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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.)

Studies of reproductive biology in insects often require quantification of sperm production, transfer or storage. Here, we develop a quantitative real-time PCR-based assay using a Y-specific marker for quantification of sperm from spermathecae of female Queensland fruit fly ('Q-fly'), overcoming constraints typical of traditional sperm quantification methods. The assay enables accurate and reliable quantification of as few as 50 sperms and provides a means to analyse large numbers of samples with flexible timing. The real-time PCR method enables revised understanding of how many sperms are stored by female Q-flies, the distribution of storage between the two spermathecae and the relationship between copula duration and sperm storage. Real-time PCR assays based on Y-specific markers provide an effective solution for sperm quantification in tephritid flies, as well as in other insects and potentially other animals with sperm storage organs., (© 2021 The Royal Entomological Society.)

Seven new genera, 26 new species, 10 new combinations, two epitypes, one new name, and 20 interesting new host and / or geographical records are introduced in this study. New genera are: Italiofungus (based on Italiofungus phillyreae ) on leaves of Phillyrea latifolia (Italy); Neolamproconium (based on Neolamproconium silvestre ) on branch of Tilia sp. (Ukraine); Neosorocybe (based on Neosorocybe pini ) on trunk of Pinus sylvestris (Ukraine); Nothoseptoria (based on Nothoseptoria caraganae ) on leaves of Caragana arborescens (Russia); Pruniphilomyces (based on Pruniphilomyces circumscissus ) on Prunus cerasus (Russia); Vesiculozygosporium (based on Vesiculozygosporium echinosporum ) on leaves of Muntingia calabura (Malaysia); Longiseptatispora (based on Longiseptatispora curvata ) on leaves of Lonicera tatarica (Russia). New species are: Barrmaelia serenoae on leaf of Serenoa repens (USA); Chaetopsina gautengina on leaves of unidentified grass (South Africa); Chloridium pini on fallen trunk of Pinus sylvestris (Ukraine); Cadophora fallopiae on stems of Reynoutria sachalinensis (Poland); Coleophoma eucalyptigena on leaf litter of Eucalyptus sp. (Spain); Cylindrium corymbiae on leaves of Corymbia maculata (Australia); Diaporthe tarchonanthi on leaves of Tarchonanthus littoralis (South Africa); Elsinoe eucalyptorum on leaves of Eucalyptus propinqua (Australia); Exophiala quercina on dead wood of Quercus sp., (Germany); Fusarium californicum on cambium of budwood of Prunus dulcis (USA); Hypomyces gamsii on wood of Alnus glutinosa (Ukraine); Kalmusia araucariae on leaves of Araucaria bidwillii (USA); Lectera sambuci on leaves of Sambucus nigra (Russia); Melanomma populicola on fallen twig of Populus canadensis (Netherlands), Neocladosporium syringae on branches of Syringa vulgarishorus (Ukraine); Paraconiothyrium iridis on leaves of Iris pseudacorus (Ukraine); Pararoussoella quercina on branch of Quercus robur (Ukraine); Phialemonium pulveris from bore dust of deathwatch beetle (France); Polyscytalum pinicola on needles of Pinus tecunumanii (Malaysia); Acervuloseptoria fraxini on Fraxinus pennsylvanica (Russia); Roussoella arundinacea on culms of Arundo donax (Spain); Sphaerulina neoaceris on leaves of Acer negundo (Russia); Sphaerulina salicicola on leaves of Salix fragilis (Russia); Trichomerium syzygii on leaves of Syzygium cordatum (South Africa); Uzbekistanica vitis-viniferae on dead stem of Vitis vinifera (Ukraine); Vermiculariopsiella eucalyptigena on leaves of Eucalyptus sp. (Australia)., (© 2020 Westerdijk Fungal Biodiversity Institute.)

Species of eucalypts are commonly cultivated for solid wood and pulp products. The expansion of commercially managed eucalypt plantations has chiefly been driven by their rapid growth and suitability for propagation across a very wide variety of sites and climatic conditions. Infection of foliar fungal pathogens of eucalypts is resulting in increasingly negative impacts on commercial forest industries globally. To assist in evaluating this threat, the present study provides a global perspective on foliar pathogens of eucalypts. We treat 110 different genera including species associated with foliar disease symptoms of these hosts. The vast majority of these fungi have been grown in axenic culture, and subjected to DNA sequence analysis, resolving their phylogeny. During the course of this study several new genera and species were encountered, and these are described. New genera include: Lembosiniella ( L. eucalyptorum on E. dunnii, Australia), Neosonderhenia ( N. eucalypti on E. costata, Australia), Neothyriopsis ( N. sphaerospora on E. camaldulensis , South Africa), Neotrichosphaeria ( N. eucalypticola on E. deglupta , Australia), Nothotrimmatostroma ( N. bifarium on E. dalrympleana , Australia), Nowamyces (incl. Nowamycetaceae fam. nov ., N. globulus on E. globulus , Australia), and Walkaminomyces ( W. medusae on E. alba , Australia). New species include (all from Australia): Disculoides fraxinoides on E. fraxinoides, Elsinoe piperitae on E. piperita, Fusculina regnans on E. regnans, Marthamyces johnstonii on E. dunnii , Neofusicoccum corticosae on E. corticosa , Neotrimmatostroma dalrympleanae on E. dalrympleana, Nowamyces piperitae on E. piperita , Phaeothyriolum dunnii on E. dunnii , Pseudophloeospora eucalyptigena on E. obliqua , Pseudophloeospora jollyi on Eucalyptus sp., Quambalaria tasmaniae on Eucalyptus sp., Q. rugosae on E. rugosa , Sonderhenia radiata on E. radiata , Teratosphaeria pseudonubilosa on E. globulus and Thyrinula dunnii on E. dunnii . A new name is also proposed for Heteroconium eucalypti as Thyrinula uruguayensis on E. dunnii , Uruguay. Although many of these genera and species are commonly associated with disease problems, several appear to be opportunists developing on stressed or dying tissues. For the majority of these fungi, pathogenicity remains to be determined. This represents an important goal for forest pathologists and biologists in the future. Consequently, this study will promote renewed interest in foliar pathogens of eucalypts, leading to investigations that will provide an improved understanding of the biology of these fungi., (© 2019 Westerdijk Fungal Biodiversity Institute. Production and hosting by ELSEVIER B.V.)

This paper represents the third contribution in the Genera of Phytopathogenic Fungi (GOPHY) series. The series provides morphological descriptions, information about the pathology, distribution, hosts and disease symptoms for the treated genera, as well as primary and secondary DNA barcodes for the currently accepted species included in these. This third paper in the GOPHY series treats 21 genera of phytopathogenic fungi and their relatives including: Allophoma , Alternaria , Brunneosphaerella , Elsinoe , Exserohilum , Neosetophoma , Neostagonospora , Nothophoma , Parastagonospora , Phaeosphaeriopsis , Pleiocarpon , Pyrenophora , Ramichloridium , Seifertia , Seiridium , Septoriella , Setophoma , Stagonosporopsis , Stemphylium , Tubakia and Zasmidium . This study includes three new genera, 42 new species, 23 new combinations, four new names, and three typifications of older names., (© 2019 Westerdijk Fungal Biodiversity Institute. Production and hosting by ELSEVIER B.V.)

