Germ Tube Formation Research Articles

The Potential Association of Human ABO Blood Group in Candida albicans Germination

Candidiasis is an infection caused by Candida spp., mostly Candida albicans. Infection with Candida spp. in immunocompetent individuals is often limited to the mouth, genital area or skin. However, patients who are immunocompromised due to infections or medications including corticosteroids or anticancer drugs are more susceptible to systemic candidiasis and candidemia. Upon C. albicans infections, the pathogenic yeasts enter the bloodstream and disseminate to the remote organs, leading to the formation of germ tube. Dimorphic adaptation is one of contributing virulent factors in C. albicans, which is responsible for disease pathogenesis and severity. However, little is known about whether differences among human ABO blood groups facilitate the development of C. albicans germ tube. Therefore, this study investigated the germ tube formation of C. albicans that grown in human serums from different ABO blood groups. Human pool serums varying ABO blood groups from healthy individuals were tested for C. albicans germ tube induction. After incubation for 2 h at 37 °C, the rate of germ tube formation at 15-min intervals was determined. Serum from human blood group A exhibited the highest rate of germ tube formation. Humans with blood group A exhibited the unique serum biochemical structure that possibly helps facilitate germ tube formation during C. albicans infection. HIGHLIGHTS The association of blood groups of Thais to germination and invasion of Candida albicans were more common in Thai people. This research study focused on changes in Candida albicans germination related to the blood group. Human with blood group type A exhibited the highest rate of germ tube formation. GRAPHICAL ABSTRACT

Antimicrobial effect of oral care gel containing hinokitiol and 4-isopropyl-3-methylphenol against intraoral pathogenic microorganisms.

Deterioration of oral hygiene is closely related to an increase in severity and mortality of corona virus disease-19 (COVID-19), and also contributes to the development of various diseases such as aspiration pneumonia or Alzheimer's. Oral care is attracting high interest in Japan, which has entered a super-aging society. In this study, we aimed to investigate whether commercially available Hinora® (HO), an oral care gel containing hinokitiol and 4-isopropyl-3-methylphenol (IPMP), has biofilm formation inhibitory and antimicrobial activities against various intraoral pathogen microorganisms. Candida species, Aggregatibacter actinomycetemcomitans, Staphylococcus aureus, and Pseudomonas aeruginosa were selected during the study period, all of which were analyzed using antimicrobial disc, microorganism turbidity, and crystal violet assays. In addition, the germ tube test using Candida albicans (C. albicans) was performed with a modification of Mackenzie's method. Images for morphological observation of the germ tubes were acquired using an inverted microscope. For comparison between products, we used Refrecare® (RC), which only contains hinokitiol (not containing IPMP). All the intraoral pathogenic microorganisms showed drug susceptibility against undiluted forms of HO and/or RC. In particular, HO was more effective at lower concentrations than RC. In the HO-added group, inhibition circles were observed in all bacteria except P. aeruginosa when added at a concentration of 0.5 g/mL or more. The optical density values at 590 nm (crystal violet) and/or 600 nm (microorganism turbidity) of all the fungi and bacteria were significantly lower when cultured in medium with HO. Inhibition of growth or biofilm formation was observed when HO was added at a concentration of 0.05 g/mL or higher. To investigate the action mechanism of HO, germ tube tests were performed in C. albicans. The results showed that culturing C. albicans in soybean-casein digest broth with HO (0.05 g/mL) significantly suppressed germ tube formation. These data suggest that oral care gel-containing hinokitiol and IPMP has strong biofilm formation inhibitory activity, as well as antifungal and antimicrobial effects against Candida fungi and multiple intraoral pathogenic microorganisms. Therefore, it may be a promising treatment option for oral infections.

Detection of Candida species isolated from immunocompromised patients in AL-Najaf province

ABSTRACT Study was carried out to examine phenotyping as a means of identifying of fungal species isolated by those who suffer from Thalassemia in the province of Al-Najaf and to assess function Inhibition of Some Antifungal Agents (Nystatin and Amphotericin B). Fifty samples, including oral swabs, were taken from the AL-Zahraa Teaching Hospitel in the AL Najaf province between October and November 2021, but only 14 showed good growth. All samples were cultured in the Microbiology Department of the Faculty of Medical Technologies at the Islamic college of Najaf, I plated them first on Sabouraud dextrose agar, and then on chromagar medium. to identify the species of Candida based on colony color. On chromagar medium, the results read as follows: C.albicans 10, C.parapsilosis 3, and C. tropicalis 1. Candida albicans' virulence factor production, including the formation of germ tubes, the formation of biofilms, and the production of phospholipase enzymes, was examined in this research. All virulence factors, such as germ tubes, biofilms, and the phospholipase enzyme, are produced by C. albicans, as evidenced by the culture findings.
 This research used a disk diffusion technique to describe the effects of the antifungal drugs nystatin and amphotercin B on the yeast species Candida. Based on these results, Amphotercin B is preferable to Nystatin for treating Candida spp..

