Association between sleep pattern, salivary cariogenic bacteria and fungi populations, pH and buffering capacity in children: A comparative study.

BACKGROUND: Sleep quality has a significant impact on a child's health and is linked to oral and systemic diseases. It affects the circadian rhythm, which plays a crucial role in regulating the balance of the endocrine and hormonal systems. Current research has focused on exploring its role in the development of caries, which is influenced by inherent oral factors such as the composition of the oral microbiome and pH levels. OBJECTIVES: This study aimed to investigate the relationship between bacterial population, pH, and buffering properties of saliva and sleep patterns in 8- to 12-year-old children. MATERIAL AND METHODS: This cross-sectional study was conducted on 85 elementary school children aged 8-12 years. After obtaining written consent, non-stimulating saliva samples were collected using the spitting method. The participants' sleep pattern information was obtained with the use of the Persian version of the Children's Sleep Habits Questionnaire (CSHQ). Based on the results of the CSHQ, the participants were divided into 2 groups: those with appropriate sleep patterns; and those with inappropriate sleep patterns. The study compared the bacterial population of Streptococcus mutans, Lactobacillus spp. and Candida albicans, as well as the buffering capacity and pH of the saliva between the 2 groups. The statistical analysis employed the χ2 test, the independent samples t-test and Spearman's correlation. RESULTS: The group with inappropriate sleep patterns had significantly lower pH and buffering capacity (p < 0.001) and significantly higher colony counts of Lactobacillus and S. mutans (p < 0.001 and p = 0.012, respectively). There was no association between C. albicans and sleep patterns (p = 0.121). CONCLUSIONS: Inappropriate sleep patterns increase the population of caries-causing bacteria and reduce salivary pH and buffering capacity. This can be a significant factor in the development of dental caries in children aged 8-12 years.

In vitro evaluation of albendazole nanocrystals against Echinococcus granulosus protoscolices.

Echinococcus granulosus is a zoonotic parasite causing hydatidosis in humans and animals. This study has been done in order to investigate the effect of albendazole nanocrystals on the viability of E. granulosus protoscolices. The average size and hydrodynamic diameter of albendazole nanocrystals were 976±218 and 1334±502 nm, respectively. Fertile hydatid cysts were isolated from the liver of slaughtered sheep. The isolated cysts were further identified using morphological and molecular techniques. The nucleotide sequence analysis indicated that the genotype of the protoscolices was E. granulosus sensu stricto with 100% similarity. The parasites were examined precisely for susceptibility to albendazole nanocrystals. The results revealed that albendazole nanocrystals are effective in removing protoscolices. It was observed that 1 µg/ml albendazole nanocrystals and albendazole completely inhibited the viability of the protoscolices within 17 and 23 days, respectively. The results suggested that albendazole nanocrystals can be used as an alternative effective treatment for E. granulosus infection.

Construction of metronidazole capped in gold nanoparticles against Helicobacter pylori: antimicrobial activity improvement.

INTRODUCTION: Helicobacter pylori is considered a major agent causing gastritis and peptic ulcer disease. Unfortunately, the occurrence of increasing drug resistance to this bacterium would result in some difficulties in its treatment. Therefore, the application of nanotechnology has been suggested to resolve such problems. Nanoparticles usage in medical research has been expanded in recent years. Among nanometals, gold nanoparticles have exclusive features that can be used in such applications. Using nanotechnology in medical science could help mankind to solve this problem in the future. AIM: Our aim in this research was to investigate the antimicrobial effect of gold nanoparticles on H. pylori strains. MATERIALS AND METHODS: Gold nanoparticles were synthesized by the Turkevich method. Then, their size and dispersion were investigated using spectrophotometry, DLS, and TEM microscopy. Subsequently, the combination of metronidazole and gold nanoparticles was obtained by mixing method, and then the anti-helicobacter effects of the two were evaluated according to CLSI. RESULTS: The highest size of gold nanoparticles was between 12 and 9 nm, and the maximum absorbance was 522 nm; however, in conjugated state, the maximum absorbance was 540 nm, which indicated the accumulation of drug-conjugated nanoparticles in the conjugate state. Some changes indicated the binding of metronidazole to gold nanoparticles. Antimicrobial testing of gold nanoparticles and metronidazole did not affect the Helicobacter pylori. Therefore, the combination of gold nanoparticles and metronidazole had a 17-mm growth inhibition zone. CONCLUSIONS: The anti-helicobacter effects of metronidazole significantly increased in conjugation with gold nanoparticles.

