pH-triggered morphological change in a self-assembling amphiphilic peptide used as an antitumor drug carrier.

The geometry of nanoparticles plays an important role in the process of drug encapsulation and release. In this study, an acid-responsive amphiphilic polypeptide consisting of lysine and leucine was prepared. In neutral media, the amphiphilic peptide L6K4 self-assembled to form spherical nanoparticles and encapsulated fat-soluble antitumor drugs. The intratumoral accumulation of the drug-loaded nanoparticles was improved in HeLa cells compared with normal cells. Compared to a neutral environment, increasingly acidic solutions changed the secondary structure of the peptide. In addition, the drug-loaded nanoparticles expanded and decomposed, rapidly releasing the poorly soluble antitumor drug doxorubicin (DOX). In addition, the amphiphilic peptide L6K4 had antitumor properties, and the antitumor performance of the combination of L6K4 and DOX was better than that of free DOX. Our results indicate that the use of acid responsiveness to induce geometric changes in drug-loaded peptide nanoparticles could be a promising strategy for antitumor drug delivery.

A case of Tinea Faciei caused by Trichophyton benhamiae: first report in China.

BACKGROUND: Trichophyton benhamiae is a zoophilic dermatophyte that can cause tinea in humans and animals. Lesions caused by T. benhamiae tend to be highly inflammatory, and patients are often infected by animals or other patients infected with T. benhamiae. In this paper, we report the first case of tinea faciei caused by T. benhamiae in a Chinese girl who might be transmitted from a fox. CASE PRESENTATION: A 4-year-old girl from HaiNing city developed an itchy, erythematous, and annular plaque on her right face for the past 2 months. Before the lesion appeared, she was in close contact with the fur of a fox for almost 1 week. Septate hyaline hyphae were detected by direct mycological examination of the scales. Cultures grew on Sabouraud's dextrose agar (SDA) at 26 °C for 2 weeks revealed the presence of T. mentagrophytes. A molecular sequencing test confirmed that the isolate was consistent with reference strains to T. benhamiae. Then, the diagnosis of tinea faciei due to T. benhamiae was made. Treatment with terbinafine (oral 125 mg/d) and sertaconazole nitrate cream (topical, twice daily) for 4 weeks was initiated and achieved significant improvement of the skin lesions. CONCLUSIONS: This rare dermatophytosis case highlights the importance of ITS sequencing in helping to recognize rare pathogenic fungi that can be easily misdiagnosed with a conventional morphological diagnosis.

Critical role of caveolin-1 in ocular neovascularization and multitargeted antiangiogenic effects of cavtratin via JNK.

Ocular neovascularization is a devastating pathology of numerous ocular diseases and is a major cause of blindness. Caveolin-1 (Cav-1) plays important roles in the vascular system. However, little is known regarding its function and mechanisms in ocular neovascularization. Here, using comprehensive model systems and a cell permeable peptide of Cav-1, cavtratin, we show that Cav-1 is a critical player in ocular neovascularization. The genetic deletion of Cav-1 exacerbated and cavtratin administration inhibited choroidal and retinal neovascularization. Importantly, combined administration of cavtratin and anti-VEGF-A inhibited neovascularization more effectively than monotherapy, suggesting the existence of other pathways inhibited by cavtratin in addition to VEGF-A. Indeed, we found that cavtratin suppressed multiple critical components of pathological angiogenesis, including inflammation, permeability, PDGF-B and endothelial nitric oxide synthase expression (eNOS). Mechanistically, we show that cavtratin inhibits CNV and the survival and migration of microglia and macrophages via JNK. Together, our data demonstrate the unique advantages of cavtratin in antiangiogenic therapy to treat neovascular diseases.

Subcutaneous granuloma caused by Kodamaea ohmeri in an immunocompromised patient in China.

We present a case of subcutaneous granuloma caused by Kodamaea ohmeri and describe the histopathological characteristics and skin lesions caused by this pathogen. A 55-year-old woman, with a history of rheumatoid arthritis, presented with red, painless plaque on her forehead, persistent for 3 months; based on the histopathological and mycological findings and gene sequencing, the patient was diagnosed with subcutaneous granuloma caused by K. ohmeri. This report highlights that early identification and diagnosis and optimal regimens are vital in the management of this intractable infection.

A novel murine antibody and an open sandwich immunoassay for the detection of clenbuterol.

