Background Cystic echinococcosis, caused by the tapeworm Echinococcus granulosus sensu stricto (ss), is a globally distributed, zoonotic disease that is recognised by WHO as a neglected tropical disease. Despite its clinical and economic importance, nuclear genomic variation in this parasite has not been systematically characterised across global populations. In this study, we aimed to characterise the genome-wide nuclear genetic diversity and population structure of E granulosus ss across globally distributed populations. Methods We conducted a genomic study of 137 E granulosus ss samples from endemic regions across five continents, derived from previously collected parasite material from livestock, wildlife, and human infections. Using a chromosome-scale reference genome, we applied population genomic approaches to investigate genome-wide nuclear genetic diversity, population structure, and patterns of evolutionary constraint. Findings We identified 1 071 085 nuclear single-nucleotide polymorphisms across 137 samples, with heterozygosity ranging from 46% to 93% per sample. Genome-wide analyses identified two major clades associated with geographical origin. Distinct regions of genetic differentiation were observed, particularly on chromosome 9. Conserved genes under purifying selection included those involved in glycan biosynthesis and core cellular functions, whereas variable genes were enriched in pathways such as ribosome biogenesis. Mitochondrial genotypes (G1 and G3) did not align with the nuclear genomic structure. Interpretation To the best of our knowledge, this study provides the first broad atlas of nuclear genomic diversity in E granulosus ss, uncovering genetic diversity and population structure. The findings have important implications for molecular epidemiology, genomic surveillance, and translational development of diagnostics and vaccines. Incorporating genomic data into cystic echinococcosis control programmes could enhance WHO-aligned efforts to reduce the burden of this neglected tropical disease. Funding Australian Research Council and the Estonian Ministry of Education and Research.

Trichophyton indotineae has recently emerged as a major etiological agent of treatment-resistant dermatophytosis, particularly in India and Iran. Resistance to terbinafine is predominantly associated with point mutations in the squalene epoxidase (SQLE) gene. This study investigated the antifungal susceptibility of clinical dermatophyte isolates from Iran and characterized SQLE mutations in terbinafine-resistant strains. Over a 6-month period, 176 clinical dermatophyte isolates comprising 134 T. indotineae and 42 non-indotineae species were collected and identified by internal transcribed spacer (ITS) sequencing. In vitro antifungal susceptibility testing to eight antifungal agents was assessed according to CLSI M38 guidelines. Minimum inhibitory concentration (MIC) ranges, MIC₅₀, MIC₉₀, and geometric mean MICs were calculated. SQLE gene sequencing was performed for isolates exhibiting reduced terbinafine susceptibility. Out of 176 dermatophyte isolates, 66 (37.5%) demonstrated terbinafine MICs ≥ 0.5 µg/ml. Among these 66 isolates, 64 were T. indotineae (out of 134 total T. indotineae isolates, 47.76%), and 2 were T. tonsurans (out of 21 total T. tonsurans isolates, 9.52%). High-level terbinafine resistance (MIC ≥ 16 µg/ml) was observed in 50% (32/64) of T. indotineae isolates. Among the 66 resistant isolates, only 3 were wild-type: one T. indotineae and two T. tonsurans. The most prevalent SQLE alterations were the double mutation Phe397Leu + Ala448Thr (34.38%) and the single mutation Leu393Ser (32.81%), followed by the 1192 C→A substitution (10.94%). Notably, Ala448Thr also occurred in phenotypically susceptible isolates, yet correlated with increased MICs of itraconazole and voriconazole. Fluconazole showed poor activity (geometric mean MIC: 43.5072 µg/ml), whereas posaconazole exhibited the highest in vitro activity. Terbinafine remained effective for non-T. indotineae species, but is ineffective for T. indotineae-associated dermatophytosis due to widespread resistance. None of the isolates identified as T. interdigitale/mentagrophytes, T. rubrum, Microsporum canis, and Trichophyton benhamiae were resistant to TRB. This multicenter survey reveals high terbinafine resistance in Iranian T. indotineae isolates, linked to SQLE mutations and azole cross-resistance. Routine antifungal susceptibility testing, molecular screening, and alternative therapies are essential, prioritizing non-terbinafine options for T. indotineae (while it remains effective against other dermatophytes). Continued surveillance, accurate species identification, and tailored management are critical to control recalcitrant dermatophytosis.

