Surveying potentially antagonistic fungi to myrtle rust (Austropuccinia psidii) in Brazil: fungicolous Cladosporium spp.

The myrtle rust (MR), caused by Austropuccinia psidii, is a worldwide threat to the cultivated and wild Myrtaceae. Originally from the neotropics, it has spread to North America, Africa, and Asia and has reached geographically isolated areas in the Pacific and Australasia. It is attacking native species in those new ranges and is still spreading and causing great concern for the damage caused to endemic Myrtaceae, and to the environment. Classical biological control is regarded as the most sustainable management option for mitigating such biological invasions. However, there are no examples of introductions of host-specific co-evolved natural enemies of plant pathogens, from their native range, as a management strategy for plant pathogens. In order to explore this neglected approach, a survey of potential fungal natural enemies of A. psidii was initiated recently in the state of Minas Gerais (Brazil). Several purported mycoparasites have been collected from A. Psidii pustules formed on myrtaceous hosts. This included some isolates of dematiaceous fungi recognized as having a Cladosporium-like morphology. Here we present the results of the investigation aimed at elucidating their identity through a polyphasic taxonomic approach. Besides morphological and cultural features, molecular analyses using sequences of translation elongation factor 1-α (EF1) and actin (ACT) were performed. The combination of data generated is presented herein and placed all Cladosporium-like isolates in six species of Cladosporium, namely, Cladosporium angulosum, C. anthropophilum, C. bambusicola, C. benschii, C. guizhouense, and C. macadamiae. None of these have ever been recorded in association with A. psidii. Now, with the identification of these isolates at hand, an evaluation of biocontrol potential of these fungi will be initiated. In contrast with the ready finding of fungicolous (possibly mycoparasitic) fungi on MR in this study, no evidence of those was recorded from Australasia until now.

Assessing the Biocontrol Potential of Clonostachys Species Isolated as Endophytes from Coffea Species and as Mycoparasites of Hemileia Rusts of Coffee in Africa.

During surveys conducted in South America and Africa to identify natural fungal enemies of coffee leaf rust (CLR), Hemileia vastatrix, over 1500 strains were isolated, either as endophytes from healthy tissues of Coffea species or as mycoparasites growing on rust pustules. Based on morphological data, eight isolates-three isolated from wild or semiwild coffee and five from Hemileia species on coffee, all from Africa-were provisionally assigned to the genus Clonostachys. A polyphasic study of their morphological, cultural and molecular characteristics-including the Tef1 (translation elongation factor 1 alpha), RPB1 (largest subunit of RNA polymerase II), TUB (ß-tubulin) and ACL1 (ATP citrate lyase) regions-confirmed these isolates as belonging to three species of the genus Clonostachys: namely C. byssicola, C. rhizophaga and C. rosea f. rosea. Preliminary assays were also conducted to test the potential of the Clonostachys isolates to reduce CLR severity on coffee under greenhouse conditions. Foliar and soil applications indicated that seven of the isolates had a significant effect (p < 0.05) in reducing CLR severity. In parallel, in vitro tests that involved conidia suspensions of each of the isolates together with urediniospores of H. vastatrix resulted in high levels of inhibition of urediniospore germination. All eight isolates showed their ability to establish as endophytes in C. arabica during this study, and some proved to be mycoparasites of H. vastatrix. In addition to reporting the first records of Clonostachys associated with healthy coffee tissues and with Hemileia rusts of coffee, this work provides the first evidence that Clonostachys isolates have potential as biological control agents against CLR.

Synthesis of bis(ylidene) cyclohexanones and their antifungal activity against selected plant pathogenic fungi.