Synergistic effects of Thymus vulgaris essential oil in combination with antifungal agents and inhibition of virulence factors of Candida albicans

BackgroundCombination antifungal therapy has become a prominent approach in medical practice as it takes advantage of synergistic interactions by interfering with multiple targets of the pathogen, broadening the spectrum of activity, reducing the development of resistance, and improving treatment outcomes. Therefore, combining conventional antifungal drugs with natural products can increase antifungal activity, reduce side effects, and optimize therapeutic effects. PurposeThe present study aimed to evaluate the antifungal activity of thyme essential oil (ThyEO) in combination with amphotericin B, caspofungin, fluconazole, itraconazole, and posaconazole (PSZ), as well as its inhibitory effect on Candida albicans virulence factors. Materials and MethodsThe microbroth dilution assay was employed to determine the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC). Interactions were assessed using the microdilution checkerboard assay. Ergosterol and methylene blue assays were used to detect effects on fungal membrane, while the sorbitol assay was used to detect effects on fungal cell wall. Inhibition of yeast virulence factors (adherence to epithelial cells, germ tube and pseudomycelium formation, secretion of hydrolytic enzymes, and biofilm formation) was assessed with previously reported methods. ResultsAlmost all combinations showed additivism against C. albicans, except ThyEO/PSZ (31.25/0.0039 µg/ml), which showed partial synergism. Furthermore, all mixtures were fungicidal against C. albicans strains. ThyEO/PSZ, its components alone, and thymol were shown to disrupt the fungal cytoplasmic membrane, increasing its permeability. ThyEO/PSZ, at sub-inhibitory concentrations, significantly decreased the ability of C. albicans to adhere to buccal epithelial cells. ThyEO/PSZ, ThyEO, and PSZ were able to reduce the pseudomycelium production of C. albicans while thymol completely inhibited its formation. ThyEO/PSZ, each combination component on its own, and thymol inhibited biofilm formation and preformed biofilm of C. albicans. Notably, ThyEO/PSZ showed synergistic and fungicidal activity against a resistant strain of C. albicans, reducing the PSZ dose by 4-fold. ConclusionThese findings make ThyEO and ThyEO/PSZ mixture valuable candidates for the development of alternative antifungals with a lower incidence of adverse effects.

The Zanthoxylum armatum fruit's oil exterminates Candida cells by inhibiting ergosterol biosynthesis without generating reactive oxygen species.

Candida spp. is a significant cause of topical and fungal infections in humans. In addition to Candida albicans, many non-albicans species such as C. krusei, C. glabrata, C. parapsilosis, C. tropicalis, C. guilliermondii cause severe infections. The main antifungal agents belong to three different classes, including azoles, polyenes, and echinocandins. However, resistance to all three categories of drugs has been reported. Therefore, there is an urgent need to search for other alternatives with antifungal activity. Many herbal extracts and compounds from natural sources show excellent antifungal activity. In this study, we used an oil extract from the fruits of Zanthoxylum armatum, which showed significant antifungal activity against various Candida spp. by two different methods-minimum inhibitory concentration (MIC) and agar diffusion. In addition, we attempted to explore the possible mechanism of action in C. albicans. It was found that the antifungal activity of Z. armatum oil is fungicidal and involves a decrease in the level of ergosterol in the cell membrane. The decrease in ergosterol level resulted in increased passive diffusion of a fluorescent molecule, rhodamine6G, across the plasma membrane, indicating increased membrane fluidity. The oil-treated cells showed decreased germ tube formation, an important indicator of C. albicans' virulence. The fungal cells also exhibited decreased attachment to the buccal epithelium, the first step toward invasion, biofilm formation, and damage to oral epithelial cells. Interestingly, unlike most antifungal agents, in which the generation of reactive oxygen species is responsible for killing, no significant effect was observed in the present study.

K-aurein: A notable aurein 1.2-derived peptide that modulates Candida albicans filamentation and reduces biofilm biomass.