BackgroundFungal infections are increasingly recognized as a global health concern, contributing to considerable morbidity and mortality in hospital settings. This underscores the urgent need for infection prevention and control in healthcare facilities to protect vulnerable patients from the risk of acquiring invasive fungal diseases (IFDs). Given the critical role of transmission-based precautions in limiting the spread of filamentous fungi responsible for IFDs, this study was conducted to explore the potential role of the hospital environment in the dissemination of these infections.MethodsA total of 83 samples were collected from the air and surface of exhaust vents in the intensive care units (ICUs) of hospitals in Isfahan, Iran, to assess the presence and diversity of fungal species. Susceptibility testing against antifungal agents, including commonly used drugs and disinfectants, was performed on the identified fungal isolates. Furthermore, the antifungal resistance profiles of isolates from clinical IFD cases were compared with those of environmental isolates.ResultsFungi were detected in 45% of air samples and 100% of exhaust vent samples, with Aspergillus species being the most commonly identified genus. Mucorales were also found in 17% of exhaust vent samples. Aspergillus spp. and Rhizopus spp. showed the highest resistance to Amphotericin B, and a considerable proportion of these isolates exhibited simultaneous resistance to disinfectants. A similar antifungal resistance profile was noted between A. flavus and some R. arrhizus isolates from both environmental and clinical samples.ConclusionsThe findings of this study indicate that the hospital environment, particularly exhaust vents, may act as a significant reservoir for causative agents of IFDs. This highlights the importance of environmental surveillance in preventing and controlling nosocomial fungal infections.

Background Posaconazole is an effective antifungal agent used for many fungal diseases, and its application has grown recently. Posaconazole was studied as a treatment option after the increase in mucormycosis cases linked to COVID-19, and promising results were obtained. However, despite its effectiveness, researchers are looking into new approaches to drug delivery due to problems including restricted bioavailability and short half-life. To address these issues, this project intends to create and synthesize posaconazole-containing liposomes. Methods In this study, according to the synthesis of Posaconazole-loaded liposomes, their physical and chemical properties were assessed using SEM, DLS, and FTIR techniques. The drug loading capacity in the nanoparticles, its release in vitro, and the antifungal activity against mucormycosis-causing fungi were evaluated. Finally, the cytotoxicity of these nanoparticles was assessed using the MTT Assay. Results The synthesized nanoparticles were spherical with an average size of approximately 60 nm. The drug loading efficiency was found to be 78.41%. The results indicated that the posaconazole nanoparticles demonstrated a slow and continuous drug release over five days. These nanoparticles demonstrated higher efficacy than posaconazole individually in antifungal tests. Additionally, cytotoxicity tests revealed that, up to a 400 µg/ml concentration, posaconazole-loaded nanoparticles were less toxic than free posaconazole. Conclusions These nanoparticles may provide a useful therapeutic option for the treatment of mucormycosis while circumventing the disadvantages of posaconazole due to their improved antifungal activities and decreased toxicity compared to free posaconazole. ### Competing Interest Statement The authors have declared no competing interest. Iran University of Medical Sciences, 24953

