scholarly journals Genetic Changes in Experimental Populations of a Hybrid in the Cryptococcus neoformans Species Complex

Pathogens ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 3 ◽  
Author(s):  
Kelly Dong ◽  
Man You ◽  
Jianping Xu
Keyword(s):  
Flow Cytometry ◽  
Cryptococcus Neoformans ◽  
Species Complex ◽  
Mitotic Division ◽  
Culture Conditions ◽  
Chromosome 1 ◽  
Allelic Loss ◽  
Genetic Changes ◽  
Susceptible Allele ◽  
Standard Culture

Hybrids between Cryptococcus neoformans and Cryptococcus deneoformans are commonly found in patients and the environment. However, the genetic stability of these hybrids remains largely unknown. Here, we established mutation accumulation lines of a diploid C. neoformans × C. deneoformans laboratory hybrid and analyzed the genotypes at 33 markers distributed across all 14 chromosomes. Our analyses found that under standard culture conditions, heterozygosity at most loci was maintained over 800 mitotic generations, with an estimated 6.44 × 10−5 loss-of-heterozygosity (LoH) event per mitotic division. However, under fluconazole stress, the observed LoH frequency increased by > 50 folds for the two markers on Chromosome 1, all due to the loss of the fluconazole susceptible allele on this chromosome. Flow cytometry analyses showed that after the 40th transfer (120 days), 19 of the 20 lines maintained the original ploidy level (2N), while one line was between 2N and 3N. The combined flow cytometry, genotyping at 33 markers, and quantitative PCR analyses showed the allelic loss was compensated for by amplification of the resistant ERG11 allele in eight of the ten fluconazole-stress lines. Our results suggest that hybrids in C. neoformans species complex are generally stable but that they can undergo rapid adaptation to environmental stresses through LoH and gene duplication.

scholarly journals The genetics of catalase in Drosophila melanogaster: isolation and characterization of acatalasemic mutants.

Genetics ◽  
1989 ◽  
Vol 122 (3) ◽  
pp. 643-652 ◽  
Author(s):  
W J Mackay ◽  
G C Bewley
Keyword(s):  
Drosophila Melanogaster ◽  
Free Radical ◽  
Catalase Activity ◽  
Oxygen Toxicity ◽  
Threshold Effect ◽  
Culture Conditions ◽  
Free Radical Damage ◽  
Isolation And Characterization ◽  
Standard Culture ◽  
Hypomorphic Alleles

Abstract Activated oxygen species have been demonstrated to be the important agents in oxygen toxicity by disrupting the structural and functional integrity of cells through lipid peroxidation events, DNA damage and protein inactivation. The biological consequences of free radical damage have long been hypothesized to be a causal agent in many aging-related diseases. Catalase (H2O2:H2O2 oxidoreductase; EC 1.15.1.1) is one of several enzymes involved in the scavenging of oxygen free radicals and free radical derivatives. The structural gene for catalase in Drosophila melanogaster has been localized to region 75D1-76A on chromosome 3L by dosage responses to segmental aneuploidy. This study reports the isolation of a stable deficiency, Df(3L)CatDH104(75C1-2;75F1), that uncovers the catalase locus and the subsequent isolation of six acatalasemic mutants. All catalase mutants are viable under standard culture conditions and recessive lethal mutations within the 75Cl-F1 interval have been shown not to affect catalase activity. Two catalase mutations are amorphic while four are hypomorphic alleles of the Cat+ locus. The lack of intergenic complementation between the six catalase mutations strongly suggests that there is only one functional gene in Drosophila. One acatalesemic mutation was mapped to position 3-47.0 which resides within the catalase dosage sensitive region. While complete loss of catalase activity confers a severe viability effect, residual levels are sufficient to restore viability to wild type levels. These results suggest a threshold effect for viability and offer an explanation for the general lack of phenotypic effects associated with the known mammalian acatalasemics.

scholarly journals Low Glucose Mediated Fluconazole Tolerance in Cryptococcus neoformans

2021 ◽  
Vol 7 (6) ◽  
pp. 489
Author(s):  
Somanon Bhattacharya ◽  
Natalia Kronbauer Oliveira ◽  
Anne G. Savitt ◽  
Vanessa K. A. Silva ◽  
Rachel B. Krausert ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Mitochondrial Function ◽  
Efflux Pump ◽  
Nile Red ◽  
Fold Increase ◽  
Growth Conditions ◽  
Chromosome 1 ◽  
Normal Glucose ◽  
Low Glucose ◽  
Synthetic Media