Clenbuterol (CLEN) is a sympathomimetic amine used as a decongestant and bronchodilator while treating breathing disorders. It is also used in food-producing animals as it improves the rate of red meat production. However, it is prohibited in many countries nowadays due to human health and safety concerns. Unfortunately, the illegal use of CLEN is still rampant. Thus, monitoring it in food and livestock is important. Here, we report a novel murine antibody and an open sandwich enzyme linked immunosorbent assay (OS-ELISA) to detect CLEN based on antigen-antibody reactions. The genes of antibody variable regions in mice immunized with CLEN conjugated with bovine serum albumin were cloned into a phagemid (pDong1/Fab) to construct a phage-display antibody library, from which a novel antibody, A12, was selected. Then, an OS-ELISA was developed to detect CLEN using separated variable regions of the A12 antibody. The limit of detection of the assay was found to be 8 ng/mL, which was useful for monitoring CLEN usage.

Characterization of Skin Microbiome in Tinea Pedis.

Tinea pedis has been associated with Trichophyton rubrum infection. However, it's not clear whether other microbes were implicated in disease pathogenesis. The composition of microbial communities of patients with tinea pedis and healthy controls were analyzed to identify the characteristics of tinea pedis and differences associated with clinical patterns. We found that microbial community structures were different in patients with tinea pedis compared with healthy controls. Moreover, skin microbiome varied in different forms of tinea pedis. Healthy controls exhibited greater fungal diversity than patients with tinea pedis. In patients with tinea pedis, the dominant bacterial and fungal genera were Staphylococcus and Trichophyton. Compared with healthy controls, Corynebacterium tuberculostearicum was decreased and T. rubrum was increased. C. tuberculostearicum was more abundant in vesicular tinea pedis than in hyperkeratotic and interdigital tinea pedis. Interdigital tinea pedis had a higher detection rate of Corynebacterium minutissimum and T. rubrum than the other forms. These results indicated that bacterial microbes may take part in the development of tinea pedis.

ZNF185-derived peptide induces fertility suppression in mice.

Zinc finger protein 185 (ZNF185) belongs to the ZNF family and is involved in male reproduction. However, it is unclear whether ZNF185 may be a target candidate for contraceptive vaccines. In this study, antigenic peptides derived from ZNF185 were prepared, and their immune contraceptive effects were investigated using mice. Results from enzyme-linked immunosorbent assay (ELISAs) showed that peptide immunization induced an antibody titre increase that reached a peak in week 12. Peptide-3 and peptide-4 were then chosen for subsequent experiments. The results of the fertility assays showed that peptide immunization inhibited the mating and fertility rates of the mice, whereas there were no obvious changes in the number of pups per litter. Subsequently, epididymal sperm was analysed. The results demonstrated that the sperm count and sperm motility were significantly decreased in the peptide group, while the amount of abnormal sperm was significantly increased in the peptide-3 group. The male reproductive organs were also evaluated. There were no obvious differences in testis or epididymal weights, in the diameters of the seminiferous tubules, or in the thicknesses of the seminiferous epithelium between the peptide group and the phosphate buffer saline (PBS) group. In addition, histological analysis indicated that there were no obvious pathologic changes in testis and epididymal histology in the peptide group; however, the number of spermatozoa present in the epididymal lumen of the peptide group was significantly decreased when compared with the PBS group. Our study demonstrates for the first time that peptides derived from ZNF185 may induce fertility suppression in mice without damaging reproductive organs. These peptides have the potential to be used as a male contraceptive vaccine.

Subcutaneous Phaeohyphomycosis Caused by Exophiala oligosperma in an Immunocompetent Host: Case Report and Literature Review.

We report a case of subcutaneous infection caused by Exophiala oligosperma. Erythematous ulcerated plaque with exudate was major clinical features. Histopathological examination showed yeast-like cells and fungal hyphae. Mycological and molecular identification revealed E. oligosperma as etiologic agent. Local debridement and oral itraconazole were effective. To the best of our knowledge, this is the first report of phaeohyphomycosis caused by E. oligosperma in mainland China. This report highlights the potential role of E. oligosperma as an emerging cause of infection in immunocompetent patients.

Successful Sequential Treatment with Itraconazole and ALA-PDT for Cutaneous Granuloma by Candida albicans: A Case Report and Literature Review.