Candida albicans is considered one of the most important opportunistic fungi due to the large arsenal of virulence factors that help throughout the progress of the infection. In this sense, antimicrobial peptides (AMPs) appear as an alternative, with great antifungal action. Among these, aurein 1.2 has been widely explored, becoming the basis for the discovery of new AMPs, such as K-aurein (K-au). Thus, this study evaluated the anti-C. albicans potential of K-au against virulence factors, planktonic growth, and biofilm formation of clinical isolates. Firstly, K-au antifungal activity was determined by the microdilution method and time-kill curve. The inhibition of hydrolytic enzyme secretion (proteinase, phospholipase, and hemolysin) and germ tube formation was tested. Then, the antibiofilm potential of K-au was verified through biomass quantification and scanning electron microscopy (SEM). All tests were compared with the classical antifungal drug, amphotericin B (AmB). The outcomes showed fungicidal action of K-au at 62.50µgmL-1 for all isolates, with a time of action around 150-180min, determined by the time-kill curve. K-au-treated cells decreased by around 40% of the germinative tube compared to the control. Additionally, K-au inhibited the biofilm formation by more than 90% compared to AmB and the control group. SEM images show apparent cellular disaggregation without the formation of filamentous structures. Therefore, the findings suggest a promising anti-C. albicans effect of K-au due to its fungicidal activity against planktonic cells, or its ability to inhibit important virulence factors like germ tube and biofilm formation. Thus, this peptide could be explored as a useful compound against C. albicans-related infection.

Antifungal activity of Allyl isothiocyanate by targeting signal transduction pathway, ergosterol biosynthesis, and cell cycle in Candida albicans.

In recent years, the inclusion of Candida albicans on the list of infections that pose a threat due to drug resistance has urged researchers to look into cutting-edge and effective antifungal medications. In this regard, the current study investigated the probable mode of action of allyl isothiocyanate (AITC) against Candida albicans. In this study, planktonic assay, germ tube inhibition assay, adhesion, and biofilm formation assay were performed to check the growth and virulence factors. Furthermore, ergosterol assay, reactive oxygen production analysis, cell cycle analysis, and quantitative real-time polymerase chain reaction analysis were performed with the aim of finding the mode of action. A biomedical model organism, like a silkworm, was used in an in vivo study to demonstrate AITC anti-infective ability against C. albicans infection. Allyl isothiocyanate completely inhibited ergosterol biosynthesis in C. albicans at 0.125 mg/ml. Allyl isothiocyanate produces reactive oxygen species in both planktonic and biofilm cells of C. albicans. At 0.125 mg/ml concentration, AITC arrested cells at the G2/M phase of the cell cycle, which may induce apoptosis in C. albicans. In quantitative real-time polymerase chain reaction analysis, it was found that AITC inhibited virulence factors, like germ tube formation, at 0.125 mg/ml concentration by downregulation of PDE2, CEK1, TEC1 by 2.54-, 1.91-, and 1.04-fold change, respectively, and upregulation of MIG1, NRG1, and TUP1 by 9.22-, 3.35-, and 7.80-fold change, respectively. The in vivo study showed that AITC treatment successfully protected silkworms against C. albicans infections and increased their survival rate by preventing internal colonization by C. albicans. In vitro and in vivo studies revealed that AITC can be an alternative therapeutic option for the treatment of C. albicans infection.

The Involvement of the Laccase Gene Cglac13 in Mycelial Growth, Germ Tube Development, and the Pathogenicity of Colletotrichum gloeosporioides from Mangoes

Colletotrichum gloeosporioides is one of the most serious diseases that causes damage to mangoes. Laccase, a copper-containing polyphenol oxidase, has been reported in many species with different functions and activities, and fungal laccase could be closely related to mycelial growth, melanin and appressorium formation, pathogenicity, and so on. Therefore, what is the relationship between laccase and pathogenicity? Do laccase genes have different functions? In this experiment, the knockout mutant and complementary strain of Cglac13 were obtained through polyethylene glycol (PEG)-mediated protoplast transformation, which then determined the related phenotypes. The results showed that the knockout of Cglac13 significantly increased the germ tube formation, and the formation rates of appressoria significantly decreased, delaying the mycelial growth and lignin degradation and, ultimately, leading to a significant reduction in the pathogenicity in mango fruit. Furthermore, we observed that Cglac13 was involved in regulating the formation of germ tubes and appressoria, mycelial growth, lignin degradation, and pathogenicity of C. gloeosporioides. This study is the first to report that the function of laccase is related to the formation of germ tubes, and this provides new insights into the pathogenesis of laccase in C. gloeosporioides.

