scholarly journals In Vivo Efficacy of Voriconazole in a Galleria mellonella Model of Invasive Infection Due to Azole-Susceptible or Resistant Aspergillus fumigatus Isolates

2021 ◽  
Vol 7 (12) ◽  
pp. 1012
Author(s):  
Sana Jemel ◽  
Jacques Guillot ◽  
Kalthoum Kallel ◽  
Grégory Jouvion ◽  
Elise Brisebard ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Galleria Mellonella ◽  
Lethal Dose ◽  
Dose Assessment ◽  
Suitable Model ◽  
Invasive Infection ◽  
Larval Hemolymph ◽  
Life Threatening ◽  
Dose Dependent

Aspergillus fumigatus is an environmental filamentous fungus responsible for life-threatening infections in humans and animals. Azoles are the first-line treatment for aspergillosis, but in recent years, the emergence of azole resistance in A. fumigatus has changed treatment recommendations. The objective of this study was to evaluate the efficacy of voriconazole (VRZ) in a Galleria mellonella model of invasive infection due to azole-susceptible or azole-resistant A. fumigatus isolates. We also sought to describe the pharmacokinetics of VRZ in the G. mellonella model. G. mellonella larvae were infected with conidial suspensions of azole-susceptible and azole-resistant isolates of A. fumigatus. Mortality curves were used to calculate the lethal dose. Assessment of the efficacy of VRZ or amphotericin B (AMB) treatment was based on mortality in the lethal model and histopathologic lesions. The pharmacokinetics of VRZ were determined in larval hemolymph. Invasive fungal infection was obtained after conidial inoculation. A dose-dependent reduction in mortality was observed after antifungal treatment with AMB and VRZ. VRZ was more effective at treating larvae inoculated with azole-susceptible A. fumigatus isolates than larvae inoculated with azole-resistant isolates. The concentration of VRZ was maximal at the beginning of treatment and gradually decreased in the hemolymph to reach a Cmin (24 h) between 0.11 and 11.30 mg/L, depending on the dose. In conclusion, G. mellonella is a suitable model for testing the efficacy of antifungal agents against A. fumigatus.

scholarly journals Streptococcus pyogenes evades adaptive immunity through specific IgG glycan hydrolysis

2019 ◽  
Vol 216 (7) ◽  
pp. 1615-1629 ◽  
Author(s):  
Andreas Naegeli ◽  
Eleni Bratanis ◽  
Christofer Karlsson ◽  
Oonagh Shannon ◽  
Raja Kalluru ◽  
...  
Keyword(s):  
Streptococcus Pyogenes ◽  
Vaccine Development ◽  
Group A Streptococcus ◽  
Invasive Infection ◽  
Invasive Infections ◽  
Life Threatening ◽  
Group A ◽  
Specific Igg

Streptococcus pyogenes (Group A streptococcus; GAS) is a human pathogen causing diseases from uncomplicated tonsillitis to life-threatening invasive infections. GAS secretes EndoS, an endoglycosidase that specifically cleaves the conserved N-glycan on IgG antibodies. In vitro, removal of this glycan impairs IgG effector functions, but its relevance to GAS infection in vivo is unclear. Using targeted mass spectrometry, we characterized the effects of EndoS on host IgG glycosylation during the course of infections in humans. Substantial IgG glycan hydrolysis occurred at the site of infection and systemically in the severe cases. We demonstrated decreased resistance to phagocytic killing of GAS lacking EndoS in vitro and decreased virulence in a mouse model of invasive infection. This is the first described example of specific bacterial IgG glycan hydrolysis during infection and thereby verifies the hypothesis that EndoS modifies antibodies in vivo. This mechanisms of immune evasion could have implications for treatment of severe GAS infections and for future efforts at vaccine development.