Marine recreational waters harbor various microorganisms that may pose health risks to users. However, fungal contamination, particularly by Candida species, has received limited attention despite evidence of their presence and potential health concerns. This study aimed to assess the microbial quality of water at five popular recreational beaches along the Persian Gulf, with a specific focus on Candida species. Fecal indicator bacteria (FIB), including Escherichia coli and Enterococci, were enumerated in marine water samples. Additionally, the presence of Candida auris and other clinically relevant Candida spp. as well as Meyerozyma guilliermondii was investigated using nested, multiplex, and conventional PCR assays, respectively. FIB were highly prevalent across most of the sampling locations. Additionally, C. albicans, C. tropicalis, C. glabrata, C. krusei, and M. guilliermondii were detected in a few samples. C. albicans specifically found at locations with high FIB concentrations, suggesting an association with anthropogenic activities. However, C. auris was not detected in this study. The presence of Candida spp. in recreational waters suggests a potential public health risk. These findings highlight the importance of further monitoring for fungal contamination to protect public health and guide future water quality management policies.

Accurate Leishmania species identification is crucial for epidemiological aspects and disease management. This study aimed to detect and identify the main causative species of human cutaneous and visceral leishmaniasis in Iran using nested polymerase chain reaction (PCR) based on size polymorphism of the ribosomal DNA (rDNA) intergenic region. In total, 180 clinical samples, including 156 confirmed positive cases, 24 confirmed negative cases, and 33 suspected cases that tested negative by microscopy, were collected from skin lesions, bone marrow, and lymph nodes across five provinces in Iran. DNA was extracted from the samples, and the nested PCR-amplified fragments were differentiated with agarose gel electrophoresis. The nested PCR assay successfully identified Leishmania major, Leishmania tropica, and Leishmania infantum with high accuracy. Among 156 microscopy-positive samples, L. major was detected in 84 cases, L. tropica was detected in 64 cases, and L. infantum was detected in 8 cases. The results obtained were fully consistent with those from a commercial real-time PCR diagnostic kit. Phylogenetic analysis supported the differentiation of Leishmania species. However, among 33 suspected cases, 14 samples were negative by all three methods (microscopy, commercial PCR kit, and nested PCR), 7 samples tested positive by both molecular methods, 8 samples were positive only with the commercial PCR kit, and 4 samples were positive exclusively by nested PCR. The findings indicate that the nested PCR assay targeting size polymorphisms in the rDNA region is a cost-effective method for detection and identification of Leishmania species in clinical specimens, particularly when microscopy results are inconclusive.

ABSTRACTBackgroundSince 2017, dermatophytosis caused by the newly introduced species Trichophyton indotineae has gained new interest worldwide due to the rise in terbinafine resistance and difficulty in the treatment of recalcitrant infections. Distinguishing T. indotineae from other Trichophyton species based on morphological features is impossible and DNA sequencing is necessary for accurate identification. Though early identification of the species is not solely sufficient for the treatment of infected cases, it is important for clinicians to take the next appropriate modalities such as antifungal susceptibility testing especially when the patients have extensive skin lesions recalcitrant to therapy by terbinafine. Here, we developed a rapid diagnostic scheme using SYBR Green real‐time PCR for the specific detection/identification of T. indotineae.MethodsDNA was extracted from 397 dermatophyte isolates and two SYBR Green real‐time PCR assays targeting the C120‐287 and E054‐58 intergenic loci were developed. Using a collection of 132 T. indotineae and 128 non‐T. indotineae strains, all had already been identified by ITS‐PCR‐sequencing and 137 unknown dermatophyte isolates, the assays were evaluated.ResultsIn both real‐time PCR assays, 130 out of 132 T. indotineae strains were positive while all non‐T. indotineae species were negative. Among 137 unknown tested isolates, 72 were identified as T. indotineae based on two real‐time PCR assays, while 65 showed no peak and were considered non‐T. indotineae. Based on PCR‐sequencing as the reference standard, the SYBR Green real‐time PCR assays demonstrated a sensitivity of 98.48% and a specificity of 100%.ConclusionThe developed diagnostic assays using SYBR Green real‐time PCR provided a rapid and accurate method for the distinction of cultured T. indotineae isolates and can be considered to evaluate for the detection of T. indotineae directly from clinical samples.