Crinum latifolium mediated biosynthesis of gold nanoparticles and their anticandidal, antibiofilm and antivirulence activity

Candida spp. is one of the most common opportunistic human fungal pathogens, responsible for 90–100% of mucosal infections. Germ tube formation, hyphal morphogenesis, the production of tissue-damaging extracellular enzymes, and drug-resistant biofilm formation contribute to their pathogenicity, which can lead to systemic infections in the worst scenarios. Thus, there is an urgent need to discover new therapeutic agents to overcome the above virulence factors. Therefore, we aimed to prepare Crinum latifolium leaves-mediated biosynthesis of gold nanoparticles (AuNPs) that was characterized by various sophisticated techniques, and further their antifungal, antibiofilm, and anti-virulence activities was investigated. The AuNPs show a zone of inhibition between 19 and 22 mm for test strains of Candida spp. at 1000 µg/ml, whereas the MIC values were ranged from 250 to 500 µg/ml. AuNPs inhibit germ tube formation in C. albicans by 93.3% at 50 µg/ml. Furthermore, exposure to AuNPs significantly reduced the secretions of phospholipase, proteinase, hemolysin, esterase, and lipase. Confocal laser scanning microscopy (CLSM) investigation shows that 25 µg/ml of AuNPs significantly inhibit colonization and biofilm formation. Lastly, the interaction of Candidal cells with AuNPs revealed, ultrastructural changes in the cell wall and cell membranes as visualized by a transmission electron microscope (TEM).

Comparison of Several Types of Plasma as Media in the Germ Tube Test for Identification of Candida Albicans

Candida albicans is a commensal on the oral mucosal surfaces of healthy individuals but can be pathogenic in immunocompromised individuals. The germ tube test is an inexpensive, faster, and easier method used to identify and differentiate it from other yeast species. Generally, the test is carried out using serum as the medium. Plasma overall has the same characteristics as serum and differs only in the content of clotting factors. In addition, plasma is easier to obtain than serum in most healthcare services. This study aims to demonstrate three types of plasma as alternative media for germ tube test. CPDA (Citrate Phosphate Dextrose Adenine), EDTA (Ethylenediaminetetraacetic), and Sodium Citrate plasma were compared to serum in producing of germ tubes. Each plasma and serum was tested in five repetitions. The experimental method was carried out in Mycology Laboratory at Health Polytechnic of Jambi. The data were observed based on the germ tube formation time and analyzed with one-way ANOVA test. The result showed the lengths of germ tube formation time were significantly different. Although CPDA plasma was also significantly different compared to serum, it showed the average time which is at the minimum limit of the incubation time range that is recommended in the standard protocol of the test. This study concluded that plasma can be used as a substitute for serum in the germ tube tes, especially CPDA plasma, as it is faster in formating time, easier to obtain, and considered safer to use than others.

Germ Tube Induction Test Comparing Total of Six Liquid and Three Solid Media in Candida albicans

Invasive candidiasis (IC) has a high mortality rate of 70%, thus diagnosis should be established without delay. Given its fast result, serological test such as β-d-glucan (BDG) test is one alternative diagnosis modalities. However, it lacks specificity. Candida albicans germ tube antibody (CAGTA) test is an alternative serological test which has a high sensitivity of 76.2% and specificity of 80.3%. Manufacturing CAGTA serological test requires provision of specific germ tube antigen. In this study, various culture media were tested to find the best media for germ tube induction. This study was an experimental in vitro study. The number and length of the germ tube were recorded in two- and three-hour incubation periods. A total of six samples containing one C. albicans ATCC 90028, four C. albicans wild type strains, and one C. krusei wild type strain were used. Nine media were tested to induce germ tube formation: human and sheep serum, fetal bovine serum, mueller hinton agar and broth, tryptic soy agar and broth, brain heart infusion agar and broth. At both incubation periods, the medium with the highest number of germ tube was human serum (p=0.001 and p=0). The longest germ tube was found in sheep serum at two-hour incubation period (p=0.005). Mueller hinton broth (MHB) showed comparable results with human and sheep serum (p>0.05). Human serum is a superior inducer of morphogenesis. However, the use of MHB is recommended in this study, since provision of fresh human and sheep serum on a regular basis is impractical.

