Caenorhabditis elegans DAF-16 regulates lifespan and immune responses to Cryptococcus neoformans and Cryptococcus gattii infections.

BACKGROUND: Cryptococcosis is a life-threatening infection is primarily caused by two sibling species Cryptococcus neoformans and Cryptococcus gattii. Several virulence-related factors of these cryptococci have been widely investigated in Caenorhabditis elegans, representing a facile in vivo model of host-pathogen interaction. While recent studies elucidated cryptococcal virulence factors, intrinsic host factors that affect susceptibility to infections by cryptococci remain unclear and poorly investigated. RESULTS: Here, we showed that defects in C. elegans insulin/insulin-like growth factor-1 (IGF-1) signaling (IIS) pathway influenced animal lifespan and mechanisms of host resistance in cryptococcal infections, which required the activation of aging regulator DAF-16/Forkhead box O transcription factor. Moreover, accumulation of lipofuscin, DAF-16 nuclear localization, and expression of superoxide dismutase (SOD-3) were elevated in C. elegans due to host defenses during cryptococcal infections. CONCLUSION: The present study demonstrated the relationship between longevity and immunity, which may provide a possibility for novel therapeutic intervention to improve host resistance against cryptococcal infections.

Regulation of DAF-16-mediated longevity and immune response to Candida albicans infection in Caenorhabditis elegans.

Candida albicans can cause infections ranging from superficial skin infections to life-threateningsystemic infections in immunocompromised hosts. Although several C. albicans virulence factorsare widely discussed in great detail, intrinsic host determinants that are critical for C. albicanspathogenesis remain less interested and poorly understood. In view of this, a model of Caenorhabditiselegans was used to study host longevity and immunity in response to C. albicans pathogenesis.The influence of C. albicans in pathological and survival aspects was evaluated using C. elegans.C. albicans hyphal formation in different C. elegans genetic backgrounds was evaluated. Moreover,several C. elegans fluorescent proteins as gene expression markers upon C. albicans infectionswere evaluated. C. albicans is pathogenic to C. elegans and reduces the lifespan of C. elegans inassociation with repression of the insulin/IGF-1-like signaling (IIS) pathway. Moreover, repressionof DAF-16/forkhead transcription factor increases aggressiveness of C. albicans by enhancing hyphalformation. In addition, infection of C. albicans increases lipofuscin accumulation, promotes DAF-16nuclear translocation, increases superoxide dismutase (SOD-3) expression, which coordinately linksbetween aging and innate immunity. Thus, we demonstrate here the strategy to utilize C. elegans asa model host to elucidate host genetic determinants that provide insights into the pathogenesis ofC. albicans infections.

Isolation of fungal communities and identification of Scedosporium species complex with pathogenic potentials from a pigsty in Phra Nakhon Si Ayutthaya, Thailand.

Soil fungal communities play an important role in regulating biogeochemical transformations, yet soil-related fungal pathogens are emerging threats to humans. Our previous studies have revealed the pathogenic Scedosporium species in soils samples from public parks with high human activities in Thailand. However, measurement and survey of soil fungal communities in other areas with high human/animal activities, such as the pigsty, are poorly determined. In this study, soil fungal pathogens from a pigsty were isolated and identified. Soil samples were collected from the surrounding drainage areas. Fungal species were identified using morphological and molecular analyses. Isolation of soil samples from the pigsty revealed at least 11 species that have been identified. The most abundant fungal species belonged to genera Aspergillus and Penicillium. Moreover, Scedo-Select III culturing and phylogenetic analysis with ß-tubulin gene sequencing revealed the three environmental isolates of Scedosporium species, which were consistent with the S.apiospermum. These three Scedosporium isolates were susceptible to voriconazole and caused pathological characteristics of scedosporiosis similar to S. apiospermum in vivo. In conclusion, our findings contribute towards a better understanding of soil-borne pathogenic fungi in the pigsty. The isolation of Scedosporium species with pathogenic potentials in the present study can be beneficial for the management of public health surveillance, epidemiologists, as well as physicians to reduce the risk of soil fungal contamination among pigsty workers.

Utilization of an in vitro biofabricated 3D skin as a pathological model of cutaneous candidiasis.

