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Seven Fusarium species complexes are treated, namely F. aywerte species complex (FASC) (two species), F. buharicum species complex (FBSC) (five species), F. burgessii species complex (FBURSC) (three species), F. camptoceras species complex (FCAMSC) (three species), F. chlamydosporum species complex (FCSC) (eight species), F. citricola species complex (FCCSC) (five species) and the F. concolor species complex (FCOSC) (four species). New species include Fusicolla elongata from soil (Zimbabwe), and Neocosmospora geoasparagicola from soil associated with Asparagus officinalis (Netherlands). New combinations include Neocosmospora akasia, N. awan, N. drepaniformis, N. duplosperma, N. geoasparagicola, N. mekan, N. papillata, N. variasi and N. warna. Newly validated taxa include Longinectria gen. nov. , L. lagenoides , L. verticilliforme , Fusicolla gigas and Fusicolla guangxiensis . Furthermore, Fusarium rosicola is reduced to synonymy under N. brevis. Finally, the genome assemblies of Fusarium secorum (CBS 175.32) , Microcera coccophila (CBS 310.34), Rectifusarium robinianum (CBS 430.91), Rugonectria rugulosa (CBS 126565), and Thelonectria blattea (CBS 952.68) are also announced here. Citation: Crous PW, Sandoval-Denis M, Costa MM, Groenewald JZ, van Iperen AL, Starink-Willemse M, Hernández-Restrepo M, Kandemir H, Ulaszewski B, de Boer W, Abdel-Azeem AM, Abdollahzadeh J, Akulov A, Bakhshi M, Bezerra JDP, Bhunjun CS, Câmara MPS, Chaverri P, Vieira WAS, Decock CA, Gaya E, Gené J, Guarro J, Gramaje D, Grube M, Gupta VK, Guarnaccia V, Hill R, Hirooka Y, Hyde KD, Jayawardena RS, Jeewon R, Jurjević Ž, Korsten L, Lamprecht SC, Lombard L, Maharachchikumbura SSN, Polizzi G, Rajeshkumar KC, Salgado-Salazar C, Shang Q-J, Shivas RG, Summerbell RC, Sun GY, Swart WJ, Tan YP, Vizzini A, Xia JW, Zare R, González CD, Iturriaga T, Savary O, Coton M, Coton E, Jany J-L, Liu C, Zeng Z-Q, Zhuang W-Y, Yu Z-H, Thines M (2022). Fusarium and allied fusarioid taxa (FUSA). 1. Fungal Systematics and Evolution 9 : 161-200. doi: 10.3114/fuse.2022.09.08., Competing Interests: Conflict of interest: The authors declare that there is no conflict of interest., (© 2022 Westerdijk Fungal Biodiversity Institute.)

Recent publications have argued that there are potentially serious consequences for researchers in recognising distinct genera in the terminal fusarioid clade of the family Nectriaceae . Thus, an alternate hypothesis, namely a very broad concept of the genus Fusarium was proposed. In doing so, however, a significant body of data that supports distinct genera in Nectriaceae based on morphology, biology, and phylogeny is disregarded. A DNA phylogeny based on 19 orthologous protein-coding genes was presented to support a very broad concept of Fusarium at the F1 node in Nectriaceae . Here, we demonstrate that re-analyses of this dataset show that all 19 genes support the F3 node that represents Fusarium sensu stricto as defined by F. sambucinum (sexual morph synonym Gibberella pulicaris ). The backbone of the phylogeny is resolved by the concatenated alignment, but only six of the 19 genes fully support the F1 node, representing the broad circumscription of Fusarium. Furthermore, a re-analysis of the concatenated dataset revealed alternate topologies in different phylogenetic algorithms, highlighting the deep divergence and unresolved placement of various Nectriaceae lineages proposed as members of Fusarium . Species of Fusarium s. str. are characterised by Gibberella sexual morphs, asexual morphs with thin- or thick-walled macroconidia that have variously shaped apical and basal cells, and trichothecene mycotoxin production, which separates them from other fusarioid genera. Here we show that the Wollenweber concept of Fusarium presently accounts for 20 segregate genera with clear-cut synapomorphic traits, and that fusarioid macroconidia represent a character that has been gained or lost multiple times throughout Nectriaceae . Thus, the very broad circumscription of Fusarium is blurry and without apparent synapomorphies, and does not include all genera with fusarium-like macroconidia, which are spread throughout Nectriaceae ( e.g. , Cosmosporella , Macroconia , Microcera ). In this study four new genera are introduced, along with 18 new species and 16 new combinations. These names convey information about relationships, morphology, and ecological preference that would otherwise be lost in a broader definition of Fusarium . To assist users to correctly identify fusarioid genera and species, we introduce a new online identification database, Fusarioid-ID, accessible at www.fusarium.org. The database comprises partial sequences from multiple genes commonly used to identify fusarioid taxa ( act1 , CaM , his3 , rpb1 , rpb2 , tef1 , tub2 , ITS, and LSU). In this paper, we also present a nomenclator of names that have been introduced in Fusarium up to January 2021 as well as their current status, types, and diagnostic DNA barcode data. In this study, researchers from 46 countries, representing taxonomists, plant pathologists, medical mycologists, quarantine officials, regulatory agencies, and students, strongly support the application and use of a more precisely delimited Fusarium (= Gibberella ) concept to accommodate taxa from the robust monophyletic node F3 on the basis of a well-defined and unique combination of morphological and biochemical features. This F3 node includes, among others, species of the F. fujikuroi, F. incarnatum-equiseti, F. oxysporum, and F. sambucinum species complexes, but not species of Bisifusarium [ F. dimerum species complex (SC)], Cyanonectria ( F. buxicola SC), Geejayessia ( F. staphyleae SC), Neocosmospora ( F. solani SC) or Rectifusarium ( F. ventricosum SC). The present study represents the first step to generating a new online monograph of Fusarium and allied fusarioid genera (www.fusarium.org)., (© 2021 Westerdijk Fungal Biodiversity Institute. Production and hosting by ELSEVIER B.V.)

Plant-soil feedbacks (PSFs) give a mechanistic understanding on how soil properties established by previous plant species go on to influence the performance of the same or different species in monoculture, intercropping or crop rotation systems. We hypothesized that different dryland crops such as Zea mays L., Helianthus annuus L., Phaseolus vulgaris L., and Glycine max L. (Merr.) will have soil legacies that are related to the crop type. We used a two-phase experiment to test plant performance in soils previously cultivated with the same or different plant species under greenhouse conditions. The positive plant growth for all species in their own soil microbiota suggests that mutualists had a greater impact on plant performance than pathogens. The consistent positive soil-feedback results of P. vulgaris were strongly associated with their own beneficial soil microbiota, meaning that the conditioning phase legacy of mutualists and decomposers were more significant than pathogens under monoculture. Despite successful nodulation in sterilized and inoculated soils, G. max unexpectedly showed neutral and insignificant positive plant feedbacks, respectively. Helianthus annuus was superior to other crop species in creating active carbon stocks and an enzymatically active soil for the next crop. Microbial biomass results suggest that raising fungal relative to bacterial biomass can be achieved by increasing the frequency of H. annuus in rotation sequences. However, more studies are necessary to evaluate whether these elevated ratios promote or depress plant performance under field conditions. This study showed that relative to other dryland crops, H. annuus seems to have the potential of increasing fungal to bacterial ratios, raising legacies in active carbon stocks and soil microbial activity that may be crucial to successional planting in dryland systems., Competing Interests: The authors have no conflict of interest to declare., (© 2020 The Authors. Journal of Plant‐Environment Interactions Published by John Wiley & Sons Ltd.)