This paper represents the second contribution in the Genera of Phytopathogenic Fungi (GOPHY) series. The series provides morphological descriptions and information regarding the pathology, distribution, hosts and disease symptoms for the treated genera. In addition, primary and secondary DNA barcodes for the currently accepted species are included. This second paper in the GOPHY series treats 20 genera of phytopathogenic fungi and their relatives including: Allantophomopsiella, Apoharknessia, Cylindrocladiella, Diaporthe, Dichotomophthora, Gaeumannomyces, Harknessia, Huntiella, Macgarvieomyces, Metulocladosporiella, Microdochium, Oculimacula, Paraphoma, Phaeoacremonium, Phyllosticta, Proxypiricularia, Pyricularia, Stenocarpella , Utrechtiana and Wojnowiciella . This study includes the new genus Pyriculariomyces , 20 new species, five new combinations, and six typifications for older names.

Novel species of fungi described in this study include those from various countries as follows: Australia : Banksiophoma australiensis (incl. Banksiophoma gen. nov.) on Banksia coccinea , Davidiellomyces australiensis (incl. Davidiellomyces gen. nov.) on Cyperaceae , Didymocyrtis banksiae on Banksia sessilis var . cygnorum , Disculoides calophyllae on Corymbia calophylla , Harknessia banksiae on Banksia sessilis , Harknessia banksiae-repens on Banksia repens , Harknessia banksiigena on Banksia sessilis var . cygnorum , Harknessia communis on Podocarpus sp., Harknessia platyphyllae on Eucalyptus platyphylla , Myrtacremonium eucalypti (incl. Myrtacremonium gen. nov.) on Eucalyptus globulus , Myrtapenidiella balenae on Eucalyptus sp., Myrtapenidiella eucalyptigena on Eucalyptus sp., Myrtapenidiella pleurocarpae on Eucalyptus pleurocarpa , Paraconiothyrium hakeae on Hakea sp., Paraphaeosphaeria xanthorrhoeae on Xanthorrhoea sp., Parateratosphaeria stirlingiae on Stirlingia sp., Perthomyces podocarpi (incl. Perthomyces gen. nov.) on Podocarpus sp., Readeriella ellipsoidea on Eucalyptus sp., Rosellinia australiensis on Banksia grandis , Tiarosporella corymbiae on Corymbia calophylla , Verrucoconiothyrium eucalyptigenum on Eucalyptus sp., Zasmidium commune on Xanthorrhoea sp., and Zasmidium podocarpi on Podocarpus sp. Brazil : Cyathus aurantogriseocarpus on decaying wood, Perenniporia brasiliensis on decayed wood, Perenniporia paraguyanensis on decayed wood, and Pseudocercospora leandrae-fragilis on Leandra fragilis. Chile : Phialocephala cladophialophoroides on human toe nail. Costa Rica : Psathyrella striatoannulata from soil. Czech Republic : Myotisia cremea (incl. Myotisia gen. nov.) on bat droppings. Ecuador : Humidicutis dictiocephala from soil, Hygrocybe macrosiparia from soil, Hygrocybe sangayensis from soil, and Polycephalomyces onorei on stem of Etlingera sp. France : Westerdykella centenaria from soil. Hungary : Tuber magentipunctatum from soil. India : Ganoderma mizoramense on decaying wood, Hodophilus indicus from soil, Keratinophyton turgidum in soil, and Russula arunii on Pterigota alata. Italy : Rhodocybe matesina from soil. Malaysia : Apoharknessia eucalyptorum , Harknessia malayensis , Harknessia pellitae , and Peyronellaea eucalypti on Eucalyptus pellita , Lectera capsici on Capsicum annuum , and Wallrothiella gmelinae on Gmelina arborea. Morocco : Neocordana musigena on Musa sp. New Zealand : Candida rongomai-pounamu on agaric mushroom surface, Candida vespimorsuum on cup fungus surface, Cylindrocladiella vitis on Vitis vinifera , Foliocryphia eucalyptorum on Eucalyptus sp., Ramularia vacciniicola on Vaccinium sp., and Rhodotorula ngohengohe on bird feather surface. Poland : Tolypocladium fumosum on a caterpillar case of unidentified Lepidoptera. Russia : Pholiotina longistipitata among moss. Spain : Coprinopsis pseudomarcescibilis from soil, Eremiomyces innocentii from soil, Gyroporus pseudocyanescens in humus, Inocybe parvicystis in humus, and Penicillium parvofructum from soil. Unknown origin : Paraphoma rhaphiolepidis on Rhaphiolepsis indica. USA : Acidiella americana from wall of a cooling tower, Neodactylaria obpyriformis (incl. Neodactylaria gen. nov.) from human bronchoalveolar lavage, and Saksenaea loutrophoriformis from human eye. Vietnam : Phytophthora mekongensis from Citrus grandis , and Phytophthora prodigiosa from Citrus grandis. Morphological and culture characteristics along with DNA barcodes are provided.