Candida albicans is an opportunistic fungal infectious agent that can cause cutaneous candidiasis in humans. Biofilms formation of C. albicans is thought to be the major cause of antifungal drug resistance. Despite numerous studies conducted on C. albicans biofilms, a comprehensive understanding of how C. albicans biofilms induced cutaneous candidiasis in humans and the development of a more effective targeted therapy remain poorly investigated. Available animal models of cutaneous candidiasis and in vitro human skin cell cultures do not fully reflect the actual human skin microenvironment or the disease pathogenesis. We investigated the molecular pathology of C. albicans infection using an in vitro biofabricated 3D skin. This in vitro biofabricated 3D skin comprises a fully humanized three-dimensional (3D) skin equivalent, consisting of a stratified terminally differentiated epidermis and an underlying dermal compartment. Antifungal drug susceptibility testing, histological and electron microscopy study, biofilms study, and pro-inflammatory cytokines analysis were conducted in C. albicans infected skin. Histological results revealed that C. albicans covered and produced biofilm on the in vitro biofabricated 3D skin, invading the skin compartments including epidermis and dermis. Elevation of proinflammatory cytokines including MMP-9, IL-1ß, TNF-α, and IL-5 were examined in the C. albicans infected skin. However, treatment with itraconazole reduced the pathology of C. albicans infection. This study provides an alternative pathological model of cutaneous candidiasis, which can physiologically represent a close-up event during C. albicans. Moreover, it is rapid, cost-effective, and reproducible of the in vitro biofabricated 3D skin model, and may further highlight the importance of utilizing in vivo-like conditions to improve high-throughput screening for drug discovery against several antifungal drug resistant pathogens.

The impact of Zika virus infection on human neuroblastoma (SH-SY5Y) cell line.

BACKGROUND & OBJECTIVES: An increase in Zika virus (ZIKV) epidemic during the last decade has become a major global concern as the virus affects both newborns and adult humans. Earlier studies have shown the impact of ZIKV infection in developing human foetus. However, effective in vitro model of target cells for studying the ZIKV infection in adult human neurons is not available. This study aimed to establish the use of human neuroblastoma cell line (SH-SY5Y) for studying an infection of ZIKV in vitro. METHODS: ZIKV growth kinetics, viral toxicity, and SH-SY5Y cell vialibity were determined after ZIKV infection in SH-SY5Y cells in vitro. ZIKV-infected SH-SY5Y cells were morphologically analysed and compared with nonhuman primate Vero cells. Furthermore, the susceptibility of SH-SY5Y cells to ZIKV infection was also determined. RESULTS: The results showed that ZIKV efficiently infects SH-SY5Y cell lines in vitro. Gradual changes of several cellular homeostasis parameters including cell viability, cytotoxicity, and cell morphology were observed in ZIKVinfected SH-SY5Y cells when compared to mock-treated or non-human primate cells. Interestingly, ZIKV particles were detected in the nucleoplasmic compartment of the infected SH-SY5Y cells. INTERPRETATION & CONCLUSION: The results suggest that ZIKV particle can be detected in the nucleoplasmic compartment of the infected SH-SY5Y cells beside the known viral replicating cytoplasmic area. Hence, SH-SY5Y cells can be used as an in vitro adult human neuronal cell-based model, for further elucidating the ZIKV biology, and highlight other possible significance of Zika virus distribution through nuclear localization, which may correlate to the neuropathological defects in ZIKV-infected adult humans.

INDUCTION BY EPIDERMOPHYTON FLOCCOSUM OF HUMAN FIBROBLAST MATRIX METALLOPROTEINASE-9 SECRETION IN VITRO.

Skin infection from pathogenic dermatophyte, Epidermophytonfloccosum, can cause serious health complications, especially in immuno-compromised patients. Proteolytic enzymes secreted from E. floccosum are required for host tissue degradation, facilitating fungal invasion. However, little is known regarding host matrix metalloproteinase (MMP) expression during E. floccosum infection. In this study human foreskin fibroblast (HFF) cell line was used to determine MMP-9 protease activity by gelatin zymography and amount by ELISA. E. floccosum-induced HFF secretion of MMP-9 in a time dependent manner, but HFF cell viability decreased. Treatment with an MMP inhibitor (SB-3CT) caused reduction in E. floccosum-induced secreted MMP-9 and improvement in HFF cell viability. These findings indicate a possible control measure for protecting skin from E. floccosum infection.

The study of tryptophol containing emulgel on fungal reduction and skin irritation.

