The development and multiple uses of a standardised bioassay method to select hypocrealean fungi for biological control of aphids

A technically standardised bioassay method was designed, evaluated and used to assess virulence and host range of hypocrealean fungi against aphids. A track mounted sprayer was used to apply conidia because hand held versions of the same sprayer can be used for field applications, thereby allowing the outcome from laboratory experiments to predict activity in the field accurately. Eighteen fungal isolates were assessed in single concentration bioassays against the black bean aphid Aphis fabae Scopoli. Isolates comprised commercially available mycoinsecticides (based on Beauveria bassiana and Lecanicillium longisporum) and isolates of B. bassiana, Lecanicillium spp., Paecilomyces fumosoroseus and Metarhizium anisopliae. Aphid mortality was in excess of 80% for 15 isolates, and HRI 1.72 (L. longipsorum), Z11 (P. fumosoroseus), Mycotech strain GHA (B. bassiana) and ARSEF 2879 (B. bassiana) were studied further. Multiple concentration bioassays identified HRI 1.72 as the most virulent isolate against A. fabae with significantly smaller LC₅₀ and LT₅₀ values compared to other isolates. A precise LC₅₀ value (2.95 × 10² conidia ml⁻¹) was calculated for HRI 1.72 using a second multiple concentration assay with smaller concentrations of conidia. The four isolates were applied at a single concentration (1 × 10⁸ conidia ml⁻¹) against Myzus persicae, A. fabae, Acyrthosiphon pisum, Metopolophium dirhodum, Sitobion avenae and Rhopalosiphum padi. A ranking of aphid susceptibility was obtained, such that S. avenae > M. persicae, A. pisum, A. fabae > R. padi. Results indicate the importance of standardising bioassay methods to reduce bioassay variability without compromising the ability to use the bioassay to investigate fungus–host interactions under varying abiotic and biotic conditions.     

Chaetosphaeria tortuosa, the newly discovered teleomorph of Menispora tortuosa, with a key to known Menispora species

Chaetosphaeria tortuosa is described as the newly discovered teleomorph of Menispora tortuosa, based on specimens from Canada and the Czech Republic, and single spore isolations from both morphs. The fungus produces superficial, more or less globose, papillate, dark brown to black smooth perithecia (200–)220–250 × (220–)230–260 μm. The asci are unitunicate, 8-spored, cylindrical-fusiform, (110–)120–133(–145) × 12–14 with a distinct apical, nonamyloid annulus 1–1.5 μm high, 3.5–4 μm wide. The ascospores are fusiform, 19–24 × 5–6 μm, hyaline, 3-septate, smooth, and 2-seriate in the ascus. The morphology of the teleomorph and anamorph are similar to that of C. ovoidea (anamorph: M. glauca), differing in dimensions of asci and ascospores, and in the disposition and morphology of the phialides of the anamorphs. The generic concept and phylogeny of Menispora is briefly discussed, and a key to the 11 species currently accepted in the genus is provided.     

Selection of Beauveria bassiana isolates for control of the whiteflies Bemisia tabaci and Trialeurodes vaporariorum on the basis of their virulence, thermal requirements, and toxicogenic activity

As part of a 3-fold approach to select potential mycoinsecticides for whitefly control, we evaluated infectivity, thermal requirements, and toxicogenic activity of the entomopathogenic fungus Beauveria bassiana (Ascomycota: Clavicipitaceae) under laboratory conditions. Twenty-five native B. bassiana isolates and a commercially available mycoinsecticide (based on B. bassiana) were evaluated for virulence to fourth instar nymphs of sweetpotato whitefly, Bemisia tabaci, and greenhouse whitefly, Trialeurodes vaporariorum, at a concentration of 1 × 10⁷ conidia/ml. All isolates were pathogenic for both whitefly species, whereas mortality rates varied from 3 to 85%. A second series of bioassays was conducted on 10 selected isolates using four 10-fold concentrations ranging from 1 × 10⁵ to 1 × 10⁸ conidia/ml. Median lethal concentrations (LC₅₀) of the four most virulent isolates varied from 1.1 × 10⁵ to 6.2 × 10⁶ conidia/ml and average survival time (AST) of treated nymphs from 5.9 to 7.4 days. T. vaporariorum were significantly more susceptible to all B. bassiana isolates than B. tabaci. The thermal biology of the eight most virulent isolates to both whitefly species was investigated at six temperatures (10–35 °C). The colony radial growth rate was estimated from the slope of the linear regression of colony radius on time and data were then fitted to a modified generalized β function that accounted for 90.5–99.3% of the data variance. Optimum temperatures for extension rate ranged from 23.1 to 27.1 °C, whereas maximum temperatures for fungal growth varied from 31.8 to 36.6 °C. On the basis of their virulence and thermal requirements, three isolates showed promise as candidates for whitefly management in Mediterranean greenhouses. Whilst in vitro production of macromolecular compounds toxic to Galleria mellonella larvae was not a requisite for virulence, ASTs of larvae injected with Sephadex G-25 fractions from candidate isolates ranged from 1.4 to 3.7 days compared with 5–6 days for non-toxic G-25 fractions. In addition, proteinase K treatment significantly reduced their toxic activity suggesting that they were proteins and revealing the potential of these isolates to be further improved through biotechnology to kill the pest more quickly.     