Novel species of fungi described in the present study include the following from Australia: Vermiculariopsiella eucalypti, Mulderomyces natalis (incl. Mulderomyces gen. nov.), Fusicladium paraamoenum, Neotrimmatostroma paraexcentricum, and Pseudophloeospora eucalyptorum on leaves of Eucalyptus spp., Anungitea grevilleae (on leaves of Grevillea sp.), Pyrenochaeta acaciae (on leaves of Acacia sp.), and Brunneocarpos banksiae (incl. Brunneocarpos gen. nov.) on cones of Banksia attenuata. Novel foliicolous taxa from South Africa include Neosulcatispora strelitziae (on Strelitzia nicolai), Colletotrichum ledebouriae (on Ledebouria floridunda), Cylindrosympodioides brabejum (incl. Cylindrosympodioides gen. nov.) on Brabejum stellatifolium, Sclerostagonospora ericae (on Erica sp.), Setophoma cyperi (on Cyperus sphaerocephala), and Phaeosphaeria breonadiae (on Breonadia microcephala). Novelties described from Robben Island (South Africa) include Wojnowiciella cissampeli and Diaporthe cissampeli (both on Cissampelos capensis), Phaeotheca salicorniae (on Salicornia meyeriana), Paracylindrocarpon aloicola (incl. Paracylindrocarpon gen. nov.) on Aloe sp., and Libertasomyces myopori (incl. Libertasomyces gen. nov.) on Myoporum serratum. Several novelties are recorded from La Réunion (France), namely Phaeosphaeriopsis agapanthi (on Agapanthus sp.), Roussoella solani (on Solanum mauritianum), Vermiculariopsiella acaciae (on Acacia heterophylla), Dothiorella acacicola (on Acacia mearnsii), Chalara clidemiae (on Clidemia hirta), Cytospora tibouchinae (on Tibouchina semidecandra), Diaporthe ocoteae (on Ocotea obtusata), Castanediella eucalypticola, Phaeophleospora eucalypticola and Fusicladium eucalypticola (on Eucalyptus robusta), Lareunionomyces syzygii (incl. Lareunionomyces gen. nov.) and Parawiesneriomyces syzygii (incl. Parawiesneriomyces gen. nov.) on leaves of Syzygium jambos. Novel taxa from the USA include Meristemomyces arctostaphylos (on Arctostaphylos patula), Ochroconis dracaenae (on Dracaena reflexa), Rasamsonia columbiensis (air of a hotel conference room), Paecilomyces tabacinus (on Nicotiana tabacum), Toxicocladosporium hominis (from human broncoalveolar lavage fluid), Nothophoma macrospora (from respiratory secretion of a patient with pneumonia), and Penidiellopsis radicularis (incl. Penidiellopsis gen. nov.) from a human nail. Novel taxa described from Malaysia include Prosopidicola albizziae (on Albizzia falcataria), Proxipyricularia asari (on Asarum sp.), Diaporthe passifloricola (on Passiflora foetida), Paramycoleptodiscus albizziae (incl. Paramycoleptodiscus gen. nov.) on Albizzia falcataria, and Malaysiasca phaii (incl. Malaysiasca gen. nov.) on Phaius reflexipetalus. Two species are newly described from human patients in the Czech Republic, namely Microascus longicollis (from toenails of patient with suspected onychomycosis), and Chrysosporium echinulatum (from sole skin of patient). Furthermore, Alternaria quercicola is described on leaves of Quercus brantii (Iran), Stemphylium beticola on leaves of Beta vulgaris (The Netherlands), Scleroderma capeverdeanum on soil (Cape Verde Islands), Scleroderma dunensis on soil, and Blastobotrys meliponae from bee honey (Brazil), Ganoderma mbrekobenum on angiosperms (Ghana), Geoglossum raitviirii and Entoloma kruticianum on soil (Russia), Priceomyces vitoshaensis on Pterostichus melas (Carabidae) (Bulgaria) is the only one for which the family is listed, Ganoderma ecuadoriense on decaying wood (Ecuador), Thyrostroma cornicola on Cornus officinalis (Korea), Cercophora vinosa on decorticated branch of Salix sp. (France), Coprinus pinetorum, Coprinus littoralis and Xerocomellus poederi on soil (Spain). Two new genera from Colombia include Helminthosporiella and Uwemyces on leaves of Elaeis oleifera. Two species are described from India, namely Russula intervenosa (ectomycorrhizal with Shorea robusta), and Crinipellis odorata (on bark of Mytragyna parviflora). Novelties from Thailand include Cyphellophora gamsii (on leaf litter), Pisolithus aureosericeus and Corynascus citrinus (on soil). Two species are newly described from Citrus in Italy, namely Dendryphiella paravinosa on Citrus sinensis, and Ramularia citricola on Citrus floridana. Morphological and culture characteristics along with ITS nrDNA barcodes are provided for all taxa.

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.

The sterile insect technique has been routinely used to eradicate fruit fly Bactrocera tryoni (Froggatt) incursions. This study considers whether fly quality in a mass-rearing facility can be improved by reducing irradiation doses, without sacrificing reproductive sterility. Pupae were exposed to one of five target irradiation dose ranges: 0, 40-45, 50-55, 60-65, and 70-75 Gy. Pupae were then assessed using routine quality control measures: flight ability, sex ratio, longevity under nutritional stress, emergence, and reproductive sterility. Irradiation did not have a significant effect on flight ability or sex ratio tests. Longevity under nutritional stress was significantly increased at 70-75 Gy, but no other doses differed from 0 Gy. Emergence was slightly reduced in the 50-55, 60-65, and 70-75 Gy treatments, but 40-45 Gy treatments did not differ from 0 Gy, though confounding temporal factors complicate interpretation. Reproductive sterility remained acceptable (> 99.5%) for all doses--40-45 Gy (99.78%), 50-55 Gy (100%), 60-65 Gy (100%), and 70-75 Gy (99.99%). We recommend that B. tryoni used in sterile insect technique releases be irradiated at a target dose of 50-55 Gy, providing improved quality and undiminished sterility in comparison with the current 70-75 Gy standard while also providing a substantial buffer against risk of under dosing.

The sterile insect technique (SIT) is a non-chemical approach used to control major pests from several insect families, including Tephritidae, and entails the mass-release of sterile insects that reduce fertility of wild populations. For SIT to succeed, released sterile males must mature and compete with wild males to mate with wild females. To reach sexual maturity, the Queensland fruit fly, Bactrocera tryoni (Froggatt) (Diptera: Tephritidae), must obtain adequate nutrition after adult emergence; however, in current SIT programs sterile B. tryoni receive a pre-release diet that lacks key nutrients required to sustain sexual development. The chief objective of this study was to determine whether pre-release yeast hydrolysate (YH) supplements affect the persistence and abundance of sexually mature sterile male B. tryoni under field conditions. Experiments were run in outdoor cages under conditions of low and high environmental stress that differed markedly in temperature and humidity, and in the field. Under low environmental stress conditions, survival of sterile B. tryoni was monitored in cages under three diet treatments: (i) sugar only, (ii) sugar plus YH or (iii) sugar plus YH for 48 h and sugar only thereafter. Under high environmental stress conditions survival of sterile B. tryoni was monitored in cages under four diet treatments: (i) white sugar only, (ii) brown sugar only, (iii) white sugar plus YH and (iv) brown sugar plus YH. In a replicated field study, we released colour-marked sterile B. tryoni from two diet regimes, YH-supplemented or YH-deprived, and monitored abundance of sexually mature males. In the low-stress cage study, there was no effect of diet, although overall females lived longer than males. In the high stress cage study, mortality was lower for YH-fed flies than YH-deprived flies and females lived longer than males. In the field, YH supplementation resulted in higher abundance of sexually mature sterile males, with 1.2 YH-fed flies trapped for every YH-deprived fly trapped. Under field conditions, YH supplementation can increase over-flooding ratios and hence may improve the effectiveness of SIT programmes.

In pest management programmes that incorporate the sterile insect technique (SIT), the ability of mass-reared insects to tolerate dry conditions may influence their survival after release in the field. In the present study, desiccation resistance of adult mass-reared Queensland fruit flies, Bactrocera tryoni (Frogatt) (Diptera: Tephritidae), that are routinely released in SIT programmes was compared with that of wild flies at 1, 10 and 20 days after adult eclosion. Under dry conditions without access to food or water, longevity of mass-reared B. tryoni was significantly less than that of their wild counterparts. Desiccation resistance of mass-reared flies declined monotonically with age, but this was not the case for wild flies. The sharp decline in desiccation resistance of mass-reared flies as they aged was likely explained by decreased dehydration tolerance. As in an earlier study, desiccation resistance of females was significantly lower than that of males but this was particularly pronounced in mass-reared females. Female susceptibility to dry conditions corresponded with declining dehydration tolerance with age and associated patterns of reproductive development, which suggests that water content of their oocyte load is not available for survival during periods of water stress.

