Browsing by Author "Höckerstedt, Layla"

Maize (Zea mays L.) is one of the most significant crops worldwide and the most common crop in Latin American countries. In Nicaragua, maize is produced mainly by small-scale farmers, who have no access to agrochemicals to control diseases. Fungi cause the majority of infectious diseases of maize. The two causal agents of the Tar spot complex do not cause major damage independently but together the oblicate parasite Phyllachora maydis Maubl. and the endophytic fungal Monographella maydis Müller & Samuels cause a lethal complex disease which causes significant yield losses in maize. The aim of this study was to determine the presence of the poorly known Tar spot disease in Nicaragua. The causal agents have been detected elsewhere in Latin American countries and now they are expected to occur also in Nicaragua, where 15 maize fields in three different municipalities (seven fields in Jalapa, one field in Pueblo Nuevo and seven fields in Nueva Guinea) were examined. Semi-structured theme interview was carried out to determine agricultural history and current situation of the maize fields. Twenty leaf samples from each field were collected randomly and the severity of Tar spot complex disease symptoms was estimated using a 9-grade scale. According to the prevalence of symptoms all seven fields from Jalapa were included in comparison of infection rate and the differences were tested by ANOVA and Tukey´s test. The fungal isolates in leaf samples were identified morphologically and isolated as pure cultures. The incidence of different fungal species in each area was examined by using logistic regression analysis. Risk odds-ratio was calculated by MedCalc calculator. To ensure the accuracy of morphological identification of the species, DNA from pure cultures was isolated with the CTAB-method. DNA was amplified with universal ITS (Internal Spacer Transcriber) primers ITS1 and ITS4, which copy the ITS1-, 5.8S- and ITS2 regions. ITS-regions of P. maydis were cloned in pGEM-T Easy vector and sequenced. All analyzed sequences were aligned with sequences of known species obtained from GenBank. In addition, ITS-regions of putative P. maydis and M. maydis isolates were annotated and their relationships in phylogenetic Neighbour-Joining analysis were determined with MEGA 5.2. The causal agents of Tar spot complex disease were tested for virulence by pathogenicity tests. Healthy maize leaves were used to determine the virulence of pathogens. Tests were carried out under laboratory conditions and repeated twice. Results suggested that Tar spot disease occurs in all sampled areas in Nicaragua and the prevalence of symptoms was affected by plant age, previous infections, nitrogen fertilization and the used maize varieties. The differences in incidence of symptoms in Jalapa were statistically significant. According the morphological and molecular identification, a total of 21 different fungi were observed in collected samples. The prevalence of species varied by region and regression analysis highlighted the occurrence of Xylaria sp. in Jalapa and Fusarium equiseti in Pueblo Nuevo. There is no previous evidence that Xylaria sp. could infect maize plants, however, the results suggested it is capable to infect maize as a secondary pathogen under favorable conditions. Sequence searches showed that ITS-sequences of P. maydis or M. maydis have not been added to GenBank previously. Phylogenetic analysis showed that both pathogens formed their own branch among the already known species. Results can be used when developing species-specific PCR primers to be used in seed inspection, predicting the spread of the disease, and studies on epidemiology, ecology and control of the fungal pathogens of maize in Nicaragua.