Molecular and morphological characterization of Leveillula (Ascomycota: Erysiphales) on monocotyledonous plants

Leveillula on monocotyledonous plants have been recorded as L. taurica by several authors, whereas the fungus on Allium has been described as an independent species, namely L. allii, by some authors. We sequenced ca 600 bp of the rDNA ITS region for two Leveillula specimens from Allium and Polianthes (both from monocotyledons) and compared them with several already published sequences from Leveillula isolates from dicotyledons. Pair-wise percentages of sequence divergences were calculated for all Leveillula isolates. The ITS sequence of the Polianthes isolate was identical to L. taurica on Helianthus and Vicia. The sequence of the Allium isolate was 99.5 % identical to L. taurica on Euphorbia, Haplophylum, Peganum, etc. These results suggest close relationships between monocot and dicot pathogenic Leveillula species. The identity between two monocot isolates was 98.4 %. Phylogenetic analysis revealed that the two monocot isolates do not group into a clade together. This result suggests that Leveillula acquired parasitism to monocots at least twice independently.     

Role of symbiotic and non-symbiotic bacteria in carbon dioxide production from hosts infected with Steinernema riobrave

Entomopathogenic nematodes of the family Steinernematidae and their mutualistic bacteria (Xenorhabdus spp.) are lethal endoparasites of insects. We hypothesized that growth of the nematode’s mutualistic bacteria in the insect host may contribute to the production of cues used by the infective juveniles (IJs) in responding to potential hosts for infection. Specifically, we tested if patterns of bacterial growth could explain differences in CO₂ production over the course of host infection. Growth of Xenorhabdus cabanillasii isolated from Steinernema riobrave exhibited the characteristic exponential and stationary growth phases. Other non-nematode symbiotic bacteria were also found in infected hosts and exhibited similar growth patterns to X. cabanillasii. Galleria mellonella larvae infected with S. riobrave produced two distinct peaks of CO₂ occurring at 25.6–36 h and 105–161 h post-infection, whereas larvae injected with X. cabanillasii alone showed only one peak of CO₂, occurring at 22.8–36.2 h post-injection. Tenebrio molitor larvae infected with S. riobrave or injected with bacteria alone exhibited only one peak of CO₂ production, which occurred later during S. riobrave infection (41.4–64.4 h post-infection compared to 20.4–35.9 h post-injection). These results indicate a relationship between bacterial growth and the first peak of CO₂ in both host species, but not for the second peak exhibited in G. mellonella.     

Novel hosts of the Eucalyptus canker pathogen Chrysoporthe cubensis and a new Chrysoporthe species from Colombia