Key essential oil components delocalize Candida albicans Kar3p and impact microtubule structure

BackgroundTreatment of Candida albicans associated infections is often ineffective in the light of resistance, with an urgent need to discover novel antimicrobials. Fungicides require high specificity and can contribute to antifungal resistance, so inhibition of fungal virulence factors is a good strategy for developing new antifungals. ObjectivesExamine the impact of four plant-derived essential oil components (1,8-cineole, α-pinene, eugenol, and citral) on C. albicans microtubules, kinesin motor protein Kar3 and morphology. MethodsMicrodilution assays were used to determine minimal inhibitory concentrations, microbiological assays assessed germ tube, hyphal and biofilm formation, confocal microscopy probed morphological changes and localization of tubulin and Kar3p, and computational modelling was used to examine the theoretical binding of essential oil components to tubulin and Kar3p. ResultsWe show for the first time that essential oil components delocalize the Kar3p, ablate microtubules, and induce psuedohyphal formation with reduced biofilm formation. Single and double deletion mutants of kar3 were resistant to 1,8-cineole, sensitive to α-pinene and eugenol, but unimpacted by citral. Strains with homozygous and heterozygous Kar3p disruption had a gene‐dosage effect for all essential oil components, resulting in enhanced resistance or susceptibility patterns that were identical to that of cik1 mutants. The link between microtubule (αβ-tubulin) and Kar3p defects was further supported by computational modeling, showing preferential binding to αβ-tubulin and Kar3p adjacent to their Mg2+-binding sites. ConclusionThis study highlights how essential oil components interfere with the localization of the kinesin motor protein complex Kar3/Cik1 and disrupt microtubules, leading to their destabilization which results in hyphal and biofilm defects.

The influence of temperature and vapour pressure deficit on conidia germination and germ tube production in an Australian Podosphaera xanthii isolate

Powdery mildew of cucurbits, caused by Podosphaera xanthii, is an economic constraint in cucurbit production worldwide. This study examined the influence of temperature and vapour pressure deficit (VPD) on the rate of conidial germination and the formation of germ tubes in an Australian P. xanthii isolate 12 – 48 h after inoculation. Two experiments were prepared by inoculating cucumber, cv. crystal salad, leaf-discs using a spore settling tower. The first experiment incubated inoculated cucumber leaf-discs at eight temperatures between 8 and 35 °C under saturated vapour pressure (SVP), the second compared 18 VPD conditions between 0.038 and 1.797 kPa, in six humidity chambers (33% – 99% relative humidity) and three temperatures (22 °C, 25 °C, 28 °C). Leaf-discs were cleared, stained and microscopically inspected for conidial germination and the number of germ tubes. The optimal temperature for germination was 28ºC at SVP, where more than 50% of conidia had germinated by 12 h, and 85% by 48 h. Fewer germinated conidia were recorded after 12 h at other temperature treatments between 17 °C and 31 °C. The germination percentage and germination rate were significantly lower when vapour pressure was between 0.13 and 2.5 kPa, with germination in response to VPD varying by approximately 10%, indicating difficulty associating conidial germination to VPD above 0.13 kPa. Germ tube production was highest between 25 ºC and 28 ºC at the lowest VPD treatment at near SVP, with more than 50% of the germinated conidia producing at least three germ tubes. Germination and formation of germ tubes significantly reduced when VPD increased. P. xanthii conidia were able to produce a primary germ tube under relatively dry conditions, such as 2.53 kPa, but these results show infection would be less likely and require longer incubation. This study provides the first crucial step in simulating powdery mildew infections on cucurbit plants and may lead to a model capable of providing risk forecasts or fungicide management decision support tools.

Evaluation of germ tube in Candida albicans identification by different media

Background: Candida albicans is a common pathogenic, and the germ tube test is one of the identification techniques for this fungal species. Objectives: 1. To experiment with the germ tube test of C. albicans in various media according to the different testing time. 2. To evaluate capacity ofthe germ tube test in C. albicans identification in various media. Methods: This study was carried out with 36 C. albicans isolates and 10 C. tropicalis isolates stored at the Department of Parasitology. Germ tube test was performed using various media, including human sera, 1% peptone medium, BHI broth, egg white, egg yolk, and fetal bovine serum. These experiments were checked after two hours, three hours, and four hours of incubation. Results: The highest rate of germ tube formation of C.albicans was recognized after 3 hours of incubation. The germ tube rates on human serum, 1% peptone medium, egg white, egg yolk, and fetal bovine serum were 91.67%; 58.33%; 25%; 33.33%; 91.67%; and 100%, respectively. The sensitivity, specificity, positive predictive value, and negative predictive value of the germ tube test on serum and egg yolk medium were 91.67%, 100%, 100%, and 76.92%. Conclusions. The capacity to identify C. albicans of the germ tube test performed on fetal bovine serum, human serum and yolk egg could identify 100%, 91.67%, and 91.67%, respectively. It suggested that egg yolk could be used in diagnostic labs as well as mycology researches and training labs to perform germ tube test production to replace human serum since it is a simple and low cost procedure. Key words: Candida albicans, germ tube, fetal bovine serum, yolk egg