Mucormycosis, a fungal emergency, poses a serious threat to both COVID-19 and non-COVID-19 individuals due to its invasive nature, rapid progression, and high rates of morbidity and mortality. This underscores the crucial need for timely detection and management. In this study, we investigated the utility of real-time PCR (RT-qPCR) assays for detecting Mucorales in clinical specimens, and assessed the performance of both SYBR Green and TaqMan probe RT-qPCR in amplifying Mucorales-specific 18S rDNA genes. We conducted accuracy analyses using direct examination with KOH as a standard for the laboratory diagnosis of mucormycosis. Additionally, we compared the results with culture and duplex PCR. Both SYBR Green and TaqMan RT-qPCR were optimized using Mucorales-specific oligonucleotides to amplify the conserved 18S rDNA targets. DNAs extracted from 120 rhino sinus specimens, which all were collected from COVID-19 patients upon suspicion of invasive fungal infections, were used for molecular diagnosis. The results of both RT-qPCR assays were compared with the result of direct microscopy, culture, and duplex Mucorales-specific PCR assay. SYBR Green real-time PCR produced a distinct melting temperature (Tm) pattern (80.24 ± 0.70 °C) and detected Mucorales in 51 out of 120 clinical samples. When compared to direct examination with KOH, the standard method for diagnosing mucormycosis, SYBR Green PCR demonstrated a sensitivity of 91.67

BACKGROUND:Lipofilling is a natural, low-risk, and long-lasting method for filling, reconstructing, and improving soft tissues such as the face, with minimal discomfort for patients. Many plastic surgeons prefer autologous fat grafting in aesthetic surgery due to its availability, cost-effectiveness, biocompatibility, and absence of allergic and carcinogenic concerns. Despite the advantages of autologous fat injection, one of the main drawbacks is the variable persistence of injected fat tissue. Given the significant implications of this issue in advanced countries, this study aims to investigate the survival of fat cells after freezing at different time intervals (1, 3, and 6 months). METHODS:Thirty female participants were enlisted for this research, and the viability of fat cell specimens was assessed at intervals of 0, 1, 3, and 6 months post-freezing at -18°C. The evaluation of viable adipocytes was conducted using the XTT assay, a live/dead staining method using fluorescence microscopy after staining with fluorescein diacetate (FDA) and propidium iodide (PI), along with histological analysis of fat tissue after freezing at the indicated time intervals. RESULTS:The results showed that the viability of frozen fat samples decreases by 34%, 60%, and 80% after 1, 3, and 6 months, respectively, compared to non-frozen samples on Day 0. CONCLUSIONS:The findings of this study underscore a rapid decline in adipocyte viability after storage at -18°C at different time intervals (1, 3, and 6 months), at which points only around 60%, 40%, and 20% of fat cells remained viable, respectively. These results suggest that current fat preservation techniques utilizing either a -18°C freezer are not sufficient for maintaining the long-term viability of adipocytes, and alternative cryopreservation methods are needed to preserve fat cells.

BACKGROUND:Trichophyton indotineae, formerly described as T. mentagrophytes rDNA-ITS genotype VIII, has recently been identified as a novel species within the T. mentagrophytes complex. It has rapidly replaced T. rubrum as the predominant dermatophyte. In this study, skin dermatophyte isolates collected from patients in Iran were sequence-analysed for species identification. Additionally, the current prevalence of T. indotineae was compared with data from the previous decade. METHODS:A total of 194 dermatophyte isolates were collected from patients in four cities across Iran between July and December 2023, with 73 isolates of the T. mentagrophytes complex from the past decade also included. DNA was extracted from fresh colonies, and the internal transcribed spacer (ITS) 1-5.8S rDNA-ITS2 region was PCR-amplified and sequenced, followed by bioinformatic sequence analysis. RESULTS:Out of the 194 dermatophyte isolates, 132 samples (68.04%) were identified as T. indotineae, followed by T. tonsurans (14.43%), T. rubrum (7.22%), Microsporum canis (4.64%), T. interdigitale (3.61%), T. mentagrophytes (1.55%) and Arthroderma benhamiae (0.51%). Sequence analysis of 73 isolates from the past decade showed T. indotineae as the most frequently identified species (43.83%), followed by T. interdigitale (32.88%), T. mentagrophytes (21.92%) and Nannizzia fulva (1.37%). These findings indicate an increasing prevalence of T. indotineae in Iran in recent years. We analysed 214 T. mentagrophytes/T. interdigitale isolates, identifying 164 as T. indotineae, including 26 with nucleotide variations. A phylogenetic tree highlighted the genetic diversity within the species complex. CONCLUSION:The alarmingly high prevalence of the potentially drug-resistant species T. indotineae signals the necessity of continuous surveillance of skin dermatophytosis in the community.