Chronic meningoencephalitis is caused by Cryptococcus neoformans and is treated in many parts of the world with fluconazole (FLC) monotherapy, which is associated with treatment failure and poor outcome. In the host, C. neoformans propagates predominantly under low glucose growth conditions. We investigated whether low glucose, mimicked by growing in synthetic media (SM) with 0.05% glucose (SMlowglu), affects FLC-resistance. A > 4-fold increase in FLC tolerance was observed in seven C. neoformans strains when minimum inhibitory concentration (MIC) was determined in SMlowglu compared to MIC in SM with normal (2%) glucose (SMnlglu). In SMlowglu, C. neoformans cells exhibited upregulation of efflux pump genes AFR1 (8.7-fold) and AFR2 (2.5-fold), as well as decreased accumulation (2.6-fold) of Nile Red, an efflux pump substrate. Elevated intracellular ATP levels (3.2-fold and 3.4-fold), as well as decreased mitochondrial reactive oxygen species levels (12.8-fold and 17-fold), were found in the presence and absence of FLC, indicating that low glucose altered mitochondrial function. Fluorescence microscopy revealed that mitochondria of C. neoformans grown in SMlowglu were fragmented, whereas normal glucose promoted a reticular network of mitochondria. Although mitochondrial membrane potential (MMP) was not markedly affected in SMlowglu, it significantly decreased in the presence of FLC (12.5-fold) in SMnlglu, but remained stable in SMlowglu-growing C. neoformans cells. Our data demonstrate that increased FLC tolerance in low glucose-growing C. neoformans is the result of increased efflux pump activities and altered mitochondrial function, which is more preserved in SMlowglu. This mechanism of resistance is different from FLC heteroresistance, which is associated with aneuploidy of chromosome 1 (Chr1).

scholarly journals Environmental Status of Cryptococcus neoformans and Cryptococcus gattii in Colombia

2021 ◽  
Vol 7 (6) ◽  
pp. 410
Author(s):  
Briggith-Nathalia Serna-Espinosa ◽  
Diomedes Guzmán-Sanabria ◽  
Maribel Forero-Castro ◽  
Patricia Escandón ◽  
Zilpa Adriana Sánchez-Quitian
Keyword(s):  
Cryptococcus Neoformans ◽  
Species Complex ◽  
Columba Livia ◽  
Systematic Investigation ◽  
Cryptococcus Gattii ◽  
Eucalyptus Species ◽  
Environmental Niche ◽  
Worldwide Distribution ◽  
Bird Droppings ◽  
The Central Nervous System

The genus Cryptococcus comprises more than 80 species, including C. neoformans and C. gattii, which are pathogenic to humans, mainly affecting the central nervous system. The two species differ in geographic distribution and environmental niche. C. neoformans has a worldwide distribution and is often isolated from bird droppings. On the contrary, C. gattii is reported in tropical and subtropical regions and is associated with Eucalyptus species. This review aims to describe the distribution of environmental isolates of the Cryptococcus neoformans species complex and the Cryptococcus gattii species complex in Colombia. A systematic investigation was carried out using different databases, excluding studies of clinical isolates reported in the country. The complex of the species of C. gattii is recovered mainly from trees of the genus Eucalyptus spp., while the complex of the species of C. neoformans is recovered mainly from avian excrement, primarily Columba livia (pigeons) excrement. In addition, greater positivity was found at high levels of relative humidity. Likewise, an association was observed between the presence of the fungus in places with little insolation and cold or temperate temperatures compared to regions with high temperatures.

Structure of vegetative organs in essential oil rose under standard culture conditions and long-term conservation in vitro

2021 ◽  
pp. 185-190
Author(s):  
I.V. Mitrofanova ◽  
V.A. Brailko ◽  
N.P. Lesnikova-Sedoshenko ◽  
N.N. Ivanova ◽  
O.V. Mitrofanova
Keyword(s):  
Essential Oil ◽  
Culture Conditions ◽  
Vegetative Organs ◽  
Standard Culture

Heteroplasmy due to coexistence of mtCOI haplotypes from different lineages of theThrips tabacicryptic species group

