scholarly journals Role of laeA in the Regulation of alb1, gliP, Conidial Morphology, and Virulence in Aspergillus fumigatus

2007 ◽  
Vol 6 (9) ◽  
pp. 1552-1561 ◽  
Author(s):  
Janyce A. Sugui ◽  
Julian Pardo ◽  
Yun C. Chang ◽  
Arno Müllbacher ◽  
Kol A. Zarember ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Survival Data ◽  
Mycelial Growth ◽  
Mrna Level ◽  
Nutritional Requirements ◽  
Wild Type ◽  
Conidial Morphology ◽  
Culture Filtrates ◽  
Data Culture

ABSTRACT The alb1 (pksP) gene has been reported as a virulence factor controlling the pigmentation and morphology of conidia in Aspergillus fumigatus. A recent report suggested that laeA regulates alb1 expression and conidial morphology but not pigmentation in the A. fumigatus strain AF293. laeA has also been reported to regulate the synthesis of secondary metabolites, such as gliotoxin. We compared the role of laeA in the regulation of conidial morphology and the expression of alb1 and gliP in strains B-5233 and AF293, which differ in colony morphology and nutritional requirements. Deletion of laeA did not affect conidial morphology or pigmentation in these strains, suggesting that laeA is not involved in alb1 regulation during conidial morphogenesis. Deletion of laeA, however, caused down-regulation of alb1 during mycelial growth in a liquid medium. Transcription of gliP, involved in the synthesis of gliotoxin, was drastically reduced in B-5233laeAΔ, and the gliotoxin level found in the culture filtrates was 20% of wild-type concentrations. While up-regulation of gliP in AF293 was comparable to that in B-5233, the relative mRNA level in AF293laeAΔ was about fourfold lower than that in B-5233laeAΔ. Strain B-5233laeAΔ caused slower onset of fatal infection in mice relative to that with B-5233. Histopathology of sections from lungs of infected mice corroborated the survival data. Culture filtrates from B-5233laeAΔ caused reduced death in thymoma cells and were less inhibitory to a respiratory burst of neutrophils than culture filtrates from B-5233. Our results suggest that while laeA is not involved in the regulation of alb1 function in conidial morphology, it regulates the synthesis of gliotoxin and the virulence of A. fumigatus.

The role of the α7nAChR-mediated cholinergic anti-inflammatory pathway in Hirschsprung associated enterocolitis

2020 ◽  
Author(s):  
Feng Chen ◽  
Xiaoyu Wei ◽  
Xiaohua Chen ◽  
Lei Xiang ◽  
Xinyao Meng ◽  
...  
Keyword(s):  
Western Blotting ◽  
Mrna Level ◽  
Underlying Mechanism ◽  
Mrna Levels ◽  
Wild Type ◽  
Inflammatory Pathway ◽  
Anti Inflammatory ◽  
Phosphorylated Stat3 ◽  
Morphology Changes

Abstract Background To investigate the role and the underlying mechanism of the α7nAChR-mediated cholinergic anti-inflammatory pathway in the pathogenesis of Hirschsprung(HSCR) associated enterocolitis（HAEC）. Methods Experimental group:twenty-one-day-old Ednrb-/- mice were selected (n=10), with comparable-age wild type(Ednrb+/+) mice controls (n=10). Intestinal samples were collected. The experimental colons were divided into narrow and dilated segments according to morphology changes. The control colons were divided into distal and proximal segments.Colon HE staining was used to judge HAEC.Acetylcholine levels in colon was measured using enzyme-linked immunosorbent assays. Detected phosphorylated Jak2 (p-Jak2), Jak2, phosphorylated Stat3 (p-Stat3), Stat3, phosphorylated IκBα (p-IκBα) and IκBα were studied by Western blotting; mRNA levels of Jak2, Stat3, and IκBα were detected by RT-qPCR. Results Colon HE staining indicated that HAEC mainly occured in the dilated segments of HSCR mice (Ednrb-/- mice) (EDNRB-P).Acetylcholine content in EDNRB-P was significantly lower than that in the narrow segments (EDNRB-D) (P<0.05). Western blotting showed that the Jak2, p-Jak2, Stat3 and p-Stat3 levels in EDNRB-D were significantly higher than those in EDNRB-P (P<0.05). The p-IκBα and IκBα levels in EDNRB-P were significantly higher than those in EDNRB-D(P<0.05). The mRNA levels of Jak2 and Stat3 in EDNRB-D were higher than those in EDNRB-P, but the IκBα mRNA level was significantly lower than that in EDNRB-P (P<0.05). Conclusions During HAEC, the inflammation in the dilated segment was more severe ,while in the narrow segment there was no obvious inflammatory reaction and the content of acetylcholine was higher, which was associated with the α7nAChR-mediated cholinergic anti-inflammatory pathway.