Yersinia ruckeri is the causative agent of enteric redmouth disease (ERM), a common pathogen affecting aquaculture facilities and implicated in large losses of cultured fish. Fisheries scientists continue to gain a greater understanding of the disease and the pathogen by investigating methods of identification and pre- and post-infection treatment. In this study, a real-time PCR probe set for Y. ruckeri was developed to detect daily changes in the bacterial load during pathogen challenges. Two species of fish, Chinook salmon, Oncorhynchus tshawytscha, and steelhead trout, Oncorhynchus mykiss, were exposed to two strains of Y. ruckeri (Hag and SC) during bath challenges. A subset of fish was killed daily for 14 days, and the kidney tissue was biopsied to enumerate copies of pathogen DNA per gram of tissue. While Chinook exposed to either the Hag or SC strains exhibited similar pathogen loads, those exposed to the Hag strain displayed higher mortality (∼66%) than fish exposed to the SC strain (∼24% mortality). Steelhead exposed to the Hag strain exhibited a greater pathogen load and higher mortality (∼42%) than those exposed to the SC strain (<1% mortality). Steelhead challenged with either strain showed lower pathogen loads than Chinook. The study illustrates the efficacy of the probe set to enumerate Y. ruckeri bacterial growth in the kidneys of fish. Also, strains of Y. ruckeri display species-specific growth patterns that result in differential mortality and pathogen load., (Published 2011. This article is a US Government work and is in the public domain in the USA.)

Aims: To identify enzymes associated with bacteriophages infecting cystic fibrosis (CF) strains of Pseudomonas aeruginosa that are able to degrade extracellular alginic acids elaborated by the host bacterium., Methods and Results: Plaques produced by 21 Ps. aeruginosa-specific phages were screened for the presence of haloes, an indicator of capsule hydrolytic activity. Four phages produced haloed plaques, and one (PT-6) was investigated further. PT-6 was shown by electron microscopy to belong to Podoviridae family C1, to reduce the viscosity of four alginate preparations using a rolling ball viscometer and to release uronic acid-containing fragments from the polymers, as judged by spectrophotometry and thin layer chromatography. The alginase was partially purified by gel filtration chromatography and shown to be a 37 kDa polypeptide., Conclusions: Infection of CF strains of Ps. aeruginosa by phage PT-6 involves hydrolysis of the exopolysaccharide secreted by the host., Significance and Impact of the Study: The alginase produced by PT-6 has the potential to increase the well-being of CF suffers by improving the surface properties of sputum, accelerating phagocytic uptake of bacteria and perturbing bacterial growth in biofilms.

The current study is an important step toward calibrating, validating, and improving irradiation methods used for Bactrocera tryoni (Froggatt) sterile insect technique (SIT). We used routine International Atomic Energy Agency/U.S. Department of Agriculture/Food and Agriculture Organization quality control tests assessing percentage of emergence, flight ability, sex ratio, mortality under stress, reproductive sterility, and sexual competitiveness, as well as a nonstandard test of longevity under nutritional stress to assess the impact of a range of target irradiation doses (60, 65, 70, 75, and 80 Gy) on the product quality of mass reared B. tryoni used in SIT. Sterility induction remained adequate (>99.5%) for sterile male-fertile female crosses, and 100% sterility was achieved in fertile male-sterile female crosses and sterile male-sterile female crosses for each irradiation doses tested. There was significant increase in mortality under stress as irradiation dose increased, and reduced participation in mating by males irradiated at higher doses. The current target-sterilizing dose for SIT of 70-75 Gy is associated with significant reduction in fly product "quality". Our data suggest that adequate sterility and improved fly quality could be achieved through a small reduction in target sterilizing dose.

Resistance to anthracnose, caused by Colletotrichum capsici and C. acutatum, was investigated in Capsicum baccatum PBC80 and PBC1422 and C. chinense PBC932. Mature green and ripe fruit were inoculated with 13 isolates of the two Colletotrichum species PBC80 contained the broadest spectrum of resistance to both Colletotrichum species because none of the isolates were able to infect the genotype. At both fruit maturity stages, PBC1422 was infected by only Colletotrichum acutatum. PBC932 at ripe fruit stage was infected by both C. capsici and C. acutatum, except for one isolate, 158ci, that did not infect PBC932. PBC932 at the mature green fruit stage was infected by only C. acutatum. An intraspecific cross between PBC80 and PBC1422 was developed to determine inheritance of resistance to C. acutatum. Anthracnose resistance was assessed at mature green and ripe fruit stages using 0 to 9 disease severity scores. Frequency distribution of the disease scores in the F(2) and BC(1) populations suggested a single recessive gene responsible for the resistance at mature green fruit stage and a single dominant gene for the resistance at ripe fruit stage. Linkage analysis between the two genes identified in both fruit maturity stages showed the genes to be independent. Based on phenotypic data, the two newly identified genes, co4 and Co5, from PBC80 appeared to be different loci from the co1 and co2 previously identified from PBC932 and will be valuable sources of resistance to anthracnose in chili breeding programs.

Genomic libraries enriched for microsatellites from Colletotrichum capsici, one of the major causal agents of anthracnose disease in chilli pepper (Capsicum spp.), were developed using a modified hybridization procedure. Twenty-seven robust primer pairs were designed from microsatellite flanking sequences and were characterized using 52 isolates from three countries India, Sri Lanka and Thailand. Highest gene diversity of 0.857 was observed at the CCSSR1 with up to 18 alleles among all the isolates whereas the differentiation ranged from 0.05 to 0.45. The sequence-tagged microsatellite site markers developed in this study will be useful for genetic analyses of C. capsici populations., (© 2009 Blackwell Publishing Ltd.)

Aims: To determine the capacity of extracts of Pelargonium reniforme and Pelargonium sidoides, plants of the Geraniaceae family, to stimulate the uptake and killing of mycobacteria by murine macrophages and to identify the constituents that are responsible., Methods and Results: Bioassay-guided fractionation of aqueous P. reniforme extracts yielded five chemically distinct structures with the capacity to increase the rate of intracellular killing by macrophages. These were: gallic acid, methyl gallate, myricetin and quercitin-3-O-beta-d-glucoside, in addition to the previously unrecognized constituent 1-O-(2-(4-methoxyphenyl)ethyl-6-O-galloyl-glucopyranoside. Kinetics of intracellular accumulation of Mycobacterium tuberculosis and Mycobacterium fortuitum by macrophages were indistinguishable; pure preparations of the four previously known plant constituents stimulated macrophage killing, but not uptake, of M. tuberculosis and M. fortuitum equally well., Conclusions: A number of distinct molecular species are present in the medicinal plant P. reniforme that stimulate the killing of the intracellular pathogen M. tuberculosis., Significance and Impact of the Study: These observations support the view that Pelargonium extracts may have utility in the treatment of tuberculosis.