Novel species of fungi described in this study include those from various countries as follows: Antarctica , Cladosporium arenosum from marine sediment sand. Argentina , Kosmimatamyces alatophylus (incl. Kosmimatamyces gen. nov.) from soil. Australia , Aspergillus banksianus , Aspergillus kumbius , Aspergillus luteorubrus , Aspergillus malvicolor and Aspergillus nanangensis from soil, Erysiphe medicaginis from leaves of Medicago polymorpha , Hymenotorrendiella communis on leaf litter of Eucalyptus bicostata , Lactifluus albopicri and Lactifluus austropiperatus on soil, Macalpinomyces collinsiae on Eriachne benthamii , Marasmius vagus on soil, Microdochium dawsoniorum from leaves of Sporobolus natalensis , Neopestalotiopsis nebuloides from leaves of Sporobolus elongatus , Pestalotiopsis etonensis from leaves of Sporobolus jacquemontii , Phytophthora personensis from soil associated with dying Grevillea mccutcheonii. Brazil , Aspergillus oxumiae from soil, Calvatia baixaverdensis on soil, Geastrum calycicoriaceum on leaf litter, Greeneria kielmeyerae on leaf spots of Kielmeyera coriacea . Chile , Phytophthora aysenensis on collar rot and stem of Aristotelia chilensis. Croatia , Mollisia gibbospora on fallen branch of Fagus sylvatica. Czech Republic , Neosetophoma hnaniceana from Buxus sempervirens. Ecuador , Exophiala frigidotolerans from soil. Estonia , Elaphomyces bucholtzii in soil. France , Venturia paralias from leaves of Euphorbia paralias. India , Cortinarius balteatoindicus and Cortinarius ulkhagarhiensis on leaf litter. Indonesia , Hymenotorrendiella indonesiana on Eucalyptus urophylla leaf litter. Italy , Penicillium taurinense from indoor chestnut mill. Malaysia , Hemileucoglossum kelabitense on soil, Satchmopsis pini on dead needles of Pinus tecunumanii. Poland , Lecanicillium praecognitum on insects' frass. Portugal , Neodevriesia aestuarina from saline water. Republic of Korea , Gongronella namwonensis from freshwater. Russia , Candida pellucida from Exomias pellucidus , Heterocephalacria septentrionalis as endophyte from Cladonia rangiferina , Vishniacozyma phoenicis from dates fruit, Volvariella paludosa from swamp. Slovenia , Mallocybe crassivelata on soil. South Africa , Beltraniella podocarpi , Hamatocanthoscypha podocarpi , Coleophoma podocarpi and Nothoseiridium podocarpi (incl. Nothoseiridium gen. nov.) from leaves of Podocarpus latifolius , Gyrothrix encephalarti from leaves of Encephalartos sp., Paraphyton cutaneum from skin of human patient, Phacidiella alsophilae from leaves of Alsophila capensis , and Satchmopsis metrosideri on leaf litter of Metrosideros excelsa. Spain , Cladophialophora cabanerensis from soil, Cortinarius paezii on soil, Cylindrium magnoliae from leaves of Magnolia grandiflora , Trichophoma cylindrospora (incl. Trichophoma gen. nov.) from plant debris, Tuber alcaracense in calcareus soil, Tuber buendiae in calcareus soil. Thailand , Annulohypoxylon spougei on corticated wood, Poaceascoma filiforme from leaves of unknown Poaceae. UK , Dendrostoma luteum on branch lesions of Castanea sativa , Ypsilina buttingtonensis from heartwood of Quercus sp. Ukraine , Myrmecridium phragmiticola from leaves of Phragmites australis. USA , Absidia pararepens from air, Juncomyces californiensis (incl. Juncomyces gen. nov.) from leaves of Juncus effusus , Montagnula cylindrospora from a human skin sample, Muriphila oklahomaensis (incl. Muriphila gen. nov.) on outside wall of alcohol distillery, Neofabraea eucalyptorum from leaves of Eucalyptus macrandra , Diabolocovidia claustri (incl. Diabolocovidia gen. nov.) from leaves of Serenoa repens , Paecilomyces penicilliformis from air, Pseudopezicula betulae from leaves of leaf spots of Populus tremuloides . Vietnam , Diaporthe durionigena on branches of Durio zibethinus and Roridomyces pseudoirritans on rotten wood. Morphological and culture characteristics are supported by DNA barcodes., (© 2020 Naturalis Biodiversity Center & Westerdijk Fungal Biodiversity Institute.)

Novel species of fungi described in this study include those from various countries as follows: Antarctica , Apenidiella antarctica from permafrost, Cladosporium fildesense from an unidentified marine sponge. Argentina , Geastrum wrightii on humus in mixed forest. Australia , Golovinomyces glandulariae on Glandularia aristigera, Neoanungitea eucalyptorum on leaves of Eucalyptus grandis, Teratosphaeria corymbiicola on leaves of Corymbia ficifolia, Xylaria eucalypti on leaves of Eucalyptus radiata. Brazil , Bovista psammophila on soil, Fusarium awaxy on rotten stalks of Zea mays, Geastrum lanuginosum on leaf litter covered soil, Hermetothecium mikaniae-micranthae (incl. Hermetothecium gen. nov.) on Mikania micrantha, Penicillium reconvexovelosoi in soil, Stagonosporopsis vannaccii from pod of Glycine max. British Virgin Isles , Lactifluus guanensis on soil. Canada , Sorocybe oblongispora on resin of Picea rubens. Chile , Colletotrichum roseum on leaves of Lapageria rosea. China , Setophoma caverna from carbonatite in Karst cave. Colombia , Lareunionomyces eucalypticola on leaves of Eucalyptus grandis. Costa Rica , Psathyrella pivae on wood. Cyprus , Clavulina iris on calcareous substrate. France , Chromosera ambigua and Clavulina iris var. occidentalis on soil. French West Indies , Helminthosphaeria hispidissima on dead wood. Guatemala , Talaromyces guatemalensis in soil. Malaysia , Neotracylla pini (incl. Tracyllales ord. nov. and Neotracylla gen. nov.) and Vermiculariopsiella pini on needles of Pinus tecunumanii. New Zealand , Neoconiothyrium viticola on stems of Vitis vinifera, Parafenestella pittospori on Pittosporum tenuifolium, Pilidium novae-zelandiae on Phoenix sp. Pakistan , Russula quercus-floribundae on forest floor. Portugal , Trichoderma aestuarinum from saline water. Russia , Pluteus liliputianus on fallen branch of deciduous tree, Pluteus spurius on decaying deciduous wood or soil. South Africa , Alloconiothyrium encephalarti, Phyllosticta encephalarticola and Neothyrostroma encephalarti (incl. Neothyrostroma gen. nov.) on leaves of Encephalartos sp., Chalara eucalypticola on leaf spots of Eucalyptus grandis × urophylla, Clypeosphaeria oleae on leaves of Olea capensis, Cylindrocladiella postalofficium on leaf litter of Sideroxylon inerme , Cylindromonium eugeniicola (incl. Cylindromonium gen. nov.) on leaf litter of Eugenia capensis , Cyphellophora goniomatis on leaves of Gonioma kamassi , Nothodactylaria nephrolepidis (incl. Nothodactylaria gen. nov. and Nothodactylariaceae fam. nov.) on leaves of Nephrolepis exaltata , Falcocladium eucalypti and Gyrothrix eucalypti on leaves of Eucalyptus sp., Gyrothrix oleae on leaves of Olea capensis subsp. macrocarpa , Harzia metrosideri on leaf litter of Metrosideros sp., Hippo pota myces phragmitis (incl. Hippo pota myces gen. nov.) on leaves of Phragmites australis , Lectera philenopterae on Philenoptera violacea , Leptosillia mayteni on leaves of Maytenus heterophylla , Lithohypha aloicola and Neoplatysporoides aloes on leaves of Aloe sp., Millesimomyces rhoicissi (incl. Millesimomyces gen. nov.) on leaves of Rhoicissus digitata , Neodevriesia strelitziicola on leaf litter of Strelitzia nicolai , Neokirramyces syzygii (incl. Neokirramyces gen. nov.) on leaf spots of Syzygium sp., Nothoramichloridium perseae (incl. Nothoramichloridium gen. nov. and Anungitiomycetaceae fam. nov.) on leaves of Persea americana , Paramycosphaerella watsoniae on leaf spots of Watsonia sp., Penicillium cuddlyae from dog food, Podocarpomyces knysnanus (incl. Podocarpomyces gen. nov.) on leaves of Podocarpus falcatus , Pseudocercospora heteropyxidicola on leaf spots of Heteropyxis natalensis , Pseudopenidiella podocarpi, Scolecobasidium podocarpi and Ceramothyrium podocarpicola on leaves of Podocarpus latifolius , Scolecobasidium blechni on leaves of Blechnum capense , Stomiopeltis syzygii on leaves of Syzygium chordatum , Strelitziomyces knysnanus (incl. Strelitziomyces gen. nov.) on leaves of Strelitzia alba , Talaromyces clemensii from rotting wood in goldmine, Verrucocladosporium visseri on Carpobrotus edulis. Spain , Boletopsis mediterraneensis on soil, Calycina cortegadensisi on a living twig of Castanea sativa , Emmonsiellopsis tuberculata in fluvial sediments, Mollisia cortegadensis on dead attached twig of Quercus robur , Psathyrella ovispora on soil, Pseudobeltrania lauri on leaf litter of Laurus azorica , Terfezia dunensis in soil, Tuber lucentum in soil, Venturia submersa on submerged plant debris. Thailand , Cordyceps jakajanicola on cicada nymph, Cordyceps kuiburiensis on spider, Distoseptispora caricis on leaves of Carex sp., Ophiocordyceps khonkaenensis on cicada nymph. USA , Cytosporella juncicola and Davidiellomyces juncicola on culms of Juncus effusus , Monochaetia massachusettsianum from air sample, Neohelicomyces melaleucae and Periconia neobrittanica on leaves of Melaleuca styphelioides × lanceolata , Pseudocamarosporium eucalypti on leaves of Eucalyptus sp . , Pseudogymnoascus lindneri from sediment in a mine, Pseudogymnoascus turneri from sediment in a railroad tunnel, Pulchroboletus sclerotiorum on soil, Zygosporium pseudomasonii on leaf of Serenoa repens. Vietnam , Boletus candidissimus and Veloporphyrellus vulpinus on soil. Morphological and culture characteristics are supported by DNA barcodes., (© 2019 Naturalis Biodiversity Center & Westerdijk Fungal Biodiversity Institute.)

Three new Fusarium species, F. convolutans , F. fredkrugeri , and F. transvaalense (Ascomycota, Hypocreales, Nectriaceae) are described from soils collected in a catena landscape on a research supersite in the Kruger National Park, South Africa. The new taxa, isolated from the rhizosphere of three African herbaceous plants, Kyphocarpa angustifolia , Melhania acuminata , and Sida cordifolia , are described and illustrated by means of morphological and multilocus molecular analyses based on sequences from five DNA loci (CAL, EF-1 α, RPB1, RPB2 and TUB). According to phylogenetic inference based on Maximum-likelihood and Bayesian approaches, the newly discovered species are distributed in the Fusarium buharicum , F. fujikuroi , and F. sambucinum species complexes.