scholarly journals Toll-Like Receptor 9 Modulates Immune Responses to Aspergillus fumigatus Conidia in Immunodeficient and Allergic Mice

2008 ◽  
Vol 77 (1) ◽  
pp. 108-119 ◽  
Author(s):  
Hemanth Ramaprakash ◽  
Toshihiro Ito ◽  
Theodore J. Standiford ◽  
Steven L. Kunkel ◽  
Cory M. Hogaboam
Keyword(s):  
Immune Responses ◽  
Aspergillus Fumigatus ◽  
Inflammatory Responses ◽  
Fungal Growth ◽  
Lower Airway ◽  
Interleukin 17 ◽  
Toll Like Receptor ◽  
Wild Type ◽  
Immunodeficient Mice

ABSTRACT The role of Toll-like receptor 9 (TLR9) in antifungal responses in the immunodeficient and allergic host is unclear. We investigated the role of TLR9 in murine models of invasive aspergillosis and fungal asthma. Neutrophil-depleted TLR9 wild-type (TLR9+/+) and TLR9-deficient (TLR9−/−) mice were challenged with resting or swollen Aspergillus fumigatus conidia and monitored for survival and lung inflammatory responses. The absence of TLR9 delayed, but did not prevent, mortality in immunodeficient mice challenged with resting or swollen conidia compared to TLR9+/+ mice. In a fungal asthma model, TLR9+/+ and TLR9−/− mice were sensitized to soluble A. fumigatus antigens and challenged with resting or swollen A. fumigatus conidia, and both groups of mice were analyzed prior to and at days 7, 14, and 28 after the conidium challenge. When challenged with resting conidia, TLR9−/− mice exhibited significantly lower airway hyper-responsiveness compared to the TLR9+/+ groups. In contrast, A. fumigatus-sensitized TLR9−/− mice exhibited pulmonary fungal growth at days 14 and 28 after challenge with swollen conidia, a finding never observed in their allergic wild-type counterparts. Increased fungal growth in allergic TLR9−/− mice correlated with markedly decreased dectin-1 expression in whole lung samples and isolated dendritic cell populations. Further, whole lung levels of interleukin-17 were lower in allergic TLR9−/− mice compared to similar TLR9+/+ mice. Together, these data suggest that TLR9 modulates pulmonary antifungal immune responses to swollen conidia, possibly through the regulation of dectin-1 expression.

Role of α2-macroglobulin in fever and cytokine responses induced by lipopolysaccharide in mice

2002 ◽  
Vol 283 (1) ◽  
pp. R218-R226 ◽  
Author(s):  
Alexander V. Gourine ◽  
Valery N. Gourine ◽  
Yohannes Tesfaigzi ◽  
Nathalie Caluwaerts ◽  
Fred Van Leuven ◽  
...  
Keyword(s):  
Mrna Level ◽  
Physiological Role ◽  
Mrna Levels ◽  
Wild Type ◽  
Cytokine Responses ◽  
Α2 Macroglobulin ◽  
Tnf Α ◽  
Factor Α ◽  
Necrosis Factor

