Simplex and triplex polymerase chain reaction (PCR) for identification of three medically important Candida species

Harmal, Nabil S.; Khodavandi, Alireza; Alshawsh, Mohammed Abdullah; Jamal, Farida; Peng, Ng Kee; Chong, Pei Pei

doi:10.5897/ajb12.1708

Cited by 5 publications

(5 citation statements)

References 11 publications

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“…17 Thus, it is possible through designing a specific set of primers to target the unique sequence of PLB gene. Being used the same set of primers as in the current study Harmal et al 9 proved that species-specific PCR assay could identify and differentiate between the four most common Candida species isolated from clinical specimens namely, C. albicans, C. glabrata, C. parapsilosis and C. tropicalis. Distinctive product size for each of these 4 species allow specific identification directly from the gel e ectrophoresis without the need for further genotyping.…”

Section: Molecular Methods For the Detection Of Candida Speciesmentioning

confidence: 79%

“…8 (Freezing-thawing technique was added to the protocol as an efficient step and enhance the cell lysis)*. The primer sequences were used for the amplification for PLB genes of Candida species were selected according to Nabil S. Harmal et al, 9 (Table 1). An internal control has been used to measure the efficiency of the DNA extraction process fluids, as well as the impact of external and internal factors on gene amplification process.…”

Section: Molecular Methods For Diagnosis Of Candida Speciesmentioning

confidence: 99%

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Molecular and conventional methods for detection of Candida species isolated from a sample of immunocompromised Iraqi patients with pulmonary symptoms

2017

J. contemp. med. sci.

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Objectives Candida species has emerged as a potentially pathogenic fungus rather than benefit mucosal commensal in patients with pulmonary diseases. Therefore, our study was carried out to detect Candida species in sputum samples from patients with pulmonary diseases using conventional and molecular methods. Methods A total of 100 sputum samples obtained from patients with pulmonary symptoms such as chronic productive cough, shortness of breath, wheezing and fever were included in this study. Sputum samples were dispensed into three specimen parts; the first one was applicated for cultured on Sabouraud dextrose agar at 37°C for 48 h and then the purified colony of Candida underwent biochemical tests including API, Candida strips, and germ tube. The second part was undergone direct gram stain, while the third part was applicated for DNA extraction and then molecular diagnosis with PCR technique using specific primers. Results Culture result revealed 43 positive samples for Candida species out of 100 samples. Among these positive samples, 23 (53.5%) were positive for C. albicans in each of culture and germ tube. API 20 Candida found that (40) samples were positive for Candida species as, 23 (57.5%) represent Candida albicans, 8 (20.0%) Candida glabrata, 4 (10.0%) Candida parapslosis, 4(10.0%) Candida tropicalis and only one (2.5%) as Candida krusei. Molecular test revealed that forty one samples out of forty three culture isolates of Candida species were positive as follow twenty three (53.48%), belong Candida albicans, Nine (20.93%) belong Candida glabrata, Six (13.95%) Candida parapslosis, Four (9.30%) belong Candida tropicalis. Conclusion Candida albicans is highly prevalent among patients suffering from bronchopulmonary symptoms. The molecular and conventional methods gave concomitant results as detection tools for the diagnosis of such microorganisms.

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Section: Molecular Methods For the Detection Of Candida Speciesmentioning

confidence: 79%

Section: Molecular Methods For Diagnosis Of Candida Speciesmentioning

confidence: 99%

Molecular and conventional methods for detection of Candida species isolated from a sample of immunocompromised Iraqi patients with pulmonary symptoms

2017

J. contemp. med. sci.

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“…Isolates were cultured in Sabouraud dextrose broth (SDB, Difco Laboratories, Detroit, Michigan). The identities of the clinical isolates of Candida were confirmed by phenotypic (16) and molecular techniques (17). Briefly, clinical isolates were plated on CHROMagar Candida medium (CHROMagar Company, France) to check for purity and the reliability of Candida species were confirmed by DNA sequencing.…”