Background: Exposure to particulate matter air pollution (PM) has been associated with cardiovascular diseases., Objectives: In this study we evaluated whether annual exposure to ambient air pollution is associated with systemic inflammation, which is hypothesized to be an intermediate step to cardiovascular disease., Methods: Six cohorts of adults from Central and Northern Europe were used in this cross-sectional study as part of the larger ESCAPE project (European Study of Cohorts for Air Pollution Effects). Data on levels of blood markers for systemic inflammation-high-sensitivity C-reactive protein (CRP) and fibrinogen-were available for 22,561 and 17,428 persons, respectively. Land use regression models were used to estimate cohort participants' long-term exposure to various size fractions of PM, soot, and nitrogen oxides (NOx). In addition, traffic intensity on the closest street and traffic load within 100 m from home were used as indicators of traffic air pollution exposure., Results: Particulate air pollution was not associated with systemic inflammation. However, cohort participants living on a busy (> 10,000 vehicles/day) road had elevated CRP values (10.2%; 95% CI: 2.4, 18.8%, compared with persons living on a quiet residential street with < 1,000 vehicles/day). Annual NOx concentration was also positively associated with levels of CRP (3.2%; 95% CI: 0.3, 6.1 per 20 μg/m3), but the effect estimate was more sensitive to model adjustments. For fibrinogen, no consistent associations were observed., Conclusions: Living close to busy traffic was associated with increased CRP concentrations, a known risk factor for cardiovascular diseases. However, it remains unclear which specific air pollutants are responsible for the association.

Novel species of fungi described in the present study include the following from South Africa: Alanphillipsia aloeicola from Aloe sp., Arxiella dolichandrae from Dolichandra unguiscati, Ganoderma austroafricanum from Jacaranda mimosifolia, Phacidiella podocarpi and Phaeosphaeria podocarpi from Podocarpus latifolius, Phyllosticta mimusopisicola from Mimusops zeyheri and Sphaerulina pelargonii from Pelargonium sp. Furthermore, Barssia maroccana is described from Cedrus atlantica (Morocco), Codinaea pini from Pinus patula (Uganda), Crucellisporiopsis marquesiae from Marquesia acuminata (Zambia), Dinemasporium ipomoeae from Ipomoea pes-caprae (Vietnam), Diaporthe phragmitis from Phragmites australis (China), Marasmius vladimirii from leaf litter (India), Melanconium hedericola from Hedera helix (Spain), Pluteus albotomentosus and Pluteus extremiorientalis from a mixed forest (Russia), Rachicladosporium eucalypti from Eucalyptus globulus (Ethiopia), Sistotrema epiphyllum from dead leaves of Fagus sylvatica in a forest (The Netherlands), Stagonospora chrysopyla from Scirpus microcarpus (USA) and Trichomerium dioscoreae from Dioscorea sp. (Japan). Novel species from Australia include: Corynespora endiandrae from Endiandra introrsa, Gonatophragmium triuniae from Triunia youngiana, Penicillium coccotrypicola from Archontophoenix cunninghamiana and Phytophthora moyootj from soil. Novelties from Iran include Neocamarosporium chichastianum from soil and Seimatosporium pistaciae from Pistacia vera. Xenosonderhenia eucalypti and Zasmidium eucalyptigenum are newly described from Eucalyptus urophylla in Indonesia. Diaporthe acaciarum and Roussoella acacia are newly described from Acacia tortilis in Tanzania. New species from Italy include Comoclathris spartii from Spartium junceum and Phoma tamaricicola from Tamarix gallica. Novel genera include (Ascomycetes): Acremoniopsis from forest soil and Collarina from water sediments (Spain), Phellinocrescentia from a Phellinus sp. (French Guiana), Neobambusicola from Strelitzia nicolai (South Africa), Neocladophialophora from Quercus robur (Germany), Neophysalospora from Corymbia henryi (Mozambique) and Xenophaeosphaeria from Grewia sp. (Tanzania). Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

Background: Long-term exposure to air pollution has been hypothesized to elevate arterial blood pressure (BP). The existing evidence is scarce and country specific., Objectives: We investigated the cross-sectional association of long-term traffic-related air pollution with BP and prevalent hypertension in European populations., Methods: We analyzed 15 population-based cohorts, participating in the European Study of Cohorts for Air Pollution Effects (ESCAPE). We modeled residential exposure to particulate matter and nitrogen oxides with land use regression using a uniform protocol. We assessed traffic exposure with traffic indicator variables. We analyzed systolic and diastolic BP in participants medicated and nonmedicated with BP-lowering medication (BPLM) separately, adjusting for personal and area-level risk factors and environmental noise. Prevalent hypertension was defined as ≥ 140 mmHg systolic BP, or ≥ 90 mmHg diastolic BP, or intake of BPLM. We combined cohort-specific results using random-effects meta-analysis., Results: In the main meta-analysis of 113,926 participants, traffic load on major roads within 100 m of the residence was associated with increased systolic and diastolic BP in nonmedicated participants [0.35 mmHg (95% CI: 0.02, 0.68) and 0.22 mmHg (95% CI: 0.04, 0.40) per 4,000,000 vehicles × m/day, respectively]. The estimated odds ratio (OR) for prevalent hypertension was 1.05 (95% CI: 0.99, 1.11) per 4,000,000 vehicles × m/day. Modeled air pollutants and BP were not clearly associated., Conclusions: In this first comprehensive meta-analysis of European population-based cohorts, we observed a weak positive association of high residential traffic exposure with BP in nonmedicated participants, and an elevated OR for prevalent hypertension. The relationship of modeled air pollutants with BP was inconsistent.

Background: Few studies have investigated effects of air pollution on the incidence of cerebrovascular events., Objectives: We assessed the association between long-term exposure to multiple air pollutants and the incidence of stroke in European cohorts., Methods: Data from 11 cohorts were collected, and occurrence of a first stroke was evaluated. Individual air pollution exposures were predicted from land-use regression models developed within the European Study of Cohorts for Air Pollution Effects (ESCAPE). The exposures were: PM2.5 [particulate matter (PM) ≤ 2.5 μm in diameter], coarse PM (PM between 2.5 and 10 μm), PM10 (PM ≤ 10 μm), PM2.5 absorbance, nitrogen oxides, and two traffic indicators. Cohort-specific analyses were conducted using Cox proportional hazards models. Random-effects meta-analysis was used for pooled effect estimation., Results: A total of 99,446 study participants were included, 3,086 of whom developed stroke. A 5-μg/m3 increase in annual PM2.5 exposure was associated with 19% increased risk of incident stroke [hazard ratio (HR) = 1.19, 95% CI: 0.88, 1.62]. Similar findings were obtained for PM10. The results were robust to adjustment for an extensive list of cardiovascular risk factors and noise coexposure. The association with PM2.5 was apparent among those ≥ 60 years of age (HR = 1.40, 95% CI: 1.05, 1.87), among never-smokers (HR = 1.74, 95% CI: 1.06, 2.88), and among participants with PM2.5 exposure < 25 μg/m3 (HR = 1.33, 95% CI: 1.01, 1.77)., Conclusions: We found suggestive evidence of an association between fine particles and incidence of cerebrovascular events in Europe, even at lower concentrations than set by the current air quality limit value.

Novel species of microfungi described in the present study include the following from South Africa: Camarosporium aloes, Phaeococcomyces aloes and Phoma aloes from Aloe, C. psoraleae, Diaporthe psoraleae and D. psoraleae-pinnatae from Psoralea, Colletotrichum euphorbiae from Euphorbia, Coniothyrium prosopidis and Peyronellaea prosopidis from Prosopis, Diaporthe cassines from Cassine, D. diospyricola from Diospyros, Diaporthe maytenicola from Maytenus, Harknessia proteae from Protea, Neofusicoccum ursorum and N. cryptoaustrale from Eucalyptus, Ochrocladosporium adansoniae from Adansonia, Pilidium pseudoconcavum from Greyia radlkoferi, Stagonospora pseudopaludosa from Phragmites and Toxicocladosporium ficiniae from Ficinia. Several species were also described from Thailand, namely: Chaetopsina pini and C. pinicola from Pinus spp., Myrmecridium thailandicum from reed litter, Passalora pseudotithoniae from Tithonia, Pallidocercospora ventilago from Ventilago, Pyricularia bothriochloae from Bothriochloa and Sphaerulina rhododendricola from Rhododendron. Novelties from Spain include Cladophialophora multiseptata, Knufia tsunedae and Pleuroascus rectipilus from soil and Cyphellophora catalaunica from river sediments. Species from the USA include Bipolaris drechsleri from Microstegium, Calonectria blephiliae from Blephilia, Kellermania macrospora (epitype) and K. pseudoyuccigena from Yucca. Three new species are described from Mexico, namely Neophaeosphaeria agaves and K. agaves from Agave and Phytophthora ipomoeae from Ipomoea. Other African species include Calonectria mossambicensis from Eucalyptus (Mozambique), Harzia cameroonensis from an unknown creeper (Cameroon), Mastigosporella anisophylleae from Anisophyllea (Zambia) and Teratosphaeria terminaliae from Terminalia (Zimbabwe). Species from Europe include Auxarthron longisporum from forest soil (Portugal), Discosia pseudoartocreas from Tilia (Austria), Paraconiothyrium polonense and P. lycopodinum from Lycopodium (Poland) and Stachybotrys oleronensis from Iris (France). Two species of Chrysosporium are described from Antarctica, namely C. magnasporum and C. oceanitesii. Finally, Licea xanthospora is described from Australia, Hypochnicium huinayensis from Chile and Custingophora blanchettei from Uruguay. Novel genera of Ascomycetes include Neomycosphaerella from Pseudopentameris macrantha (South Africa), and Paramycosphaerella from Brachystegia sp. (Zimbabwe). Novel hyphomycete genera include Pseudocatenomycopsis from Rothmannia (Zambia), Neopseudocercospora from Terminalia (Zambia) and Neodeightoniella from Phragmites (South Africa), while Dimorphiopsis from Brachystegia (Zambia) represents a novel coelomycetous genus. Furthermore, Alanphillipsia is introduced as a new genus in the Botryosphaeriaceae with four species, A. aloes, A. aloeigena and A. aloetica from Aloe spp. and A. euphorbiae from Euphorbia sp. (South Africa). A new combination is also proposed for Brachysporium torulosum (Deightoniella black tip of banana) as Corynespora torulosa. Morphological and culture characteristics along with ITS DNA barcodes are provided for all taxa.

