Rational design of a survey protocol for avocado sunblotch viroid in commercial orchards to demonstrate pest freedom.

Avocado sunblotch viroid (ASBVd) is a subcellular pathogen of avocado that reduces yield from a tree, diminishes the appearance of the fruit by causing unsightly scarring and impedes trade because of quarantine conditions that are imposed to prevent spread of the pathogen via seed-borne inoculum. For countries where ASBVd is officially reported, permission to export fruit to another country may only be granted if an orchard can be demonstrated to be a pest free production site. The survey requirements to demonstrate pest freedom are usually defined in export protocols that have been mutually agreed upon by the trading partners. In this paper, we introduce a flexible statistical protocol for use in optimizing sampling strategies to establish pest free status from ASBVd in avocado orchards. The protocol, which is supported by an interactive app, integrates statistical considerations of multistage sampling of trees in orchards with a RT-qPCR assay allowing for detection of infection in pooled samples of leaves taken from multiple trees. While this study was motivated by a need to design a survey protocol for ASBVd, the theoretical framework and the accompanying app have broader applicability to a range of plant pathogens in which hierarchical sampling of a target population is coupled with pooling of material prior to diagnosis.

Diversity of gall-forming rusts (Uromycladium, Pucciniales) on Acacia in Australia.

Uromycladium tepperianum has been reported on over 100 species of Acacia, as well as on the closely related plant genera, Falcataria, Racosperma and Paraserianthes. Previous studies have indicated that U. tepperianum may represent a complex of host-specific, cryptic species. The phylogenetic relationships between 79 specimens of Uromycladium were determined based on a concatenated dataset of the Small Subunit, the Internal Transcribed Spacer and the Large Subunit regions of nuclear ribosomal DNA, and the mitochondrial cytochrome c oxidase subunit 3. This study showed that the host range of U. tepperianum s.str. was restricted to species of Acacia in the 'A. bivenosa group' sensu Chapman & Maslin (1992). An epitype of U. tepperianum on A. ligulata is designated to create a stable taxonomy for the application of this name. Sixteen novel species of Uromycladium are described, based on host preference, morphology and a phylogenetic species concept.

Fungal Planet description sheets: 320-370.

Novel species of fungi described in the present study include the following from Malaysia: Castanediella eucalypti from Eucalyptus pellita, Codinaea acacia from Acacia mangium, Emarcea eucalyptigena from Eucalyptus brassiana, Myrtapenidiella eucalyptorum from Eucalyptus pellita, Pilidiella eucalyptigena from Eucalyptus brassiana and Strelitziana malaysiana from Acacia mangium. Furthermore, Stachybotrys sansevieriicola is described from Sansevieria ehrenbergii (Tanzania), Phacidium grevilleae from Grevillea robusta (Uganda), Graphium jumulu from Adansonia gregorii and Ophiostoma eucalyptigena from Eucalyptus marginata (Australia), Pleurophoma ossicola from bone and Plectosphaerella populi from Populus nigra (Germany), Colletotrichum neosansevieriae from Sansevieria trifasciata, Elsinoë othonnae from Othonna quinquedentata and Zeloasperisporium cliviae (Zeloasperisporiaceae fam. nov.) from Clivia sp. (South Africa), Neodevriesia pakbiae, Phaeophleospora hymenocallidis and Phaeophleospora hymenocallidicola on leaves of a fern (Thailand), Melanconium elaeidicola from Elaeis guineensis (Indonesia), Hormonema viticola from Vitis vinifera (Canary Islands), Chlorophyllum pseudoglobossum from a grassland (India), Triadelphia disseminata from an immunocompromised patient (Saudi Arabia), Colletotrichum abscissum from Citrus (Brazil), Polyschema sclerotigenum and Phialemonium limoniforme from human patients (USA), Cadophora vitícola from Vitis vinifera (Spain), Entoloma flavovelutinum and Bolbitius aurantiorugosus from soil (Vietnam), Rhizopogon granuloflavus from soil (Cape Verde Islands), Tulasnella eremophila from Euphorbia officinarum subsp. echinus (Morocco), Verrucostoma martinicensis from Danaea elliptica (French West Indies), Metschnikowia colchici from Colchicum autumnale (Bulgaria), Thelebolus microcarpus from soil (Argentina) and Ceratocystis adelpha from Theobroma cacao (Ecuador). Myrmecridium iridis (Myrmecridiales ord. nov., Myrmecridiaceae fam. nov.) is also described from Iris sp. (The Netherlands). Novel genera include (Ascomycetes): Budhanggurabania from Cynodon dactylon (Australia), Soloacrosporiella, Xenocamarosporium, Neostrelitziana and Castanediella from Acacia mangium and Sabahriopsis from Eucalyptus brassiana (Malaysia), Readerielliopsis from basidiomata of Fuscoporia wahlbergii (French Guyana), Neoplatysporoides from Aloe ferox (Tanzania), Wojnowiciella, Chrysofolia and Neoeriomycopsis from Eucalyptus (Colombia), Neophaeomoniella from Eucalyptus globulus (USA), Pseudophaeomoniella from Olea europaea (Italy), Paraphaeomoniella from Encephalartos altensteinii, Aequabiliella, Celerioriella and Minutiella from Prunus (South Africa). Tephrocybella (Basidiomycetes) represents a novel genus from wood (Italy). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

A co-evolutionary relationship exists between Endoraecium (Pucciniales) and its Acacia hosts in Australia.