α2-Macroglobulin (α2M) is not only a proteinase inhibitor in mammals, but it is also a specific cytokine carrier that binds pro- and anti-inflammatory cytokines implicated in fever, including interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). To define the role of α2M in regulation of febrile and cytokine responses, wild-type mice and mice deficient in α2M (α2M −/−) were injected with lipopolysaccharide (LPS). Changes in body temperature as well as plasma levels of IL-1β, IL-6, and TNF-α and hepatic TNF-α mRNA level during fever in α2M −/− mice were compared with those in wild-type control mice. The α2M −/− mice developed a short-term markedly attenuated (ANOVA, P < 0.05) fever in response to LPS (2.5 mg/kg ip) compared with the wild-type mice. At 1.5 h after injection of LPS, the plasma concentration of TNF-α, but not IL-1β or IL-6, was significantly lower (by 58%) in the α2M −/− mice compared with their wild-type controls (ANOVA, P < 0.05). There was no difference in hepatic TNF-α mRNA levels between α2M −/− and wild-type mice 1.5 h after injection of LPS. These data support the hypotheses that 1) α2M is important for the normal development of LPS-induced fever and 2) a putative mechanism of α2M involvement in fever is through the inhibition of TNF-α clearance. These findings indicate a novel physiological role for α2M.

scholarly journals The Role of Leptin in the Development of the Cerebral Cortex in Mouse Embryos

Endocrinology ◽  
2006 ◽  
Vol 147 (2) ◽  
pp. 647-658 ◽  
Author(s):  
Jun Udagawa ◽  
Ryuju Hashimoto ◽  
Hiroaki Suzuki ◽  
Toshihisa Hatta ◽  
Yusuke Sotomaru ◽  
...  
Keyword(s):  
Neuronal Development ◽  
Mrna Level ◽  
Mouse Embryos ◽  
Neural Progenitors ◽  
Cortical Plate ◽  
High Dose ◽  
Wild Type ◽  
Leptin Receptors ◽  
Astrocyte Differentiation

Leptin is detected in the sera, and leptin receptors are expressed in the cerebrum of mouse embryos, suggesting that leptin plays a role in cerebral development. Compared with the wild type, leptin-deficient (ob/ob) mice had fewer cells at embryonic day (E) 16 and E18 and had fewer 5-bromo-2′-deoxyuridine+ cells at E14 and E16 in the neuroepithelium. Intracerebroventricular leptin injection in E14 ob/ob embryos increased the number of neuroepithelium cells at E16. In cultured neurosphere cells, leptin treatment increased Hes1 mRNA expression and maintained neural progenitors. Astrocyte differentiation was induced by low-dose (0.1 μg/ml) but not high-dose (1 μg/ml) leptin. High-dose leptin decreased Id mRNA and increased Ngn1 mRNA in neurosphere cells. The neuropeptide Y mRNA level in the cortical plate was lower in ob/ob than the wild type at E16 and E18. These results suggest that leptin maintains neural progenitors and is related to glial and neuronal development in embryos.

scholarly journals Aspergillus fumigatus LaeA-Mediated Phagocytosis Is Associated with a Decreased Hydrophobin Layer

2009 ◽  
Vol 78 (2) ◽  
pp. 823-829 ◽  
Author(s):  
Taylor R. T. Dagenais ◽  
Steve S. Giles ◽  
Vishukumar Aimanianda ◽  
Jean-Paul Latgé ◽  
Christina M. Hull ◽  
...  
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Wild Type ◽  
Macrophage Receptors ◽  
Macrophage Phagocytosis ◽  
Life Threatening ◽  
Carbohydrate Ligands ◽  
Cell Wall Carbohydrates ◽  
Molecular Patterns