The pathogen Chrysoporthe cubensis (formerly Cryphonectria cubensis) is best known for the important canker disease that it causes on Eucalyptus species. This fungus is also a pathogen of Syzygium aromaticum (clove), which is native to Indonesia, and like Eucalyptus, is a member of Myrtaceae. Furthermore, C. cubensis has been found on Miconia spp. native to South America and residing in Melastomataceae. Recent surveys have yielded C. cubensis isolates from new hosts, characterized in this study based on DNA sequences for the ITS and β-tubulin gene regions. These hosts include native Clidemia sericea and Rhynchanthera mexicana (Melastomataceae) in Mexico, and non-native Lagerstroemia indica (Pride of India, Lythraceae) in Cuba. Isolates from these hosts and areas group in the sub-clade of C. cubensis accommodating the South American collections of the fungus. This sub-clade also includes isolates recently collected from Eucalyptus in Cuba, which are used to epitypify C. cubensis. New host records from Southeast Asia include exotic Tibouchina urvilleana from Singapore and Thailand and native Melastoma malabathricum (Melastomataceae) in Sumatra, Indonesia. Consistent with their areas of occurrence isolates from the latter collections group in the Asian sub-clade of C. cubensis. DNA sequence comparisons of isolates from Tibouchina lepidota in Colombia revealed that they represent a new sub-clade within the greater Chrysoporthe clade. Isolates in this clade are described as Chrysoporthe inopina sp. nov., based on distinctive morphological differences.     

Field trials against Hoplia philanthus (Coleoptera: Scarabaeidae) with a combination of an entomopathogenic nematode and the fungus Metarhizium anisopliae CLO 53

The white grub, Hoplia philanthus Füessly (Coleoptera: Scarabaeidae), is a major pest of turf and ornamental plants in Belgium. Previously, the combination of lethal concentration of the entomopathogenic nematodes Heterorhabditis megidis or Steinernema glaseri with the entomopathogenic fungus Metarhizium anisopliae (strain CLO 53) caused additive or synergistic mortality to third-instar H. philanthus in the laboratory and greenhouse. In this present study, we examined this interaction under field conditions and compared a combination of a commercial formulation of Heterorhabditis bacteriophora (Nema-green^®) and M. anisopliae. Controls were M. anisopliae, chlorpyrifos (Dursban 5 Granules) and H. bacteriophora. Field applications (surface or subsurface) were made against a mixed population of second/third-instar H. philanthus at a sport field and lawn infested in the province of West-Flanders. In both trials, the combination of M. anisopliae with H. bacteriophora at 5 × 10¹² conidia/ha +2.5 × 10⁹ infective juveniles/ha resulted in additive or synergistic effects, causing more than 95% grub mortality when the nematodes was applied 4 weeks after the application of fungus. However, application of nematode, chlorpyrifos or fungus alone provided 39–66%, 42–60% (surface) and 33–76%, 82–100% or 37–65%, (subsurface) control of H. philanthus. We concluded that the pathogen combinations we tested are compatible elements of integrated pest management and are likely to improve control of H. philanthus larvae and perhaps other insect pests beyond what is expected from single application of the pathogen.     

Effect of Host Plant on the Potential ofPaecilomyces fumosoroseus(Deuteromycotina: Hyphomycetes) for Controlling the Silverleaf Whitefly,Bemisia argentifolii(Homoptera: Aleyrodidae) in Greenhouses

We determined host plant effect on susceptibility of the silverleaf whitefly,Bemisia argentifolii, to the entomopathogenic fungusPaecilomyces fumosoroseus. Whiteflies were reared on three vegetable species (cucumber, cabbage, and tomato) and three cultivars of tomato (Heatwave, Better Boy, and Rutgers). Second instars were sprayed with 5 × 10⁴conidia/cm²ofPfr97, aP. fumosoroseusstrain, used as a microbial control agent of whiteflies. Trials were conducted in an experimental greenhouse, where temperature and relative humidity were adjusted to favor infection (22–33°C, and 68–100% RH). Larval susceptibility to fungal infection was high and not significantly affected by the host plant. Mortality was > 70% 1 week after treatment and increased further during the second week. Percentages of cadavers with subsequent production of conidia observed in the greenhouse did not vary significantly either with the host vegetable species (85–93% 7 days after treatment and 99–100% 14 days after treatment), or with the cultivar of tomato (96–97% 7 days after treatment and 99–100% 14 days after treatment). After incubation under optimal laboratory conditions, the percentages based on the total number of sporulating cadavers (includingin situsporulating individuals and cadavers sporulating afterin vitroincubation) were not significantly influenced either by host vegetable or cultivar of tomato. According to the conditions prevailing in the series of experiments with the three vegetable species or in the series of experiments with the three cultivars of tomatoes, the production of newly formed conidia varied from approximately 10,000 to 18,000 conidia/cadaver. However, in both series, there was no significant influence of the host vegetable species or cultivar. The survival of the newly formed conidia harvested 7 days following treatment reached more than 50% but was not affected by host plant. These results indicate thatP. fumosoroseusshows potential as a microbial control agent for controllingB. argentifoliion greenhouse crops.     