Possible Synergistic Effect of Combined Metarhizium anisopliae and Fenitrothion for Control of German Cockroach (Blattella germanica) as a Novel Approach.

Background: Cockroaches are one of the most common pests in many residential areas. In this study, the simultaneous effects of fungi, Metarhizium anisopliae and fenitrothion-coated baits on the mortality rate of the German cockroach nymphs were investigated. Methods: To determine the lethal level of fenitrothion insecticide, a bioassay test was performed on the last instar nymphs of the German cockroach reared at insectarium conditions. Various toxic concentrations of fenitrothion (0.1%, 0.3%, 0.5%, 0.7%, 0.9%, 1.5%, and 2%) were used. Different concentrations of M. anisopliae (1×104, 1×105, 1×106, 1×107, 1×108 Conidia/ml) were also applied to nymphs. Eventually, we combined the effective dose of fenitrothion (0.93%) with the effective concentration of M. anisopliae (6.6 ×106 Conidia/ml) to provide the fungus-coated bait to attract insects. Mortality was recorded 24-96 hours after exposure to the toxic bait. The resulting data were subjected to Probit analysis. Results: The results of applying M. anisopliae spores with fenitrothion composition showed that the mortality rate of German cockroach nymphs was significant. Therefore, the optimal dose of fenitrothion used in combination with M. anisopliae seems essential to reduce the German cockroach nymphs. Conclusion: The results of this study can be considered a suitable method as a mixture with low cost and minimal damage to the environment and other organisms.

Isolation of arsenic accumulating bacteria from garbage leachates for possible application in bioremediation.

BACKGROUND AND OBJECTIVES: Bioremediation is a process to reduce toxic heavy-metals, such as arsenic, in the environment using microorganisms. This study aimed to isolate arsenic remediating microbial strains from garbage leachates and to evaluate the effects of several factors on bioremediation by isolated strains. MATERIALS AND METHODS: After isolating arsenic-resistant bacteria from garbage leachates and determining their MIC values, Taguchi design of experiments was used to evaluate the effect of arsenic concentration, pH solution, temperature, and contact time on arsenic bioremediation by isolated bacteria. RESULTS: The results revealed that 3 arsenic-resistant strains of genus Bacillus characterized as KL1, KL4, and KL6 had arsenic bioremediation activity. Based on the results, the highest bioremediation of arsenic by Bacillus sp. KL1 was obtained as 77% after 24 hours at 40°C, pH 5, and 150 ppm concentration. However, the maximum bioremediation of arsenic by KL4 (91.66%) and KL6 (88%) was achieved after 24 hours at 40°C, pH 5, and 60 ppm concentration and at 35°C, 90 ppm concentration, pH 5 after 36 hours, respectively. CONCLUSION: The results presented here may facilitate improvements in the eliminating arsenic from contaminated sites and reducing environmental pollutions.

In vivo identification of novel TGIF2LX target genes in colorectal adenocarcinoma using the cDNA-AFLP method.

BACKGROUND AND STUDY AIMS: Homeobox-containing genes are composed of a group of regulatory genes encoding transcription factors involved in the control of developmental processes. The homeodomain proteins could activate or repress the expression of downstream target genes. This study was conducted to in vivo identify the potential target gene(s) of TGIF2LX in colorectal adenocarcinoma. METHODS: A human colorectal adenocarcinoma cell line, SW48, was transfected with the recombinant pEGFPN1-TGIF2LX. The cells were injected subcutaneously into the flank of the three groups of 6-week-old female athymic C56BL/6 nude mice (n = 6 per group). The transcript profiles in the developed tumours were investigated using the cDNA amplified fragment length polymorphism (cDNA-AFLP) technique. RESULTS: The real-time RT-PCR and DNA sequencing data for the identified genes indicated that the N-terminal domain-interacting receptor 1 (Nir1) gene was suppressed whereas Nir2 and fragile histidine triad (FHIT) genes were upregulated followed by the overexpression of TGIF2LX gene. CONCLUSION: Downregulation of Nir1 and upregulation of Nir2 and FHIT genes due to the overexpression of TGIF2LX suggests that the gene plays an important role as a suppressor in colorectal adenocarcinoma.

The impact of genetic variation in IL28B, IFNL4 and HLA genes on treatment responses against chronic hepatitis C virus infection.