The synergistic effects of hydroxychavicol and amphotericin B towards yeast-hyphae transition and the germination of Candida albicans

ObjectivesDimorphic transformation from yeast cells to hyphae is considered one of the major virulence factors of candidal species. The development of antifungal resistance against several candida diseases has led researchers to find plant derived alternatives. We aimed to determine the effect of hydroxychavicol (HC), Amphotericin B (AMB), and their combination (HC + AMB) on the transition and germination of oral Candida species. MethodsThe antifungal susceptibility of hydroxychavicol (HC) and Amphotericin B (AMB) separately and in a mixture (HC + AMB) against Candida albicans ATCC 14053, Candida parapsilosis ATCC 22019, Candida tropicalis ATCC 13803, and Candida dubliniensis ATCC MYA-2975 was determined by broth microdilution technique. Minimal Inhibitory Concentration was calculated based on the CLSI protocols. The MIC50, fractional inhibitory concentration (FIC) index, and IC50 were also determined. The IC50 values were used as the treatment concentration of HC, AMB, and HC + AMB to study the effect of antifungal inhibition on yeast hypha transition (gemination). The germ tube formation percentage of candida species was calculated at several intervals using a colorimetric assay. ResultsThe MIC50 range of HC alone against Candida species was between 120-240 µg per mL while that of AMB was between 2-8 µg per mL, respectively. The combination of HC + AMB at 1:1 and 2:1 demonstrated the strongest synergistic activity against C. albicans with an FIC index of 0.07. Moreover, within the first hour of treatment, the total percentage of germinating cells was significantly reduced by 79% (p < 0.05). ConclusionThe combination of HC + AMB displayed synergism and inhibited C. albicans hyphal growth. HC + AMB combination slowed the germination process and exhibited consistent prolonged effect up to 3 h post-treatment. The results of this study will pave the way for potential in vivo studies.

Detection of Virulence Factors from Candida spp. Isolated from Oral and Vaginal Candidiasis in Iraqi Patients.

Yeast-like fungi (YLF) of the genus Candida are unicellular microorganisms of relatively large size and rounded shape, aerobes, and belong to conditionally pathogenic microorganisms. The genus Candida includes approximately 150 species, which are classified as Deuteromycetes due to the absence of a sexual stage of development. This study aimed to identify virulence factors from Candida spp. isolated from oral and vaginal candidiasis. Fifty-eight oral and vaginal swab specimens were collected from patients, including (28) oral swabs from children and (30) vaginal swabs from different infected women. All isolates were subjected to direct examination, Morphological tests, Germ tube formation, growth at 45ºC, CHROM agar Candida culture, and VITEK 2 Compact system to ensure this diagnosis. (31) isolates were identified as Candida spp., including (21) (C. albicans (14), C. glabrata (1), C. guilliermondii (2), C. dubliniensis (3), C. parapsilosis (1)) were isolated from oral swabs and (10) isolates includedC. parapsilosis (4), C. albicans (6) were isolated from vaginal swabs. Moreover, these isolates had been detected to have some virulence factors, including phospholipase, esterase, proteinase, coagulase, hemolysin, and biofilm formation. Different species of Candida were isolated and identified from oral and vaginal. Phospholipase (Pz), Esterase (Ez), and Proteinase (Prz) were produced by 19 (61.29%), 16 (51.61%), and 26(83.87%), respectively, out of 31 isolates, whereas. All isolates produce coagulase enzyme except C. dubliniensis, which did not produce coagulase enzyme. All Candida spp. isolates produce hemolysin and biofilm formation in different percentages.

Cladosporium cladosporioides causes leaf blight on garden pea in Telangana, India.