Photodynamic therapy (PDT) is a process that combines a photosensitizing drug and light and promotes phototoxic responses in target cells, mainly via oxidative damage. Antifungal photodynamic therapy has been successfully employed against Candida species, dermatophytes, and deep mycoses. We present a case of a cutaneous granuloma caused by C. albicans treated with 5-aminolevulinic acid (ALA)-PDT. A 64-year-old man presented with two plaques on his right hand and wrist for 2 years. The diagnosis was made based on histopathology, mycology, and molecular identification of paraffin-embedded tissues. The patient was treated with itraconazole for 1 month and two sessions of ALA-PDT. After 2 months of follow-up, the patient was cured and has not experienced any recurrence to date. ALA-PDT was well tolerated in this patient with little pain. In general, application of PDT in mycoses is safe and effective in most cases. ALA-PDT is a good choice for inactivation of C. albicans.

Vasoprotective effect of PDGF-CC mediated by HMOX1 rescues retinal degeneration.

Blood vessel degeneration is critically involved in nearly all types of degenerative diseases. Therefore strategies to enhance blood vessel protection and survival are highly needed. In this study, using different animal models and cultured cells, we show that PDGF-CC is a potent vascular protective and survival factor. PDGF-CC deficiency by genetic deletion exacerbated blood vessel regression/degeneration in various animal models. Importantly, treatment with PDGF-CC protein not only increased the survival of retinal blood vessels in a model of oxygen-induced blood vessel regression but also markedly rescued retinal and blood vessel degeneration in a disease model of retinitis pigmentosa. Mechanistically, we revealed that heme oxygenase-1 (HMOX1) activity is critically required for the vascular protective/survival effect of PDGF-CC, because blockade of HMOX1 completely abolished the protective effect of PDGF-CC in vitro and in vivo. We further found that both PDGF receptors, PDGFR-ß and PDGFR-α, are required for the vasoprotective effect of PDGF-CC. Thus our data show that PDGF-CC plays a pivotal role in maintaining blood vessel survival and may be of therapeutic value in treating various types of degenerative diseases.

miR-93 promotes TGF-ß-induced epithelial-to-mesenchymal transition through downregulation of NEDD4L in lung cancer cells.

The level of microRNA-93 (miR-93) in tumors has been recently reported to be negatively correlated with survival of lung cancer patients. Considering that the most devastating aspect of lung cancer is metastasis, which can be promoted by transforming growth factor-ß (TGF-ß)-induced epithelial-to-mesenchymal transition (EMT), we sought to determine whether miR-93 is involved in this process. Here, we report that a previously unidentified target of miR-93, neural precursor cell expressed developmentally downregulated gene 4-like (NEDD4L), is able to mediate TGF-ß-mediated EMT in lung cancer cells. miR-93 binds directly to the 3'-UTR of the NEDD4L messenger RNA (mRNA), leading to a downregulation of NEDD4L expression at the protein level. We next demonstrated that the downregulation of NEDD4L enhanced, while overexpression of NEDD4L reduced TGF-ß signaling, reflected by increased phosphorylation of SMAD2 in the lung cancer cell line after TGF-ß treatment. Furthermore, overexpression of miR-93 in lung cancer cells promoted TGF-ß-induced EMT through downregulation of NEDD4L. The analysis of publicly available gene expression array datasets indicates that low NEDD4L expression correlates with poor outcomes among patients with lung cancer, further supporting the oncogenic role of miR-93 in lung tumorigenesis and metastasis.

pH-Responsive Hyaluronic Acid-Based Mixed Micelles for the Hepatoma-Targeting Delivery of Doxorubicin.

The tumor targetability and stimulus responsivity of drug delivery systems are crucial in cancer diagnosis and treatment. In this study, hepatoma-targeting mixed micelles composed of a hyaluronic acid-glycyrrhetinic acid conjugate and a hyaluronic acid-l-histidine conjugate (HA-GA/HA-His) were prepared through ultrasonic dispersion. The formation and characterization of the mixed micelles were confirmed via ¹H-NMR, particle size, and ζ potential measurements. The in vitro cellular uptake of the micelles was evaluated using human liver carcinoma (HepG2) cells. The antitumor effect of doxorubicin (DOX)-loaded micelles was investigated in vitro and in vivo. Results indicated that the DOX-loaded HA-GA/HA-His micelles showed a pH-dependent controlled release and were remarkably absorbed by HepG2 cells. Compared with free DOX, the DOX-loaded HA-GA/HA-His micelles showed a higher cytotoxicity to HepG2 cells. Moreover, the micelles effectively inhibited tumor growth in H22 cell-bearing mice. These results suggest that the HA-GA/HA-His mixed micelles are a good candidate for drug delivery in the prevention and treatment of hepatocarcinoma.

miR-125b-1-3p inhibits trophoblast cell invasion by targeting sphingosine-1-phosphate receptor 1 in preeclampsia.