In vivo protection studies of bis-quaternary 2-(hydroxyimino)-N-(pyridin-3-yl) acetamide derivatives against sarin poisoning in mice

2016 ◽  
Vol 36 (1) ◽  
pp. 23-32 ◽  
Author(s):  
Devyani Swami ◽  
Hitendra N Karade ◽  
Jyotiranjan Acharya ◽  
Pravin Kumar
Keyword(s):  
Time Course ◽  
Lethal Dose ◽  
Nerve Agent ◽  
Ache Inhibition ◽  
Inhibition Concentration ◽  
Time Course Study ◽  
Dose Dependent ◽  
Protection Index ◽  
Lethal Doses

In vivo antidotal efficacy of new bis- quaternary 2-(hydroxyimino)- N-(pyridin-3yl) acetamide derivatives (HNK series), to counter multiples of lethal doses of nerve agent sarin (GB) and reactivation of acetylcholinesterase (AChE), was evaluated in Swiss albino mice. [Protection index PI; median lethal dose (LD50) of sarin with treatment/LD50 of sarin] was estimated, using 0.05, 0.10, and 0.20 LD50 as treatment doses of all the oximes with atropine against sarin poisoning. Dose-dependent time course study was conducted at 0.2, 0.4 and 0.8 LD50 dose of sarin for estimating maximum AChE inhibition. At optimized time (15 min), in vivo enzyme half inhibition concentration (IC50) was calculated. AChE reactivation efficacy of HNK series and pralidoxime (2-PAM) were determined by plotting shift of log IC50 doses. HNK-102 with atropine showed three fold higher PI compared to 2-PAM. In vivo IC50 of sarin for brain and serum AChE was found to be 0.87 LD50 (139.2 µg/kg) and 0.48 LD50 (77.23 µg/kg), respectively. Treatment with HNK-102 and HNK-111 (equal to their 0.20LD50) significantly reactivated sarin-intoxicated AChE ( p < 0.05) at 2× IC50 dose of sarin, compared to 2-PAM. The study revealed that HNK-102 oxime was three times more potent as antidote, for acute sarin poisoning compared to 2-PAM in vivo.

scholarly journals The Immunomodulatory Effects of AlbuminIn VitroandIn Vivo

2011 ◽  
Vol 2011 ◽  
pp. 1-7 ◽  
Author(s):  
Derek S. Wheeler ◽  
John S. Giuliano ◽  
Patrick M. Lahni ◽  
Alvin Denenberg ◽  
Hector R. Wong ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lethal Dose ◽  
Intraperitoneal Administration ◽  
Peritoneal Macrophages ◽  
Luciferase Reporter Assay ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Dose Dependent

Albumin appears to have proinflammatory effectsin vitro. We hypothesized that albumin would induce a state of tolerance to subsequent administration of lipopolysaccharide (LPS)in vitroandin vivo. RAW264.7 and primary peritoneal macrophages were treated with increasing doses of bovine serum albumin (BSA) and harvested for NF-κB luciferase reporter assay or TNF-αELISA. In separate experiments, RAW264.7 cells were preconditioned with 1 mg/mL BSA for 18 h prior to LPS (10 μg/mL) treatment and harvested for NF-κB luciferase reporter assay or TNF-αELISA. Finally, C57Bl/6 mice were preconditioned with albumin via intraperitoneal administration 18 h prior to a lethal dose of LPS (60 mg/kg body wt). Blood was collected at 6 h after LPS administration for TNF-αELISA. Albumin produced a dose-dependent and TLR-4-dependent increase in NF-κB activation and TNF-αgene expressionin vitro. Albumin preconditioning abrogated the LPS-mediated increase in NF-κB activation and TNF-αgene expressionin vitroandin vivo. The clinical significance of these findings remains to be elucidated.

scholarly journals THE CHEMISTRY OF INSECT HEMOLYMPH

1956 ◽  
Vol 39 (6) ◽  
pp. 853-868 ◽  
Author(s):  
G. R. Wyatt ◽  
T. C. Loughheed ◽  
S. S. Wyatt
Keyword(s):  
Amino Acids ◽  
Free Amino Acids ◽  
Free Amino ◽  
Developmental Stages ◽  
Galleria Mellonella ◽  
Similar Distribution ◽  
Total Acid ◽  
Protein Nitrogen ◽  
Larval Hemolymph