Stromatinia cryptomeriae sp. nov., the teleomorph ofGloeosporidina cryptomeriae causing twig blight of Japanese cedar

A new species,Stromatinia cryptomeriae, is described based on a specimen collected in Iwate Prefecture, Japan. It was found on fallen dead branches and twigs of Japanese cedar,Cryptomeria japonica. The morphology of isolates on potato-dextrose agar (PDA), which were obtained from single ascospores ofS. cryptomeriae, was identical withGloeosporidina cryptomeriae, the causal fungus of Japanese cedar twig blight. In an inoculation test using single ascospore isolates, many minute black spots (sclerotioid bodies; sclerotules) and acervuli ofG. cryptomeriae were formed on the necrotic lesions, which developed into typical symptoms of Japanese cedar twig blight. These results show thatStromatinia cryptomeriae is the teleomorph ofG. cryptomeriae. On PDA, the fungus grew over a range of about 1 to 25°C, with the optimum growth at about 15–20°C.     

In vitro evaluation of combination of Trichoderma harzianum and chitosan for the control of sapstain fungi

In vitro assays were undertaken to evaluate the control of two sapstain fungi, Leptographium procerum and Sphaeropsis sapinea by a combination of chitosan or chitosan oligomer and an albino strain of Trichoderma harzianum. Spore germination and hyphal growth of the test fungi were assessed on media amended with chitosan or chitosan oligomer with and without T. harzianum using either simultaneous inoculation with test fungus or inoculation 1, 2, or 3 days after pre-infection with test fungus.There was no mycelial growth of the test fungi regardless of chitosan concentrations used when either L. procerum or S. sapinea was simultaneously inoculated with T. harzianum. However, the dose–response of chitosan or chitosan oligomer on the test fungi was apparent when T. harzianum was not simultaneously inoculated with test fungus but introduced later. There was a greater growth reduction at higher concentrations (0.075–0.1% v/v) of chitosan, and overall chitosan oligomer was more effective than chitosan aqueous solution.Chitosan alone was able to restrict or delay the germination of spores but the combination of chitosan and T. harzianum inhibited spore germination and hence colony formation of test fungi regardless of time delay.     

Differential sensitivity of fungi to lithium chloride in culture media

Forty species of fungi, representing a range of ecological and taxonomic groups, were tested for their ability to grow on agar media amended with lithium chloride (LiCl) at 1.5, 3 and 6 g l⁻¹. Species of Trichoderma varied considerably in their sensitivity to LiCl; at one week on 6 g l⁻¹ LiCl medium, the growth of seven species of Trichoderma was considerably inhibited; however, by three weeks at this level, four of the species tested were able to attain ≥30 % of control growth. Of the seven species tested, an isolate of T. viride was the most sensitive to LiCl in agar. Eleven other imperfect fungi also showed a range of ability to grow on agar amended with LiCl, from total inhibition to complete lack of inhibition. Six ascomycete fungi were greatly inhibited by LiCl at all levels; however, an isolate of Chaetomium globosum was highly tolerant of LiCl. Seven basidiomycete wood-decay fungi were quite sensitive to LiCl in agar, showing total to nearly total inhibition even at the lowest level; however, after three weeks, an isolate of Postia placenta was nearly uninhibited except at 6 g l⁻¹. Five ectomycorrhizal basidiomycete fungi were totally inhibited by all levels of LiCl; however, one ectomycorrhizal imperfect fungus (Cenococcum graniforme) was able to grow at 3 g l⁻¹ and was uninhibited at 1.5 g l⁻¹. Four zygomycete fungus isolates were nearly unaffected in their growth by all levels of LiCl.     

Redheadia quercus gen. et sp. nov., the teleomorph of Mycopappus quercus, the frosty mildew fungus in Quercus acutissima

The teleomorph of Mycopappus quercus causing frosty mildew in Quercus acutissima is described as a new genus and species, Redheadia quercus, in the Sclerotiniaceae. Apothecia sprout from sclerotia on the fallen infected leaves kept for 10 months at 5°C and subsequent incubation at 15°C under diffused room light. Typical zonate lesions and multicellular propagules of M. quercus are produced on Q. acutissima, by mycelial inoculation using an isolate from a single ascospore, confirming the teleomorph–anamorphic connection. No significant differences are observed between cultured colonies of isolates from the ascospore and those from the propagule. Sclerotia and microconidia of the fungus are produced on culture media.     