Preeclampsia (PE) is the leading cause of maternal and perinatal mortality and morbidity. Understanding the molecular mechanisms underlying placentation facilitates the development of better intervention of this disease. MicroRNAs are strongly implicated in the pathogenesis of this syndrome. In current study, we found that miR-125b-1-3p was elevated in placentas derived from preeclampsia patients. Transfection of miR-125b-1-3p mimics significantly inhibited the invasiveness of human trophoblast cells, whereas miR-125b-1-3p inhibitor enhanced trophoblast cell invasion. Luciferase assays identified that S1PR1 was a novel direct target of miR-125b-1-3p in the placenta. Overexpression of S1PR1 could reverse the inhibitory effect of miR-125b-1-3p on the invasion of trophoblast cells. These findings suggested that abnormal expression of miR-125b-1-3p might contribute to the pathogenesis of preeclampsia.

[Association of 15 short tandem repeats loci with aggressive behavior].

OBJECTIVE: To assess the association between aggressive behaviors and 15 short tandem repeats (STRs) loci. METHODS: Peripheral blood samples from 541 army men with aggressive behaviors and 459 healthy individuals were collected. All sample were amplified with a AmpFlSTR Identifiler(TM) system and separated by electrophoresis to compare the genotypic and allelic frequencies of 15 STRs (CSF1PO, D2S1338, D3S1358, D5S818, D7S820, D8S1179, D13S317, D16S539, D18S51, D19S433, D21S11, FGA, THO1, TPOX and vWA) in the two groups. RESULTS: A significant difference was found in allelic and genotypic frequencies at loci D2S1338 and D19S433 (P< 0.01) between the two groups, but not for the remaining 13 STR loci (P> 0.05). Univarite analysis also showed a significant difference for allele 16, genotypes 19-22, 22-24 on D2S1338 and genotypes 13-14.2 on D19S433 between the two groups (P= 0.0018, P= 0.0001, P= 0.0003, P= 0.0000), with the OR values being 7.380 (95%CI: 1.701-32.028), 0.051(95%CI: 0.007-0.388), 13.933(95%CI: 1.845-105.717), 0.349 (95%CI: 0.216-0.564), respectively. CONCLUSION: D2S1338 and D19S433 may be associated with aggressive behavior. Allele 16 and genotype 22-24 on D2S1338 may be susceptible factors for the disease, whilst genotypes 19-22 on D2S1338 and 13-14.2 on D19S433 may confer a protective effect on it.

Enhancing the Inhibition of Breast Cancer Growth Through Synergistic Modulation of the Tumor Microenvironment Using Combined Nano-Delivery Systems.

Purpose: Breast cancer is a prevalent malignancy among women worldwide, and malignancy is closely linked to the tumor microenvironment (TME). Here, we prepared mixed nano-sized formulations composed of pH-sensitive liposomes (Ber/Ru486@CLPs) and small-sized nano-micelles (Dox@CLGs). These liposomes and nano-micelles were modified by chondroitin sulfate (CS) to selectively target breast cancer cells. Methods: Ber/Ru486@CLPs and Dox@CLGs were prepared by thin-film dispersion and ethanol injection, respectively. To mimic actual TME, the in vitro "condition medium of fibroblasts + MCF-7" cell model and in vivo "4T1/NIH-3T3" co-implantation mice model were established to evaluate the anti-tumor effect of drugs. Results: The physicochemical properties showed that Dox@CLGs and Ber/Ru486@CLPs were 28 nm and 100 nm in particle size, respectively. In vitro experiments showed that the mixed formulations significantly improved drug uptake and inhibited cell proliferation and migration. The in vivo anti-tumor studies further confirmed the enhanced anti-tumor capabilities of Dox@CLGs + Ber/Ru486@CLPs, including smaller tumor volumes, weak collagen deposition, and low expression levels of α-SMA and CD31 proteins, leading to a superior anti-tumor effect. Conclusion: In brief, this combination therapy based on Dox@CLGs and Ber/Ru486@CLPs could effectively inhibit tumor development, which provides a promising approach for the treatment of breast cancer.