ABSTRACT Aspergillus fumigatus is the causal agent of the life-threatening disease invasive aspergillosis. A. fumigatus laeA deletants, aberrant in toxin biosynthesis and spore development, are decreased in virulence. Among other characteristics, the decreased virulence is associated with increased spore susceptibility to macrophage phagocytosis. Three characteristics, cell wall microbe-associated molecular patterns (MAMPs), secreted metabolites, and rodlet content, thought to be important in macrophage-Aspergillus spore interactions were examined. Flow cytometry analysis of wild-type and ΔlaeA spores did not reveal any differences in surface-accessible MAMPs, including β-(1,3)-glucan, α-mannose, chitin, and other carbohydrate ligands. Blocking experiments with laminarin and mannan supported the conclusion that differences in cell wall carbohydrates were not responsible for enhanced ΔlaeA spore phagocytosis. Aspergillus spores have been reported to secrete metabolites affecting phagocytosis. Neither spent culture exchange, transwell, nor coincubation internalization experiments supported a role for secreted metabolites in the differential uptake of wild-type and ΔlaeA spores. However, sonication assays implicated a role for surface rodlet protein/hydrophobin (RodAp) in differential spore phagocytosis. A possible role of RodAp in enhanced ΔlaeA spore uptake was further assessed by RodAp extraction and quantification, where wild-type spores were found to contain 60% more RodAp than ΔlaeA spores. After removal of the surface rodlet layer, wild-type spores were phagocytosed at similar rates as ΔlaeA spores. We conclude that increased uptake of ΔlaeA resting spores is not associated with changes in secreted metabolite production of this mutant or surface carbohydrate availability but, rather, due to a decrease in the surface RodAp content of ΔlaeA spores. We theorize that RodAp acts as an antiphagocytic molecule, possibly via physicochemical means and/or by impeding MAMP recognition by macrophage receptors.

scholarly journals Defects in Conidiophore Development and Conidium-Macrophage Interactions in a Dioxygenase Mutant of Aspergillus fumigatus

2008 ◽  
Vol 76 (7) ◽  
pp. 3214-3220 ◽  
Author(s):  
Taylor R. T. Dagenais ◽  
DaWoon Chung ◽  
Steven S. Giles ◽  
Christina M. Hull ◽  
David Andes ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Aspergillus Fumigatus ◽  
Essential Role ◽  
Octadecadienoic Acid ◽  
Wild Type ◽  
Host Interactions ◽  
Host Interaction ◽  
Hydroperoxyoctadecadienoic Acid

ABSTRACT Oxygenated fatty acids, or oxylipins, play an essential role in physiological signaling and developmental processes in animals, plants, and fungi. Previous characterization of three Aspergillus fumigatus dioxygenases (PpoA, PpoB, and PpoC), similar in sequence to mammalian cyclooxygenases, showed that PpoA is responsible for the production of the oxylipins 8R-hydroperoxyoctadecadienoic acid and 5S,8R-dihydroxy-9Z,12Z-octadecadienoic acid and that PpoC is responsible for 10R-hydroxy-8E,12Z-hydroperoxyoctadecadienoic acid. Here, Δppo mutants were characterized to elucidate the role of fungal dioxygenases in A. fumigatus development and host interactions. The ΔppoC strain displayed distinct phenotypes compared to those of other Δppo mutants and the wild type, including altered conidium size, germination, and tolerance to oxidative stress as well as increased uptake and killing by primary alveolar macrophages. These experiments implicate oxylipins in pathogen development and suggest that ΔppoC represents a useful model for studying the A. fumigatus-host interaction.

scholarly journals Role of a Cytotoxic Enterotoxin in Aeromonas-Mediated Infections: Development of Transposon and Isogenic Mutants

1998 ◽  
Vol 66 (8) ◽  
pp. 3501-3509 ◽  
Author(s):  
Xin-J. Xu ◽  
M. R. Ferguson ◽  
V. L. Popov ◽  
C. W. Houston ◽  
J. W. Peterson ◽  
...  
Keyword(s):  
Lethal Dose ◽  
Sublethal Dose ◽  
Toxin Gene ◽  
Cytotoxic Activities ◽  
Wild Type ◽  
Chromosomal Dna ◽  
Culture Filtrates ◽  
Transposon Mutants ◽  
Isogenic Mutants