Eleven isolates of Colletotrichum capsici were screened on nine chili genotypes derived from four cultivated species of Capsicum: Capsicum annuum, C. baccatum, C. chinense, and C. frutescens. Host reactions were assessed 9 days after inoculation by microinjection of spores into the pericarp of red fruit. A set of disease scales, with 0 to 9 scores, were developed for anthracnose infection of each Capsicum sp. based on percent lesion size in relation to fruit size, appearance of necrotic or water-soaked tissue, and presence of acervuli. Three pathotypes, PCc1, PCc2, and PCc3, were identified according to differential qualitative infection of fruit of C. chinense genotypes PBC932 and C04714. PCc1 was the most virulent pathotype, infecting all genotypes of C. annuum, C. chinense, and C. frutescens, whereas PCc3 was the least virulent pathotype, infecting only the genotypes C. annuum and C. frutescens. Quantitative infection occurred in all chili genotypes except for genotypes of C. baccatum, where no infection occurred, demonstrating various levels of aggressiveness of isolates within pathotypes.

Aims: To measure antibacterial activity of the semi-synthetic flavonoid 3-O-octanoyl-(-)-epicatechin and investigate the mechanism of action., Methods and Results: MICs determined by the broth microdilution method were 50 microg ml(-1) for beta-lactam sensitive and resistant Staphylococcus aureus, and 100 microg ml(-1) for vancomycin sensitive and resistant enterococci. In time-kill studies, 100 microg ml(-1) 3-O-octanoyl-(-)-epicatechin reduced colony forming unit numbers of antibiotic sensitive and methicillin-resistant Staph. aureus below detectable levels within 120 min. Bacterial aggregation was not observed when cells exposed to 3-O-octanoyl-(-)-epicatechin were examined by light microscopy. It was also shown that 50 microg ml(-1) 3-O-octanoyl-(-)-epicatechin is capable of reducing colony forming unit numbers of high cell density Staph. aureus populations by 80-fold within 60 min incubation, and inducing leakage of 50% of their internal potassium within just 10 min., Conclusions: 3-O-Octanoyl-(-)-epicatechin is active against Gram-positive bacteria, has bactericidal activity against both antibiotic sensitive and resistant strains, and is likely to exert its primary antibacterial effect by damaging the cytoplasmic membrane., Significance and Impact of the Study: 3-O-Octanoyl-(-)-epicatechin has significant antibacterial activity and additional structural modification and/or formulation studies may allow this to be potentiated.

Aim: (-)-epicatechin gallate (ECg) modifies the morphology, cell wall architecture and beta-lactam antibiotic susceptibility of Staphylococcus aureus. As these effects result primarily from intercalation into the bacterial cytoplasmic membrane, the capacity of ECg to modulate the secretion of two key staphylococcal virulence factors, coagulase and alpha-toxin, was examined., Methods and Results: Bioassays were used to determine coagulase and haemolysin activity in culture supernatants of a number of S. aureus isolates grown in the presence and absence of ECg; alpha-toxin secretion was also evaluated by immunoblotting. Growth in ECg reduced the levels of activity of both proteins in culture supernatants; the effects could only be partly explained by ECg-mediated inhibition of bioactivity and by induction of secreted proteases., Conclusion: ECg suppresses the secretion of coagulase and alpha-toxin by clinical isolates of S. aureus., Significance and Impact of the Study: The observation that secretion of key components of staphylococcal virulence can be compromised by a naturally occurring polyphenol supports the notion that ECg and related compounds may have therapeutic utility for the control of infections that are currently difficult to treat due to the propensity of methicillin-resistant S. aureus to accumulate antibiotic resistance genes.

Aim: The flavonol galangin, an antimicrobial constituent of the traditional medicines propolis and Helichrysum aureonitens, is being assessed as part of an ongoing investigation into the antibacterial activity of flavonoids. The present study sought to establish whether galangin has any aggregatory effect on bacterial cells., Methods and Results: In preparatory time-kill assays, 50 microg ml(-1) of galangin was found to reduce colony counts of c. 5 x 10(7) CFU ml(-1)Staphylococcus aureus NCTC 6571 by approximately 15 000-fold during 60 min of incubation. Subsequent light microscopy studies demonstrated significant increases in the number of large clusters of bacterial cells in populations treated with the flavonol., Conclusion: Data presented here show that galangin causes aggregation of bacterial cells., Significance and Impact of the Study: The finding that galangin causes bacterial cells to clump together may implicate the cytoplasmic membrane as a target site for this compound's activity. More importantly, this observation indicates that decreases in CFU numbers detected in time-kill and minimum bactericidal concentration (MBC) assays in previous investigations were at least partially attributable to this aggregatory effect. This raises the possibility that galangin is not genuinely bactericidal in action, and calls into question the suitability of time-kill and MBC assays for determining the nature of activity of naturally occurring flavonoids.

Female Mediterranean fruit flies, Ceratitis capitata (Wiedemann), from the sterile-male rearing facility in El Pino, Guatemala, were exposed to broadcasts of wing-fanning vibrations recorded from males engaged in calling behavior to investigate the feasibility of developing a female-selective acoustic trap. The recorded signals had frequent amplitude fluctuations and peak frequencies approximately 350 Hz, typical of signals observed in previous studies of Mediterranean fruit fly acoustic behavior. Females did not exhibit long-distance phonotaxis, but remained near a speaker significantly longer when the sounds were broadcast at 103-107 dB than when the speaker was silent. In addition, significantly higher percentages of females were captured by yellow adhesive traps next to a broadcasting speaker than by traps next to a silent mimic. Additional bioassays were conducted with synthetic, 350-Hz tones produced by a thermoacoustic tube as well as with silent mimics of the different sound sources to examine the relative responsiveness of female Mediterranean fruit flies to traps with different acoustic and visual features. The visual attributes of the different sound source assemblies significantly affected capture rates. The range over which the broadcast significantly increased the percentage of female captures was <0.5 m, which may limit the utility of these acoustic cues in large-scale trapping programs. However, the findings of this study do justify further testing of whether optimized short-range acoustic signals could be used to augment longer range pheromonal and visual cues to improve the efficacy of female-selective traps.

Aims: To isolate bacteriophages lytic for oral pathogens from human saliva, dental plaque and mature biofilms constituted from saliva-derived bacteria., Methods and Results: Saliva and dental plaque samples from healthy volunteers and from patients with gingivitis and periodontitis were examined for the presence of lytic bacteriophage using a panel of oral pathogens and bacteria isolated from the samples. Samples were also enriched for bacteriophage using static culture techniques and mature biofilms. A limited number of samples contained bacteriophage particles that were visualized using electron microscopy. Cultures yielded phage infecting non-oral bacteria (Proteus mirabilis) but no bacteriophage specific for recognized oral pathogens were found. Some micro-organisms from the oral microflora elaborated antibacterial substances that inhibited growth of other residents of the oral cavity., Conclusions: Unlike other ecosystems, the composition of the oral cavity does not appear to be heavily influenced by interactions between bacteriophages and their hosts., Significance and Impact of the Study: Bacteriophage for control of oral infections may need to be obtained from other sources. Antibacterial substances derived from some members of the oral microflora warrant investigation as potential antibiotics.