Duodenal-jejunal bypass improves hypothalamic oxidative stress and inflammation in diabetic rats via glucagon-like peptide 1-mediated Nrf2/HO-1 signaling.

BACKGROUND: Type 2 diabetes mellitus (T2DM) is often accompanied by impaired glucose utilization in the brain, leading to oxidative stress, neuronal cell injury and infla-mmation. Previous studies have shown that duodenal jejunal bypass (DJB) surgery significantly improves brain glucose metabolism in T2DM rats, the role and the metabolism of DJB in improving brain oxidative stress and inflammation condition in T2DM rats remain unclear. AIM: To investigate the role and metabolism of DJB in improving hypothalamic oxidative stress and inflammation condition in T2DM rats. METHODS: A T2DM rat model was induced via a high-glucose and high-fat diet, combined with a low-dose streptozotocin injection. T2DM rats were divided into DJB operation and Sham operation groups. DJB surgical intervention was carried out on T2DM rats. The differential expression of hypothalamic proteins was analyzed using quantitative proteomics analysis. Proteins related to oxidative stress, inflammation, and neuronal injury in the hypothalamus of T2DM rats were analyzed by flow cytometry, quantitative real-time PCR, Western blotting, and immunofluorescence. RESULTS: Quantitative proteomics analysis showed significant differences in proteins related to oxidative stress, inflammation, and neuronal injury in the hypothalamus of rats with T2DM-DJB after DJB surgery, compared to the T2DM-Sham groups of rats. Oxidative stress-related proteins (glucagon-like peptide 1 receptor, Nrf2, and HO-1) were significantly increased (P < 0.05) in the hypothalamus of rats with T2DM after DJB surgery. DJB surgery significantly reduced (P < 0.05) hypothalamic inflammation in T2DM rats by inhibiting the activation of NF-κB and decreasing the expression of interleukin (IL)-1ß and IL-6. DJB surgery significantly reduced (P < 0.05) the expression of factors related to neuronal injury (glial fibrillary acidic protein and Caspase-3) in the hypothalamus of T2DM rats and upregulated (P < 0.05) the expression of neuroprotective factors (C-fos, Ki67, Bcl-2, and BDNF), thereby reducing hypothalamic injury in T2DM rats. CONCLUSION: DJB surgery improve oxidative stress and inflammation in the hypothalamus of T2DM rats and reduce neuronal cell injury by activating the glucagon-like peptide 1 receptor-mediated Nrf2/HO-1 signaling pathway.

Improved efficacy of cisplatin delivery by peanut agglutinin­modified liposomes in non­small cell lung cancer.

Globally, non­small cell lung cancer (NSCLC) is a significant threat to human health, and constitutes >80% of lung cancer cases. Cisplatin (CDDP), a commonly used drug in clinical treatment, has been the focus of research aiming to mitigate its potent toxicity through encapsulation within liposomes. However, challenges, such as a reduced drug loading efficiency and nonspecific release, have emerged as obstacles. The present study aimed to improve the encapsulation efficiency of CDDP within liposomes by pre­preparation of CDDP and modifying the liposome surface through the incorporation of peanut agglutinin (PNA) as a ligand [CDDP­loaded PNA­modified liposomes (CDDP­PNA­Lip)]. This strategy was designed to enhance the delivery of CDDP to tumour tissues, thereby reducing associated side effects. The effect of CDDP­PNA­Lip on the proliferation and migration of NSCLC cell lines with high MUC1 expression was elucidated through in vitro studies. Additionally, the capacity of PNA modification to augment the targeted anti­tumour efficacy of liposomes was assessed through xenograft tumour experiments. The results indicated that in an in vitro uptake assay Rhodamine B (RhB)­loaded PNA­modified liposomes were taken up by cells with ~50% higher efficiency compared with free RhB. In addition, CDDP­PNA­Lip resulted in a 2.65­fold enhancement of tumour suppression in vivo compared with free CDDP. These findings suggested that the encapsulation of CDDP within ligand­modified liposomes may significantly improve its tumour­targeting capabilities, providing valuable insights for clinical drug development.