Tryptophol (TOH), a fungal quorum-sensing molecule, that possesses anti-fungal activities for controlling the growth of human pathogenic fungi. In the present study, we developed TOH-containing emulgel formulations and examined the antifungal activities and potential use as topical treatments on the skin. The results showed that TOH-containing emulgel at 1000 µM has excellent physical characteristics as homogenous, stability, and inhibits the growth of 30 species of human pathogenic fungi in vitro. TOH-containing emulgel did not cause skin irritation in mouse model of irritation and in healthy human volunteers. Moreover, an increase in skin hydration and a decrease in trans-epidermal water loss (TEWL) were observed after TOH-containing emulgel treatment on human skin. Our findings indicated that TOH-containing emulgel can be utilize as an antifungal agent for topical treatment against fungal infections on the skin.

Infections of Cryptococcus species induce degeneration of dopaminergic neurons and accumulation of α-Synuclein in Caenorhabditis elegans.

Cryptococcosis in the central nervous system (CNS) can present with motor declines described as Parkinsonism. Although several lines of evidence indicate that dopaminergic (DA) neuron degeneration and α-synuclein accumulation contribute to the hallmark of Parkinsonism and Parkinson's disease (PD), little is known about cryptococcal infections associated with neuronal degeneration. In this study, the effects of Cryptococcus neoformans and C. gattii infections on dopaminergic neuron degeneration, α-synuclein accumulation, and lifespan in Caenorhabditis elegans were investigated. The results showed that cryptococcal infections significantly (P<0.05) induced DA neuron degeneration similar to a selective cathecholamine neurotoxin 6-hydroxydopamine (6-OHDA) in C. elegans (BZ555 strain) when compared to mock infected controls. Cryptococcal infections also significantly (P< 0.05) induced α-synuclein aggregation in C. elegans (NL5901 strain). Moreover, lifespan of the infected worms was significantly decreased (P<0.0001). In conclusion, DA neurodegeneration and α-synuclein accumulation are associated with lifespan reduction during cryptococcal infection in C elegans.

Development and efficacy of tryptophol-containing emulgel for reducing subcutaneous fungal nodules from Scedosporium apiospermum eumycetoma.

Background and purpose: Subcutaneous infections caused by Scedosporium apiospermum present as chronic eumycetomatous manifestations in both immunocompromised and immunocompetent individuals. Serious adverse effects/toxicities from the long-term use of antifungal drugs and antifungal resistance have been reported in patients with S. apiospermum infections. The present study aimed to determine the anti-S. apiospermum activities of fungal quorum sensing molecule known as tryptophol (TOH) and to develop a TOH-containing emulgel for treating S. apiospermum eumycetoma. Experimental approach: Anti-S. apiospermum activities of TOH were determined and compared with voriconazole. Effects of TOH on S. apiospermum biofilm formation and human foreskin fibroblast (HFF)-1 cell cytotoxicity were determined. Moreover, TOH-containing emulgel was developed and physical properties, in vitro, and in vivo antifungal activities against S. apiospermum eumycetoma were evaluated. Findings/Results: The minimal concentration of TOH at 100 µM exhibited anti-S. apiospermum activities by reducing growth rate, germination rate, and biofilm formation with less cytotoxicity to HFF-1 cells than voriconazole. Further study on the development of an emulgel revealed that TOH-containing emulgel exhibited excellent physical properties including homogeneity, consistency, and stability. Treatment by TOH-containing emulgel significantly reduced subcutaneous mass in a mouse model of S. apiospermum eumycetoma. The histopathological assessment showed marked improvement after 14 days of TOH-containing emulgel treatment. Conclusion and implications: TOH could be used as an anti-fungal agent against S. apiospermum infections. A novel and stable TOH-containing emulgel was developed with excellent anti-S. apiospermum activities suggesting the utilization of TOH-containing emulgel as an innovative therapeutic approach in the treatment of S. apiospermum eumycetoma.

Erythroderma and Skin Desquamation in Paederus Dermatitis.

Exfoliative erythroderma is rare but serious condition, which requires close supervision. We report a rare case of 28-year-old man with kissing lesions of Paederus dermatitis at his right side of neck. The rash caused by Paederus beetle was improved after treatments. However, the patient developed generalized erythema with desquamation and scaling. The patient was successfully treated topically with moisturizing liquid soap and topical moisturizer with emollients and humectants, and triamcinolone lotion was applied on the bright red lesion. The patient was also treated with oral replacement solution and tropical azelaic cream was applied on the hyperpigmented kissing lesion. This case report shows the importance of a diagnostic practice with follow-up examination.

Tryptophol Coating Reduces Catheter-Related Cerebral and Pulmonary Infections by Scedosporium apiospermum.