In both controlled environment and the field, six QTLs for ascochyta blight resistance were identified in three regions of the genome of an intraspecific population of chickpea using the IDS and AUDPC disease scoring systems. One QTL-region was detected from both environments, whereas the other two regions were detected from each environment. All the QTL-regions were significantly associated with ascochyta blight resistance using either of the disease scoring systems. The QTLs were verified by multiple interval mapping, and a two-QTL genetic model with considerable epistasis was established for both environments. The major QTLs generally showed additive gene action, as well as dominance inter-locus interaction in the multiple genetic model. All the QTLs were mapped near a RGA marker. The major QTLs were located on LG III, which was mapped with five different types of RGA markers. A CLRR-RGA marker and a STMS marker flanked QTL 6 for controlled environment resistance at 0.06 and 0.04 cM, respectively. Other STMS markers flanked QTL 1 for field resistance at a 5.6 cM interval. After validation, these flanking markers may be used in marker-assisted selection to breed for elite chickpea cultivars with durable resistance to ascochyta blight. The tight linkage of RGA markers to the major QTL on LG III will allow map-based cloning of the underlying resistance genes.

The first intraspecific linkage map of the lentil genome was constructed with 114 molecular markers (100 RAPD, 11 ISSR and three RGA) using an F(2) population developed from a cross between lentil cultivars ILL5588 and ILL7537 which differed in resistance for ascochyta blight. Linkage analysis at a LOD score of 4.0 and a maximum recombination fraction of 0.25 revealed nine linkage groups comprising between 6 and 18 markers each. The intraspecific map spanned a total length of 784.1 cM. The markers were distributed throughout the genome, however markers were clustered in the middle or near the middle of the linkage groups, suggesting the location of centromeres. Of 114 mapped markers, 16 (14.0%) were distorted, usually at the end or middle of the linkage groups. The utility of ISSR and RGA markers for mapping in lentil was explored, and the primer with an (AC) repeat motif was found to be useful.

SUMMARY A genome linkage map was developed for Ascochyta rabiei (Pass.) Labrousse, (teleomorph) Didymella rabiei (Kovachevski), an important pathogen causing ascochyta blight in chickpea (Cicer arietinum L.). The map was constructed using 96 progeny generated from a single pseudothecium produced from a cross between a USA MAT-2 isolate and an Australian MAT-1 isolate. The map comprised 126 molecular markers of which 69 were random amplified polymorphic DNA (RAPD) markers, 46 were amplified fragment length polymorphic (AFLP) markers, 10 were sequence-tagged microsatellite site (STMS) markers, and one was a sequence characterized amplified region (SCAR) marker. Eighteen large and 10 small linkage groups (LG) were characterized and the mating-type locus was mapped on to LGd. The map spanned 1271 cM with an average spacing between markers of 15.1 cM. The SCAR marker, specific for mating type 2, was designed to amplify a region of the MAT locus and was used to identify the mating type of A. rabiei isolates. One AFLP marker, derived from the MAT-1 parent, was closely linked to the mating-type locus (9.6 cM). The linkage map provides a framework for the future identification of the locations of other important traits such as virulence/avirulence and fungicide resistance. Findings from this study suggest that the MAT-2 isolates of D. rabiei should be renamed to MAT-1 isolates because the alpha-box, specific for MAT-1 from other ascomycetes, was amplified from A. rabiei MAT-2 isolates.

Accessions from Cicer echinospermum, a wild relative of chickpea (Cicer arietinum L.), contain resistance to the fungal disease ascochyta blight, a devastating disease of chickpea. A linkage map was constructed based on an interspecific F(2) population, derived from a cross between a susceptible chickpea cultivar (Lasseter) and a resistant C. echinospermum accession (PI 527930). The linkage map incorporated 83 molecular markers, that included RAPD, ISSR, STMS and RGA markers; eight markers remained unlinked. The map comprised eight linkage groups and covered a map distance of 570 cM. Six out of the eight linkage groups were correlated to linkage groups from the integrated Cicer map using STMS markers. Quantitative trait loci (QTLs) associated with ascochyta blight resistance were detected using interval mapping and single-point analysis. The F(2) population was evaluated for seedling and stem resistance in glasshouse trials. At least two QTLs were identified for seedling resistance, both of which were located within linkage group 4. Five markers were associated with stem resistance, four of which were also associated with seedling resistance. QTLs from previous studies also mapped to LG 4, suggesting that this linkage group is an important region of the Cicer genome for resistance to ascochyta blight.

An intraspecific linkage map of the chickpea genome based on STMS as anchor markers, was established using an F(2) population of chickpea cultivars with contrasting disease reactions to Ascochyta rabiei (Pass.) Lab. At a LOD-score of 2.0 and a maximum recombination distance of 20 cM, 51 out of 54 chickpea-STMS markers (94.4%), three ISSR markers (100%) and 12 RGA markers (57.1%) were mapped into eight linkage groups. The chickpea-derived STMS markers were distributed throughout the genome, while the RGA markers clustered with the ISSR markers on linkage groups LG I, II and III. The intraspecific linkage map spanned 534.5 cM with an average interval of 8.1 cM between markers. Sixteen markers (19.5%) were unlinked, while l1 chickpea-STMS markers (20.4%) deviated significantly ( P < 0.05) from the expected Mendelian segregation ratio and segregated in favor of the maternal alleles. However, ten of the distorted chickpea-STMS markers were mapped and clustered mostly on LG VII, suggesting the association of these loci in the preferential transmission of the maternal germ line. Preliminary comparative mapping revealed that chickpea may have evolved from Cicer reticulatum, possibly via inversion of DNA sequences and minor chromosomal translocation. At least three linkage groups that spanned a total of approximately 79.2 cM were conserved in the speciation process.

An aberrant random amplified polymorphic DNA (RAPD) marker in genomic DNA of tissue culture plantlets was frequently observed during a comparison of DNA fingerprints derived from potato germplasm grown in tissue culture and the field. The RAPD marker was cloned, sequenced and determined to be of bacterial origin. A bacterial contaminant was isolated from the tissue culture plants and identified as a Bacillus pumilus. A set of sequence characterised amplified region (SCAR) primers were designed from the sequence of the cloned fragment and tested for the specific detection of B. pumilus. Polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) were also used to generate B. pumilus profiles specific to our isolate in order to test and confirm the sequence homology of amplified markers generated from a range of DNA samples isolated from tissue culture plants and pure isolates of B. pumilus-like bacteria.

The question of why receivers accept a selfish signaller's message as reliable or 'honest' has fuelled ample controversy in discussions of communication. The handicap mechanism is now widely accepted as a potent constraint on cheating. Handicap signals are deemed reliable by their costs: signallers must choose between investing in the signal or in other aspects of fitness. Accordingly, resources allocated to the signal come to reflect the signaller's fitness budget and, on average, cheating is uneconomic. However, that signals may also be deemed reliable by their design, regardless of costs, is not widely appreciated. Here we briefly describe indices and amplifiers, reliable signals that may be essentially cost free. Indices are reliable because they bear a direct association with the signalled quality rather than costs. Amplifiers do not directly provide information about signaller quality, but they facilitate assessment by increasing the apparency of pre-existing cues and signals that are associated with quality. We present results of experiments involving a jumping spider (Plexippus paykulli) to illustrate how amplifiers can facilitate assessment of cues associated with physical condition without invoking the costs required for handicap signalling.

