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The classification of two root-infecting fungi, Magnaporthe garrettii and M. griffinii , was examined by phylogenetic analysis of multiple gene sequences. This analysis demonstrated that M. garrettii and M. griffinii were sister species that formed a well-supported separate clade in Papulosaceae ( Diaporthomycetidae , Sordariomycetes ), which clusters outside of the Magnaporthales . Wongia gen. nov, is established to accommodate these two species which are not closely related to other species classified in Magnaporthe nor to other genera, including Nakataea, Magnaporthiopsis and Pyricularia , which all now contain other species once classified in Magnaporthe .

This work reports the complete genome sequence, production of a polyclonal antiserum, and host range of a Brazilian strain of johnsongrass mosaic virus (JGMV) found infecting Panicum maximum in the state of São Paulo, Brazil. The complete genome sequence of this potyvirus, comprising 9874 nucleotides, showed 82 % amino acid sequence identity in the polyprotein to that of an isolate of JGMV from Australia. The experimental host range of this virus included mainly fodder species. Cultivated species such as rice, oats, sugarcane, rye, corn and wheat were not infected, suggesting that current isolates of this potyvirus do not represent a threat to these crops in Brazil.

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.

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.

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.

The extent and importance of endogenous viral elements have been extensively described in animals but are much less well understood in plants. Here we describe a new genus of Caulimoviridae called 'Florendovirus', members of which have colonized the genomes of a large diversity of flowering plants, sometimes at very high copy numbers (>0.5% total genome content). The genome invasion of Oryza is dated to over 1.8 million years ago (MYA) but phylogeographic evidence points to an even older age of 20-34 MYA for this virus group. Some appear to have had a bipartite genome organization, a unique characteristic among viral retroelements. In Vitis vinifera, 9% of the endogenous florendovirus loci are located within introns and therefore may influence host gene expression. The frequent colocation of endogenous florendovirus loci with TA simple sequence repeats, which are associated with chromosome fragility, suggests sequence capture during repair of double-stranded DNA breaks.

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., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Efficient and reliable diagnostic tools for the routine indexing and certification of clean propagating material are essential for the management of pospiviroid diseases in horticultural crops. This study describes the development of a true multiplexed diagnostic method for the detection and identification of all nine currently recognized pospiviroid species in one assay using Luminex bead-based suspension array technology. In addition, a new data-driven, statistical method is presented for establishing thresholds for positivity for individual assays within multiplexed arrays. When applied to the multiplexed array data generated in this study, the new method was shown to have better control of false positives and false negative results than two other commonly used approaches for setting thresholds. The 11-plex Luminex MagPlex-TAG pospiviroid array described here has a unique hierarchical assay design, incorporating a near-universal assay in addition to nine species-specific assays, and a co-amplified plant internal control assay for quality assurance purposes. All assays of the multiplexed array were shown to be 100% specific, sensitive and reproducible. The multiplexed array described herein is robust, easy to use, displays unambiguous results and has strong potential for use in routine pospiviroid indexing to improve disease management strategies.

Ambrosia beetle fungiculture represents one of the most ecologically and evolutionarily successful symbioses, as evidenced by the 11 independent origins and 3500 species of ambrosia beetles. Here we document the evolution of a clade within Fusarium associated with ambrosia beetles in the genus Euwallacea (Coleoptera: Scolytinae). Ambrosia Fusarium Clade (AFC) symbionts are unusual in that some are plant pathogens that cause significant damage in naïve natural and cultivated ecosystems, and currently threaten avocado production in the United States, Israel and Australia. Most AFC fusaria produce unusual clavate macroconidia that serve as a putative food source for their insect mutualists. AFC symbionts were abundant in the heads of four Euwallacea spp., which suggests that they are transported within and from the natal gallery in mandibular mycangia. In a four-locus phylogenetic analysis, the AFC was resolved in a strongly supported monophyletic group within the previously described Clade 3 of the Fusarium solani species complex (FSSC). Divergence-time estimates place the origin of the AFC in the early Miocene ∼21.2 Mya, which coincides with the hypothesized adaptive radiation of the Xyleborini. Two strongly supported clades within the AFC (Clades A and B) were identified that include nine species lineages associated with ambrosia beetles, eight with Euwallacea spp. and one reportedly with Xyleborus ferrugineus, and two lineages with no known beetle association. More derived lineages within the AFC showed fixation of the clavate (club-shaped) macroconidial trait, while basal lineages showed a mix of clavate and more typical fusiform macroconidia. AFC lineages consisted mostly of genetically identical individuals associated with specific insect hosts in defined geographic locations, with at least three interspecific hybridization events inferred based on discordant placement in individual gene genealogies and detection of recombinant loci. Overall, these data are consistent with a strong evolutionary trend toward obligate symbiosis coupled with secondary contact and interspecific hybridization., (Copyright © 2013 Elsevier Inc. All rights reserved.)