Botrytis cinerea, Rhizoctonia solani and Hemileia vastatrix are three species of phytopathogenic fungi behind major crop losses worldwide. These have been selected as target models for testing the fungicide potential of a series of bis(ylidene) cyclohexanones. Although some compounds of this chemical class are known to have inhibitory activity against human pathogens, they have never been explored for the control of phytopathogens until now. In the present work, bis(ylidene) cyclohexanones were synthesized through simple, fast and low-cost base- or acid-catalyzed aldol condensation reaction and tested in vitro against B. cinerea, R. solani and H. vastatrix. bis(pyridylmethylene) cyclohexanones showed the highest activity against the target fungi. When tested at 200 nmol per mycelial plug against R. solani., these compounds completely inhibited the mycelial growth, and the most active bis(pyridylmethylene) cyclohexanone compound had an IC50 of 155.5 nmol plug-1. Additionally, bis(pyridylmethylene) cyclohexanones completely inhibited urediniospore germination of H. vastatrix, at 125 µmol L-1. The most active bis(pyridylmethylene) cyclohexanone had an IC50 value of 4.8 µmol L-1, which was estimated as approximately 2.6 times lower than that found for the copper oxychloride-based fungicide, used as control. Additionally, these substances had a low cytotoxicity against the mammalian Vero cell line. Finally, in silico calculations indicated that these compounds present physicochemical parameters regarded as suitable for agrochemicals. Bis(ylidene) cyclohexanones may constitute promising candidates for the development of novel antifungal agents for the control of relevant fungal diseases in agriculture.

Synthesis of Tyrosol 1,2,3-Triazole Derivatives and Their Phytotoxic Activity against Euphorbia heterophylla.

The synthesis and phytotoxic activity of a series of tyrosol 1,2,3-triazole derivatives are reported herein. Target compounds were synthesized through the copper(I)-catalyzed azide-alkyne cycloaddition reaction (CuAAC), known as click reaction, and these were tested for phytotoxic activity on leaves of wild poinsettia (Euphorbia heterophylla), fleabane (Conyza sumatrensis), and tropical spiderwort (Commelina benghalensis). These are three highly noxious agricultural weeds that challenge available weed control methods, including the use of chemical herbicides. Twenty-five compounds were synthesized and tested. None of the compounds showed phytotoxic activity against C. benghalensis and C. sumatrensis, but almost all of them produced yellowing, bleaching, and necrosis on leaves of E. heterophylla. Two of the tyrosol 1,2,3-triazole derivatives produced more extensive lesions than those produced by the commercial herbicide diquat, used as a positive control (p ≤ 0.05). When applied on leaves of E. heterophylla, these compounds interfered with the stomatal conductance, net photosynthesis, internal carbon concentration, transpiration rate, water-use efficiency, and chlorophyll A and B contents. The interference of such compounds on such photosynthesis-related variables indicates that tyrosol 1,2,3-triazole derivatives may be capable of lowering the competitiveness of E. heterophylla and acting as additional tools for managing this competitive weed in agricultural lands.

Daylight LED promotes photochemical ring contraction of 2-amine-4H-pyran-3-carbonitriles with consequent loss of their antifungal activity.

The initial objective of our work was to synthesize a series of 2-amino-4H-pyran-3-carbonitriles to be tested for their antifungal activities against economically relevant phytopathogenic fungi. Fourteen compounds were prepared in up to 94% yield and shown percentages of Botrytis cinerea inhibition above 70%. Despite the promising biological results, we observed that stock solutions prepared for biological tests showed color changing when kept for a few days on the laboratory bench, under room conditions, illuminated by common LED daylight tubes (4500-6000 k). This prompted us to investigate the possible photo-induced degradation of our compounds. FT-IR ATR experiments evidenced variations in the expected bands for functional of -amino-4H-pyran-3-carbonitriles stored under LED daylight. Following, HPLC-UV analysis showed reductions in the intensity of chromatographic peaks of 2-amino-4H-pyran-3-carbonitriles, and but not for solutions kept in the dark. A solution of (E)-2-amino-8-(4-nitrobenzylidene)-4-(4-nitrophenyl)-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile underwent 84.4% of conversion after 72 h of exposure to continuous LED daylight in a BOD chamber, and the reaction product was isolated in 36% yield and characterized as (E)-7-cyano-5-(4-nitrobenzylidene)-8-(4-nitrophenyl)bicyclo[4.2.0]oct-1(6)-ene-7-carboxamide (7*). Despite freshly prepared solutions of 2-amino-4H-pyran-3-carbonitriles produced antifungal activities, these solutions lost biological activity when left on the bench for a week. Besides, compound 7* formed from photo-induced degradation of 7 also showed no antifungal activity. With this, we hope to bring two contributions: (1) production of cyclobutenes through photochemical reactions of 2-amino-4H-pyran-3-carbonitriles can be carried out through exposure to simple white LED daylight; (2) biological applications of such 2-amino-4H-pyran-3-carbonitriles may be impaired by their poor photostability.