Case report: Subcutaneous Mycobacterium haemophilum infection in an immunocompetent patient after lipolysis injections.

Mycobacterium haemophilum is a slow-growing, aerobic mycobacterium that acts as a pathogen in immunocompromised adult patients and immunocompetent children. There are only a few rare cases in the literature describing this species as a cause of subcutaneous infections. Here, we describe a subcutaneous infection caused by M. haemophilum in an immunocompetent female after lipolysis injections at an unqualified beauty salon, suggesting that this bacteria can also be a potential causative agent of adverse events in medical aesthetics. In addition, M. haemophilum caused lesions not only at the injection sites and adjacent areas but also invaded distant sections through the subcutaneous sinus tracts. Thus, early diagnosis and appropriate treatment are vital to prevent further deterioration and improve prognosis.

Quantitative SERS detection of multiple breast cancer miRNAs based on duplex specific nuclease-mediated signal amplification.

Simultaneous and ultrasensitive detection of multiple microRNA (miRNA) biomarkers is an essential precondition for early cancer diagnosis and treatment. Here we developed a sandwich surface-enhanced Raman scattering (SERS) sensor based on Au@Ag core-shell nanorods combined with duplex specific nuclease-mediated signal amplification (DSNSA) for quantitative detection of multiple breast cancer miRNA biomarkers. The DSNSA strategy enables quantitative detection of target miRNA through rehybridizing the capture probe DNA-SERSnanotag conjugates to trigger signal amplification. The Au@Ag core-shell nanorods coated with an Ag shell exhibit excellent SERS performance, implying that molecules can be concentrated by the Ag shell at the hot spots. By monitoring the Raman signal attenuation of hot spots in the presence of target miRNAs, three breast cancer associated miRNAs (miR-21, miR-155, and let 7b) were simultaneously determined using the sandwich SERS sensor, and their detection limits (LODs) were 0.05 fM, 0.063 fM and 0.037 fM, respectively. These results indicated that our sandwich SERS sensor combined with the DSNSA strategy holds remarkable promise for multiplex detection of cancer biomarkers and contributes to early diagnosis of cancer.

DNA microarray chip assay in new use: early diagnostic value in cutaneous mycobacterial infection.

Introduction: The clinical practicability of DNA microarray chip in detecting the presence of mycobacterial species/isolates directly in the skin tissues has not been evaluated, nor the efficacy of DNA microarray chip as a novel diagnostic tool for the early diagnosis of cutaneous mycobacterial infections is known. Methods: The present study analyzed the incidence of cutaneous mycobacterial infections in Shanghai and explored the efficacy of a novel DNA microarray chip assay for the clinical diagnosis of the disease from skin tissue specimens compared to traditional detection methods. A total of 60 participants fulfilling the defined diagnostic criteria and confirmed positive for cutaneous mycobacterial infections from 2019 to 2021 were enrolled in the study. Subsequent to recording the participants' medical history and clinical characteristics, the skin tissue specimens were collected for analyses. The specimens underwent histopathological analyses, skin tissue culture, and DNA microarray chip assay. Results: Increased incidence of cutaneous mycobacterial infection was detected from 2019 to 2021. The most common infecting pathogen was M. marinum followed by M. abscessus. The sensitivity, specificity and accuracy of the skin tissue culture method were 70%, 100% and 76.62%, respectively, while that of the DNA microarray chip assay were 91.67%, 100% and 93.51%, respectively. The sensitivity and accuracy of the DNA microarray chip assay were significantly higher than those of the skin tissue culture method. The positive likelihood and diagnostic odds ratio were >10 and >1, respectively for both the methods. The negative likelihood ratio was significantly higher (30% vs 8.33%) and the Youden's index was significantly lower (70.00% vs 91.67%) in the skin culture method compared to that of the DNA microarray chip assay. There was a significant association of false negative results with a history of antibiotic use in the skin tissue culture method. Discussion: Given the increasing incidence of cutaneous mycobacterial infections, early diagnosis remains a prime clinical focus. The DNA microarray chip assay provides a simple, rapid, high-throughput, and reliable method for the diagnosis of cutaneous mycobacterial infections with potential for clinical application.