Septoria represents a genus of plant pathogenic fungi with a wide geographic distribution, commonly associated with leaf spots and stem cankers of a broad range of plant hosts. A major aim of this study was to resolve the phylogenetic generic limits of Septoria, Stagonospora, and other related genera such as Sphaerulina, Phaeosphaeria and Phaeoseptoria using sequences of the the partial 28S nuclear ribosomal RNA and RPB2 genes of a large set of isolates. Based on these results Septoria is shown to be a distinct genus in the Mycosphaerellaceae, which has mycosphaerella-like sexual morphs. Several septoria-like species are now accommodated in Sphaerulina, a genus previously linked to this complex. Phaeosphaeria (based on P. oryzae) is shown to be congeneric with Phaeoseptoria (based on P. papayae), which is reduced to synonymy under the former. Depazea nodorum (causal agent of nodorum blotch of cereals) and Septoria avenae (causal agent of avenae blotch of barley and rye) are placed in a new genus, Parastagonospora, which is shown to be distinct from Stagonospora (based on S. paludosa) and Phaeosphaeria. Partial nucleotide sequence data for five gene loci, ITS, LSU, EF-1α, RPB2 and Btub were generated for all of these isolates. A total of 47 clades or genera were resolved, leading to the introduction of 14 new genera, 36 new species, and 19 new combinations., Taxonomic Novelties: New genera - Acicuseptoria Quaedvlieg, Verkley & Crous, Cylindroseptoria Quaedvlieg, Verkley & Crous, Kirstenboschia Quaedvlieg, Verkley & Crous, Neoseptoria Quaedvlieg, Verkley & Crous, Neostagonospora Quaedvlieg, Verkley & Crous, Parastagonospora Quaedvlieg, Verkley & Crous, Polyphialoseptoria Quaedvlieg, R.W. Barreto, Verkley & Crous, Ruptoseptoria Quaedvlieg, Verkley & Crous, Septorioides Quaedvlieg, Verkley & Crous, Setoseptoria Quaedvlieg, Verkley & Crous, Stromatoseptoria Quaedvlieg, Verkley & Crous, Vrystaatia Quaedvlieg, W.J. Swart, Verkley & Crous, Xenobotryosphaeria Quaedvlieg, Verkley & Crous, Xenoseptoria Quaedvlieg, H.D. Shin, Verkley & Crous. New species - Acicuseptoria rumicis Quaedvlieg, Verkley & Crous, Caryophylloseptoria pseudolychnidis Quaedvlieg, H.D. Shin, Verkley & Crous, Coniothyrium sidae Quaedvlieg, Verkley, R.W. Barreto & Crous, Corynespora leucadendri Quaedvlieg, Verkley & Crous, Cylindroseptoria ceratoniae Quaedvlieg, Verkley & Crous, Cylindroseptoria pistaciae Quaedvlieg, Verkley & Crous, Kirstenboschia diospyri Quaedvlieg, Verkley & Crous, Neoseptoria caricis Quaedvlieg, Verkley & Crous, Neostagonospora caricis Quaedvlieg, Verkley & Crous, Neostagonospora elegiae Quaedvlieg, Verkley & Crous, Paraphoma dioscoreae Quaedvlieg, H.D. Shin, Verkley & Crous, Parastagonospora caricis Quaedvlieg, Verkley & Crous, Parastagonospora poae Quaedvlieg, Verkley & Crous, Phlyctema vincetoxici Quaedvlieg, Verkley & Crous, Polyphialoseptoria tabebuiae-serratifoliae Quaedvlieg, Alfenas & Crous, Polyphialoseptoria terminaliae Quaedvlieg, R.W. Barreto, Verkley & Crous, Pseudoseptoria collariana Quaedvlieg, Verkley & Crous, Pseudoseptoria obscura Quaedvlieg, Verkley & Crous, Sclerostagonospora phragmiticola Quaedvlieg, Verkley & Crous, Septoria cretae Quaedvlieg, Verkley & Crous, Septoria glycinicola Quaedvlieg, H.D. Shin, Verkley & Crous, Septoria oenanthicola Quaedvlieg, H.D. Shin, Verkley & Crous, Septoria pseudonapelli Quaedvlieg, H.D. Shin, Verkley & Crous, Setophoma chromolaenae Quaedvlieg, Verkley, R.W. Barreto & Crous, Setoseptoria phragmitis Quaedvlieg, Verkley & Crous, Sphaerulina amelanchier Quaedvlieg, Verkley & Crous, Sphaerulina pseudovirgaureae Quaedvlieg, Verkley & Crous, Sphaerulina viciae Quaedvlieg, H.D. Shin, Verkley & Crous, Stagonospora duoseptata Quaedvlieg, Verkley & Crous, Stagonospora perfecta Quaedvlieg, Verkley & Crous, Stagonospora pseudocaricis Quaedvlieg, Verkley, Gardiennet & Crous, Stagonospora pseudovitensis Quaedvlieg, Verkley & Crous, Stagonospora uniseptata Quaedvlieg, Verkley & Crous, Vrystaatia aloeicola Quaedvlieg, Verkley, W.J. Swart & Crous, Xenobotryosphaeria calamagrostidis Quaedvlieg, Verkley & Crous, Xenoseptoria neosaccardoi Quaedvlieg, H.D. Shin, Verkley & Crous. New combinations - Parastagonospora avenae (A.B. Frank) Quaedvlieg, Verkley & Crous, Parastagonospora nodorum (Berk.) Quaedvlieg, Verkley & Crous, Phaeosphaeria papayae (Speg.) Quaedvlieg, Verkley & Crous, Pseudocercospora domingensis (Petr. & Cif.) Quaedvlieg, Verkley & Crous, Ruptoseptoria unedonis (Roberge ex Desm.) Quaedvlieg, Verkley & Crous, Septorioides pini-thunbergii (S. Kaneko) Quaedvlieg, Verkley & Crous, Sphaerulina abeliceae (Hiray.) Quaedvlieg, Verkley & Crous, Sphaerulina azaleae (Voglino) Quaedvlieg, Verkley & Crous, Sphaerulina berberidis (Niessl) Quaedvlieg, Verkley & Crous, Sphaerulina betulae (Pass.) Quaedvlieg, Verkley & Crous, Sphaerulina cercidis (Fr.) Quaedvlieg, Verkley & Crous, Sphaerulina menispermi (Thüm.) Quaedvlieg, Verkley & Crous, Sphaerulina musiva (Peck) Quaedvlieg, Verkley & Crous, Sphaerulina oxyacanthae (Kunze & J.C. Schmidt) Quaedvlieg, Verkley & Crous, Sphaerulina patriniae (Miura) Quaedvlieg, Verkley & Crous, Sphaerulina populicola (Peck) Quaedvlieg, Verkley & Crous, Sphaerulina quercicola (Desm.) Quaedvlieg, Verkley & Crous, Sphaerulina rhabdoclinis (Butin) Quaedvlieg, Verkley & Crous, Stromatoseptoria castaneicola (Desm.) Quaedvlieg, Verkley & Crous. Typifications: Epitypifications - Phaeosphaeria oryzae I. Miyake, Phaeoseptoria papayae Speg.; Neotypification - Hendersonia paludosa Sacc. & Speg.

Traditionally the San people of southern Africa used Hoodia species as an appetite suppressant and various medicinal purposes (1). Hoodia gordonii (Masson) Sweet ex Decne therefore became a commercially sought-after species due to the claim of its anorectic activity. During 2004, extensive wilting was observed on H. gordonii in commercial plantings near Kakamas and Pofadder (northern Cape, South Africa). The wilting gradually increased, which caused stems to rot at the base and shrivel up, causing plants to collapse and die. Affected plants exhibited discoloration in the stems' vascular tissues. Vascular tissue excised from stems and excisions of roots were surface sterilized for 3 min in 70% ethanol followed by 3 min in 1% NaOCl, rinsed with sterile water and plated onto Van Wyks Agar, a Fusarium-selective medium (3). Isolates were grown on potato dextrose agar (PDA) and carnation leaf agar (CLA) for 14 days at 25°C. The morphological features were examined (2); identification was based on colony and sporodochia color as well as conidial morphology from single-spore colonies. The conidial morphology includes the presence or absence of macro- and microconidia and chlamydospores as well as the shape, number of septa, and basal cell of the macroconidia. The shape, characteristics, and phialides of the microconidia was also included in this analysis. To confirm pathogenicity, 18 1-year-old H. gordonii plants, 18 H. pilifera (L.f.) Plowes subsp. annulata (N.E.Br.) Bruyns plants, and 18 carnation seedlings were planted into autoclaved soil amended with 1% finely grounded oats inoculated with isolate CBS 132482 (PREM 11783), while control plants were planted in sterile soil. After 30 days, tissue was dissected from each stem, surface sterilized, rinsed, and plated on CLA and PDA for recovery of fungi. Control plants and carnations remained healthy and no fungi were recovered. All Hoodia plants displayed wilt symptoms and F. oxysporum were reisolated from the infected plants. DNA was extracted from the representative isolate (CBS 132482) and a fragment of the translation elongation factor 1-alpha (EF-1α) gene was amplified using primers EF-1/EF-2 by the polymerase chain reaction assay (4). After the isolate was sequenced and aligned, BLAST analysis of the 603-bp fragment (GenBank Accession No. JX003858) showed a 100% homology with F. oxysporum (GenBank Accession No. GU226828). The beta tubulin gene sequenced (GenBank Accession No. JX003859) was amplified using the primers Bt-2a/Bt-2b. BLAST searches with the resulting 311-bp fragment showed a 99.4% homology with several isolates of F. oxysporum in the GenBank database (JQ265753; FR828825; DQ092480). The fungus had a specific host preference because it did not infect carnations as well as previously tested plants, which included beans, pumpkin, tomato, and watermelon. To our knowledge, this is the first report of F. oxysporum causing wilt in H. gordonii in South Africa. References: (1) B. Hargreaves and Q. Turner. Askelpios 86:11, 2002. (2) J. F. Leslie and B. A. Summerell. Page 369 in: The Fusarium Laboratory Manual, Blackwell Professional, Ames, IA, 2006. (3) P. S. van Wyk et al. Phytophylactica 18:67, 1986. (4) P. Vos et al. Nucleic Acids Res. 23:4407, 1995.