2017 ◽  
Vol 107 (4) ◽  
pp. 534-542 ◽  
Author(s):  
S.J. Gawande ◽  
S. Anandhan ◽  
A.A. Ingle ◽  
Alana Jacobson ◽  
R. Asokan
Keyword(s):  
Flow Cytometry ◽  
Mitochondrial Dna ◽  
Real Time Pcr ◽  
Copy Number ◽  
Species Complex ◽  
Species Group ◽  
Thrips Tabaci ◽  
Mitochondrial Cytochrome ◽  
Quantitative Real Time Pcr ◽  
Putative Species

AbstractHeteroplasmy is the existence of multiple mitochondrial DNA haplotypes within the cell. Although the number of reports of heteroplasmy is increasing for arthropods, the occurrence, number of variants, and origins are not well studied. In this research, the occurrence of heteroplasmy was investigated inThrips tabaci, a putative species complex whose lineages can be distinguished by their mitochondrial DNA haplotypes. The results from this study showed that heteroplasmy was due to the occurrence of mitochondrial cytochrome oxydase I (mtCOI) haplotypes from two differentT. tabacilineages. An assay using flow cytometry and quantitative real-time PCR was then used to quantify the per cell copy number of the two mtCOI haplotypes present in individuals exhibiting heteroplasmy from nine geographically distant populations in India. All of theT. tabaciindividuals in this study were found to exhibit heteroplasmy, and in every individual the per cell copy number of mtCOI from lineage 3 comprised 75–98% of the haplotypes detected and was variable among individuals tested. There was no evidence to suggest that the presense of lineage-specific haplotypes was due to nuclear introgression; however, further studies are needed to investigate nuclear introgression and paternal leakage during rare interbreeding between individuals from lineages 2 and 3.

Experimental studies on Lecanora rupicola (L.) Zahlbr.: chemical and microscopical investigations of the mycobiont and re-synthesis stages

2006 ◽  
Vol 38 (6) ◽  
pp. 577-585 ◽  
Author(s):  
Georg BRUNAUER ◽  
Armin HAGER ◽  
Wolf Dietrich KRAUTGARTNER ◽  
Roman TÜRK ◽  
Elfie STOCKER-WÖRGÖTTER
Keyword(s):  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Sequence Data ◽  
Experimental Studies ◽  
Culture Conditions ◽  
Voucher Specimen ◽  
Dna Sequence Data ◽  
Standard Culture ◽  
Voucher Specimens ◽  
Scanning Electron

Culture experiments that trigger the axenically grown mycobionts of Lecanora rupicola to produce the polyketide chemosyndrome typical of the naturally grown lichen are reported. This chemosyndrome comprises lecanoric, haematommic and orsellinic acids, sordidone, eugenitol and atranorin, all of which were hardly produced under standard culture conditions. The only exception was arthothelin that was only present in the voucher specimen. It has been shown that almost the complete acetyl-polymalonyl-pathway leading to depsides and chromones can be induced in culture, but apparently not the xanthones. The mycobiont was also successfully re-synthesized with its original photobiont, as confirmed by Scanning Electron Microscope studies (SEM). Cultures of the resynthesised lichen biosynthesized additional satellite substances, which were not detected either in the voucher specimens or in the aposymbiontically (without the photobiont) grown mycobiont cultures. The identity of cultured mycobionts of L. rupicola was confirmed by comparing ITS-DNA-sequence data from the original lichen with publicly available (GeneBank) sequences of that species.

scholarly journals Microbe Profile: Cryptococcus neoformans species complex

Microbiology ◽  
2020 ◽  
Vol 166 (9) ◽  
pp. 797-799 ◽  
Author(s):  
Yong-Sun Bahn ◽  
Sheng Sun ◽  
Joseph Heitman ◽  
Xiaorong Lin
Keyword(s):  
Cryptococcus Neoformans ◽  
Species Complex ◽  
Carbon Sources ◽  
The Brain

Cryptococcus neoformans is a lethal fungus disguised in a polysaccharide coat. It can remain dormant in the host for decades prior to reactivation, causing systemic cryptococcosis in humans and other mammals. Cryptococcus deploys a multitude of traits to adapt to and survive within the host, including immunosuppression, an ability to replicate intra- and extra-cellularly in phagocytes, changes in morphology and ploidy, a predilection to infect the CNS, and the capacity to utilize neurotransmitters and unique carbon sources available in the brain. These pathogenic strategies displayed by this fungus might have evolved through its interactions with microbial predators in the environment.