Backgrounds: Leishmaniasis is one of the neglected tropical diseases, distributed across 89 countries in both the Old and New Worlds. Among the 54 identified Leishmania species, 21 are known to be pathogenic to humans. Cutaneous leishmaniasis (CL) is primarily caused by L. major and L. tropica, while visceral leishmaniasis (VL) in Iran is caused by L. infantum. Accurate detection and species identification of Leishmania spp. are crucial for more effective treatment, epidemiology, and control strategies for the disease. Among the molecular targets recently used for detecting Leishmania species, the heat-shock protein 70 (Hsp70) gene has proven to be highly suitable. Methods: This study aimed to establish and evaluate a SYBR Green real-time PCR targeting the Hsp70 gene to identify and differentiate three Leishmania species: L. major, L. tropica, and L. infantum in clinical specimens. A total of 219 microscopic smears, consisting of both positive and negative leishmaniasis cases diagnosed by microscopy, were subjected to DNA extraction and the Hsp70 real-time PCR assay designed in this study. Results: Based on the analysis of the melting temperature (Tm) of the amplified Hsp70 target, 115 microscopy-positive smears were identified, comprising 70.4% L. major, 23.5% L. tropica, and 6.1% L. infantum. All results were confirmed using a commercial diagnostic kit. Sanger sequencing of selected positive amplicons unequivocally confirmed the accuracy of this method in identifying and distinguishing the three Leishmania species. Conclusions: The Hsp70 real-time PCR can be considered an effective method for detecting and identifying Leishmania species from microscopic slides prepared from CL and VL cases in different regions of Iran.

Trichophyton indotineae has emerged as a significant global public health concern due to its role in recalcitrant dermatophytosis and antifungal treatment failure. Precise identification of T. indotineae is essential for timely and effective therapy, and for curbing the spread of antifungal resistance. However, current routine diagnostic methods are limited to reliably distinguish T. indotineae from other closely related dermatophytes. This study aimed to develop and evaluate three simple, cost-effective molecular methods for the accurate differentiation of T. indotineae. In silico analyses were performed to identify specific restriction enzyme cut sites within the internal transcribed spacer (ITS) region and the topoisomerase II gene of T. indotineae. A total of 430 dermatophyte isolates, including 267 previously identified by ITS sequencing and 163 clinical isolates of unknown identity, were subjected to PCR amplification of ITS and topoisomerase II followed by restriction fragment length polymorphism (PCR-RFLP) analysis using EarI (Eam11041) and BsuRI (HaeIII), respectively. Additionally, a previously described T. indotineae-specific PCR assay was evaluated. The enzyme EarI digested the ITS region of T. indotineae, producing a distinct PCR-RFLP pattern; likewise, BsuRI digested the topoisomerase II gene, enabling accurate differentiation of T. indotineae. The isolates previously identified by ITS sequencing were correctly classified by both methods, achieving high sensitivity and specificity. The T. indotineae-specific PCR assay demonstrated high sensitivity, although faint cross-reactivity was observed with T. tonsurans isolates. The ITS-PCR-RFLP and topoisomerase II-PCR-RFLP methods demonstrated high accuracy, affordability, and speed for the reliable identification of T. indotineae, making them suitable for routine use in clinical laboratories, especially in resource-limited settings. Although the T. indotineae-specific PCR assay showed high sensitivity, occasional cross-reactivity with T. tonsurans suggests that it should be interpreted with caution and ideally used alongside confirmatory tests.