Novel species of microfungi described in the present study include the following from Australia: Catenulostroma corymbiae from Corymbia, Devriesia stirlingiae from Stirlingia, Penidiella carpentariae from Carpentaria, Phaeococcomyces eucalypti from Eucalyptus, Phialophora livistonae from Livistona, Phyllosticta aristolochiicola from Aristolochia, Clitopilus austroprunulus on sclerophyll forest litter of Eucalyptus regnans and Toxicocladosporium posoqueriae from Posoqueria. Several species are also described from South Africa, namely: Ceramothyrium podocarpi from Podocarpus, Cercospora chrysanthemoides from Chrysanthemoides, Devriesia shakazului from Aloe, Penidiella drakensbergensis from Protea, Strelitziana cliviae from Clivia and Zasmidium syzygii from Syzygium. Other species include Bipolaris microstegii from Microstegium and Synchaetomella acerina from Acer (USA), Brunneiapiospora austropalmicola from Rhopalostylis (New Zealand), Calonectria pentaseptata from Eucalyptus and Macadamia (Vietnam), Ceramothyrium melastoma from Melastoma (Indonesia), Collembolispora aristata from stream foam (Czech Republic), Devriesia imbrexigena from glazed decorative tiles (Portugal), Microcyclospora rhoicola from Rhus (Canada), Seiridium phylicae from Phylica (Tristan de Cunha, Inaccessible Island), Passalora lobeliae-fistulosis from Lobelia (Brazil) and Zymoseptoria verkleyi from Poa (The Netherlands). Valsalnicola represents a new ascomycete genus from Alnus (Austria) and Parapenidiella a new hyphomycete genus from Eucalyptus (Australia). Morphological and culture characteristics along with ITS DNA barcodes are also provided.

Kenaf (Malvaceae; Hibiscus cannabinus L.) is being commercially cultivated in Winterton, South Africa for its high-quality cellulose fibers with approximately 2,000 ha currently under cultivation. In 2004, 25% of 1-month-old kenaf plants grown from seed were observed in the field with severe wilting followed by lodging and mortality within 1 week. Isolations from diseased stem and root tissue on malt extract agar (MEA) consistently yielded Fusarium verticillioides (Sacc.) Nirenberg (2). Pathogenicity tests were conducted by inoculating kenaf seedlings with inoculum prepared from barley grains that had been colonized by the pathogen in vitro for 2 weeks prior to being finely ground in a laboratory mill. Fifty seeds from each of eight kenaf cultivars were incubated at 25°C on sterile filter paper to ensure germination and the absence of pathogens. Germinated seeds were sown in pots (400 cm 3 ) containing steam sterilized loam soil (200 g) by placing 20 germinated seeds from each cultivar, with four replicates (5 seeds per pot), on the soil in each pot and covering them with 100 g of the same soil. Inoculum powder was sprinkled on the surface of the soil in each pot and covered by 100 g of soil. Pots were maintained in a glasshouse at an ambient temperature of 25°C. Sterile ground barley seeds served as the control treatment. Pots were watered daily with 20 ml of water and observed periodically for seedling emergence. The percentage of diseased seedlings was recorded after 3 weeks and the experiment was repeated. Wilting had occurred in 85% of seedlings when they were approximately 4 cm high and all diseased seedlings had died within 1 week thereafter. Subsequent examination revealed dark brown lesions girdling the stem and decayed roots in all instances. No symptoms developed on control plants. From means of combined data, the greatest seedling mortality was observed for cv. Gregg (65%) and the least for cv. Cuba108 (5%). Mean mortalities for the remaining six cultivars ranged from 30 to 55%. The pathogen was reisolated on MEA from all diseased seedlings. To our knowledge, this is the first report of F. verticillioides occurring on kenaf in South Africa. The only other report of Fusarium sp. causing serious damping-off of kenaf is from Iran (1). The potential impact of the pathogen on kenaf production in South Africa must be considered in the implementation of disease control measures. References: (1) J. M. Dempsey. Kenaf. In: Fiber Crops. The University Press of Florida, Gainesville, 1975. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006.

The cashew plant (Anacardium occidentale L.) (family Anacardiaceae) is native to Brazil. It was introduced in East Africa by the Portuguese in the 16th century where it is now widely cultivated, especially in Tanzania, Kenya, and Mozambique. The processed kernels are the most important product derived from the plant, although in Brazil and India, juices, jam, and alcoholic and soft drinks are also made from the pear-shaped edible receptacle. The plant is currently being evaluated in South Africa for commercial production. During May 2002, at least 25% of 5-year-old cashew trees grown from seed in the northern KwaZulu-Natal Province of South Africa were infected with powdery mildew. Signs included extensive growth of white, superficial mycelium bearing upright conidiophores on young shoots with tender leaves, inflorescences, and young receptacles. In severely affected trees, approximately 35% of young shoots and 45% of young receptacles displayed signs of powdery mildew. Severely infected young leaves were brown and deformed in contrast to older leaves that were unaffected. Microscopic examination of diseased tissue revealed hyaline, cylindrical-to-slightly doliform, single-celled conidia (10 to 17.5 × 2.5 to 5 μm) borne in chains. The pathogen was subsequently identified as Oidium anacardii Noack on the basis of morphology (1). No other species of powdery mildew fungi have been reported on cashew. A pathogenicity test was conducted by gently pressing a heavily diseased leaf onto two healthy leaves of each of 10 cashew plants maintained in pots on open benches in the glasshouse at 22 to 25°C and mean relative humidity of 65%. Control treatments entailed pressing an asymptomatic leaf onto each of two healthy leaves per plant. The experiment was conducted three times. After 14 days, at least one powdery mildew colony had developed on 80% of inoculated leaves but were absent from all replications of the control treatment. The source of inoculum for this reported outbreak is unknown, although O. anacardii is known to occur in southern Mozambique less than 100 km from the infected site. Cashew powdery mildew was first officially reported in Tanzania in 1979 where significant crop losses, partially attributable to the pathogen, have been recorded since (3). No significant damage to production has been recorded in Brazil (2). To our knowledge, this is the first report of O. anacardii occurring on cashew in South Africa. References: (1) E. Castellani and F. Casulli. Rivista di Agricoltura Subtropicale e Tropicale 75:211, 1981. (2) F. C. O. Freire et al. Crop Prot. 21:489, 2002. (3) P. J. Martin et al. Crop Prot. 16:5, 1996.

Discoloration, cankers, and decay in branches, stems, and root collars of Amaranthus hybridus were observed in Bloemfontein, South Africa. Examination of symptomatic stems revealed larval galleries of the pigweed weevil (Hypolixus haerens). The objectives of this study were to: identify the most common fungal species associated with this damage, determine if the adult pigweed weevil might be a vector for the fungi, and test if the associated fungi can cause the stem canker disease observed in the field. The most common fungal species isolated were Fusarium subglutinans from discolored tissues adjacent to insect galleries (42%), F. subglutinans from weevil larvae (29%), the Alternaria tenuissima group from adult weevils (31%), and the A. tenuissima group from cankered stems (40%). Three of the seven most common fungal species produced cankers following wounding and inoculation, with F. sambucinum and F. oxysporum being the most aggressive. Although fungal species compositions differed (P < 0.01) among the four tissue/insect stage combinations tested, all four had the same major fungal species, suggesting the pigweed weevil as a vector for the Fusarium pathogens. There is significant potential for yield loss affiliated with this insect-fungal association. The identification of this insect-fungal relationship and the pathogens involved in disease set the stage for further research on the etiology and disease management of this important insect-fungal relationship.

Kenaf, Hibiscus cannabinus L. (Malvaceae), is being planted commercially in South Africa for the high quality cellulose fibers that it produces. In a January 2001 survey of 3-month-old kenaf plants grown from seed in experimental plots near Rustenburg, Northwest Province, 30% of plants were observed with severe wilting. Stems at ground level of all infected plants had sunken tan lesions, white mycelial strands, and small, dark brown, 1 to 2 mm diameter sclerotia. Isolations from diseased stem tissue on malt extract agar (MEA) consistently yielded a fungus conforming to the description of Sclerotium rolfsii Sacc. (teleomorph Athelia rolfsii (Curzi) Tu & Kimbrough). Pathogenicity tests were conducted by applying toothpick tips (5 mm) colonized by S. rolfsii on MEA to the stems of 120-day-old potted plants of 10 kenaf cultivars in the greenhouse. Five plants of each cultivar were wounded once using a sharp dissecting needle, and a colonized toothpick tip was placed on top of each wound. Control treatments consisted of five plants per cultivar each wounded and inoculated with sterile toothpick tips. All inoculation points were wrapped using Parafilm, and the experiment was conducted twice. Lesions were measured after 10 days. Mean lesion lengths for the 10 cultivars were as follows: Dowling (34.9 mm), Cuba 108 (38.6 mm), Gregg (41.1 mm), Everglades 41 (44.2 mm), SF459 (44.9 mm), Tainung 2 (45.8 mm), El Salvador (45.9 mm), Whitton (46.1 mm), Everglades 71 (46.4 mm), and Endora (54.0 mm). The Newman-Keuls multiple comparison test revealed that cvs. Dowling and Endora were significantly more resistant and more susceptible (P < 0.05), respectively, than the other cultivars. Lesions did not develop on control plants. The fungus was reisolated on MEA from all artificially inoculated plants. The pathogen is reported to cause serious losses in yield and fiber quality of kenaf (1). To our knowledge, this is the first report of S. rolfsii on kenaf in South Africa. Commercial plantings of kenaf in South Africa are expected to exceed 500 ha during the next 2 years, so its potential impact on kenaf production in this country will be significant if efficient disease control measures are not practiced. References: (1) J. M. Dempsey. Kenaf. Pages 203-304 in: Fiber Crops. The University Press of Florida, Gainesville, 1975.

