Fusarium oxysporum f. sp. cubense Tropical Race 4 (Foc TR4) is threatening banana production worldwide. Despite quarantine efforts, the pathogen continues to spread; thus, early diagnosis plays an essential role for the proper execution of contingency plans. Here, we assess the accuracy of four PCR-based molecular methods described in the literature for the identification and detection of race 4 strains, including Subtropical (Foc STR4) and Tropical Race 4 causing Fusarium wilt of banana. We screened a total of 302 isolates using these four markers, and performed phylogenetic analyses, Vegetative Compatibility Group (VCG) testing, sequence comparison, and pathogenicity tests for selected isolates. Our results show that three out of the four markers tested are not reliable for identification of Foc STR4 and TR4, as DNA from isolates from Ecuador, pathogenic and nonpathogenic to banana, obtained from different banana cultivars, displayed cross-reaction with these methods; that is, false positives can occur during the diagnostic process for race 4. Phylogenetic analyses, VCG testing, sequence comparison, and pathogenicity tests suggest the presence of non-target F. oxysporum isolates that share genomic regions with pathogenic strains but lack true pathogenicity to banana. The findings of this work are of foremost importance for international regulatory agencies performing surveillance tests in pathogen-free areas using the current diagnostic methods. We suggest the use of a genetic locus possibly related to virulence, previously identified by T-DNA, and amplified with primers W2987F/ W2987R, for diagnosis of Foc TR4 as the most reliable alternative. We urge the adoption of a more holistic view in the study of F. oxysporum as a plant pathogen that considers the biology and diversity of the species for the development of better diagnostic tools.

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The root-knot nematode, Meloidogyne spp., is an endoparasite that infects plants' root system and causes yield losses in several important crops. Meloidogyne is one of the most devastating pests, so searching for effective biological agents is needed to mitigate its damage. In this study, the predatory activity of Arthrobotrys oligosporus Fresen strain C-2197, obtained from a tropical dry forest of Ecuador, was evaluated as a biocontrol alternative for root-knot caused by Meloidogyne spp. Our results showed that A. oligosporus C-2197 has predatory activity against juvenile nematodes, 72.31%, and 79% efficacy, for in vitro and greenhouse conditions. Besides, the studied strain showed growth-promoting activity, increasing leaf and root area of inoculated plants. Growth promoting activity was also observed in field tests. The present study validates the potential use of A. oligosporus as a biocontrol of Meloidogyne spp. in tomato production systems under greenhouse. It also presents useful information on the use of different cultivation media and substrates for massive A. oligosporus spore concentrates.

One of the main problems in the production of cacao in Ecuador is the disease caused by the fungus Moniliophthora roreri (frosty pod rot) which affects the pods. Here, we evaluate the genetic diversity of this fungus in Ecuador, and its sensitivity to the fungicide azoxystrobin. We evaluated 76 monosporic cultures from the Amazon and the Pacific coast regions. In vitro sensitivity assays tested several doses of the fungicide azoxystrobin to determine the percent of growth inhibition and the IC50. Concentrations of 1 to 0.1 µg mL−1 inhibited the growth of at least 91% of the isolates. Three isolates were the less sensitive (IC50 = 0.0220–0.0364 µg mL−1), two from Guayas (Pacific coast) and one from Sucumbíos (Amazon) provinces. However, M. roreri is highly sensitive, and it could be used in integrated management of the disease. Genetic analyses were carried out by amplifying microsatellite markers (SSR). All the genetic diversity statistics show a higher diversity in the Amazon compared to samples of the coast region; however, the molecular variance was low (FST = 0.11). Discriminant analysis clearly distinguishes three clusters concurrent with the provinces (Sucumbíos, Orellana and El Oro) and a group with the rest of the provinces. Minimum spanning networks shows, unexpectedly, that M. roreri from the coast were derived from at least three independent introductions from the Amazon. Findings are discussed in light of previous Pan-American genetic studies and available historical reports.

Ecuador is responsible for up to 70 % of the world's production of fine flavor cocoa.Although fungal diseases have reduced cocoa production, eco-friendly alternatives have been implemented to control the spread of phytopathogens. This study aimed to explore the potential of endophytic strains isolated from fine flavor Nacional cocoa trees to synthetize compounds with antimicrobial and surface active properties. It was determined that under in vitro conditions, seven strains (DS03, DS07, DS18, DS23, DS31, DS34 and DS50) exhibited antifungal activity that inhibited the mycelial growth of Moniliophthora roreri and Moniliophthora perniciosa. The strains were identified based on their morphological (pattern forming Bacilli) and genetic characteristics using the 16S SSU rRNA gene sequences. Furthermore, we determined that the endophytic strains contained genes that encoded the lipopeptides Fengycin (fenA, fenB, fenC, fenE), Iturin (ituB, ituC, ituD), and Bacyllomicin D (bamC, bamD). The production of biosurfactant compounds was determined by a reduction in surface tension in the mineral medium. Additionally, the biosurfactant crude extract (BCE) was tested for its inhibitory activity against M. perniciosa and M. roreri. The minimal inhibitory concentration (MIC) was determined to be 0.07 and 0.035 mg mL -1 , respectively, in both cases in a fungistatic action mode. The swelling granulation and fragmentation of both phytopathogens' hyphae were observed using the BCE treatment.These results suggested the biocontrol capacity of the endophytic strains, which under in vitro conditions, have the ability to synthesize antifungal and surface active compounds that might be used as an ecological alternative in cocoa plants to prevent infection by phytopathogens.

La adaptabilidad de los hongos endófitos a sus hospedantes, los beneficios ecológicos que le brinda y los diversos mecanismos antagónicos contra plagas que poseen los convierten  en una alternativa para el control biológico de enfermedades. Se determinó el potencial de 17 cepas de hongos endofíticos foliares (FEF) obtenidas de tejido sano de Theobroma cacao como candidatas para el control biológico de Moniliophthora roreri (MR) y M. perniciosa (MP). Se evaluaron: i) el micoparasitismo de los FEF frente a colonias de Moniliophthora spp., ii) la acción de los metabolitos crudos de los FEF en el crecimiento, y iii) la habilidad para recolonizar hojas sanas del hospedante mediante ensayos de hojas sueltas. Tres cepas de Lasiodiplodia theobromae fueron las más promisorias: Ec098, Ec151 and Ec157. Estas cepas inhibieron el crecimiento de MR y MP, tanto en el enfrentamiento de las colonias como mediante sus metabolitos y, adicionalmente, recolonizaron el hospedante entre el 80-100 % de las veces. Otras cepas mostraron valores destacados en un indicador, y no deseables en otros. Por ejemplo, la Ec035 (L. theobromae) mostró los niveles más altos de micoparasitismo contra ambos patógenos en la interacción de las colonias, y el segundo mejor por sus metabolitos, pero no pudo reinfectar el hospedante. La cepa Ec059 (Xylaria feejeensis) reinfectó 100 %, pero no mostró los atributos deseados de antagonismo. Por su parte, los metabolitos de Ec107 (Colletotrichum gloeosporioides s.l.) inhibieron a MR en un 60 %, pero también estimularon el crecimiento de MP. Ninguna cepa logró todas las características deseables para un agente de control biológico.

No abstract

The new species Zanclospora bicolorata, found on a rotten leaf of an unidentified plant in Ecuador, is described and illustrated. The fungus is characterized by setiform bicolored conidiophores and amygdaliform to suballantoid unicellular hyaline conidia.