrela odorata and Swietenia macrophylla, B) Septate mycelium and microconidia of F. solani, C) Macroconidia (3-4 septa) and microconidia (0-1 septum) of F. solani observed at 100X magnification, E) Macroconidia and microconidia observed at 40X magnification, and F) Sporodochia
Figure 3. Fungicidal effect of Prochloraz and Mancozeb on the spore solution of Fusarium solani obtained from the microdilution plate after two days of evaluation. A) Distribution in Petri dishes with PDA medium of five MIC obtained from Prochloraz, B and C) No mycelial growth of F. solani in the MIC of Prochloraz evaluated after 24 and 48 h, D) Distribution in Petri dishes with PDA medium of five MIC obtained from Mancozeb, E and F) No mycelial growth of F. solani in the MIC of Mancozeb evaluated after 24 and 48 h
Table 1. Conventional fungicides evaluated for their effect on spore germination and mycelial growth of Fusarium solani isolated from Cedrela odorata and Swietenia macrophylla.
Table 2. Minimum inhibitory concentration (MIC) and minimum lethal concentration (MLC) of conventional fungicides against Fusarium solani isolated from Cedrela odorata and Swietenia macrophylla