Section: Methodsmentioning

confidence: 99%

“…The universal fungal primers ITS1 (5'-TCC GTA GGT GAA CCT GCG G-3') and ITS4 (5'-TCC TCC GCT TAT TGA TAT GC-3') were used for DNA amplification (17). The amplified PCR products were confirmed by DNA sequencing.…”

Section: Dna-sequencing Techniques For Clinical Isolates Identificationmentioning

confidence: 99%

Analysis of ergosterol and gene expression profiles of sterol Δ5,6-desaturase (ERG3) and lanosterol 14α-demethylase (ERG11) in Candida albicans treated with carvacrol

Alizadeh¹,

Khodavandi²,

Esfandyari³

et al. 2018

J Herbmed Pharmacol

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“…All the isolates were identified by conventional 19 and molecular methods. 20 The reliability of C. tropicalis colonies were confirmed by CHROMagar Candida medium (CHROMagar Company, France) and DNA sequencing. All isolate of C. tropicalis were kept in sterile 20% (v/v) glycerol stocks at −80 °C.…”

Section: Candida Tropicalis Isolates and Growth Conditionsmentioning

confidence: 99%

Synergistic Interaction of Fluconazole/Amphotericin B on Inhibition of Enzymes Contributes to the Pathogenesis of Candida Tropicalis

2018

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A B S T R A C TBackground: Candidiasis has gained much attention in recent decades due to its increasing prevalence in immunocompromised patients. Usually, antifungals such as fluconazole and amphotricin B are used for treatment of candidiasis, but one of the major clinical problems is the emergence of antifungal resistance. Combination antifungal therapy is one of the most commonly used methods to alleviate the problem of antifungal resistance. Methods: The effect of fluconazole alone and in combination with amphotericin B on C. tropicalis isolates were performed using the Clinical and Laboratory Standards Institute (CLSI) reference method. Eventually hypha formation, time kill study, proteinase and phospholipase activity and expression of PLB and SAP2 genes were carried out to investigate the enzymes inhibitory properties of antifungal tested against C. tropicalis. Results: Results showed the significant synergic effect of fluconazole in combination with amphotericin B in inhibiting the growth of C. tropicalis isolates, with fractional inhibitory concentration indices ranging from 0.06 to 0.5. The combination of fluconazole with amphotericin B reduced the number of yeast form and inhibited the yeast to hyphae transition in C. tropicalis. The antifungals tested were able to show the effect of down regulating expression of the selected genes significantly in fluconazole/amphotericin B ranging from 1.42-to 2.27-fold. Conclusion: Our results demonstrated that the synergistic interaction of fluconazole/amphotericin B would be worth exploring for the management of candidiasis. In addition, PLB and SAP2 genes could be probable molecular targets in the synergistic interaction of fluconazole/amphotericin B in C. tropicalis. Pharmaceutical Sciences, December 2018, 24, 280-290 | 281 Khodavandi A, et al. Pharmaceutical Sciences, December 2018, 24, 280-290 | 285 Khodavandi A, et al.Table 4. Results of phospholipase activity assay (colony diameter/ precipitation zone diameter) of Candida tropicalis isolates treated with fluconazole alone and in combination with amphotericin B in different concentration based on MIC.a-e Means ± S.D in each treatment and column with different superscript differ significantly (Tukey's HSD, P < 0.05). The results were performed in three independent experiments.

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Simplex and triplex polymerase chain reaction (PCR) for identification of three medically important Candida species

Cited by 5 publications

References 11 publications

Molecular and conventional methods for detection of Candida species isolated from a sample of immunocompromised Iraqi patients with pulmonary symptoms

Molecular and conventional methods for detection of Candida species isolated from a sample of immunocompromised Iraqi patients with pulmonary symptoms

Analysis of ergosterol and gene expression profiles of sterol Δ5,6-desaturase (ERG3) and lanosterol 14α-demethylase (ERG11) in Candida albicans treated with carvacrol

Synergistic Interaction of Fluconazole/Amphotericin B on Inhibition of Enzymes Contributes to the Pathogenesis of Candida Tropicalis

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