INTRODUCTION: Central venous catheter (CVC) is a medical device that is used to administer medication for a long duration. Colonization by an emerging opportunistic pathogen Scedosporium apiospermum in the CVC lumen is frequently reported to cause severe complications in patients. Here, we describe the effect of fungal quorum-sensing molecule (QSM) known as tryptophol (TOH) to control S. apiospermum colonization in catheter tube lumens in both in vitro and in vivo models. METHODS: Antifungal susceptibility of TOH against S. apiospermum was compared with voriconazole, and the colony diameter was determined on days 2, 4, and 6. Experimental catheterization rat model was conducted with pre-coating of TOH and voriconazole or an uncoated control and an infection with S. apiospermum. Biofilm formation on the catheter luminal surface was assessed using the scanning electron microscopy, crystal violet, and 2,3-bis(2-methoxy-4-ni-tro-5-sulfophenyl)-5-(phenylamino)-carbonyl-2H-tetra-zolium hydroxide (XTT) reduction assays. Brain and lung samples of catheterized rats were histopathologically assessed. Serum samples from catheterized rats were injected into Galleria mellonella larvae. Survival of catheterized rats and G. mellonella was determined. RESULTS: TOH impeded the growth of S. apiospermum by reducing the colony diameter in a dose-dependent manner. TOH coating remarkably lessened S. apiospermum biofilm formation and fungal cell viability on the catheter luminal surface. Additionally, TOH coating lessens cerebral edema that is associated with abscess and invasive pulmonary damages due to S. apiospermum catheter-related infection. Furthermore, TOH coating also lessened the virulence of S. apiospermum in sera of experimental catheterized rats and extended the survival rate of larvae Galleria mellonella infection model. CONCLUSION: An alternative modification of catheter by coating with TOH is effective in preventing S. apiospermum colonization in vivo. Our study gives a new strategy to control catheter contamination and prevents nosocomial diseases due to S. apiospermum infection.

Saponin-rich extracts from Holothuria leucospilota mediate lifespan extension and stress resistance in Caenorhabditis elegans via daf-16.

Saponins are secondary metabolite compounds that can be found in sea cucumbers (Holothuroidea spp.). However, little is known about how saponin-rich extracts from Holothuria leucospilota can delay and prolong the lifespan of the whole organism. In this study, anti-aging effects of H. leucospilota extracts were studied on Caenorhabditis elegans. NMR analysis revealed that body wall n-butanol-extract of H. leucospilota (BW-BU) is saponin-rich. BW-BU extracts exhibited antioxidant activities by 2,2'-diphenyl-2-picrylhydrazyl assay (EC50  = 10.23 ± 0.12 mg/ml) and 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid assay (EC50  = 3.91 ± 0.04 mg/ml). BW-BU extracts increased lifespan of L4 and L1 C. elegans (5.92% and 15.76%, respectively), which also increased worm growth, stress resistance, and reduced biomarkers for aging. BW-BU extracts activated DAF-16 nuclear localization and upregulated daf-16 and DAF-16 target genes expression. Taken together, this study revealed the evidences on anti-aging activities of saponin-rich extracts from H. leucospilota, which can extend lifespan of C. elegans via daf-16. PRACTICAL APPLICATIONS: In recent years, age-associated chronic diseases have had a significant impact on quality of life. Many natural compounds exhibit anti-aging activities, especially in sea cucumber, H. leucospilota. Our results indicated that H. leucospilota is good for health. Extracts from H. leucospilota contain a bioactive compound that can be potentially used to promote longevity and disease prevention in aging.

In vitro screening of 10 edible thai plants for potential antifungal properties.

Growing rates of fungal infections and increasing resistance against standard antifungal drugs can cause serious health problems. There is, therefore, increasing interest in the potential use of medicinal plants as novel antifungal agents. This study investigates the antifungal properties of crude plant extracts from ten medicinal plant species. Crude samples were extracted using the hot water extraction process. The minimum inhibitory concentrations (MIC) and diameter zone of inhibition were determined in each extract against ten fungal strains, and fluconazole was used as a positive control. The cytotoxicity of crude extracts on in vitro human skin fibroblast (HSF) cell models was determined by MTT assay. Of the ten crude extracts, Psidium guajava L. exhibited the highest antifungal activity, diameter zone of inhibition, and percentage HSF cell viability. Although all extracts exhibited antifungal activity, Psidium guajava L. had the greatest potential for developing antifungal treatments.