With the increased use of Amaranthus hybridus as a leafy-vegetable crop in Africa and the recent identification of Alternaria leaf spot on this host in southern Africa, the role of this potentially damaging pathogen was investigated. The goals of this study were to test the pathogenicity of the Alternaria tenuissima group, determine how these fungi infect Amaranthus hybridus leaves, and examine the colonization pattern within host tissues. Asymptomatic leaves of Amaranthus hybridus were collected from two field sites in South Africa. Eight A. tenuissima group isolates collected from these leaves were used in inoculation experiments conducted in both greenhouse and growth chamber studies. Scanning electron microscopy revealed A. tenuissima-like conidia germinating on leaf surfaces and mycelia entering leaves only through stomata of both field-collected and artificially inoculated leaves. Unwounded, inoculated leaves had no symptoms, and light-microscopy observations of both asymptomatic field-collected and unwounded and inoculated leaves revealed hyphae in mesophyll tissue growing intercellularly with no host cell penetration or host-cell response. Seven of the eight isolates produced brown to black, circular to oval, necrotic lesions only at the wound site of injured and inoculated leaves. These results confirm that isolates of the A. tenuissima group can infect and colonize Amaranthus hybridus leaves in a manner consistent with other endophytic fungi, and suggest that these fungi can act as latent leaf pathogens when the host is altered by wounding.

The commercial cultivation of spineless cactus (Opuntia ficus-indica (L.) Miller) for its fruit is a relatively recent undertaking in South Africa but has been shown to possess huge export potential. To date, only one fungal pathogen, Didymosphaeria opulenta (De Not.) Sacc., has been officially reported on the genus Opuntia in South Africa, but the report is from O. stricta Haw. and not O. ficus-indica (1). The need for research on diseases of O. ficus-indica in South Africa has recently become important since local growers are increasingly reporting disease-related yield losses. Surveys conducted over a period of 3 years indicated that stems or cladodes are particularly prone to various forms of tissue necrosis, caused primarily by three fungi, which can ultimately lead to death of entire cladodes. Alternaria tenuissima was isolated from a dry superficial necrosis of the cuticle and underlying tissue as much as 3 mm deep. Symptoms include small chlorotic spots on the cuticle, which coalesce to form raised gray scabs. Fusarium sporotrichoides was isolated more commonly from dry necrotic lesions that were darker, larger, and less superficial, sometimes extending through the tissue to the opposite side of the cladode. Lasiodiplodia theobromae (teleomorph Botryosphaeria rhodina) was isolated from roundish black cankers (15 to 50 mm diameter) on cladodes and characterized by black gum exudation from the perimeter of the canker. Pycnidia were often evident on the surface of the canker. The fulfillment of Koch's postulates demonstrated that an isolate of each respective species was very aggressive in colonizing cladodes following artificial inoculations in the glasshouse. Mean lesion diameters measuring 15, 27, and 44 mm for A. tenuissima, F. sporotrichoides, and L. theobromae, respectively, were recorded 14 days after inserting wooden toothpick tips that had been colonized by the three pathogens into each of five cladodes of 18-month-old potted plants of O. ficus-indica (cv. Morado). Alternaria sp. and B. rhodina have been reported on Opuntia sp. in the United States (2), but no records of the above three fungi occurring on O. ficus-indica were found. References: (1) P. W. Crous et al. Phytopathogenic Fungi from South Africa. University of Stellenbosch, Department of Plant Pathology Press, Stellenbosch, South Africa, 2000. (2) D. F. Farr et al. Fungi on Plants and Plant Products in the United States. The American Phytopathological Society, St. Paul, MN, 1989.

Kenaf (Hibiscus cannabinus L. Malvaceae) presents a source of high-quality cellulose fibers and is being investigated in South Africa for commercial production. In March 2001, 12 5-month-old kenaf plants grown from seed in experimental plots near Bloemfontein, South Africa, displayed large, black, sunken lesions (10 to 20 cm long) at the base of the stem, and severe root rot. A study was undertaken to characterize the pathogen, and to determine the relative susceptibilities of five kenaf genotypes being considered for commercial cultivation. Isolations from diseased tissue on malt extract agar consistently yielded a fungus identified as Pythium group G (1). Four-month-old kenaf plants were artificially inoculated in the field by inserting wooden toothpick tips colonized by the pathogen approximately 25 cm above soil level into the stems of 10 plants of each of five genotypes. Inoculation points were wrapped using Parafilm. The fungus was highly virulent to all five kenaf genotypes in two experiments, with mean cambial lesion lengths of 117, 119, 120, 122, and 139 mm at 7 days after inoculation for Tainung-2, Cuba 108, SF-459, El Salvador, and Everglades 41, respectively. Lesions ranged from 44 to 164 mm, with an overall mean of 124 mm for all five genotypes. No lesions developed in control plants. Although Everglades had the longest lesions, there were no significant differences (P < 0.05) among genotypes. Koch's postulates were completed by reisolating the fungus from all inoculated plants. To our knowledge, there are no published reports of Pythium group G causing stem or root rot of kenaf. References: (1) M. W. Dick. Keys to Pythium. University of Reading Press, Reading, UK, 1990.

Thirty isolates of Fusarium oxysporum were recovered from six substrates: (i) tap roots of Amaranthus hybridus showing symptoms of root rot, (ii) side roots of A. hybridus showing symptoms of root rot, (iii) soil surrounding plants of A. hybridus, (iv) pigweed weevils (Hypolixus haerens) that feed on A. hybridus, (v) rhizosphere of maize plants growing adjacent to a field of amaranth, and (vi) rhizosphere of dry bean plants growing adjacent to a field of amaranth. The isolates were characterized by means of pathogenicity tests, isozyme analysis, and vegetative compatibility group (VCG) tests. In the pathogenicity tests, toothpick tips were infested with F. oxysporum and inserted into amaranth stems. All 30 isolates were pathogenic on A. hybridus, with significant differences in pathogenicity based on lesion length measured 4 weeks after inoculation. The isolates were grouped into nine VCGs by complementation tests using nitrate nonutilizing mutants. Self-incompatibility was not observed for any of the isolates. The most common VCG was VCG1, which comprised 20 of the 30 isolates tested. The second most common group was VCG3, which included three isolates, while the remaining seven VCGs each consisted of a single isolate. The results indicate that the population of the amaranth root rot pathogen examined in this study is relatively homogeneous. Results of the isozyme analysis supported the results of VCG tests. Three major groups were delineated within the 30 isolates of F. oxysporum following cluster analysis of electrophoretic phenotypic values for seven isozymes (EST, IDH, G6PDH, ACP, PEP1, PEP2, and PEP3) tested. No relationship was found between isozyme phenotype and the substrate from which the isolates were recovered.

Kenaf (Hibiscus cannabinus L.) (Malvaceae) is a source of high-quality cellulose fibers and is being investigated in South Africa with a view to commercial production. In April 2001, 20 to 30% of 5-month-old kenaf plants grown from seed in experimental plots near Rustenburg, Northwest Province, South Africa, were affected by gray mold caused by Botrytis cinerea Pers.:Fr. Infected plants displayed brown necrotic areas that girdled the stem, resulting in wilting and lodging in at least 50% of observed cases. Symptoms included extensive growth of mycelia and gray conidia on stem lesions. Microscopic examination revealed hyaline, one-celled conidia and conidiophores conforming to the description of B. cinerea. Plating of diseased stem tissue on malt extract agar (MEA) consistently yielded B. cinerea. Koch's postulates were satisfied by applying toothpick tips (5 mm) colonized by B. cinerea on MEA to the stems of 10 120-day-old greenhouse-grown plants of each of five kenaf cultivars. A colonized toothpick tip was placed on the stem of each of five plants per cultivar at a point ≍15 cm above soil level. Another five plants of each cultivar were wounded once using a sharp dissecting needle, and a colonized toothpick tip was placed on top of each wound. Corresponding control treatments consisted of five additional plants per cultivar, each wounded and mock-inoculated with sterile toothpick tips. Inoculation points were wrapped in Parafilm. The experiment was conducted twice. Developing lesions were measured after 7 days. Mean lesion lengths for the two treatments, nonwounded and wounded, on the five cultivars were, respectively: 32.4 and 35.2 mm for Everglades 41; 14.9 and 53.8 mm for Cuba 108; 39.5 and 55.8 mm for El Salvador; 19.0 and 44.3 mm for SF459; and 12.4 and 43.9 mm for Tainung 2. The Newman-Keuls multiple comparison test revealed no significant difference (P < 0.05) in means among cultivars for the wounded treatment. For the nonwounded treatment, Everglades 41 and El Salvador were significantly more susceptible (P < 0.05) than the three remaining cultivars. No lesions developed on control treatments. The fungus was reisolated on MEA from all artificially inoculated plants. The pathogen is reported to cause serious losses in yield and fiber quality of kenaf in Spain (1). This is the first report of B. cinerea on kenaf in South Africa, and its potential impact on kenaf production in this country should be taken seriously. Reference: (1) A. De Cal and P. Melgarejo. Plant Dis. 76:539, 1992.