Endoraecium is a genus of rust fungi that infects several species of Acacia in Australia, South-East Asia and Hawaii. This study investigated the systematics of Endoraecium from 55 specimens in Australia based on a combined morphological and molecular approach. Phylogenetic analyses were conducted on partitioned datasets of loci from ribosomal and mitochondrial DNA. The recovered molecular phylogeny supported a recently published taxonomy based on morphology and host range that divided Endoraecium digitatum into five species. Spore morphology is synapomorphic and there is evidence Endoraecium co-evolved with its Acacia hosts. The broad host ranges of E. digitatum, E. parvum, E. phyllodiorum and E. violae-faustiae are revised in light of this study, and nine new species of Endoraecium are described from Australia based on host taxonomy, morphology and phylogenetic concordance.

Fungal Planet description sheets: 371-399.

Novel species of fungi described in the present study include the following from Australia: Neoseptorioides eucalypti gen. & sp. nov. from Eucalyptus radiata leaves, Phytophthora gondwanensis from soil, Diaporthe tulliensis from rotted stem ends of Theobroma cacao fruit, Diaporthe vawdreyi from fruit rot of Psidium guajava, Magnaporthiopsis agrostidis from rotted roots of Agrostis stolonifera and Semifissispora natalis from Eucalyptus leaf litter. Furthermore, Neopestalotiopsis egyptiaca is described from Mangifera indica leaves (Egypt), Roussoella mexicana from Coffea arabica leaves (Mexico), Calonectria monticola from soil (Thailand), Hygrocybe jackmanii from littoral sand dunes (Canada), Lindgomyces madisonensis from submerged decorticated wood (USA), Neofabraea brasiliensis from Malus domestica (Brazil), Geastrum diosiae from litter (Argentina), Ganoderma wiiroense on angiosperms (Ghana), Arthrinium gutiae from the gut of a grasshopper (India), Pyrenochaeta telephoni from the screen of a mobile phone (India) and Xenoleptographium phialoconidium gen. & sp. nov. on exposed xylem tissues of Gmelina arborea (Indonesia). Several novelties are introduced from Spain, namely Psathyrella complutensis on loamy soil, Chlorophyllum lusitanicum on nitrified grasslands (incl. Chlorophyllum arizonicum comb. nov.), Aspergillus citocrescens from cave sediment and Lotinia verna gen. & sp. nov. from muddy soil. Novel foliicolous taxa from South Africa include Phyllosticta carissicola from Carissa macrocarpa, Pseudopyricularia hagahagae from Cyperaceae and Zeloasperisporium searsiae from Searsia chirindensis. Furthermore, Neophaeococcomyces is introduced as a novel genus, with two new combinations, N. aloes and N. catenatus. Several foliicolous novelties are recorded from La Réunion, France, namely Ochroconis pandanicola from Pandanus utilis, Neosulcatispora agaves gen. & sp. nov. from Agave vera-cruz, Pilidium eucalyptorum from Eucalyptus robusta, Strelitziana syzygii from Syzygium jambos (incl. Strelitzianaceae fam. nov.) and Pseudobeltrania ocoteae from Ocotea obtusata (Beltraniaceae emend.). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

A bead-based suspension array for the multiplexed detection of begomoviruses and their whitefly vectors.

Bead-based suspension array systems enable simultaneous fluorescence-based identification of multiple nucleic acid targets in a single reaction. This study describes the development of a novel approach to plant virus and vector diagnostics, a multiplexed 7-plex array that comprises a hierarchical set of assays for the simultaneous detection of begomoviruses and Bemisia tabaci, from both plant and whitefly samples. The multiplexed array incorporates genus, species and strain-specific assays, offering a unique approach for identifying both known and unknown viruses and B. tabaci species. When tested against a large panel of sequence-characterized begomovirus and whitefly samples, the array was shown to be 100% specific to the homologous target. Additionally, the multiplexed array was highly sensitive, efficiently and concurrently determining both virus and whitefly identity from single viruliferous whitefly samples. The detection limit for one assay within the multiplexed array that specifically detects Tomato yellow leaf curl virus-Israel (TYLCV-IL) was quantified as 200fg of TYLCV-IL DNA, directly equivalent to that of TYLCV-specific qPCR. Highly reproducible results were obtained over multiple tests. The flexible multiplexed array described in this study has great potential for use in plant quarantine, biosecurity and disease management programs worldwide.

A review of the Ustilago-Sporisorium-Macalpinomyces complex.