Considerations and consequences of allowing DNA sequence data as types of fungal taxa.

Nomenclatural type definitions are one of the most important concepts in biological nomenclature. Being physical objects that can be re-studied by other researchers, types permanently link taxonomy (an artificial agreement to classify biological diversity) with nomenclature (an artificial agreement to name biological diversity). Two proposals to amend the International Code of Nomenclature for algae, fungi, and plants (ICN), allowing DNA sequences alone (of any region and extent) to serve as types of taxon names for voucherless fungi (mainly putative taxa from environmental DNA sequences), have been submitted to be voted on at the 11th International Mycological Congress (Puerto Rico, July 2018). We consider various genetic processes affecting the distribution of alleles among taxa and find that alleles may not consistently and uniquely represent the species within which they are contained. Should the proposals be accepted, the meaning of nomenclatural types would change in a fundamental way from physical objects as sources of data to the data themselves. Such changes are conducive to irreproducible science, the potential typification on artefactual data, and massive creation of names with low information content, ultimately causing nomenclatural instability and unnecessary work for future researchers that would stall future explorations of fungal diversity. We conclude that the acceptance of DNA sequences alone as types of names of taxa, under the terms used in the current proposals, is unnecessary and would not solve the problem of naming putative taxa known only from DNA sequences in a scientifically defensible way. As an alternative, we highlight the use of formulas for naming putative taxa (candidate taxa) that do not require any modification of the ICN.

Bipolaris marantae sp. nov., A Novel Helminthosporoid Species Causing Foliage Blight of the Garden Plant Maranta leuconeura in Brazil.

A severe leaf spot, turning to foliage blight, was observed on leaves of Maranta leuconeura growing in a garden in Brazil (state of Rio de Janeiro) in 2015. A dematiaceous hyphomycete bearing a morphology typical of a helminthosporoid fungi was regularly found in association with diseased tissues. The fungus was isolated and pathogenicity was demonstrated through the completion of Koch's postulates. A morphology and molecular analysis led to the conclusion that the fungus belonged to the genus Bipolaris, which is characterized by having fusiform conidia, externally thickened and truncate hila and a bipolar pattern of germination. Additionally, homology of internal transcribed spacer and GAPDH sequences with sequences of other Bipolaris species, confirmed its generic placement. A phylogenetic study also indicated clearly that the fungus on M. leuconeura is phylogenetically distinct from related species of this genus, leading to the proposal of the new species Bipolaris marantae.

Mycobiota of the weed Conyza canadensis (Asteraceae) in Brazil.

Conyza canadensis is a noxious and notably problematic weed in Brazil whose control is greatly challenging due to the appearance, in the last years, of herbicide resistant populations. An investigation regarding the associated pathogenic fungi of this plant was conducted in Brazil aimed at starting a biological control program to help mitigate its economic impact. Ten fungal species were yielded and described: two ascomycetes (Leptosphaerulina conyzicola sp. nov. and Wentiomyces melioloides), five hyphomycete asexual morphs (Alternaria tenuissima, Cercospora conyzicola sp. nov., Cercosporella virgaureae, and Fusarium fujikuroi), two coelomycete asexual morphs (Phoma conyzaphthora sp. nov., Septoria erigerontis), one rust fungus (Aecidium conyzicola sp. nov.), and one oomycete (Basidiophora entospora). Four among the fungi that were collected represent new taxa, and the others represented either new host-records of known fungal species or new geographic records or both, except for C. virgaureae that had already been recorded on C. canadensis in Brazil. None of the discovered species have any potential to be used as a mycoherbicide or to be introduced somewhere else, nevertheless, the knowledge about this mycobiota will help guide the selection of potential biocontrol agents to be introduced in Brazil.

Erasing the past: a new identity for the Damoclean pathogen causing South American leaf blight of rubber.