Introduction:It is believed that DNA methylation can modify disease susceptibility in response to environmental factors as early as the perinatal period. In this study, we aimed to present a streamlined DNA methylation analysis procedure for osteoporosis-related gene promoters in the umbilical cord blood. Methods:The Prospective Epidemiological Research Studies in Iran (PERSIAN) birth cohort was established in 2016. In this study, a total of 300 umbilical cord blood samples were collected at the time of delivery. For all samples, DNA was extracted and converted using sodium bisulfite. Multiple primer sets were designed for Wnt1, Wnt10b, β-catenin, OPG, and RANKL gene promoters in the online MethPrimer platform. Next, bisulfite sequencing PCR (BSP), as the gold standard method for exploring methylated and unmethylated cytosines, was performed in a gradient-controlled setting. The PCR products were then purified and directly sequenced. Subsequently, the chromatograms were interpreted. Results:For Wnt10b, β-catenin, and OPG genes, the converted DNA could be successfully amplified. The frequency of acceptable chromatograms for analysis was 195 for Wnt10b (195/300, 0.65%), 198 for β-catenin (198/300, 0.66%), and 50 for OPG (50/50, 100%). Conclusion:BSP can be efficiently used to investigate the methylation of target gene promoters in umbilical cord blood DNA.

IntroductionAlthough the existence of Candida species in the respiratory tract is often considered commensal, it is crucial to recognize the significance of Candida colonization in immunocompromised or COVID-19 patients. The emergence of Candida auris as an emerging pathogen further emphasizes the importance of monitoring yeast infection/colonization, particularly in COVID-19 patients.MethodsIn this study, respiratory samples mainly from COVID-19 patients, primarily those suspected of having a fungal infection, were cultured on Sabouraud dextrose agar plates and the yeast colonies were identified using a two-step multiplex PCR method. The samples suspected of C. auris underwent specific nested PCR followed by sequence analysis.ResultsA total of 199 respiratory samples were collected from 73 women and 126 men, ranging in age from 1.6 to 88 years. Among the patients, 141 had COVID-19, 32 had cancer, 5 were hospitalized in ICU, 2 had chronic obstructive pulmonary disease)COPD(, and others were patients with combination diseases. From these samples, a total of 334 yeast strains were identified. C. albicans (n=132, 39.52%) was the most common species, followed by C. tropicalis (n=67, 20%), C. glabrata (n=56, 16.76%), C. krusei (n=18, 5.4%), C. parapsilosis (n=17, 5.08%), Saccharomyces cerevisiae (n=10, 3%), C. kefyr (n=9, 2.6%), C. dubliniensis (n=7, 2.1%), C. lusitaniae (n=5, 1.5%), C. auris (n=3, 0.9%), C. guilliermondii (n=2, 0.6%), C. rugosa (n=1, 0.3%), C. intermedia (n=1, 0.3%), and Trichosporon spp. (n=1, 0.3%). C. auris was detected in a patient in ICU and two COVID-19 patients. While its presence was confirmed through sequence analysis, our extensive efforts to isolate C. auris were unsuccessful.ConclusionWhile C. albicans colonization remains prevalent, our study found no evidence of Candida lung infection. Since the role of Candida colonization in airway secretions remains ambiguous due to limited research, further studies are imperative to shed light on this matter.