Because the complex phenotype of human hypertension is at least in part genetically determined, how individual genes ultimately contribute to the disease is not well understood. By contrast, intermediate phenotypes are traits associated with complex disease, but which may display simpler genetic properties such as greater heritability, more consistent and earlier penetrance and bimodality, and may suggest particular candidate susceptibility genes. Because autonomic nervous system activity is altered in hypertension, we examined biochemical, physiologic, and pharmacologic autonomic traits that fulfill at least some of these properties. Such biochemical, physiologic, or pharmacologic autonomic traits may be especially valuable as phenotypic anchor points in linkage or association studies probing the genetic basis of human hypertension.

Bacterial biofilms have been observed and reported on food and food-processing surfaces and can contribute to increased risks for product quality and food safety. The colonization of fruit and vegetables by pectynolitic bacteria like Pseudonomas fluorescens attributable to conditions such as soft rot, can also manifest as biofilms. A developed biofilm structure can provide a protective environment for pathogens such as Listeria monocytogenes reducing the effectiveness of sanitisers and other inhibitory agents. Understanding the colonization of bacteria on leaf surfaces is essential to the development of a better understanding of the leaf ecology of vegetable products. Studies of microbial colonization of leaf surfaces have been conducted using SEM and more recently using confocal microsocpy techniques. In the current study, a Leica TCS NT laser scanning confocal microscope was used to investigate biofilm formation using vital fluorescence staining on intact vegetable leaves. Reflection contrast and fluorescence three-dimensional imaging successfully delineated bacterial and biofilm morphology without disturbing the bacterial or leaf surface structure. The results demonstrate the presence and development of biofilm on the surface of lettuce. The biofilms appeared to originate on the cuticle in distinct micro-environments such as in the natural depression of the stomata, or in the intercellular junction. Bacteria also adhered to and developed biofilm colonies within an hour of contact and with clean stainless steel surfaces. Our study investigates the progression of biofilm formation from leaf colonization, and will assist in characterising the critical mechanisms of plant/host interaction and facilitate the development of improved preservation, sanitising and packaging strategies for minimally processed vegetable products., (1998 Society of Applied Microbiology.)

Carbohydrate residues covalently linked to plasma membrane proteins and lipids often provide specific markers at the cell surface. Traditionally such carbohydrate structures have been identified using antibodies and lectins. However problems of affinity and lack of specificity have restricted their usefulness. Protein engineering offers a way round these difficulties. In the case of some specialised cell surface carbohydrate structures, such as polysialic acid, enzymes may be useful analytical tools. Endosialidases specific for polysialic acid have recently been cloned and sequenced.

CGP55847, liposomal zinc(II)-phthalocyanine (Zn-Pc), was administered by the intravenous route to Swiss mice bearing intramuscularly implanted Ehrlich carcinomas or to C57/BL6 mice bearing subcutaneously implanted B16 melanomas. Tumors were removed 3 h or 24 h after dosing the intratumoral distribution determined by fluorescence microscopy. Localization of the photosensitizer occurred more rapidly in the Ehrlich carcinoma than in the B16 melanoma; this difference in photosensitizer uptake may be related to a higher degree of vascularization of the carcinoma. The photosensitizer was found in association with blood vessels at 3 h but not 24 h after dosing and appeared to have a greater affinity for areas of tissue necrosis within the tumor compared to viable tumor tissue. Little or no Zn-Pc was detected in the muscle tissue invaded by the Ehrlich carcinoma and was associated with the membranes and the cytosol, but not the nucleus, of cells in both tumors.

Objective: To characterize the etiologic agents (MO1) of the first reported case of babesiosis acquired in Missouri., Design: Case report, serologic testing, animal inoculations, and molecular studies., Setting: Southeastern Missouri., Patient: A 73-year-old man who had had a splenectomy and had a fatal case of babesiosis., Measurements: Serum specimens from the patient were assayed by indirect immunofluorescent antibody testing and immunoprecipitation for reactivity with antigens from various Babesia species. Whole blood obtained from the patient before treatment was inoculated into hamsters and jirds and into calves and bighorn sheep that had had splenectomy and were immunosuppressed with dexamethasone. Piroplasm-specific nuclear small-subunit ribosomal DNA was recovered from the patient's blood by using broad-range amplification with the polymerase chain reaction; a 144 base-pair region of the amplification product was sequenced; and phylogenetic analysis was done to compare MO1 with various Babesia species., Results: Indirect immunofluorescent antibody testing showed that the patient's serum had strong reactivity with Babesia divergens, which causes babesiosis in cattle and humans in Europe, but that it had minimal reactivity with B. microti and WA1, which are the piroplasms previously known to cause zoonotic babesiosis in the United States. Immunoprecipitations showed that MO1 is more closely related to B. divergens than to B. canis (a canine parasite). None of the experimentally inoculated animals became demonstrably parasitemic. Phylogenetic analyses, after DNA sequencing, showed that MO1 is most closely related to B. divergens (100% similarity)., Conclusions: Although MO1 is probably distinct from B. divergens, the two share morphologic, antigenic, and genetic characteristics; MO1 probably represents a Babesia species not previously recognized to have infected humans. Medical personnel should be aware that patients in the United States can have life-threatening babesiosis even though they are seronegative to B. microti and WA1 antigen.

Bacteriophage E specifically recognizes and infects strains of Escherichia coli which display the alpha-2,8-linked polysialic acid K1 capsule. Bacteriophage E endosialidase, which is thought to be responsible for initial absorption of the phage to the host bacterium, was purified, and the N-terminal amino acid sequences of the polypeptide monomer and cyanogen bromide fragments were determined. Synthetic oligonucleotide probes were designed from the N-terminal amino acid sequences and used to identify restriction fragments of bacteriophage E DNA encoding the endosialidase. The primary nucleotide sequence of the bacteriophage E endosialidase gene contains an open reading frame encoding a 90 kDa polypeptide which is processed to give a mature 74 kDa protein. The native enzyme is probably a trimer of identical 74 kDa subunits. In the bacteriophage E genome the K1E endosialidase open reading frame is preceded by a putative upstream promoter region with homology to a bacteriophage SP6 promoter. A central region of 500 amino acids of the deduced protein sequence of the K1E endosialidase was found to have 84% identity to K1F endosialidase. Both endosialidases contain two copies of a sialidase sequence motif common to many bacterial and viral sialidases. These sequences flank the region of greatest identity between the two endosialidase forms, which suggests that this central domain is involved in binding and hydrolysis of the polysialic acid substrate.