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.

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.

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.

Monocotyledonous plant infecting mastreviruses (family Geminiviridae) have been found in the Old World. The greatest diversity of these viruses has been found in Africa but this may simply reflect the more extensive sampling that has been done there. To provide a better understanding of mastrevirus diversity in Australia, we have sequenced the genomes of 41 virus isolates found in naturalised and native grasses and identified four new species in addition to the four previously characterised species. Two of these species, which were recovered from a single Sporobolus plant, are highly divergent and are most closely related to the African streak viruses. This, coupled with the discovery of divergent dicotyledonous plant infecting mastreviruses in Australia brings into question the hypothesis that mastreviruses may have originated in Africa. We found that the patterns of inter- and intra-species recombination and the recombination hotspots mirror those found in both their African monocot-infecting counterparts and dicot-infecting mastrevirus., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Three monocot-infecting mastreviruses from Australia, all found primarily in pasture and naturalised grasses, have been characterised at the molecular level. Here, we present the full genome sequence of a fourth, Paspalum striate mosaic virus (PSMV), isolated from Paspalum dilatatum from south-east Queensland. The genome was 2816 nt long and had an organisation typical of other monocot-infecting mastreviruses. Its nearest relative is Bromus cartharticus striate mosaic virus (BCSMV), with which it shares an overall genome identity of 75%. Phylogenetic analysis of the complete genome and each of the putative viral proteins places PSMV in a group with the other three Australian striate mosaic viruses. PSMV, BCSMV and Digitaria didactyla striate mosaic virus all contain a similar, small recombinant sequence in the small intergenic region.

In 1999, banana streak disease outbreaks occurred at two locations in Australia in new banana hybrids that were being screened for fusarium wilt resistance. Two different badnaviruses, banana streak GF virus and a newly discovered virus called banana streak IM virus (BSIMV), were detected in these plants. The complete nucleotide sequence of the BSIMV genome was determined and comprised 7768 nt. Three open reading frames were detected, the first beginning with a non-conventional start codon (CUG). A 55-nt repetition in the putative pregenomic RNA promoter was also identified. Phylogenetic analysis suggests that BSIMV is most closely related to banana streak VN virus.

Endogenous members of the family Caulimoviridae have now been found in the genomes of many plant species. Although these sequences are usually fragmented and rearranged and show varying degrees of decay, the genomes of the ancestral viruses can often be reassembled in silico, allowing classification within the existing viral taxonomic framework. In this paper, we describe analyses of endogenous members of the family Caulimoviridae in the genomes of Oryza sativa, Nicotiana tabacum and Solanum spp. and on the basis of phylogeny, genome organization and genetic distance within the pol gene, propose two new virus genera called Orendovirus and Solendovirus. A system of nomenclature for endogenous virus sequences in plants is also proposed.

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.

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.

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.

Summary Banana streak virus strain OL (BSV-OL) commonly infects new Musa hybrids, and this infection is thought to arise de novo from integrated virus sequences present in the nuclear genome of the plant. Integrated DNA (Musa6+8 sequence) containing the whole genome of the virus has previously been cloned from cv. Obino l'Ewai (Musa AAB group), a parent of many of the hybrids. Using a Southern blot hybridization assay, we have examined the distribution and structure of integrated BSV-OL sequences in a range of Musa cultivars. For cv. Obino l'Ewai, almost every restriction fragment hybridizing to BSV-OL was predicted from the Musa6+8 sequence, suggesting that this is the predominant type of BSV-OL integrant in the genome. Furthermore, since only two junction fragments of Musa/BSV sequence were detected, and the Musa6+8 sequence is believed to be integrated as multiple copies in a tandem array, then the internal Musa spacer sequences must be highly conserved. Similarly sized restriction fragments were detected in four BB group cultivars, but not in six AA or AAA group cultivars, suggesting that the BSV-OL sequences are linked to the B-genome of Musa. We also provide evidence that cv. Williams (Musa AAA group) contains a distinct badnavirus integrant that is closely related to the 'dead' virus integrant previously characterized from Calcutta 4 (Musa acuminata ssp. burmannicoides). Our results suggest that the virus integrant from cv. Williams is linked to the A-genome, and the complexity of the hybridization patterns suggest multiple sites of integration and/or variation in sequence and structure of the integrants.