Garden pea (Pisum sativum L.) is an important vegetable and pulse crop grown worldwide. Leaf blight symptoms with chlorotic lesions were observed on the pea genotypes, 2006-2008A and Arkel in field conditions at Asian Vegetable Research and Development Centre (AVRDC), Hyderabad (17°23' N; 78°29' E) and greenhouse conditions at University of Hyderabad, Hyderabad, India (17° 27' N; 78° 19' E) since 2010. The blight symptoms showed black lesions coated by a fine velvety layer of olive-grey conidia. The disease incidence on field and greenhouse pea plants ranged from 3-6%. The Cladosporium isolates, Gp01 and Gp02 were isolated from pea genotypes, Arkel and 2006-2008A in greenhouse and field conditions respectively. One fungus was predominantly isolated from infected leaf tissue plated onto potato dextrose agar (PDA), and it was identified as Cladosporium sp. based on colony morphology and conidial appearance. Single-spored cultures grown at 26±2 °C on PDA showed olive green mycelia on the obverse-side and dark green on reverse-side (Fig. 1 a,b). The conidia were limoniform with dimensions ranging from 5.91-7.33 x 3.19-4.58 µm (Table 1) and were produced on solitary conidiophores containing 1-2 septa, with no constriction whereas the aerial hyphae were septate and unbranched or rarely branched (Fig.1 c-d). To verify the pathogenicity of the fungus, detached leaf assay was carried on 30-day old Arkel leaves by point inoculation with 20 µL (106 spores/ml) spore suspension using a needle-less syringe to infiltrate the spores into the abaxial surface and the inoculated leaves were incubated at 26±2 °C for 3 days in petri plates containing moistened autoclaved absorbent cotton. Pea leaves inoculated with water served as controls. The experiment was repeated with a minimum of three times with five leaves per replication and three replications per experiment for each of the isolates. The appressorial germ tube formation was observed by scanning electron microscopy (SEM) at 12 hours post inoculation (hpi) (Fig.1 e-f). Thirty-day old pea plants were spray-inoculated with a spore suspension of 1-2 ml per plant with a concentration of 106 spores/ ml while the control plants were mock-inoculated (sprayed) with water. 5 plants per replicate per isolate with three replications per experiment and a minimum of three experiments were carried. The inoculated plants were covered with polythene zip-lock bags for two days post inoculation (dpi) and maintained in greenhouse at 26±2 °C with natural photoperiod of 14 hours. Wilting and necrosis symptoms accompanied by chlorotic lesions were observed on pea plants at 7 days post inoculation (dpi) (Fig. 2). Complying with Koch's postulates, the pathogen, which was re-isolated from the first appearance at 3 dpi from infected pea showed similar morphological characteristics as those used for inoculation. Sequencing analysis of the internal transcribed spacer (ITS) region using ITS1 and ITS4 primers (White et al. 1990), D1-D2 region sequencing (Kwiatkowski et al. 2012), Translation elongation factor 1-alpha (TEF) and Actin (ACT) gene sequencing (Nam et al. 2015) of Gp01 and Gp02 showed significant similarity with C. cladosporioides (Table S1). Further, concatenated phylogenetic trees constructed by maximum likelihood method using all the four (ITS, D1-D2, TEF and ACTIN) gene sequences of these isolates along with other reported Cladosporium species (Fig. S1a) and specific isolates of C. cladosporiodes which are reported as "sensu stricto" (Bensch et al. 2010) collected from different substrata (Fig. S1b) confirmed them as C. cladosporioides. Gp01 and Gp02 were also confirmed by Microbial Type Culture Collection (MTCC, India) as C. cladosporioides with Acc. No. MTCC 9994 and MTCC 9995. C. cladosporioides has been previously reported on Dalbergia sp. and Eucalyptus sp in India and on garden pea in other countries (Bensch et al. 2010). However, owing to the impact of this pathogen on growth and productivity of pea crop, further in-depth study needs to be carried. To our understanding this is the first report of Cladosporium cladosporioides infecting garden pea in India.