Random amplified polymorphic DNA (RAPD) analysis using 10-mer oligonucleotide primers efficiently differentiated sugarcane cultivars and proved suitable for detecting gross genetic change such as that which can occur in sugarcane subjected to prolonged tissue culture, for example in protoplast-derived callus. However, RAPD analysis was not sufficiently sensitive to detect smaller genetic changes that occur during sugarcane genetic transformation. The length of DNA scored for polymorphism per primer averaged 13.2 kb, or 0.0001% of the typical sugarcane genome size of 1.2 × 107 kb (2C). RAPD analysis of sugarcane plants regenerated from embryogenic callus revealed very few polymorphisms, indicating that gross genetic change is infrequent during this tissue culture procedure, although epigenetic effects result in transient morphological changes in regenerated plants. More sensitive variations on the RAPD technique may increase the practicality of DNA-based screening of regenerated plant lines to reveal somaclonal variants.

Two solid bacteriologic media were compared for cultivating Aeromonas spp. from piscine sources: the Rimler-Shotts (RS) medium and a starch-glutamate-ampicillin-penicillin-based medium (SGAP-10C) used for the recovery of Aeromonas spp. from water samples. The selective and differential capacities of the media were assessed March through October 1992 by recovery rate and phenotype of 99 isolates representing 15 genera of bacteria. Recovery frequency of Aeromonas spp. (n = 62) was similar at 97% on RS and 95% on SGAP-10C. The SGAP-10C medium proved to be more specific than RS toward Aeromonas species (P < or = 0.005). Use of SGAP-10C at 24 C for 48 hr offers a better choice for the laboratory recovery of Aeromonas spp. from clinical fish specimens.

We have studied the interaction of small unilamellar liposomes containing zinc(II)-phthalocyanine (Zn-Pc) with human plasma lipoproteins. High-, low- and very low-density lipoproteins (HDL, LDL and VLDL), were purified from plasma and combined in amounts reflecting their natural abundance in plasma. After short periods of incubation at 37 degrees C, the bulk of Zn-Pc was incorporated into HDL and LDL; very little 14C-labelled palmitoyl oleoyl phosphocholine, the most abundant phospholipid in the formulation, was associated with lipoproteins. When liposomes were incubated in pooled plasma, 73%-85% of Zn-Pc and 27%-34% of radiolabelled phospholipid were recovered with HDL and LDL, indicating a possible role for plasma lipid transfer proteins in the incorporation of phospholipid into lipoproteins. Some Zn-Pc was also found in association with VLDL. The buoyant density of Zn-Pc liposomes increased in a dose-dependent fashion when the particles were incubated with plasma, and it is suggested that this was due, at least in part, to opsonization of liposomes by plasma proteins.

The interaction of human low density lipoprotein (LDL) and small unilamellar liposomes containing the photosensitiser zinc(II)-phthalocyanine (Zn-Pc) was studied in vitro to determine if Zn-Pc could be directly incorporated into the lipoprotein in the absence of other serum components. Incubation of LDL with increasing concentrations of liposomes resulted in a progressive increase in the net negative charge of LDL as determined by agarose gel electrophoresis and both Zn-Pc and liposomal phospholipid were incorporated into the modified LDL particles. Gel chromatography experiments indicated an increase in the molecular mass of modified LDL and immunoaffinity chromatography provided evidence that apoprotein B epitopes on modified LDL were unable to bind to antibody. The study indicated that the liposomal components could be selectively incorporated into LDL by a process that did not appear to involve either aggregation or fusion of particles.

We have attempted to direct low molecular weight compounds to the liver via the internalizing asialoglycoprotein receptor on parenchymal cells by conjugation to a monomeric triantennary galactosyl cluster. Acetate and a hypolipidaemic ansamycin were derivatized and the biodistribution of the conjugates was determined 250 sec and 30 min after administration to Wistar rats. The ansamycin conjugate (CGH46) was rapidly cleared from the circulation by the liver; after 250 sec, 64% of the radiolabelled dose was found in the liver compared to 18% in the blood. However, the distribution of the conjugate did not differ significantly from that of unconjugated ansamycin (CGH45). Tris-galactosyl acetate showed no capacity to localize in the liver, with only 2% recovered from that organ 250 sec after administration compared to 38% in the blood and 13-18% in the kidneys, skin and muscle. Extraction efficiency of CGH46 by isolated perfused rat livers was almost 20% of the administered dose and this value was not significantly changed by co-administration of specific inhibitors of the uptake process. It is concluded that derivatization of low molecular weight molecules with monomeric triantennary galactosyl residues is unlikely to increase their affinity for the liver.

Certain classes of the antibacterial agent rifamycin SV have recently been shown to possess marked hypolipidaemic activity. An acyclic oxazolylrifamycin has been prepared and its hypolipidaemic properties evaluated; it was found to be significantly more potent when administered orally to Wistar and Sprague-Dawley rats than other previously described rifamycin hypolipidaemics. The plasma decay rate of a bolus of intravenously administered 125I-LDL (low density lipoprotein) was significantly greater in treated rats than in rats receiving vehicle alone, compatible with a drug-induced increase in LDL catabolism. A bolus of radiolabelled drug was rapidly removed from the circulation by the liver, the presumed target organ. Compound 3 constitutes the first example of a new class of acyclic hypolipidaemic ansamycins.

Exposure of gram-negative bacteria to a source of antibody and complement frequently results in efficient cell killing; this effect follows the deposition of C5b-9 membrane attack complexes onto the bacterial surface. This review examines the molecular and physiological events culminating in C5b-9 deposition and cell death and advances an hypothesis that may account for cytoplasmic-membrane damage following complement-mediated perturbation of the outer membrane.

Murine monoclonal antibodies specific for neoepitopes expressed by C9 incorporated into membrane attack complexes and by membrane-bound C3b and iC3b have been prepared and characterised. These reagents were used to determine the extent and locus of complement activation in synovial-tissues obtained from patients with rheumatoid arthritis and osteoarthritis. In the four rheumatoid arthritis patients there was extensive deposition of C3 activation products and C5b-9 complexes onto the synovial membrane and the pattern of deposition of both neoantigens in serial tissue sections was very similar. There was less extensive staining for C3 and, particularly, C9 neoepitopes on the apical surface of vessel endothelia. In two of four osteoarthritic patients a similar pattern of C3 and C9 neoepitope deposition was found; in the remaining patients no C5b-9 could be located. Synovial vessel walls, but not synovial cells, from both groups of patients stained extensively for the complement regulatory protein CD59. In synovial membranes from patients with osteoarthritis, C9 appeared to be present predominantly in SC5b-9 complexes whereas in rheumatoid arthritis patients no evidence of S-protein incorporation into membrane attack complexes could be demonstrated, suggesting that in rheumatoid arthritis there is damage to the synovial membrane as a result of complement activation and C5b-9 deposition.

The relationship between the phagocytic uptake of latex microspheres (1.1 microns diam.) and the mobility of rat polymorphonuclear leucocytes (PMNLs) has been investigated in-vitro. The movement of PMNLs was found to be independent of the uptake of particles but about half of the PMNLs were not receptive to the chemo-attractant N-formylmethionyleucylphenylalanine. However, the uptake of particles was greater when particle/cell ratios were high and the greatest number of particles were carried into a cellulose nitrate filter by chemotaxis at the highest particle/cell ratio of 30:1.