BACKGROUND: South American leaf blight (SALB) of rubber has been the main constraint to production in its neotropical centre of origin since commercial plantations were first established. The fungal causal agent was identified and described more than a century ago but its precise placement within the Ascomycota still remains uncertain. Indeed, such is the ambiguity surrounding the pathogen that each of the spore morphs would, according to their present classification, be placed in different ascomycete families: the Microcyclus sexual morph in the Planistromellaceae and the two purported asexual morphs--Fusicladium and Aposphaeria--in the Venturiaceae and Lophiostomataceae, respectively. Given the historical importance of the fungus and the ever-menacing threat that it poses to rubber production in the Palaeotropics--and, thus to the rubber industry and to the global economy--its phylogeny, as well as its biology, should be resolved as a matter of urgency. METHODS AND RESULTS: Here, six genomic regions (LSU rRNA, mtSSU, MCM7, EF-1α, Act and ITS) were used for reconstructing the molecular phylogeny of the SALB fungus based on material collected throughout Brazil. The analyses support the classification of the fungus in the family Mycosphaerellaceae s. str. (Capnodiales, Dothideomycetes) and place it firmly within the clade Pseudocercospora s. str., now accepted as one of the distinct genera within Mycosphaerellaceae. The new combination Pseudocercospora ulei is proposed and the life cycle of the fungus is confirmed, based on both experimental and phylogenetic evidence, with the Aposphaeria morph shown to have a spermatial rather than an infective-dispersal function. CONCLUSIONS: Because the phylogeny of the SALB fungus has now been clarified, new insights of its epidemiology and genomics can be gained following comparison with closely-related, better-researched crop pathogens.

Corynespora cassiicola f. sp. schinii, a Potential Biocontrol Agent for the Weed Schinus terebinthifolius in the United States.

Schinus terebinthifolius (Anacardiaceae), Brazilian peppertree (BP), is a major environmental weed in many tropical and subtropical areas of the globe, including Florida, Hawai'i, and Australia. This plant has been the target of a classical biocontrol project in the United States involving pathogens collected in Brazil for several years. A fungus was found in the Brazilian state of Espírito Santo causing leaf spots and severe foliage blight on BP. Examination of the morphology and internal transcribed spacer sequence analysis confirmed that the fungus is a strain of Corynespora cassiicola. Preliminary host-range tests involving 24 species, including 11 species in the family Anacardiaceae, were conducted with the fungus, and specificity toward BP was confirmed. Plants of Brazilian pepper tree from populations in Florida and Hawai'i included in the tests became severely diseased. Therefore, the recognition of a new forma specialis-Corynespora cassiicola f. sp. schinii-is proposed. The specificity of this forma specialis and the severity of the disease it caused in the field and under controlled conditions indicate that it has the potential for use as a biocontrol agent for BP in areas where it is an exotic invasive species.

Chemical characterization of volatile compounds of Lantana camara L. and L. radula Sw. and their antifungal activity.

A comparative study of the chemical composition of essential oils of two very similar species of the Verbenaceae family (Lantana camara and L. radula) revealed that the main components of essential oil of L. camara were germacrene-D (19.8%) and E-caryophyllene (19.7%), while those of L. radula were E-caryophyllene (25.3%), phytol (29.2%) and E-nerolidol (19.0%). We have hypothesized that the observed differences could contribute to the differentiated reaction of the two species of Lantana to the attack of the phytopathogenic fungi Corynespora cassiicola. An experiment, involving C. cassiicola cultivation in culture media containing volatile oils of the two species demonstrated that the oils of L. radula were more fungistatic than the oils of L. camara, in accordance with the in vivo observations. It is likely that E-nerolidol and phytol, only found in the oil of L. radula, play a significant role in the effects of L. radula on C. cassiicola.

New Meliolaceae from the Brazilian Atlantic forest 1. Species on hosts in the families Asteraceae, Burseraceae, Euphorbiaceae, Fabaceae and Sapindaceae.