Single nucleotide polymorphisms (SNPs) near the interleukin-28B (IL28B), interferon lambda 4 (IFNL4) and the human leukocyte antigen (HLA) gene are associated with treatment responses in patients with chronic hepatitis C (CHC) virus infection treated with pegylated interferon-α and ribavirin (pegIFN-α/RBV). We compared the role of IL28B SNPs (rs12979860, rs12980275, and rs8099917), IFNL4 ss469415590 and HLA rs4273729 with treatment outcomes in patients with CHC virus. A total of 520 Iranian patients with CHC infection were enrolled. SNPs in IL28B, IFNL4 ss469415590 and HLA rs4273729 were genotyped by PCR-restriction fragment length polymorphism, TaqMan® Real-Time PCR and direct sequence. Out of 520 CHC treatment-naive patients, 42.9% were infected with HCV-1a, 15.4% with HCV-1b, 9.8% with HCV-2, and 31.9% with HCV-3a. Rapid virologic response (RVR), complete early virologic response (cEVR), and sustained virologic response (SVR) were 53.3%, 73.8%, and 66.7%, respectively. Multivariate logistic regression analysis showed that IL28B rs12980275 and IFNL4 ss469415590 in all HCV genotypes were associated with RVR. In addition, IL28B rs12979860 and RVR in all HCV genotypes and IL28B rs12980275, IFNL4 ss469415590, and HLA rs4273729 in HCV subtypes 1a, 1b, and 3a correlated with cEVR. In patient's achieving-SVR, IL28B rs12980275, and RVR in all HCV genotypes and IL28B rs12979860, IFNL4 ss469415590, and HLA rs4273729 in HCV subtypes 1a, 1b, and 3a were the powerful predictor factors. As the first report of its kind published in Iran, we indicated that beside IL28B SNPs and HLA rs4273729, IFNL4 ss469415590 was a powerful predictor factor for RVR, cEVR and SVR. Genotyping these SNPs may be a helpful priority in the treatment of patients with HCV infection, especially in countries where access to triple or double therapy with a viral protease inhibitor is limited.

A comparative study of various methods for detection of IL28B rs12979860 in chronic hepatitis C.

Interleukin-28B (IL28B) single-nucleotide polymorphisms (SNPs) constitute important host-related factors influencing the response rate to Hepatitis C virus (HCV) standard antiviral therapy. In the last few years, several new technologies for SNP detection have been developed. However, the sensitivity and specificity of various methods are different and needs evaluation. Five different methods (resolution melting curve [RMC], polymerase chain reaction-restriction fragment length polymorphism [PCR-RFLP], PCR-sequencing analysis, amplification refractory mutation system [ARMS], and zip nucleic acid probe-based real-time PCR [ZNA]) were developed for genotyping rs12979860 associated with IL28B. In this study, limit of detection (LD), costs and turnaround time of these methods were compared in 350 subjects. As for IL28B rs12979860 polymorphisms, 348/350 (99.4%) samples were consistent among the five methods, while results for 2/350 (0.57%) samples were concordant by ZNAs and PCR-sequencing, and discordant by other methods. Without considering the cost of DNA extraction, the price of each reaction for ARMS-PCR, RMC, PCR-RFLP, ZNA and PCR-sequencing were respectively: US$3.10, US$5.0, US$5.50, US$8.50 and US$17.0. RMC was the fastest method, while the ZNA method was easy to use, reliable and effective. Lower LD was determined to be 50-60 copies/µL for the PCR-RFLP, RMC and ARMS-PCR assays; whilst ZNA assay was able to detect 2-3 copies/µL. In conclusion, in the current study, all four methods are suitable for IL28B rs12979860 genotyping, but the ZNA assay can be a reliable tool. Due to its lower LD for SNP identification, this method is better than others for detecting this type of polymorphism.

Molecular identification and antifungal susceptibility profile of Candida species isolated from patients with vulvovaginitis in Tehran, Iran.

BACKGROUND: Rapid and accurate identification and evaluation of antifungal susceptibility pattern of Candida isolates are crucial to determine suitable antifungal drugs for the treatment of patients with vulvovaginitis candidiasis. MATERIALS AND METHODS: Vaginal samples were collected from 150 women with suspicious vaginal candidiasis, and then cultured on Sabouraoud's Dextrose Agar with chloramphenicol to isolate Candida species. After identification of Candida isolates using polymerase chain reaction-restriction fragment length polymorphism technique, antifungal susceptibility testing of four azolic antifungal drugs was carried out using broth microdilution method according to the CLSI M27-A3. RESULTS: Candida species were isolated from eighty suspected patients (61.79%). The most common pathogen was Candida albicans (63.75%). Resistance to fluconazole and ketoconazole was observed in 27.5% and 23.75% of Candida isolates, respectively, and only 2% of Candida isolates were resistant to miconazole. Interestingly, resistance to fluconazole in C. albicans was more than other Candida species. CONCLUSION: The results indicated that therapy should be selected according to the antifungal susceptibility tests for the prevention of treatment failure and miconazole therapy can be considered as the best therapeutic choice in the management of vulvovaginitis.