ABSTRACT Transposon and marker exchange mutagenesis were used to evaluate the role of Aeromonas cytotoxic enterotoxin (Act) in the pathogenesis of diarrheal diseases and deep wound infections. The transposon mutants were generated by random insertion of Tn5-751 in the chromosomal DNA of a diarrheal isolate SSU of Aeromonas hydrophila. Some of the transposon mutants had dramatically reduced hemolytic and cytotoxic activities, and such mutants exhibited reduced virulence in mice compared to wild-typeAeromonas when injected intraperitoneally (i.p.). Southern blot data indicated that transposition in these mutants did not occur within the cytotoxic enterotoxin gene (act). The transcription of the act gene was affected drastically in the transposon mutants, as revealed by Northern blot analysis. The altered virulence of these transposon mutants was confirmed by developing isogenic mutants of the wild-type Aeromonas by using a suicide vector. In these mutants, the truncated actgene was integrated in place of a functionally active actgene. The culture filtrates from isogenic mutants were devoid of hemolytic, cytotoxic, and enterotoxic activities associated with Act. These filtrates caused no damage to mouse small intestinal epithelium, as determined by electron microscopy, whereas culture filtrates from wild-type Aeromonas caused complete destruction of the microvilli. The 50% lethal dose of these mutants in mice was 1.0 × 108 when injected i.p., compared to 3.0 × 105 for the wild-type Aeromonas. Reintegration of the native act gene in place of the truncated toxin gene in isogenic mutants resulted in complete restoration of Act’s biological activity and virulence in mice. The animals injected with a sublethal dose of wild-type Aeromonas or the revertant, but not the isogenic mutant, had circulating toxin-specific neutralizing antibodies. Taken together, these studies clearly established a role for Act in the pathogenesis of Aeromonas-mediated infections.

scholarly journals β(1-3)Glucanosyltransferase Gel4p Is Essential for Aspergillus fumigatus

2010 ◽  
Vol 9 (8) ◽  
pp. 1294-1298 ◽  
Author(s):  
Amandine Gastebois ◽  
Thierry Fontaine ◽  
Jean-Paul Latgé ◽  
Isabelle Mouyna
Keyword(s):  
Aspergillus Fumigatus ◽  
Mycelial Growth ◽  
Fungal Species ◽  
The Other ◽  
Content Type

ABSTRACT The β(1-3)glucanosyltransferase GEL family of Aspergillus fumigatus contains 7 genes, among which only 3 are expressed during mycelial growth. The role of the GEL4 gene was investigated in this study. Like the other Gelps, it encodes a glycosylphosphatidylinositol (GPI)-anchored protein. In contrast to the other β(1-3)glucanosyltransferases analyzed to date, it is essential for this fungal species.

scholarly journals GliZ, a Transcriptional Regulator of Gliotoxin Biosynthesis, Contributes to Aspergillus fumigatus Virulence

2006 ◽  
Vol 74 (12) ◽  
pp. 6761-6768 ◽  
Author(s):  
Jin Woo Bok ◽  
DaWoon Chung ◽  
S. Arunmozhi Balajee ◽  
Kieren A. Marr ◽  
David Andes ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Survival Data ◽  
Polymorphonuclear Leukocytes ◽  
Marker Gene ◽  
Flow Cytometric Analysis ◽  
Transcriptional Regulator ◽  
Multiple Copy ◽  
Wild Type ◽  
Flow Cytometric ◽  
Multiple Copies

