Aflatoxins are highly toxic carcinogens that detrimentally influence profitability of agriculture and the health of humans and domestic animals. Several phylogenetically distinct fungi within
...Aspergillus
section
Flavi
have S-morphology (average sclerotial size < 400 μm), and consistently produce high concentrations of aflatoxins in crops. S-morphology fungi have been implicated as important etiologic agents of aflatoxin contamination in the United States (US), but little is known about the diversity of these fungi. The current study characterized S-morphology fungi (
n
= 494) collected between 2002 and 2017, from soil and maize samples, in US regions where aflatoxin contamination is a perennial problem. Phylogenetic analyses based on sequences of the calmodulin (1.9 kb) and nitrate reductase (2.1 kb) genes resolved S-morphology isolates from the US into four distinct clades: (1)
Aspergillus flavus
S-morphotype (89.7%); (2)
Aspergillus agricola
sp. nov. (2.4%); (3)
Aspergillus texensis
(2.2%); and (4)
Aspergillus toxicus
sp. nov. (5.7%). All four S-morphology species produced high concentrations of aflatoxins in maize at 25, 30, and 35°C, but only the
A. flavus
S-morphotype produced unacceptable aflatoxin concentrations at 40°C. Genetic typing of
A. flavus
S isolates using 17 simple sequence repeat markers revealed high genetic diversity, with 202 haplotypes from 443 isolates. Knowledge of the occurrence of distinct species and haplotypes of S-morphology fungi that are highly aflatoxigenic under a range of environmental conditions may provide insights into the etiology, epidemiology, and management of aflatoxin contamination in North America.
Aflatoxins are carcinogenic metabolites produced primarily by fungi within
section
. These fungi infect a wide range of crops in warm regions. Molecular phylogenetic analyses of fungi with S ...morphology (average sclerotium size < 400 µm) within section
collected from across the United States (US) resulted in the discovery of a novel aflatoxin-producing species,
.
was isolated from maize grown in Arkansas, Louisiana, and Texas, and from soils cropped to maize in Texas.
produces sparse conidia and abundant sclerotia on various culture media, and on maize. Physiological studies have revealed optimal growth on culture media at 35 °C. All isolates of
produced B and G aflatoxins, cyclopiazonic acid and aspergillic acid.
and
S strain morphotypes produced similar concentrations of total aflatoxins on maize (
> 0.05). Phylogenetic analyses of aflatoxin-producers based on partial gene sequences of the β-tubulin (0.9 kb), calmodulin (1.2 kb), and nitrate reductase (2.1 kb) genes placed
in a highly supported monophyletic clade closely related to
and a previously reported unnamed lineage designated Lethal Aflatoxicosis Fungus.
Coccidioides spp. are mammalian fungal pathogens endemic to the Southwestern US and other desert regions of Mexico, Central and South America, with the bulk of US infections occurring in California ...and Arizona. In the soil, Coccidioides grows in a hyphal form that differentiates into 3-5 micron asexual spores (arthroconidia). When arthroconidia are inhaled by mammals they undergo a unique developmental transition from polar hyphal growth to isotropic expansion with multiple rounds of nuclear division, prior to segmentation, forming large spherules filled with endospores. Very little is understood about the molecular basis of spherule formation. Here we characterize the role of the conserved transcription factor Ryp1 in Coccidioides development. We show that Coccidioides Δryp1 mutants have altered colony morphology under hypha-promoting conditions and are unable to form mature spherules under spherule-promoting conditions. We analyze the transcriptional profile of wild-type and Δryp1 mutant cells under hypha- and spherule-promoting conditions, thereby defining a set of hypha- or spherule-enriched transcripts ("morphology-regulated" genes) that are dependent on Ryp1 for their expression. Forty percent of morphology-regulated expression is Ryp1-dependent, indicating that Ryp1 plays a dual role in both hyphal and spherule development. Ryp1-dependent transcripts include key virulence factors such as SOWgp, which encodes the spherule outer wall glycoprotein. Concordant with its role in spherule development, we find that the Δryp1 mutant is completely avirulent in the mouse model of coccidioidomycosis, indicating that Ryp1-dependent pathways are essential for the ability of Coccidioides to cause disease. Vaccination of C57BL/6 mice with live Δryp1 spores does not provide any protection from lethal C. posadasii intranasal infection, consistent with our findings that the Δryp1 mutant fails to make mature spherules and likely does not express key antigens required for effective vaccination. Taken together, this work identifies the first transcription factor that drives mature spherulation and virulence in Coccidioides.
Magnaporthe grisea is the most destructive pathogen of rice worldwide and the principal model organism for elucidating the molecular basis of fungal disease of plants. Here, we report the draft ...sequence of the M. grisea genome. Analysis of the gene set provides an insight into the adaptations required by a fungus to cause disease. The genome encodes a large and diverse set of secreted proteins, including those defined by unusual carbohydrate-binding domains. This fungus also possesses an expanded family of G-protein-coupled receptors, several new virulence-associated genes and large suites of enzymes involved in secondary metabolism. Consistent with a role in fungal pathogenesis, the expression of several of these genes is upregulated during the early stages of infection-related development. The M. grisea genome has been subject to invasion and proliferation of active transposable elements, reflecting the clonal nature of this fungus imposed by widespread rice cultivation.