scholarly journals Erratum: Microbe Profile: Cryptococcus neoformans species complex

Microbiology ◽  
2020 ◽  
Vol 166 (12) ◽  
pp. 1191-1191
Author(s):  
Yong-Sun Bahn ◽  
Sheng Sun ◽  
Joseph Heitman ◽  
Xiaorong Lin
Keyword(s):  
Cryptococcus Neoformans ◽  
Species Complex

scholarly journals Darunavir inhibits Cryptococcus neoformans/Cryptococcus gattii species complex growth and increases the susceptibility of biofilms to antifungal drugs

2020 ◽  
Vol 69 (6) ◽  
pp. 830-837
Author(s):  
Raimunda Sâmia Nogueira Brilhante ◽  
José Alexandre Telmos Silva ◽  
Géssica dos Santos Araújo ◽  
Vandbergue Santos Pereira ◽  
Wilker Jose Perez Gotay ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Amphotericin B ◽  
Metabolic Activity ◽  
Biofilm Formation ◽  
Species Complex ◽  
Cryptococcus Gattii ◽  
Antifungal Drugs ◽  
Planktonic Cells ◽  
Species Activity ◽  
Checkerboard Assay

Introduction. Cryptococcus species are pathogens commonly associated with cases of meningoencephalitis in individuals who are immunosuppressed due to AIDS. Aim. The aim was to evaluate the effects of the antiretroviral darunavir alone or associated with fluconazole, 5-flucytosine and amphotericin B against planktonic cells and biofilms of Cryptococcus species. Methodology. Susceptibility testing of darunavir and the common antifungals against 12 members of the Cryptococcus neoformans/Cryptococcus gattii species complex was evaluated by broth microdilution. The interaction between darunavir and antifungals against planktonic cells was tested by a checkerboard assay. The effects of darunavir against biofilm metabolic activity and biomass were evaluated by the XTT reduction assay and crystal violet staining, respectively. Results. Darunavir combined with amphotericin B showed a synergistic interaction against planktonic cells. No antagonistic interaction was observed between darunavir and the antifungals used. All Cryptococcus species strains were strong biofilm producers. Darunavir alone reduced biofilm metabolic activity and biomass when added during and after biofilm formation (P<0.05). The combination of darunavir with antifungals caused a significant reduction in biofilm metabolic activity and biomass when compared to darunavir alone (P<0.05). Conclusion. Darunavir presents antifungal activity against planktonic cells of Cryptococcus species and synergism with amphotericin B. In addition, darunavir led to reduced biofilm formation and showed activity against mature biofilms of Cryptococcus species. Activity of the antifungals against mature biofilms was enhanced in the presence of darunavir.

scholarly journals Application of Flow Cytometry to Segregated Kinetic Modeling Based on the Physiological States of Microorganisms

2007 ◽  
Vol 73 (12) ◽  
pp. 3993-4000 ◽  
Author(s):  
Covadonga Quir�s ◽  
M�nica Herrero ◽  
Luis A. Garc�a ◽  
Mario D�az
Keyword(s):  
Flow Cytometry ◽  
Culture Media ◽  
Glucose Consumption ◽  
Viable Cell ◽  
Counting Method ◽  
Batch Cultures ◽  
Viable Cells ◽  
The Status ◽  
Standard Culture

ABSTRACT Flow cytometry (FC) has been introduced to characterize and to assess the physiological states of microorganisms in conjunction with the classical plate-counting method. To show the applicability of the technique, in particular for the development of kinetic models, pure culture fermentation experiments were followed over time, using both prokaryotic (Lactobacillus hilgardii) and eukaryotic (Saccharomyces cerevisiae) microorganisms growing in standard culture media (MRS and YPD). The differences observed between the active and viable cells determined by FC and CFU, respectively, allowed us to determine that a large number of cells were in a viable but nonculturable (VBNC) state, which resulted in a subpopulation much larger than the damaged-cell (double-stained) subpopulation. Finally, the determination of the evolution of viable, the VBNC, and the dead cells allowed us to develop a segregated kinetic model to describe the yeast and the bacteria population dynamics and glucose consumption in batch cultures. This model, more complete than that which is traditionally used, based only on viable cell measurements, describes better the behavior and the functionality of the cultures, giving a deeper knowledge in real time about the status and the course of the bioprocesses.

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