1. Hemolymph was collected for analysis from the silkworm, Bombyx mori, in a series of developmental stages ranging from the second molt to the late pupa. The mean pH of larval hemolymph after collection was found to be 6.45, that of pupal hemolymph, 6.57; in vivo values may be slightly lower. Total dry solids ranged from 5.4 to 10.6 per cent. Total protein ranged from 1.2 to 5.3 per cent, increasing rapidly during the fifth instar. 2. Free amino acids were separated chromatographically and estimated. Of 19 amino acids identified, amounting collectively to 823 to 1497 mg. per 100 ml., glutamine, histidine, and lysine generally occurred in greatest amount. Tryptophan was not detected, and cystine (or cysteine) was found in only one sample. The total free amino acids account for 35 to 55 per cent of the non-protein nitrogen of the plasma. 3. Free sugars, estimated semiquantitatively on chromatograms, comprise glucose, fructose, and sucrose in total amount ranging from about 5 to 40 mg. per 100 ml. Total acid-soluble, ultrafiltrable carbohydrate, estimated as glucose by the anthrone reaction, ranged from 166 to 635 mg. per 100 ml., indicating the presence of low molecular weight sugar derivatives. 4. Inorganic phosphate amounted to 5 to 15 mg. per 100 ml., and acid-soluble organic phosphate to 100 to 200 mg. per 100 ml. The latter fraction includes several substances, of which one was tentatively identified as glucose-6-phosphate and the remainder are as yet unidentified. 5. Single samples of hemolymph were also taken from larvae of the wax moth, Galleria mellonella, and the spruce sawfly, Diprion hercyniae. These contained even higher concentrations of solutes than the silkworm samples, but with a generally similar distribution. The proportions of the free amino acids were different in each species.

scholarly journals In Vivo Analysis of Aspergillus fumigatus Developmental Gene Expression Determined by Real-Time Reverse Transcription-PCR

2008 ◽  
Vol 76 (8) ◽  
pp. 3632-3639 ◽  
Author(s):  
Fabrice N. Gravelat ◽  
Thomas Doedt ◽  
Lisa Y. Chiang ◽  
Hong Liu ◽  
Scott G. Filler ◽  
...  
Keyword(s):  
Gene Expression ◽  
Aspergillus Fumigatus ◽  
Real Time ◽  
Reverse Transcription ◽  
Invasive Pulmonary Aspergillosis ◽  
Pcr Analysis ◽  
Invasive Infection ◽  
Reverse Transcription Pcr

ABSTRACT Very little is known about the developmental stages of Aspergillus fumigatus during invasive aspergillosis. We performed real-time reverse transcription-PCR analysis on lung samples from mice with invasive pulmonary aspergillosis to determine the expression of A. fumigatus genes that are expressed at specific stages of development. In established infection, A. fumigatus exhibited mRNA expression of genes specific to developmentally competent hyphae, such as stuA. In contrast, mRNA of genes expressed by conidia and precompetent hyphae was not detected. Many genes required for mycotoxin synthesis, including aspHS, gliP, mitF, and metAP, are known to be expressed by developmentally competent hyphae in vitro. Interestingly, each of these genes was expressed at significantly higher levels during invasive infection than in vitro. The expression of gliP mRNA in vitro was found to be highly dependent on culture conditions. Furthermore, gliP expression was found to be dependent on the transcription factor StuA both in vitro and in vivo. Therefore, developmentally competent hyphae predominate during established invasive infection, and many mycotoxin genes are expressed at high levels in vivo. These results highlight the importance of the evaluation of putative virulence factors expressed by competent hyphae and analysis of gene expression levels during invasive infection rather than in vitro alone.

scholarly journals PneumococcalSurface Protein A Is Expressed In Vivo, and Antibodies to PspA AreEffective for Therapy in a Murine Model of PneumococcalSepsis

2003 ◽  
Vol 71 (12) ◽  
pp. 7149-7153 ◽  
Author(s):  
E. Swiatlo ◽  
J. King ◽  
G. S. Nabors ◽  
B. Mathews ◽  
D. E. Briles
Keyword(s):  
Northern Blot Analysis ◽  
Lethal Dose ◽  
Protein A ◽  
Surface Protein ◽  
Immune Serum ◽  
Invasive Infection ◽  
Pneumococcal Infections ◽  
Type 3