ABSTRACT Gliotoxin is a nonribosomal peptide produced by Aspergillus fumigatus. This compound has been proposed as an A. fumigatus virulence factor due to its cytotoxic, genotoxic, and apoptotic properties. Recent identification of the gliotoxin gene cluster identified several genes (gli genes) likely involved in gliotoxin production, including gliZ, encoding a putative Zn2Cys6 binuclear transcription factor. Replacement of gliZ with a marker gene (ΔgliZ) resulted in no detectable gliotoxin production and loss of gene expression of other gli cluster genes. Placement of multiple copies of gliZ in the genome increased gliotoxin production. Using endpoint survival data, the ΔgliZ and a multiple-copy gliZ strain were not statistically different from the wild type in a murine pulmonary model; however, both the wild-type and the multiple-copy gliZ strain were more virulent than ΔlaeA (a mutant reduced in production of gliotoxin and other toxins). A flow-cytometric analysis of polymorphonuclear leukocytes (PMNs) exposed to supernatants from wild-type, ΔgliZ, complemented ΔgliZ, and ΔlaeA strains supported a role for gliotoxin in apoptotic but not necrotic PMN cell death. This may indicate that several secondary metabolites are involved in A. fumigatus virulence.

scholarly journals A Novel Environmental Azole Resistance Mutation inAspergillus fumigatusand a Possible Role of Sexual Reproduction in Its Emergence

mBio ◽  
2017 ◽  
Vol 8 (3) ◽  
Author(s):  
Jianhua Zhang ◽  
Eveline Snelders ◽  
Bas J. Zwaan ◽  
Sijmen E. Schoustra ◽  
Jacques F. Meis ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Sexual Reproduction ◽  
Crop Protection ◽  
Resistance Mutation ◽  
Azole Resistance ◽  
Wild Type ◽  
Resistance Mutations ◽  
Resistance Selection ◽  
Genotype 2

ABSTRACTThis study investigated the dynamics ofAspergillus fumigatusazole-resistant phenotypes in two compost heaps with contrasting azole exposures: azole free and azole exposed. After heat shock, to which sexual but not asexual spores are highly resistant, the azole-free compost yielded 98% (49/50) wild-type and 2% (1/50) azole-resistant isolates, whereas the azole-containing compost yielded 9% (4/45) wild-type and 91% (41/45) resistant isolates. From the latter compost, 80% (36/45) of the isolates contained the TR46/Y121F/T289A genotype, 2% (1/45) harbored the TR46/Y121F/M172I/T289A/G448S genotype, and 9% (4/45) had a novel pan-triazole-resistant mutation (TR463/Y121F/M172I/T289A/G448S) with a triple 46-bp promoter repeat. Subsequent screening of a representative set of clinicalA. fumigatusisolates showed that the novel TR463mutant was already present in samples from three Dutch medical centers collected since 2012. Furthermore, a second new resistance mutation was found in this set that harbored four TR46repeats. Importantly, in the laboratory, we recovered the TR463mutation from a sexual cross between two TR46isolates from the same azole-containing compost, possibly through unequal crossing over between the double tandem repeats (TRs) during meiosis. This possible role of sexual reproduction in the emergence of the mutation was further implicated by the high level of genetic diversity of STR genotypes in the azole-containing compost. Our study confirms that azole resistance mutations continue to emerge in the environment and indicates compost containing azole residues as a possible hot spot. Better insight into the biology of environmental resistance selection is needed to retain the azole class for use in food production and treatment ofAspergillusdiseases.IMPORTANCEComposting of organic matter containing azole residues might be important for resistance development and subsequent spread of resistance mutations inAspergillus fumigatus. In this article, we show the dominance of azole-resistantA. fumigatusin azole-exposed compost and the discovery of a new resistance mutation with clinical relevance. Furthermore, our study indicates that current fungicide application is not sustainable as new resistance mutations continue to emerge, thereby threatening the use of triazoles in medicine. We provide evidence that the sexual part of the fungal life cycle may play a role in the emergence of resistance mutations because under laboratory conditions, we reconstructed the resistance mutation through sexual crossing of two azole-resistantA. fumigatusisolates derived from the same compost heap. Understanding the mechanisms of resistance selection in the environment is needed to design strategies against the accumulation of resistance mutations in order to retain the azole class for crop protection and treatment ofAspergillusdiseases.

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