Molecular characterisation of Halobacillus strains isolated from different medieval wall paintings and building materials in Austria

We previously reported on the detection and isolation of an indigenous population of Halobacillus from salt-damaged medieval wall paintings and building materials of Herberstein castle in St. Johann bei Herberstein in Styria, Austria. Several moderately halophilic, Gram-positive, endospore-forming Halobacillus-like bacteria could be again isolated by conventional enrichment from salt efflorescences collected in the medieval St. Virgil's chapel in Vienna. Comparative 16S rDNA sequence analyses showed that the St. Virgil isolates are most closely related (>98.5% sequence similarity) to Halobacillus trueperi, Halobacillus litoralis, and to our previous halobacilli strains obtained from the castle Herberstein. Based on 16S rDNA sequence analysis, the strains could be clustered in three different groups. Group I: St. Virgil strains S3, S4, S21, and S22 (99.8–100% sequence similarity); group II: Herberstein strains K3-1, I7, and the St. Virgil strain S20 (99.3–99.7% sequence similarity); and group III: Herberstein strains I3, I3A, and I3R (100% sequence similarity). Molecular typing by denaturing gradient gel electrophoresis (DGGE), random amplified polymorphic DNA (RAPD-PCR), and internal transcribed spacer-homoduplex–heteroduplex polymorphism (ITS-HHP) fingerprinting showed that all isolates are typeable by each of the methods. RAPD was the most discriminatory method. With respect to their physiological characteristics—i.e., growth in the presence of 5–20% (w/v) NaCl, no growth in the absence of NaCl, optimum growth at 37 °C in media containing 5–10% (w/v) NaCl, and optimum pH around 7.5–8.0—the St. Virgil isolates resembled our previously isolated strains. However, the St. Virgil strains showed some differences in their biochemical properties. St. Virgil isolates hydrolysed Tween 80, two isolates reduced nitrate, and no isolate liquefied gelatine. The recurrent isolation of halobacilli from salt efflorescences on historic buildings and monuments at two different geographical locations may indicate that this group of bacteria is common in salt-affected ruins.     

Conidial germination and germ tube elongation of Phomopsis amaranthicola and Microsphaeropsis amaranthi on leaf surfaces of seven Amaranthus species: Implications for biological control

Microsphaeropsis amaranthi and Phomopsis amaranthicola are potential biological control agents for several Amaranthus species. In an effort to understand the initial infection processes with these pathogens, a study was conducted of the conidial germination and germ tube length (μm) on the weed leaf surfaces at 21 °C and 28 °C. Weeds included Amaranthus rudis, A. palmeri, A. powellii, A. retroflexus, A. spinosus, A. hybridus, and A. albus. For P. amaranthicola, conidial germination and germ tube length varied among the seven weed species at both temperatures, while for M. amaranthi the differences in germ tube lengths were significant among weed species only at 21 °C. While the conidia of M. amaranthi and P. amaranthicola germinated on the leaf surfaces of all seven weed species, temperature appeared to impact the number and length of germ tubes on the leaf surfaces. The percentage of germinated conidia and the length of germ tubes at both temperatures were often greater for M. amaranthi than for P. amaranthicola. In order for the fungal pathogen to successfully infect and kill a weedy host, conidia must germinate and form a germ tube, two processes that vary with host species and temperature for M. amaranthi and P. amaranthicola. The extent to which successive infection processes, e.g., penetration, invasion and colonization, contribute to host specificity warrants study.     

Antifungal activity of strains of lactic acid bacteria isolated from a semolina ecosystem against Penicillium roqueforti, Aspergillus niger and Endomyces fibuliger contaminating bakery products