Five new species, two new varieties and three newly reported taxa belonging to the Meliolaceae were collected in fragments of Atlantic forest from Minas Gerais, Brazil, in association with native plants and are described and illustrated herein. The newly described species are Appendiculella eupatorii, Meliola cassiae-ferrugineae, M. mutisiae, M. peruiferae, M. vernaliae. The new varieties are M. garugae var. protii and M. paullinifolii var. rubiginosae. These taxa are reported in Brazil for the first time: Asteridiella cyclopoda, A. entebbeensis var. codiaei and Meliola pazschkeana var. macropoda. We studied other species belonging in Meliolaceae collected on hosts belonging to the Asteraceae, Burseraceae, Euphorbiaceae, Fabaceae (Caesalpinioideae and Papilionoideae) and Sapindaceae in Brazil.

Additions to the mycobiota of the invasive weed Miconia calvescens (Melastomataceae).

Miconia calvescens (Melastomataceae) is a shrub or small tree native to the Neotropics that has become one of the worst invaders of forest ecosystems, particularly in Pacific islands such as Hawaii and French Polynesia. It has been a target for biological control for more than 10 y, both with arthropod and pathogen natural enemies. Until now Colletotrichum gloeosporioides f. sp. miconiae was the only organism to be used in biological control against this weed. This fungus was introduced both in Hawaii and in French Polynesia in the late 1990s/early 2000s, where it has caused some damage to the weed, but it became evident that additional agents are needed to achieve adequate control. Exploratory surveys for plant pathogens as potential biocontrol agents of M. calvescens were undertaken in Brazil, Costa Rica and Ecuador and yielded a diverse list of pathogens, including one phytoplasma, two nematodes, one oomycete and numerous fungi. A study including the description of five fungal species found attacking M. calvescens was published recently. Herein the following additional fungi also belonging to the mycobiota of M. calvescens are described: Hyalosphaera ornata sp. nov, Microsphaeropsis miconiae sp. nov., Myrothecium miconiae sp. nov., Phyllachora miconiiphila sp. nov., as well as Hyalosphaera miconiae, Lembosia melastomatum and Microsphaeropsis clidemiae, which are recorded here for the first time on this host. Although preliminary our observations of damage to M. calvescens caused by these seven fungal species did not indicate any potential for use in classical biological control.

Calidion bombacis, a new combination for Uredo bombacis with the record of Bombacopsisglabra (Bombacaceae) as a new host from Brazil / Calidion bombacis, uma nova combinação para Uredo bombacis e o relato de Bombacopsisglabra (Bombacaceae), como um novo hospedeiro no Brasil

The new combination Calidion bombacis is proposed for the rust species formerly known as Uredobombacis. This nomenclatural change is based on the examination of newly collected material of this fungus from a new host, Bombacopsisglabra from Brazil, and reexamination of the isotype. Until now this fungus was only known to occur in Asia (China, India and Sri Lanka). Therefore, this is also the first record of this fungus from the Neotropics.

A combinação nova Calidion bombacis é proposta para a ferrugem anteriormente conhecida como Uredo bombacis. Esta modificação nomenclatural é proposta baseada no exame de material deste fungo coletado em um novo hospedeiro, Bombacopsisglabra (Bombacaceae) no Brasil e reexame do isotipo. Até então este fungo tinha sido relatado apenas na Ásia (China, Índia e Sri Lanka). Portanto, este é o primeiro relato deste fungo nos Neotrópicos.

Calidion bombacis, a new combination for Uredo bombacis with the record of Bombacopsis glabra (Bombacaceae) as a new host from Brazil.

The new combination Calidion bombacis is proposed for the rust species formerly known as Uredo bombacis. This nomenclatural change is based on the examination of newly collected material of this fungus from a new host, Bombacopsis glabra from Brazil, and reexamination of the isotype. Until now this fungus was only known to occur in Asia (China, India and Sri Lanka). Therefore, this is also the first record of this fungus from the Neotropics.