ABSTRACT Pneumococcal surface protein A (PspA) is an immunogenic protein expressed on the surface of all strains of Streptococcus pneumoniae (pneumococcus) and induces antibodies which protect against invasive infection in mice. Pneumococci used for infectious challenge in protection studies are typically collected from cultures grown in semisynthetic medium in vitro. The purpose of these studies is to confirm that PspA is expressed by pneumococci during growth in vivo at a level sufficient for antibodies to PspA to be protective. Mice were actively immunized with purified PspA or by passive transfer of monoclonal antibody (MAb) and challenged with a capsular type 3 strain in diluted whole blood from bacteremic mice. All were protected against challenge with 10 times the 50% lethal dose (LD50), and mice challenged with 1,000 times the LD50 had increased survival compared with controls. Additionally, nonimmune mice treated with MAbs to PspA or PspA immune serum at 6 and 12 h after infection with 10 times the LD50 also showed increased survival. Northern blot analysis of RNA from pneumococci grown either in vitro or in vivo showed similar levels of PspA mRNA. These results demonstrate that PspA is expressed in vivo in a mouse model and that immunization with PspA induces antibodies to an antigen which is expressed during the course of invasive infection. Immunotherapy with antibodies to PspA may have some utility in treating pneumococcal infections in humans.

scholarly journals Expression of a Toll Signaling Regulator Serpin in a Mycoinsecticide for Increased Virulence

2014 ◽  
Vol 80 (15) ◽  
pp. 4531-4539 ◽  
Author(s):  
Linzhi Yang ◽  
Nemat O. Keyhani ◽  
Guirong Tang ◽  
Chuang Tian ◽  
Ruipeng Lu ◽  
...  
Keyword(s):  
Entomopathogenic Fungi ◽  
Galleria Mellonella ◽  
Lethal Dose ◽  
Defense Responses ◽  
Content Type ◽  
Greater Wax Moth ◽  
Wide Range ◽  
New Strategy

ABSTRACTSerpins are ubiquitously distributed serine protease inhibitors that covalently bind to target proteases to exert their activities. Serpins regulate a wide range of activities, particularly those in which protease-mediated cascades are active. TheDrosophila melanogasterserpin Spn43Ac negatively controls the Toll pathway that is activated in response to fungal infection. The entomopathogenic fungusBeauveria bassianaoffers an environmentally friendly alternative to chemical pesticides for insect control. However, the use of mycoinsecticides remains limited in part due to issues of efficacy (low virulence) and the recalcitrance of the targets (due to strong immune responses). Since Spn43Ac acts to inhibit Toll-mediated activation of defense responses, we explored the feasibility of a new strategy to engineer entomopathogenic fungi with increased virulence by expression of Spn43Ac in the fungus. Compared to the 50% lethal dose (LD50) for the wild-type parent, the LD50ofB. bassianaexpressing Spn43Ac (strain Bb::S43Ac-1) was reduced ∼3-fold, and the median lethal time against the greater wax moth (Galleria mellonella) was decreased by ∼24%, with the more rapid proliferation of hyphal bodies being seen in the host hemolymph.In vitroandin vivoassays showed inhibition of phenoloxidase (PO) activation in the presence of Spn43Ac, with Spn43Ac-mediated suppression of activation by chymotrypsin, trypsin, laminarin, and lipopolysaccharide occurring in the following order: chymotrypsin and trypsin > laminarin > lipopolysaccharide. Expression of Spn43Ac had no effect on the activity of the endogenousB. bassiana-derived cuticle-degrading protease (CDEP-1). These results expand our understanding of Spn43Ac function and confirm that suppression of insect immune system defenses represents a feasible approach to engineering entomopathogenic fungi for greater efficacy.

scholarly journals The Aspergillus fumigatus toxin fumagillin suppresses the immune response of Galleria mellonella larvae by inhibiting the action of haemocytes