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly contagious virus that uses angiotensin converting enzyme 2 (ACE2), a pivotal member of the renin–angiotensin system (RAS), as its cell-entry receptor. Another member of the RAS, angiotensin II (Ang II), is the major biologically active component in this system. There is growing evidence suggesting that serum miRNAs could serve as prognostic biomarkers for SARS-CoV-2 infection and regulate ACE2 expression. Therefore, the aim of this study is to evaluate the changes in the serum levels of sACE2 and Ang II, as well as the expression level of miR-141-3p and miR-421 in SARS-CoV-2 positive and negative subjects. Methods In the present study, the serum levels of sACE2 and Ang II were measured in 94 SARS-CoV-2 positive patients and 94 SARS-CoV-2 negative subjects with some symptoms similar to those of SARS-CoV-2 positive patients using the ELISA method. In addition, the expression level of miR-141-3p and miR-421 as ACE2 regulators and biomarkers was evaluated using quantitative real-time PCR (qRT-PCR) method. Results The mean serum sACE2 concentration in the SARS-CoV-2-positive group was 3.268 ± 0.410 ng/ml, whereas in the SARS-CoV-2 negative group, it was 3.564 ± 0.437 ng/ml. Additionally, the mean serum Ang II level in the SARS-CoV-2 positive and negative groups were 60.67 ± 6.192 ng/L and 67.97 ± 6.837 ng/L, respectively. However, there was no significant difference in the serum levels of sACE2 (P value: 0.516) and Ang II (P value: 0.134) between the SARS-CoV-2 positive and negative groups. Meanwhile, our findings indicated that the expression levels of miR-141-3p and miR-421 in SARS-CoV-2 positive group were significantly lower and higher than SARS-CoV-2 negative group, respectively (P value < 0.001). Conclusions Taken together, the results of this study showed that the serum levels of sACE2 and Ang II in SARS-CoV-2 positive and negative subjects were not significantly different, but the expression levels of miR-141-3p and miR-421 were altered in SARS-CoV-2 positive patients which need more investigation to be used as biomarkers for COVID-19 diagnosis.

Introduction:The skin plays a crucial role as a protective barrier against external factors, but disruptions to its integrity can lead to wound formation and hinder the natural healing process. Scar formation and delayed wound healing present significant challenges in skin injury treatment. While alternative approaches such as skin substitutes and tissue engineering exist, they are often limited in accessibility and cost. Exosomes have emerged as a potential solution for wound healing due to their regenerative properties. Methods:In this study, exosomes were isolated from human blood serum using a kit. The exosomes were characterized, and their effects on cell migration were assessed in vitro. Additionally, the wound healing capacity of exosomes was evaluated in vivo using a rat full-thickness wound model. Results:Our in vitro findings revealed that exosomes significantly promoted cell migration. In vivo experiments demonstrated that the injection of exosomes at different areas of the wound accelerated the wound healing process, resulting in wound closure, collagen synthesis, vessel formation, and angiogenesis in the wound area. These results suggest that exosomes have a promising therapeutic potential for expediting wound healing and minimizing scar formation. Conclusions:The findings of this study highlight the potential of exosomes as a novel approach for enhancing wound healing. Exosomes showed positive effects on both cell migration and wound closure in in vitro and in vivo studies, suggesting their potential use as a regenerative therapy for skin injuries. Further research is needed to fully understand the mechanisms underlying the beneficial effects of exosomes on wound healing and to optimize their application in clinical settings.

Given the increasing occurrence of invasive fungal infections and the limited efficacy of modern antifungal medications, it is crucial to disseminate information regarding the potential sources of nosocomial mycoses through the One Health approach. This study investigated the presence and antifungal susceptibility of fungi in biofilm and water samples obtained from the drinking water distribution system (DWDS) of hospitals. The positivity rate for fungi in biofilm and water samples was 41% and 9%, respectively, with Aspergillus species, a significant causative agent of nosocomial mycoses, being the predominant fungi identified. Analysis of antifungal susceptibility test revelead a comparable resistance profile between some isolated species from the DWDS and those reported for certain clinical samples. While further research is required to determine the specific contribution of waterborne fungi to nosocomial fungal infections, our results emphasize the importance of controlling biofilm formation within DWDSs, particularly in high-risk hospital wards.