Kenaf (Hibiscus cannabinus L.) is a fast-growing, bamboo-like annual plant belonging to the Malvaceae. The stem, which ranges from 1.5 to 4 m, presents a source of high-quality cellulose fibers. The plant is being investigated in South Africa with a view to commercial production. In April 2001, at least 50% of 4- to 5-month-old kenaf plants grown from seed in trials near Rustenburg, Northwest Province, South Africa, were observed as having powdery mildew. Signs included extensive growth of white, superficial mycelium and emergent conidiophores on the abaxial leaf surface, followed by partial defoliation. On older leaves, the abaxial leaf surface was completely covered by powdery mildew, and chlorotic and necrotic patches were clearly visible on the adaxial surface. Symptoms were observed on all five planted cultivars (Everglades 41, Cuba 108, El Salvador, SF459, and Tainung 2), and no difference in disease severity was noted among cultivars. Leveillula taurica (Lév.) Arnaud (anamorph Oidiopsis taurica [Lév.] Salmon) was subsequently identified by the presence of endophytic mycelia, often branched conidiophores, and dimorphic conidia borne singly or in short chains (1). In 100 measurements of each type, pyriform conidia averaged 69 ± 5 × 18 ± 2 μm and cylindrical conidia averaged 62 ± 6 × 16 ± 2 μm. The teleomorph was not observed. The source of L. taurica for this reported outbreak is unknown, and powdery mildew was not observed in a field of mature cotton (Gossypium hirsutum L.) growing within 10 m of the kenaf plot. L. taurica was reported on kenaf in Texas in 1992 (2) and in Italy in 1995 (3). The pathogen can cause significant losses in seed yield and reduce seed quality in susceptible kenaf cultivars. Although L. taurica has been reported from Hibiscus sabdariffa in Egypt (4), to our knowledge this is the first report of the pathogen occurring on kenaf in Africa. References: (1) H. J. Boesewinkel. Bot Rev. 46:167, 1980. (2) C. G. Cook and J. L. Riggs. Plant Dis. 79:968, 1995. (3) S. Frisullo et al. Inf. Fitopatol. 45:37-41, 1995. (4) M. Khairy, et al. Phytopathol. Medit. 10:269-271, 1971.

Pigeonpea (Cajanus cajan [L.] Mills.) is an important legume with potential as a dryland crop with multiple uses in the semi-arid areas of South Africa. Approximately 150 tons of dry, split seeds are imported monthly to meet the needs of South Africa. In May 2000, field trials and farmer's plots with plant ages varying from 1 to 3 years old were visited in Mpumalanga and Kwazulu-Natal to assess problems associated with pigeonpea cultivation. Rust was prevalent on more than 80% of plants on young and old leaves at all sites but was most severe at sites in Mpumalanga, where severe rust was observed on all 17 ICRISAT varieties evaluated. Leaf lesions began as chlorotic flecks that expanded and developed into necrotic spots with several orange red to brown uredinia present mostly on the abaxial sides of leaves. Urediospores were 1-celled and initially hyaline, turning dark orange, minutely echinulate, spherical with 2 to 4 circular germpores and measured 20-27 × 17 to 21μ. No telia were found and all morphological characteristics therefore correspond with the CMI description of Uredo cajani Syd. (1). In Africa, pigeonpea rust has been reported from Kenya, Nigeria, Sierra Leone, Tanzania, and Uganda. In South Africa, rust, described as Uromyces dolicholi Arthur (2), has only once been reported on pigeonpea. In the United States, U. dolicholi has also once been reported on pigeonpea (3). However, since U. dolicholi, unlike U. cajani, produces telia and occurs only on Rhyncosia spp. (4), these reports can be considered incorrect. This is therefore the first report of U. cajani on pigeonpea in South Africa. References: (1) K. H. Anahosur and J. M. Waller. 1978. No. 590: Descriptions of Plant Pathogenic Fungi and Bacteria. Commonw. Mycol. Inst., Kew, England. (2) E. M. Doidge. Bothalia 5:1-1094, 1950. (3) D. F. Farr et al. 1989. Fungi on Plants and Plant Products in the United States. American Phytopathological Society, St. Paul, MN, 1989. (4) A. Sivanesan. 1970. No. 269: Descriptions of Plant Pathogenic Fungi and Bacteria. Commonw. Mycol. Inst., Kew, England.

Cultivation of English walnut (Juglans regia L.) has considerable economic potential in South Africa. In February 1999, die-back of walnut trees was observed on ≈30% of trees in an orchard in Gauteng Province, South Africa. Die-back was associated with characteristic cankers and discoloration of the cambial tissue of twigs and young branches. Fifty pieces of discolored tissue from the margins of cankers of five trees were transferred to potato dextrose agar (PDA) after surface-sterilization with 0.5% NaOCl. Fungal colonies emerged from all 50 pieces of diseased tissue. Fungi recovered included Fusarium solani (74.5%), Alternaria tenuissima (17.3%), and a Phoma sp. (6.2%). Small pieces of sterile cheesecloth (10 × 5 mm) were cultured on PDA with an isolate of F. solani from infected walnut trees. Colonized pieces of cheesecloth were applied to stems (≈5-mm diameter) of nine potted J. regia plants in a glasshouse. Each stem was wounded by lightly scraping off a length of bark (5 × 5 mm). Colonized cheesecloth was wrapped around the wound and covered with Parafilm to prevent desiccation of mycelia in the cheesecloth. Nine walnut stems were similarly treated with sterile cheesecloth pieces to serve as control treatments. After 4 weeks, die-back of foliage was observed in all plants artificially inoculated with F. solani. Cheesecloth pieces and surrounding bark were removed, and the length and width of each cambial lesion was measured. The mean area of cambial lesions (length by width) resulting from artificial inoculations was 27.3 × 8.5 mm. None of the control plants developed any symptoms. Koch's postulates were confirmed by consistently reisolating F. solani from inoculated plants. A similar disease of black walnut (J. nigra) was reported by Carlson et al. (1) from five central states in the United States. Our findings represent the first report of F. solani as a causal agent of stem canker of English walnut in South Africa. Reference: (1) J. C. Carlson et al. N. J. Appl. For. 10:1, 1993.

Amaranthus hybridus has been identified as an important alternative vegetable crop with potential for increased commercial production in South Africa (1). In summer 1999, severe losses occurred in a large plot of 2-month-old A. hybridus plants on an experimental farm near Bloemfontein, South Africa. More than 90% of the plants were severely stunted, with chlorotic foliage that was wilted in most cases. Root rot was present in all symptomatic plants and was clearly visible as an amber to brown discoloration of tap and secondary roots; in severe cases, white mycelium was clearly visible on diseased root tissue. Isolations from symptomatic roots were made on potato dextrose agar (PDA) amended with streptomycin sulfate. Isolates (N = 121) were recovered from diseased roots (n = 89). The two most frequently isolated fungi were transferred to carnation leaf agar and identified as Fusarium oxysporum (n = 90, 74%) and F. sambucinum (n = 29, 24%). Pathogenicity tests with one isolate of each species were performed in the greenhouse on 1-month-old potted A. hybridus seedlings (10 plants per treatment). A single hyphal tip of each isolate was transferred to PDA and incubated at 25°C for 7 days in the dark. Five 4-mm-diameter mycelial plugs were taken and placed directly on the taproot of each plant, halfway along the length and ≈30 mm below the soil surface. Control plants were treated with uncolonized PDA plugs. Seedlings inoculated with either fungus exhibited wilting within 7 days; stunting, chlorosis (pale green to yellow), and root necrosis after 2 weeks; and mortality after 4 weeks. Inoculated plants were removed from pots after 3 weeks, roots were washed free of potting soil, and necrotic lesion length was measured. Necrosis and discoloration of root tissue were similar to those observed in field plants. The mean length of tissue necrosis induced by the fungi was 22.5 and 34.8 mm for F. oxysporum and F. sambucinum, respectively. F. sambucinum, thus, was more pathogenic than F. oxysporum despite being recovered significantly less often from field plants. Control plants inoculated with noninfested PDA plugs remained healthy. The presence of both pathogens was confirmed by reisolation from artificially inoculated taproots of all plants. No Fusarium spp. were recovered from the 10 control treatments. F. oxysporum has been reported on diseased red root pigweed (A. retroflexus) in the United States (2), but this is the first report of both F. oxysporum and F. sambucinum as causal agents of root rot in A. hybridus. These pathogens, therefore, must be considered a potential threat to commercial production of A. hybridus in South Africa and elsewhere. References: (1) W. J. Swart et al. S. Afr. J. Sci. 93:22, 1997. (2) R. M. Harveson and C. M. Rush. Plant Dis. 81:85, 1997.