Microbiology ◽  
2011 ◽  
Vol 157 (5) ◽  
pp. 1481-1488 ◽  
Author(s):  
John P. Fallon ◽  
Emer P. Reeves ◽  
Kevin Kavanagh
Keyword(s):  
Immune Response ◽  
Candida Albicans ◽  
Aspergillus Fumigatus ◽  
Immune Cells ◽  
Antimicrobial Agents ◽  
Galleria Mellonella ◽  
Subsequent Infection ◽  
Nadph Oxidase Complex ◽  
Cellular Immune

Larvae of Galleria mellonella are widely used to evaluate microbial virulence and to assess the in vivo efficacy of antimicrobial agents. The aim of this work was to examine the ability of an Aspergillus fumigatus toxin, fumagillin, to suppress the immune response of larvae. Administration of fumagillin to larvae increased their susceptibility to subsequent infection with A. fumigatus conidia (P = 0.0052). It was demonstrated that a dose of 2 µg fumagillin ml−1 reduced the ability of insect immune cells (haemocytes) to kill opsonized cells of Candida albicans (P = 0.039) and to phagocytose A. fumigatus conidia (P = 0.016). Fumagillin reduced the oxygen uptake of haemocytes and decreased the translocation of a p47 protein which is homologous to p47phox, a protein essential for the formation of a functional NADPH oxidase complex required for superoxide production. In addition, toxin-treated haemocytes showed reduced levels of degranulation as measured by the release of a protein showing reactivity to an anti-myeloperoxidase antibody (P<0.049) that was subsequently identified by liquid chromatography-MS analysis as prophenoloxidase. This work demonstrates that fumagillin suppresses the immune response of G. mellonella larvae by inhibiting the action of haemocytes and thus renders the larvae susceptible to infection. During growth of the fungus in the larvae, this toxin, along with others, may facilitate growth by suppressing the cellular immune response.

scholarly journals Dexamethasone and Methylprednisolone Promote Cell Proliferation, Capsule Enlargement, and in vivo Dissemination of C. neoformans

2021 ◽  
Vol 2 ◽  
Author(s):  
Glauber R. de S. Araújo ◽  
Vinicius Alves ◽  
Pedro H. Martins-de-Souza ◽  
Allan J. Guimarães ◽  
Leandro Honorato ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Fungal Pathogen ◽  
Galleria Mellonella ◽  
Histopathological Analysis ◽  
Fungal Cell ◽  
Fungal Virulence ◽  
Promote Cell Proliferation ◽  
Significant Difference ◽  
Life Threatening

Cryptococcus neoformans is a fungal pathogen that causes life-threatening infections in immunocompromised individuals, who often have some inflammatory condition and, therefore, end up using glucocorticoids, such as dexamethasone and methylprednisolone. Although the effects of this class of molecules during cryptococcosis have been investigated, their consequences for the biology of C. neoformans is less explored. Here, we studied the effects of dexamethasone and methylprednisolone on the metabolism and on the induction of virulence factors in C. neoformans. Our results showed that both glucocorticoids increased fungal cell proliferation and surface electronegativity but reduced capsule and secreted polysaccharide sizes, as well as capsule compaction, by decreasing the density of polysaccharide fibers. We also tested whether glucocorticoids could affect the fungal virulence in Galleria mellonella and mice. Although the survival rate of Galleria larvae increased, those from mice showed a tendency to decrease, with infected animals dying earlier after glucocorticoid treatments. The pathogenesis of spread of cryptococcosis and the interleukin secretion pattern were also assessed for lungs and brains of infected mice. While increases in the spread of the fungus to lungs were observed after treatment with glucocorticoids, a significant difference in brain was observed only for methylprednisolone, although a trend toward increasing was also observed for dexamethasone. Moreover, increases in both pulmonary and cerebral IL-10 production, reduction of IL-6 production but no changes in IL-4, IL-17, and INF-γ were also observed after glucocorticoid treatments. Finally, histopathological analysis confirmed the increase in number of fungal cells in lung and brain tissues of mice previously subjected to dexamethasone or methylprednisolone treatments. Together, our results provide compelling evidence for the effects of dexamethasone and methylprednisolone on the biology of C. neoformans and may have important implications for future clinical treatments, calling attention to the risks of using these glucocorticoids against cryptococcosis or in immunocompromised individuals.

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