Comparison of Three Different Methods for Detection of IL28 rs12979860 Polymorphisms as a Predictor of Treatment Outcome in Patients with Hepatitis C Virus.

OBJECTIVES: This study aimed to evaluate the specificity, sensitivity, cost, and turn-around time of three methods of gene polymorphism analysis and to study the relationship between IL28B rs12979860 and SVR rate to pegIFN-α/RVB therapy among patients with chronic hepatitis C. METHODS: A total of 100 samples from chronic hepatitis C patients were analyzed in parallel using the three methods: direct sequencing, real-time polymerase chain reaction (PCR), amplification refractory mutation system (ARMS)-PCR. RESULTS: The different profiles for IL28B rs12979860 alleles (CC, CT, and TT) obtained with PCR-RFLP, ARMS-PCR, and direct sequencing were consistent among the three methods. Prevalence of rs12979860 genotypes CC, CT and TT in HCV genotype 1a was 10(19.6%), 35(68.6%), and six (11.8%), respectively, and in HCV genotype 31, it was 13(26.5%), 31(63.3%), and five (10.2%), respectively. No significant difference was seen between rs12979860 genotype and HCV genotype (p = 0.710). CONCLUSION: Screening by ARMS - PCR SNOP detection represents the most efficient and reliable method to determine HCV polymorphisms in routine clinical practice.

Identification of Azole Resistance Markers in Clinical Isolates of Candida tropicalis Using cDNA-AFLP Method.

BACKGROUND: Global reports have highlighted the increasing prevalence of Candida tropicalis infections as well as organism(') s drug resistance. This study aimed at identifying azole resistance markers in clinical isolates of C. tropicalis, which will be a great resource for developing new drugs. METHODS: Two susceptible and resistant isolates of C. tropicalis were recovered from an epidemiological investigation of candidiasis in immunocompromised patients. C. tropicalis ATCC 750 was used as reference strain. Antifungal susceptibility to fluconazole and itraconazole was determined using Clinical and Laboratory Standards Institute (CLSI) method. Complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) technology and real-time reverse-transcriptase (RT) PCR were used for identification of potential genes involved in azole resistance of C. tropicalis clinical isolates. RESULTS: Five genes encoding the following enzymes were identified as superoxide dismutase (SOD) implicated in antioxidant defense, ornithine aminotransferase (OAT), acetyl ornithine aminotransferase (ACOAT), adenosylmethionine-8-amino-7-oxononanoate aminotransferase (DAPA AT), and 4-aminobutyrate aminotransferase (ABAT)-belonging to pyridoxal phosphate (PLP) dependent enzymes and acting in an important physiological role in many fungal-cell cycles. Real-time RT-PCR confirmed mRNA level of the aforementioned genes. CONCLUSION: Our findings showed that factors such as PLP-dependent enzymes and SOD might be implicated in drug resistance in C. tropicalis clinical isolate. Therefore, further studies are required to explore the accurate biological functions of the mentioned genes that would be helpful for effective drug development.

Fungal Airborne Contamination as a Serious Threat for Respiratory Infection in the Hematology Ward.

BACKGROUND: Fungi existing in hospital departments may grow and produce micro-colonies. The spores arising from these micro-colonies circulate easily and could be inhaled by patients and cause infections in immune-compromised subjects. Due to the lack of an acceptable method of sampling and evaluation of microbiological quality of air in the isolation units, the purpose of this study was to determine the concentrations of airborne fungi through active and passive sampling and also identify fungi genera in the air of the isolation unit. MATERIALS AND METHODS: The air of the isolation unit was monitored through active and passive sampling. In passive sampling, the plates were placed in the room. The active sampling was performed in the hematology unit by using a slit-to-agar biological air sampler with a flow rate of 10 L/minute. Plates were incubated at 30°C for 10 days and were examined daily for fungal growth. Fungal species were identified on the basis of their macroscopic and microscopic morphological features. RESULTS: In active samples, Penicillium spp. was the predominant genus (66.8%), followed by Aspergillus spp. (23.9%) and Cladosporium spp. (2.5%). Yeast spp. accounted for only 2.2% of the isolated fungi. In passive samples, Penicillium spp. (94.4%) was the most frequently found fungi, followed by Aspergillus spp (2.2%), Cladosporium spp. (1.1%) and Yeast spp. (0.5%). The identified genera included Penicillium, Aspergillus, Alternaria, Mucorales, Cladosporium, Yeasts and other filamentous fungi. CONCLUSION: Active and passive sampling can be used for monitoring the fungal content of air. Assessment of fungal contamination profiles in hospitals may provide important information about the level of fungal concentration in the hospitals and for the control of nosocomial infections. In addition, installation of special ventilation systems equipped with HEPA filters in hematology wards could enhance the quality of air. Also, observing sanitary protocols for disinfection of the surfaces is imperative for infection control.