Isolation, Identification And Toxin Production From Vaginal Clinical Isolates Of Candida Albicans

Thirty two isolates of candida albican were sequestered from female patients with vaginal ulcer (lying in marjan hospital hillia city) .The vaginal swap were cultured on&nbsp; sabouraud dextrose broth and agar plate respectively . for differential identification of C. albican, hi chrome candida agar was used. Gram staining used to smear of vaginal scrapings. suspected isolates of C.albican then immunized on corn meal agar&nbsp; mediocre for chlamydospore, formation , for germ tube formation the isolates of&nbsp; Caliban was cultured in human serum .Carbohydrate assimilation and fe mentation&nbsp; tests also used for further identification . four identification s trains of C.albican namely (&nbsp; CA1, CA2, CA3, CA4) were selected according to their antagonistic activity against&nbsp; standard strain of bacteria E.coli NCTC5933 and staphylococcus aureus NCTC6571 ;&nbsp; To produced extracellular toxin of glycoprotein nature in solid media and fermentation&nbsp; liquid media . The toxin which produced (secondary metabolites) don’t effecting the producers strains of C.albican (CA1,CA2,CA3,CA4) ,but the produced toxin was&nbsp; inhibited the growth of standard strain of candida albicans ATcc8201( sensitive ).&nbsp; Simple method used for toxin extraction by using ethanol 90%. And the extracted toxin&nbsp; from free cells supernatant . cytotoxicity of toxin against human blood cells were 6 ppm also, the stability of producing toxin was determined.

Prevalence of Candidiasis among Pregnant and Non-Pregnant Women in Eleme and Okrika Local Government Areas of Rivers State

Candidiasis is a common opportunistic fungal infection caused by yeast, which affects mainly women. The prevalence of candidiasis among pregnant and non-pregnant women in two Local Government Areas of Rivers State, Nigeria was investigated. A total of ninety-five women from ages of 18 to 46 years were selected for the study. The aim of this study was to determine the prevalence of vaginal candidiasis among pregnant and non-pregnant women in two Local Government areas of Rivers State. The study population included pregnant and non-pregnant women. Vaginal specimens were collected and cultured on Sabouraud Dextrose Agar (SDA) and also sub cultured on Hi Crome Candida differential Agar. The isolates obtained were subjected to gram staining, germ-tube test and sensitivity testing. Nystatin and fluconazole are used as antifungal agent. Identified Candida species using cultured technique include Candida albicans, Candida glabrata, Candida tropicalis and Candida parapsilopsis. Results further revealed a higher prevalence rate of candidiasis in pregnant women 29.4% compared to non-pregnant women 13.68%. Results of distribution of candidiasis according to trimester of pregnancy showed that Candida albicans and Candida glabrata were more prevalent in pregnant women while the distribution for non-pregnant women showed that Candida albicans were the most implicated species that causes vaginal candidiasis. Twenty-six Candida albicans species were subjected to germ-tube test and ten isolates were confirmed positive to germ-tube formation. This study showed lower rate of candidiasiss in women due to high rate of personal hygiene and medical facilities provided and also the study showed a higher prevalence in pregnant women compare to non-pregnant women. The women at great risk were those between the ages of 26 to 35 years and in their third trimester. Candida albicans was the most prevalent vaginal Candida species across all age groups and trimesters. Appropriate health education is necessary to reduce candidiasis since poor hygiene can lead to opportunistic infection.

A Melanin-Deficient Isolate of Venturia inaequalis Reveals Various Roles of Melanin in Pathogen Life Cycle and Fitness.

Venturia inaequalis is the ascomycetous pathogen causing apple scabs and forms dark-pigmented spores and partially melanised infection structures. Although melanin is considered to be essential for the infection of host tissue, a spontaneously occurring melanin-deficient mutant was isolated from an abaxial side of an apple leaf and can be cultivated in vitro as well as in vivo. The morphology and development of the melanin-deficient-isolate SW01 on leaves of susceptible apple plants were compared to that of the corresponding wild-type isolate HS1. White conidia of SW01 were often wrinkled when dry and significantly increased their volume in suspension. Germination and formation of germtubes and appressoria were not impaired; however, the lack of melanisation of the appressorial ring structure at the interface with the plant cuticle significantly reduced the infection success of SW01. The colonisation of leaf tissue by non-melanised subcuticular hyphae was not affected until the initiation of conidiogenesis. Non-melanised conidiophores penetrated the plant cuticle from inside less successfully than the wild type, and the release of white conidia from less solid conidiophores above the cuticle was less frequent. Melanin in the outer cell wall of V. inaequalis was not required for the survival of conidia under ambient temperature or at -20 °C storage conditions, however, promoted the tolerance of the pathogen to copper and synthetic fungicides affecting the stability and function of the fungal cell wall, plasma membrane, respiration (QoIs) and enzyme secretion, but had no effect on the sensitivity to sulphur and SDHIs. The roles of melanin in different steps of the V. inaequalis life cycle and the epidemiology of apple scabs are discussed.