The fungal genera Ustilago, Sporisorium and Macalpinomyces represent an unresolved complex. Taxa within the complex often possess characters that occur in more than one genus, creating uncertainty for species placement. Previous studies have indicated that the genera cannot be separated based on morphology alone. Here we chronologically review the history of the Ustilago-Sporisorium-Macalpinomyces complex, argue for its resolution and suggest methods to accomplish a stable taxonomy. A combined molecular and morphological approach is required to identify synapomorphic characters that underpin a new classification. Ustilago, Sporisorium and Macalpinomyces require explicit re-description and new genera, based on monophyletic groups, are needed to accommodate taxa that no longer fit the emended descriptions. A resolved classification will end the taxonomic confusion that surrounds generic placement of these smut fungi.

Soral synapomorphies are significant for the systematics of the Ustilago-Sporisorium-Macalpinomyces complex (Ustilaginaceae).

The genera Ustilago, Sporisorium and Macalpinomyces are a polyphyletic complex of plant pathogenic fungi. The four main morphological characters used to define these genera have been considered homoplasious and not useful for resolving the complex. This study re-evaluates character homology and discusses the use of these characters for defining monophyletic groups recovered from a reconstructed phylogeny using four nuclear loci. Generic delimitation of smut fungi based on their hosts is also discussed as a means for identifying genera within this group. Morphological characters and host specificity can be used to circumscribe genera within the Ustilago-Sporisorium-Macalpinomyces complex.

Taxonomic revision of Ustilago, Sporisorium and Macalpinomyces.

Morphological characters within the Ustilago-Sporisorium-Macalpinomyces complex are defined explicitly. The genera Sporisorium and Anthracocystis are emended to reflect morphological synapomorphies. Three new genera, Langdonia, Stollia and Triodiomyces are described based on soral synapomorphies and host classification. The new classification of the Ustilago-Sporisorium-Macalpinomyces complex incorporates 142 new taxonomic combinations.

Development of an immunomagnetic capture-reverse transcriptase-PCR assay for three pineapple ampeloviruses.

A semi-automated, immunomagnetic capture-reverse transcription PCR (IMC-RT-PCR) assay for the detection of three pineapple-infecting ampeloviruses, Pineapple mealybug wilt-associated virus-1, -2 and -3, is described. The assay was equivalent in sensitivity but more rapid than conventional immunocapture RT-PCR. The assay can be used either as a one- or two-step RT-PCR and allows detection of the viruses separately or together in a triplex assay from fresh, frozen or freeze-dried pineapple leaf tissue. This IMC-RT-PCR assay could be used for high throughput screening of pineapple planting propagules and could easily be modified for the detection of other RNA viruses in a range of plant species, provided suitable antibodies are available.

Identification of viral and non-viral reverse transcribing elements in pineapple (Ananas comosus), including members of two new badnavirus species.

A previously published partial sequence of pineapple bacilliform virus was shown to be from a retrotransposon (family Metaviridae) and not from a badnavirus as previously thought. Two newly discovered sequence groups isolated from pineapple were associated with bacilliform virions and were transmitted by mealybugs. Phylogenetic analyses indicated that they were members of new badnavirus species. A third caulimovirid sequence was also amplified from pineapple, but available evidence suggests that this DNA is not encapsidated, but more likely derived from an endogenous virus.

Characterisation of Banana streak Mysore virus and evidence that its DNA is integrated in the B genome of cultivated Musa.

We have sequenced the complete genome of an isolate of Banana streak virus from banana cv. 'Mysore' and show that it is sufficiently different from a previously characterised isolate from cv. 'Obino l'Ewai' to warrant recognition as a distinct species, for which the name Banana streak Mysore virus (BSMysV) is proposed. The structure of the BSMysV genome was typical of badnaviruses in general, although ORF I had a non-conventional start codon. Evidence that at least part of the BSMysV genome is integrated in the B genome of cultivated Musa is presented and transmissibility by the mealybug Planococcus citri also demonstrated.

A New Badnavirus in Ribes Species, its Detection by PCR, and its Close Association with Gooseberry Vein Banding Disease.

Gooseberry vein banding disease (GVBD) affects Ribes species and cultivars worldwide. It is the second most important virus-like disease in these crops after black currant reversion disease. In this paper, we describe a bacilliform virus, Gooseberry vein banding associated virus (GVBAV), which is associated closely with GVBD, and provide evidence that GVBAV is a distinct species within the genus Badnavirus. Purified GVBAV particles were ca. 120 × 30 nm in size and contained dsDNA. The sequence of a 1.5-kb DNA fragment amplified from viral genomic DNA was similar to those of a wide range of badnaviruses and contained motifs characteristic of the RNase H domain of the badnavirus open reading frame (ORF) III polyprotein. Phylogenetic analyses suggest that GVBAV is most closely related to Spiraea yellow leaf spot virus. Using sequence derived from the polymerase chain reaction (PCR)-amplified DNA fragment, virus-specific primers were designed. These primers were used in PCR to assay for GVBAV in a range of Ribes germplasm affected with GVBD, with other unrelated virus-like diseases and viruses found in Ribes, and in healthy plants. GVBAV was detected in all of 58 GVBD-affected plants from diverse sources, but not from healthy Ribes plants nor from plants infected with other viruses.