, the primary causal agent for aflatoxin contamination on crops, consists of isolates with two distinct morphologies: isolates of the S morphotype produce numerous
mall sclerotia and lower numbers of ...conidia while isolates of the L morphotype produce fewer
arge sclerotia and abundant conidia. The morphotypes also differ in aflatoxin production with S isolates consistently producing high concentrations of aflatoxin, whereas L isolates range from atoxigenic to highly toxigenic. The production of abundant sclerotia by the S morphotype suggests adaptation for long-term survival in the soil, whereas the production of abundant conidia by the L morphotype suggests adaptation for aerial dispersal to the phyllosphere. To identify genomic changes that support differential niche adaption, the sequences of three S and three L morphotype isolates were compared. Differences in genome structure and gene content were identified between the morphotypes. A >530 kb inversion between the morphotypes affect a secondary metabolite gene cluster and a cutinase gene. The morphotypes also differed in proteins predicted to be involved in carbon/nitrogen metabolism, iron acquisition, antimicrobial defense, and evasion of host immunity. The S morphotype genomes contained more intact secondary metabolite clusters indicating there is higher selection pressure to maintain secondary metabolism in the soil and that it is not limited to aflatoxin production. The L morphotype genomes were enriched in amino acid transporters, suggesting efficient nitrogen transport may be critical in the nutrient limited phyllosphere. These findings indicate the genomes of the two morphotypes differ beyond developmental genes and have diverged as they adapted to their respective niches.
Aflatoxins (AF) are hepatocarcinogenic metabolites produced by several
species. Crop infection by these species results in aflatoxin contamination of cereals, nuts, and spices. Etiology of aflatoxin ...contamination is complicated by mixed infections of multiple species with similar morphology and aflatoxin profiles. The current study investigates variation in aflatoxin production between two morphologically similar species that co-exist in West Africa,
and
. Consistent distinctions in aflatoxin production during liquid fermentation were discovered between these species. The two species produced similar concentrations of AFB
in defined media with either urea or ammonium as the sole nitrogen source. However, production of both AFB
and AFG
were inhibited (
< 0.001) for
in a yeast extract medium with sucrose. Although production of AFG
by both species was similar in urea,
produced greater concentrations of AFG
in ammonium (
= 0.039). Based on these differences, a reliable and convenient assay for differentiating the two species was designed. This assay will be useful for identifying specific etiologic agents of aflatoxin contamination episodes in West Africa and other regions where the two species are sympatric, especially when phylogenetic analyses based on multiple gene segments are not practical.
•There is need for a vaccine to prevent coccidioidomycosis in dogs and humans.•Δcps1, an avirulent, live vaccine, appeared safe and well-tolerated in dogs.•Prime plus boost provided significant ...reduction in fungal burden and disease scores.•Efficacious and well-tolerated, Δcps1 is targeted for human studies as well.
Coccidioidomycosis is a significant health problem of dogs and humans in endemic regions, especially California and Arizona in the U.S. Both species would greatly benefit from a vaccine to prevent this disease. A live avirulent vaccine candidate, Δcps1, was tested for tolerability and efficacy to prevent pulmonary coccidioidomycosis in a canine challenge model. Vaccine injection-site reactions were transient and there were no systemic effects observed. Six of seven vaccine sites tested and all draining lymph nodes were sterile post-vaccination. Following infection with Coccidioides posadasii, strain Silveira, arthroconidia into the lungs, dogs given primary and booster vaccinations had significantly reduced lung fungal burdens (P = 0.0003) and composite disease scores (P = 0.0002) compared to unvaccinated dogs. Dogs vaccinated once had fungal burdens intermediate between those given two doses or none, but disease scores were not significantly different from unvaccinated (P = 0.675). Δcps1 was well-tolerated in the dogs and it afforded a high level of protection when given as prime and boost. These results drive the Δcps1 vaccine toward a licensed veterinary vaccine and support continued development of this vaccine to prevent coccidioidomycosis in humans.
Murine infections with most Coccidioides spp. strains are lethal by 3 weeks, limiting the study of immune responses. Coccidioides posadasii, strain 1038 (Cp1038), while slowly lethal, resulted in ...protracted survival of C57BL/6 (B6) mice. In resistant (B6D2)F1/J mice, lung fungal burdens stabilized by week 4 without progression through week 16, better modeling human coccidioidal infections after their immunologic control. Immunodeficient tumor necrosis factor (Tnf) α knockout (KO) and interferon (Ifn) γ receptor 1 (Ifn-γr1) KO mice survived a median of 22.5 and 34 days, compared with 70 days in B6 mice (P = .001 and P < .01, respectively), though 14-day lung fungal burden studies showed little difference between Ifn-γr1 KO and B6 mice. B6 mice showed peak concentrations of key inflammatory lung cytokines, including interleukin 6, 23, and 17A, Tnf-α, and Ifn-γ, only after 4 weeks of infection. The slower progression in B6 and the acquired fungal burden stability in B6D2 mice after Cp1038 infection greatly increases the array of possible immunologic studies.
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