Amaranthus hybridus is an important alternative leafy-vegetable crop with the potential for increased commercial production in southern Africa and other semi-arid regions of the world (2). In May 1998, extensive leaf spotting was observed on A. hybridus at Potchefstroom, South Africa. Many of the leaves had symptoms and most were severe. Symptoms were dark brown to black, circular to oval, necrotic lesions with a diameter ranging from <1 mm up to 7 mm. Larger lesions had tan centers. Tissues adjacent to the leaf spots remained green. Alternaria tenuissima was isolated from 43% of 40 lesions sampled making up 89% of the isolates recovered. A. tenuissima was isolated from asymptomatic leaves of 5-month-old A. hybridus plants sampled from the same site in April 1997 (1). The foliar symptoms observed on A. hybridus in Potchefstroom were reproduced by inoculating wounded leaves of A. hybridus with single-spore A. tenuissima isolates obtained from asymptomatic leaves collected at Potchefstroom. Eight isolates were selected for pathogenicity tests conducted in a growth chamber. A. hybridus leaves grown from seed in a greenhouse were placed in moist chambers and wounded with a needle (0.5 mm) at leaf center; one 5-mm-diameter, colonized potato dextrose agar plug of each of the isolates was placed on the center of each leaf. A sterile plug was used as a control. Moist chambers were placed in a growth chamber set at 25°C day and 20°C night and provided artificial light for 16 h per day. In a greenhouse (average temperature 25°C day, 17°C night), a conidial suspension (10 5 conidia per ml of sterile, distilled water) was applied to an individual leaf of each of 14 plants with an atomizer. Sterile, distilled water was applied to control leaves. Leaves were then wounded with a needle (0.5 mm) at leaf center. Treatments were assigned randomly and the experiments were repeated. Symptoms were first observed at 14 and 18 days (growth chamber and greenhouse, respectively). Seven of the eight isolates caused necrotic lesions with an average diameter of 3 mm (both growth chamber and greenhouse). Symptoms were observed on an average of 56 and 82% of the inoculated leaves (growth chamber and greenhouse, respectively). The range of symptoms was the same as that observed in the field, but symptoms were only observed at the wounds. Controls remained green and showed no symptoms. A. tenuissima was recovered from necrotic lesions of surface-disinfested, inoculated leaves (average 93 and 68%; growth chamber and greenhouse, respectively), never recovered from growth chamber controls, and seldom recovered from greenhouse controls (5%). These results suggest that A. tenuissima is a leaf-spot pathogen of A. hybridus and wounding might trigger disease expression. A minimal amount of leaf spotting of this leafy-vegetable crop can cause total crop loss. References: (1) J. T. Blodgett et al. S. Afr. J. Sci. 94:xviii, 1998. (2) W. J. Swart et al. S. Afr. J. Sci. 93:xxii, 1997.

Amaranthus hybridus (common name: amaranth) is a fast-growing crop with nutritious leaves and seeds that is cultivated in semi-arid regions throughout the world. In South Africa, cultivation of this crop as a leafy vegetable is increasing. In autumn 1997, extensive tissue discoloration and decay were observed in branches, stems, and root collars of mature A. hybridus in Bloemfontein, Free State Province. Symptoms included discolored phloem, xylem, and pith, black cankers, and weakened stems prone to wind breakage. Examination of these tissues revealed larval galleries of the pigweed weevil (Hypolixus haerens), the main insect pest of A. hybridus in South Africa (1). Six-month-old A. hybridus stems were split and small samples of discolored tissue adjacent to the larval galleries of each stem and the associated larvae were placed aseptically on corn-meal agar containing streptomycin and incubated for 4 to 7 days. The seven fungi most frequently isolated from discolored stem tissues (n = 166) were Fusarium subglutinans (46%), a Phomopsis sp. (11%), Alternaria alternata (10%), F. oxysporum (9%), F. solani (5%), a Phoma sp. (5%), and F. sambucinum (4%). The nine fungi most frequently isolated from larvae (n = 90) were F. subglutinans (46%), F. solani (8%), F. equiseti (8%), F. oxysporum (7%), A. alternata (6%), a Phomopsis sp. (4%), F. proliferatum (3%), F. sambucinum (2%), and a Phoma sp. (2%). Stems of greenhouse-grown A. hybridus were inoculated with the seven most common species isolated from the discolored stem tissues. One isolate of each species was used. Inoculations involved wounding stems by removing approximately 36 mm 2 of the epidermis 5 cm above the soil, placing a colonized water agar plug on the wound, and wrapping Parafilm around the stems at the wound site. Wounded and nonwounded (untreated) controls were also included. A noncolonized water agar plug was applied to wounded controls but not to nonwounded controls. Ten plants per isolate and 10 wounded and nonwounded control plants were used in each of two separate trials (180 total plants). Treatments were assigned randomly. Four weeks after inoculation, canker lengths were measured and stem sections were surface disinfected and transferred to water agar plates. The presence of the fungi was confirmed after 20 days. Only F. sambucinum, F. oxysporum, and F. subglutinans caused cankers with frequencies of 100, 100, and 65% (n = 20), and mean lesion lengths of 30, 26, and 10 mm, respectively. Lesions were never observed on either of the controls. Discoloration and cankers were similar to that observed in the field. F. sambucinum, F. oxysporum, and F. subglutinans were recovered from 65, 50, and 60% of the tissues, respectively, and none of the Fusarium spp. were recovered from the control treatments (n = 20 for all). In artificial inoculations, these species can act as pathogens independent of the pigweed weevil and are likely the cause of the discoloration, decay, and cankers observed in branches, stems, and root collars of mature A. hybridus. However, there are no prior reports of a Fusarium sp. causing disease on A. hybridus, and H. haerens larvae were observed in all symptomatic stems in the field. Further studies are needed to determine the potential for significant disease loss associated with this insect-fungal association and the potential role of these fungi in further weakening Amaranthus stems that are colonized by H. haerens. Reference: (1) S. vdM. Louw et al. Afr. Crop Sci. J. 3:93, 1995.

Pistachio (Pistacia vera) cultivation is a relatively new industry in South Africa with tremendous economic potential. However, disease problems could develop in time. The pathogens of pistachio found in other parts of the world plausibly will spread to South Africa and new pathogens never recorded on this host may develop here. In the summer of 1998, 2-year-old rootstocks of P. atlantica and P. integerrima displaying basal cankers and discolored phloem and xylem were observed in the Prieska district of the Northern Cape province. The objective of this study was to identify the causal agent(s). Cankered stems were split and small samples of discolored wood were aseptically placed on corn-meal agar containing streptomycin and incubated for 5 days. Fungi that grew into the agar were transferred to 1.5% water agar dishes containing pine needles to aid sporulation. Species isolated from stems included Fusarium spp. (44%), Fusicoccum aesculi (anamorph of Botryosphaeria dothidea) (23%), a Cytospora sp. (19%), a Chaetomium sp. (3%), and several nonsporulating mycelial fungi (11%). Inoculum of suspected pathogens (F. aesculi, the Cytospora sp., and two Fusarium spp.) were prepared by culturing fungi in petri dishes on potato dextrose agar overlaid with sterile cheesecloth strips (approximately 15 × 25 mm) until the strips were completely colonized. Greenhouse inoculations involved wounding P. atlantica stems by removing the bark (approximately 3 × 6 mm) approximately 10 cm above the soil, and wrapping the colonized cheesecloth followed by Parafilm around the stems at the wound site. Eight plants per isolate and eight wounded and nonwounded (untreated) control plants were used. Noncolonized cheesecloth was applied to wounded controls but not to nonwounded controls. Treatments were assigned randomly. Eight weeks after inoculation, the surrounding bark was removed from all treated shoots and the cambium was examined for discoloration. The length of each cambial lesion was measured and stem sections were surface disinfested and transferred to 1.5% water agar dishes, and the presence of the inoculated fungi was confirmed. Only F. aesculi was pathogenic to P. atlantica and produced cankers on all stems. The fungus was recovered from all tissues sampled 3 cm beyond the wound sites, 88% of those sampled from the wound sites, and none of the control treatments. Discoloration of the phloem and xylem was similar to that observed in the field. The mean canker length was 41 mm on trees with a mean stem diameter of 7 mm. The appearance of B. dothidea, in addition to B. obtusa (1), on pistachio justifies the need for establishing a disease management program for pistachio in South Africa. Reference: (1) W. J. Swart and W.-M. Botes. Plant Dis. 79:1036, 1995.

A mutant of phage T5 which is unable to induce thymidylate synthetase was isolated. T5 thy mutants synthesized less DNA than did wild-type T5, and the burst size of progeny phage was correspondingly reduced two- to threefold in thy+ Escherichia coli. No DNA or progeny phage were made in E. coli thy hosts grown in the absence of exogenous thymine. When the T5 thy mutation was recombined with a T5 dut mutation (unable to induce dUTPase), replication resulted in progeny which contained significant amounts of uracil in their DNA, and these phage failed to produce plaques unless the plating host was deficient in uracil-DNA glycosylase. T5 phage containing various amounts of uracil in their DNA were prepared and used to determine the effect of uracil on the induction of the early enzyme dTMP kinase. The presence of uracil in the parental DNA increased the rate of induction of this enzyme by about 2.5-fold. The T5 thy gene was mapped and is located near the T5 frd gene on the B region of the T5 genome.

Cultures of starch-elicited peritoneal mouse macrophages in medium containing macrophage growth factor (MGF) were infected with lactate dehydrogenase-elevating virus (LDV) and, after various times in culture, LDV production was monitored as a function of time by infectivity titrations in mice, by measuring [3H]uridine incorporation into LDV RNA and extracellular LDV, by autoradiographic analysis of the proportion of productively infected cells and by electron microscopy. Regardless of the age of the cultures when infected with LDV, only a small proportion of the macrophages (generally between 3 and 20% of the total) became productively infected after a primary infection; maximum virus RNA synthesis and virus production occurred during the first 24 h after infection and then decreased precipitously. Productively infected macrophages could be readily recognized in electron micrographs of 24-h infected macrophage cultures and in sections of spleens from 24-h infected mice by characteristic morphological alterations. These consisted of formation of clusters of double-membrane vesicles with a diameter of 100 to 300 mumol, budding of nucleocapsids into vesicles with single membranes and accumulation of mature virions in these vesicles. One to 4 days later, however, such cells were no longer found in infected cultures or spleens of infected mice and superinfection did not restimulate LDV replication. Cultures established with macrophages from 1-day LDV-infected mice also did not support LDV replication. We conclude that LDV replication in cultures or mice is limited to a single cycle in a subpopulation of macrophages and that infection leads to cell death and rapid phagocytosis of the dead cells by the resistant, uninfected macrophages.