BackgroundIdentification of the opportunistic fungus Pneumocystis jirovecii in respiratory specimens presents challenges, particularly in differentiating between colonization and active infection. The present study assessed a probe-based real time PCR (qPCR) diagnostic effectiveness in patients with diverse underlying conditions, particularly those with COVID-19 and pulmonary insufficiency.MethodsTo set up the qPCR, clinical samples from 281 patients with respiratory ailments were tested. Subsequently, a descriptive study was conducted on 112 patients with pulmonary insufficiency with and without COVID-19 suspected of P. jirovecii infection. All specimens were subjected to DNA extraction followed by nested PCR and qPCR targeting the mitochondrial large subunit (mtLSU)-rRNA gene.ResultsBased on nested PCR and qPCR, P. jirovecii was identified in 40 out of 281 patients, with slight variations in positive samples observed across dilutions. Three patients who tested positive in nested PCR yielded negative results in probe-based qPCR. Conversely, three patients who tested positive in probe-based qPCR yielded negative results in nested PCR. Considering nested PCR as the golden standard, probe-based qPCR demonstrated good diagnostic performance, with 92.5% sensitivity and 98.7% specificity. Based on cycle threshold (Ct) values, the positive cases were categorized: ≤32 as infection, >35 as colonization, and a grey zone between these values (32 < X ≤ 35). The analysis of 112 PCP-suspected patients revealed a prevalence ranging from 6.25% (nested PCR) to 7% (probe-based qPCR).ConclusionsThis study suggested Ct values to differentiate Pneumocystis pneumonia/colonization in immunocompromised patients. To further augment the diagnostic sensitivity, it is recommended to integrate qPCR results with clinical parameters and biomarkers to offer a more precise understanding of Pneumocystis-related conditions.

IntroductionEarly detection of Pneumocystis jirovecii as an opportunistic pathogen that may endanger predisposed persons, including COVID-19 patients, may help to choose the optimal management.MethodsIn this study, 585, including 530 COVID-19 patients, with clinical and radiological evidence of respiratory diseases, were investigated for P. jirovecii screening. Clinical specimens were examined by direct microscopy and PCR, and randomly selected positive PCR products were confirmed through DNA sequence analysis.ResultsThirty-one (5.3%) samples were positive in P. jirovecii-specific nested-PCR, while by direct microscopic tests, Pneumocystis was observed in 22 (3.76%) samples. Males (61.7%) and patients over 50 years old (75.6%) were more commonly affected than others, and malaise and fatigue (84%), and wheezing (75%) were the most common symptoms, followed by fever (40.48%) and dyspnea (39.51%). Among the Pneumocystis-positive patients, three cases had coinfection with Aspergillus fumigatus, A. flavus, and A. niger (each n = 1), as documented by direct microscopy, culture, and species identification by PCR-sequencing.ConclusionPneumocystis pneumonia is still a diagnostic challenge; therefore, additional large-scale studies are needed to clarify the epidemiology of the disease in immunocompromised or COVID-19 patients.

During replication, some mutations occur in SARS-CoV-2, the causal agent of COVID-19, leading to the emergence of different variants of the virus. The mutations that accrue in different variants of the virus, influence the virus' ability to bind to human cell receptors and ability to evade the human immune system, the rate of viral transmission, and effectiveness of vaccines. Some of these mutations occur in the receptor binding domain (RBD) of the spike protein that may change the affinity of the virus for the ACE2 receptor. In this study, several in silico techniques, such as MD and SMD simulations, were used to perform comparative studies to deeply understand the effect of mutation on structural and functional details of the interaction of the spike glycoprotein of SARS-CoV-2, with the ACE2 receptor. According to our results, the mutation in the RBD associated with the Omicron variant increase binding affinity of the virus to ACE2 when compared to wild type and Delta variants. We also observed that the flexibility of the spike protein of the Omicron variant was lower than in comparison to other variants. In summary, different mutations in variants of the virus can have an effect on the binding mechanism of the receptor binding domain of the virus with ACE2.