First record of Botryosphaeria ribis associated with leaf spots on Magnolia aff. candollei in Brazil / Primeiro relato de Botryosphaeria ribis associada com doença foliar em Magnolia aff. candollei no Brasil

A leaf spot disease was observed attacking some Magnolia aff. candollei plants grown in a private garden in the city of Rio de Janeiro, RJ, Brazil. Such leaf spots coalesced and led to extensive blight of foliage. A fungus was clearly associated to the disease symptoms and was identified as Botryosphaeria ribis. Its anamorph, Fusicoccum ribis, was also present, although less abundant than the teleomorph. This is first report of this fungus on this host in Brazil and the first record of any fungal disease on a member of the genus Magnolia in Brazil.

Neste trabalho são apresentadas observações feitas sobre a etiologia de uma doença foliar encontrada atacando alguns indivíduos da espécie Magnolia aff. candollei cultivados em um jardim particular na cidade do Rio de Janeiro. Esta doença se apresentava como manchas foliares que tendiam a coalescer levando à queima de áreas extensas das folhas atacadas. Um fungo estava claramente associado a estes sintomas e foi identificado como Botryosphaeria ribis. Seu anamorfo, Fusicoccum ribis, também estava presente, embora sendo menos comum que o teliomorfo. Este é o primeiro relato deste fungo neste hospedeiro no Brasil e o primeiro relato de uma doença fúngica em planta do gênero Magnolia no Brasil.

First record of Botryosphaeria ribis associated with leaf spots on Magnolia aff. candollei in Brazil.

A leaf spot disease was observed attacking some Magnolia aff. candollei plants grown in a private garden in the city of Rio de Janeiro, RJ, Brazil. Such leaf spots coalesced and led to extensive blight of foliage. A fungus was clearly associated to the disease symptoms and was identified as Botryosphaeria ribis. Its anamorph, Fusicoccum ribis, was also present, although less abundant than the teleomorph. This is first report of this fungus on this host in Brazil and the first record of any fungal disease on a member of the genus Magnolia in Brazil.

Hemibeltrania urbanodendrii sp. nov. and Pseudobeltrania angamosensis: new fungal records from the brazilian tropical seasonal semi-deciduous montane forest

The new species Hemibeltrania urbanodendrii, associated to leaf-spots on Urbanodendron verrucosum (Lauracea) and Pseudobeltrania angamosensis, associated with leaf-spots on Virola gardneri (Myristicaceae), are recorded for the first time in Brazil. They represent additions to the mycobiota of the Tropical Seasonal Semi-Deciduous Montane Forest in Viçosa (Minas Gerais, Brazil), a highly threatened ecosystem.

Novas ocorrências de fungos relacionados a manchas foliares são apresentadas: Hemibeltrania urbanodendrii sp. nov., associado a Urbanodendron verrucosum (Lauracea) e Pseudobeltrania angamosensis, associado a Virola gardneri (Myristicaceae). Eles representam adições à micobiota da Floresta Tropical Estacional Semidecidual Montana de Viçosa (Minas Gerais), um ecossistema fortemente ameaçado.

Cercospora apii causing leaf spots on two Brazilian toxic weeds: solanum glaucophyllum and Xanthium strumarium

Samples of two toxic weeds, Solanum glaucophyllum and Xanthium strumarium, bearing leaf spots symptoms were found. Cercospora apii was found associated to these leaf spots in both plants. These are the first reports of this fungus on these hosts in Brazil and its first report on S. glaucophyllum worldwide.

Foram encontrados plantas das espécies Solanum glaucophyllum e Xanthium strumaruium apresentando sintomas de manchas foliares. Cercospora apii foi encontrado associado lesões foliares em ambas as plantas. Estes são os primeiros relatos desse fungo nestes hospedeiros no Brasil e o primeiro relato de sua ocorrência em S. glaucophyllum no mundo.

Reavaliação taxonômica de Colletotrichum guaranicola Albuq. agente causal da antracnose do guaranazeiro / Taxonomic reavaluation of Colletotrichum guaranicola Albuq. causal agent of guarana anthracnose

Foi realizada uma reavaliação morfológica de estruturas reprodutivas e de infecção do agente causal da antracnose do guaranazeiro (Paullinia cupana var. sorbilis), visando confirmar a identidade do patógeno. Os resultados reforçam o reconhecimento da espécie Colletotrichum guaranicola Albuq., como um novo taxon dentro do gênero Colletotrichum.