Thirty samples of Italian durum wheat semolina and whole durum wheat semolina, generally used for the production of Southern Italy's traditional breads, were subjected to microbiological analysis in order to explore their lactic acid bacteria (LAB) diversity and to find strains with antifungal activity. A total of 125 presumptive LAB isolates (Gram-positive and catalase-negative) were characterized by repetitive extragenic palindromic-PCR (REP-PCR) and sequence analysis of the 16S rRNA gene, leading to the identification of the following species: Weissella confusa, Weissella cibaria, Leuconostoc citreum, Leuconostoc mesenteroides, Lactococcus lactis, Lactobacillus rossiae and Lactobacillus plantarum. The REP-PCR results delineated 17 different patterns whose cluster analysis clearly differentiated W. cibaria from W. confusa isolates. Seventeen strains, each characterized by a different REP-PCR pattern, were screened for their antifungal properties. They were grown in a flour-based medium, comparable to a real food system, and the resulting fermentation products (FPs) were tested against fungal species generally contaminating bakery products, Aspergillus niger, Penicillium roqueforti and Endomyces fibuliger. The results of the study indicated a strong inhibitory activity – comparable to that obtained with the common preservative calcium propionate (0.3% w/v) – of ten LAB strains against the most widespread contaminant of bakery products, P. roqueforti. The screening also highlighted the unexplored antifungal activity of L. citreum, L. rossiae and W. cibaria (1 strain), which inhibited all fungal strains to the same or a higher extent compared with calcium propionate. The fermentation products of these three strains were characterized by low pH values, and a high content of lactic and acetic acids.     

Geographic locality and host identity shape fungal endophyte communities in cupressaceous trees

Understanding how fungal endophyte communities differ in abundance, diversity, taxonomic composition, and host affinity over the geographic ranges of their hosts is key to understanding the ecology and evolutionary context of endophyte–plant associations. We examined endophytes associated with healthy photosynthetic tissues of three closely related tree species in the Cupressaceae (Coniferales): two native species within their natural ranges [Juniperus virginiana in a mesic semideciduous forest, North Carolina (NC); Cupressus arizonica, under xeric conditions, Arizona (AZ)], and a non-native species planted in each site (Platycladus orientalis). Endophytes were recovered from 229 of 960 tissue segments and represented at least 35 species of Ascomycota. Isolation frequency was more than threefold greater for plants in NC than in AZ, and was 2.5 (AZ) to four (NC) times greater for non-native Platycladus than for Cupressus or Juniperus. Analyses of ITS rDNA for 109 representative isolates showed that endophyte diversity was more than twofold greater in NC than in AZ, and that endophytes recovered in AZ were more likely to be host-generalists relative to those in NC. Different endophyte genera dominated the assemblages of each host species/locality combination, but in both localities, Platycladus harboured less diverse and more cosmopolitan endophytes than did either native host. Parsimony and Bayesian analyses for four classes of Ascomycota (Dothideomycetes, Sordariomycetes, Pezizomycetes, Eurotiomycetes) based on LSU rDNA data (ca 1.2 kb) showed that well-supported clades of endophytes frequently contained representatives of a single locality or host species, underscoring the importance of both geography and host identity in shaping a given plant's endophyte community. Together, our data show that not only do the abundance, diversity, and taxonomic composition of endophyte communities differ as a function of host identity and locality, but that host affinities of those communities are variable as well.     

Thermus kawarayensis sp. nov., a new member of the genus Thermus, isolated from Japanese hot springs

A long-rod-shaped thermophilic microorganism, strain KW11, was isolated from a hot springs located in the Kawarayu, Gunma, Japan. Cloning and preliminary sequence analysis of 16S rDNA showed that this isolate belongs to the genus Thermus. The cells were 10–20 m long, about 0.8 m in diameter, and produced no pigment in contrast with most of the Thermus species previously reported. KW11 was an aerobic heterotroph and grew at temperatures ranging from 40–73°C, with optimal growth occurring at 68°C. The pH range for growth was from 5.8–8.9, with optimal growth around pH 7. KW11 was sensitive to ampicillin, penicillin G, kanamycin, and streptomycin. The G+C content of DNA was 69 mol%. The main fatty acids were 16:0 (52.9%), iso-15:0 (22.1%), and iso-17:0 (15.6%). The 16S rDNA sequence of KW11 showed 96.0, 95.8, and 95.4% similarity with the sequences of T. aquaticus, T. igniterrae, and T. thermophilus, respectively, and less than 95% with other Thermus species. The physiological differences and phylogenetic evidence indicated that strain KW11 represents T. kawarayensis, a novel species of the genus Thermus. The type strain is isolate KW11^T (JCM12314, DSM16200).     