ABSTRACT Banana streak virus (BSV) is an important pathogen of bananas and plantains (Musa spp.) throughout the world. We have cloned and sequenced part of the genomes of four isolates of BSV from Australia, designated BSV-RD, BSV-Cav, BSV-Mys, and BSV-GF. These isolates originated from banana cvs. Red Dacca, Williams, Mysore, and Goldfinger, respectively. All clones contained a sequence covering part of open reading frame III and the intergenic region of the badnavirus genome. The sequences were compared with those of other badnaviruses, including BSV-Onne, a previously characterized isolate from Nigeria. The BSV-RD sequence was virtually identical to that of BSV-Onne, differing by only two nucleotides over 1,292 bp. However, BSV-Cav, -Mys, and -GF were divergent in nucleotide sequence. Phylogenetic analyses using conserved sequences in the ribonuclease H domain revealed that all BSV isolates were more closely related to each other than to any other badnavirus. BSV-Cav was most closely related to BSV-Onne, and there was 95.1% identity between the two amino acid sequences. Other relationships between the BSV isolates were less similar, with sequence identities ranging from 66.4 to 78.2%, which is a magnitude comparable to the distance between some of the recognized badnavirus species. Immunocapture-polymerase chain reaction assays have been developed, allowing specific detection and differentiation of the four isolates of BSV.

We have isolated a previously undescribed potexvirus from Alternanthera pungens (Amaranthaceae) in southern Queensland, Australia. This virus was shown to have a moderately wide experimental host range, infecting plants in nine of the twelve families tested. Using specific antibodies, a plate trapped antigen ELISA was developed, allowing detection of virions down to 0.8 microgram/ml of leaf extract. Virions averaged 554 nm long and had a capsid protein with a M(r) of 23.1 x 10(3). A portion of the genome containing the capsid protein ORF and 3' untranslated region was cloned and sequenced. From both serological and amino acid sequence comparisons, the virus was shown to be closely related to papaya mosaic potexvirus (PMV). To determine the taxonomic status of the virus, we assessed variation in the amino acid sequence of capsid proteins of distinct species within the potexvirus genus, as well as variation between strains of the same virus. When the core region of the capsid proteins were compared, distinct species had a maximum of 62.2% sequence identity, whereas strains had a minimum of 88.8% identity. By comparison, the core region of the capsid proteins of the Alternanthera virus and PMV had 79.8% identity. We have concluded that the Alternanthera virus is a different species from PMV, and its relationship with PMV resembles that of potyvirus subgroup members.

ABSTRACT Using biochemical, serological, and cytopathological evidence, we have confirmed that banana bract mosaic virus (BBrMV) is a distinct member of the family Potyviridae. Virions of a Philippine isolate of BBrMV were purified from field-infected banana cv. Cardaba. Particles were approximately 725-nm long, banded at a density equivalent to 1.29 to 1.31 g/ml in cesium chloride equilibrium gradients, and had an A(260/280) of 1.17. Yields of about 4 mg/kg were obtained from fresh or frozen leaf midrib or lamina tissue. Three major protein species with sizes of 31, 37, and 39 kDa were resolved from dissociated virions, and all reacted specifically with polyclonal antibodies to BBrMV. Infected leaf cells contained typical pinwheel inclusions. Virus-specific cDNA was amplified from field samples by reverse transcription-polymerase chain reaction (RT-PCR) assay using potyvirus degenerate primers. In plate-trapped antigen-enzyme-linked immunosorbent assay (ELISA), weak serological relationships were demonstrated between BBrMV and other members of the family Potyviridae, including abaca mosaic (AbaMV), dasheen mosaic, maize dwarf mosaic, sorghum mosaic, sugarcane mosaic, and wheat streak mosaic viruses. Despite similarities in the symptoms caused by the two viruses, AbaMV was serologically distinct from BBrMV and reacted only weakly, or not at all, with BBrMV antibodies in double-antibody sandwich (DAS)-ELISA. No cross reactions were observed when RT-PCR products from the two viruses were examined by Southern blot hybridization using BBrMV- and AbaMV-specific digoxigenin-labeled DNA probes. BBrMV was consistently associated with banana bract mosaic disease, as assessed by DAS-ELISA and Southern blot hybridization using DNA probes. The known geographical distribution of BBrMV was extended to include India (Kokkan disease) and Sri Lanka.

The complete nucleotide sequence of the nucleocapsid (N) gene of lettuce necrotic yellows virus (LNYV), the type member of the genus Cytorhabdovirus of plant rhabdoviruses, has been determined from cDNA clones of both the viral genomic and messenger RNAs. The previously identified N gene was adjacent to the 3' leader RNA and began at position 91 from the 3' end of the LNYV genome. Analysis of the sequence showed that the N mRNA contained a 78-nt 5' untranslated region, followed by a 1377-nt open reading frame (ORF) encoding a 459-amino-acid protein. This ORF was similar in size to the N protein ORFs of other rhabdoviruses. However, little if any direct sequence homology was found with N protein sequences of other rhabdoviruses. Short amino acid sequences were found to be conserved between the N proteins of LNYV and sonchus yellow net virus (SYNV), a member of the genus Nucleorhabdovirus of plant rhabdoviruses. Among them, a sequence of 24 amino acids which was similar between LNYV and SYNV N proteins corresponded to a region with high homology between isolates of vesicular stomatitis virus and rabies virus N protein sequences, the type members of the genera Vesiculovirus and Lyssavirus of animal rhabdoviruses, respectively.