Evaluation of mRNA Expression Levels of cyp51A and mdr1, Candidate Genes for Voriconazole Resistance in Aspergillus flavus.

BACKGROUND: Voriconazole Resistance (VRC-R) in Aspergillus flavus isolates impacts the management of aspergillosis, since azoles are the first choice for prophylaxis and therapy. However, to the best of our knowledge, the mechanisms underlying voriconazole resistance are poorly understood. OBJECTIVES: The present study was designed to evaluate mRNA expression levels of cyp51A and mdr1 genes in voriconazole resistant A. flavus by a Real-Time Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) technique. MATERIALS AND METHODS: Five A. flavus isolates with resistance to VRC were examined by a RT-PCR approach. RESULTS: Four out of five isolates revealed cyp51A and mdr1 mRNA overexpression. Interestingly, the isolate, which was negative for cyp51A and mdr1 mRNA expression showed a high voriconazole Minimum Inhibitory Concentration (MIC). Furthermore, a computational-based analysis predicted that voriconazole resistance could be mediated through cooperation with a network protein interaction. CONCLUSIONS: Our experimental and in silico findings may provide new insight in the complex molecular pathways of drug resistance and also could assist design an efficient therapeutic strategy for aspergillosis treatment.

Identification and Sequencing of Candida krusei Aconitate Hydratase Gene Using Rapid Amplification of cDNA Ends Method and Phylogenetic Analysis.

BACKGROUND: The production and development of an effective fungicidal drug requires the identification of an essential fungal protein as a drug target. Aconitase (ACO) is a mitochondrial protein that plays a vital role in tricarboxylic acid (TCA) cycle and thus production of energy within the cell. OBJECTIVES: The current study aimed to sequence Candida krusei ACO gene and determine any amino acid residue differences between human and fungal aconitases to obtain selective inhibition. MATERIALS AND METHODS: Candida krusei (ATCC: 6258) aconitase gene was determined by 5'Rapid Amplification of cDNA Ends (RACE) method and degenerate Polymerase Chain Reaction (PCR) and analyzed using bioinformatics softwares. RESULTS: One thousand-four hundred-nineteen nucleotide of C. krusei aconitase gene were clarified and submitted in Genbank as a partial sequence and then taxonomic location of C. krusei was determined by nucleotide and amino acid sequences of this gene. The comparison of nucleotide and amino acid sequences of Candida species ACO genes showed that C. krusei possessed characteristic sequences. No significant differences were observed between C. krusei and human aconitases within the active site amino acid residues. CONCLUSIONS: Results of the current study indicated that aconitase was not a suitable target to design new anti-fungal drugs that selectively block this enzyme.

Synthesis and antifungal activity of 1-[(2-benzyloxy)phenyl]-2-(azol-1-yl)ethanone derivatives: exploring the scaffold flexibility.

Based on the N-(phenethyl)azole backbone of azole antifungals, we designed 1-[(2-benzyloxy)phenyl]-2-(azol-1-yl)ethanone derivatives 2 and 3, containing benzyloxyphenyl scaffold of croconazole. Also these compounds can be considered as flexible analogs, resulted from C2-C3 disconnection of 3'-chloro-3-imidazolylflavanone 1, recently described as antifungal agent. Thus, in this report, we describe the synthesis of 1-[(2-benzyloxy)phenyl]-2-(azol-1-yl)ethanone derivatives 2 and 3 and their biological evaluation against different pathogenic fungi. By comparing the antifungal activity profile of flexible compounds 2 and 3 with that of rigid analog 1, it can be inferred that lower susceptibilities (higher minimum inhibitory concentrations) were observed with flexible compounds. However, among the synthesized compounds, 1-[2-(2,4-dichlorobenzyloxy)phenyl]-2-(1H-imidazol-1-yl)ethanone hydrochloride (2g) showed comparable or more potent antifungal activity in comparison with fluconazole as a standard drug.