Caeoma tsukubaense n. sp., a rhododendron rust fungus of Japan and southern Asia, and its relationship to Chrysomyxa rhododendri

A rust fungus found in Japan on Rhododendron kaempferi, R. kiusianum, and R. dauricum has previously been identified as Chrysomyxa rhododendri. Light and scanning electron microscopy of fresh and herbarium materials of the rust fungus, however, show that the spore surface morphology differs from the urediniospores of C. rhododendri, and the spores are slightly smaller. Furthermore, the DNA sequence of the 5′-end of the large subunit of ribosomal DNA differs from that of C. rhododendri by 3%. Telia have not been found; therefore, it is redescribed as a new anamorphic species, Caeoma tsukubaense. Several specimens from North Korea, Tibet, and Nepal bearing a similar rust fungus are also included in the species.Contribution no.193 from the Laboratory of Plant Parasitic Mycology, Graduate School of Life and Environmental Sciences, University of Tsukuba, Japan     

Pathogenicity and specificity of Neozygites tanajoae and Neozygites floridana (Zygomycetes: Entomophthorales) isolates pathogenic to the cassava green mite

The cassava green mite (CGM), Mononychellus tanajoa, a native of South America was accidentally introduced into Africa where it causes serious crop losses. The possibility of introducing classical biological agents from the native home of CGM into Africa was investigated. Thus, we conducted a series of laboratory assays of the native fungal pathogens, Neozygites tanajoae from Brazil and Neozygites floridana from Colombia and Brazil, and compared them with N. tanajoae isolates from Benin. Infectivity of both fungal species, was assayed against the twospotted spider mite, Tetranychus urticae, and against the red mite, Oligonychus gossypii. Pathogenicity against CGM and host range studies were conducted by transferring adult females of each mite species to leaf discs containing sporulated cadavers with a halo of conidia of each fungal isolate. All isolates caused some degree of infectivity to CGM. None of the isolates of N. floridana and N. tanajoae tested were pathogenic to O. gossypii, and only two isolates infected T. urticae. Most isolates from Brazil were highly virulent and infected only CGM. Sixteen N. tanajoae isolates caused more than 89% mortality and more than 62% of the CGM became mummified. A mummified CGM is characteristically a swollen, brown fungus-killed mite that has great potential to produce conidia. However, high mortality was not always associated with high mummification. The median mummification time ranged from 4.4 to 6.7 days. Five Brazilian isolates caused >75% mummification with a median mummification time <5 days. Isolates that cause high mummification in a short period of time would be more likely to cause epizootics and to establish in the new environment. Therefore, these isolates would be the best candidates for introduction to Africa.     

Growth of dinoflagellates, Ceratium furca and Ceratium fusus in Sagami Bay, Japan: The role of temperature, light intensity and photoperiod

Seasonal changes of field populations and growth rates of two dinoflagellates, Ceratium furca and Ceratium fusus, were examined in the temperate coastal water of Sagami Bay, Japan. Weekly field sampling was conducted from August 2002 to August 2003, and laboratory experiments were also carried out to investigate effects of temperature, irradiance and photoperiod on the growth rates of these two Ceratium species. In the field, the abundances of both species increased significantly from April to August 2003, were gradually decreased from November 2002 and were not observed in January 2003. C. fusus was able to increase at lower temperatures in February 2003 compared to C. furca. In the laboratory, the two species did not grow at <10 °C or >32 °C. The highest specific growth rate of C. furca was 0.72 d⁻¹ at 24 °C and 600 μmol m⁻² s⁻¹. Optimum growth rates (>0.4 d⁻¹) of C. furca were observed at temperatures from 18 to 28 °C and at irradiances from 216 to 796 μmol m⁻² s⁻¹. The highest growth rate of C. fusus was 0.56 d⁻¹ at 26 °C and 216 μmol m⁻² s⁻¹. Optimum growth rates of C. fusus were observed at the same irradiance rage of C. furca, whereas optimum temperature range was narrower (26–28 °C). The growth curves of both species indicated saturation of the growth rates when light intensity was above 216 μmol m⁻² s⁻¹, and did not show photoinhibition at irradiances up to 796 μmol m⁻² s⁻¹. The specific growth rates of both Ceratium species were clearly decreased at L:D = 10:14 relative to those at L:D = 14:10 and L:D = 12:12. The present study indicates the two Ceratium species can adapt to a wide range of temperature and irradiance.