A comparative study on the detection of Mycobacterium leprae DNA in urine samples of leprosy patients using Rlep-PCR with other conventional samples.

BACKGROUND: Mycobacterium leprae causes leprosy that is highly stigmatized and chronic infectious skin disease. Only some diagnostic tools are being used for the identification M. leprae in clinical samples, such as bacillary detection, and histopathological tests. These methods are invasive and often have low sensitivity. Currently, the PCR technique has been used as an effective tool fordetecting M. leprae DNA across different clinical samples. The current study aims to detect M. leprae DNA in urine samples of untreated and treated leprosy patients using the Rlep gene (129 bp) and compared the detection among Ridley-Jopling Classification. METHODS: Clinical samples (Blood, Urine, and Slit Skin Smears (SSS)) were collected from leprosy and Non-leprosy patients. DNA extraction was performed using standard laboratory protocol and Conventional PCR was carried out for all samples using Rlep gene target and the amplicons of urine samples were sequenced by Sanger sequencing to confirm the Rlep gene target. RESULTS: The M. leprae DNA was successfully detected in all clinical samples across all types of leprosy among all the study groups using RLEP-PCR. Rlep gene target was able to detect the presence of M. leprae DNA in 79.17% of urine, 58.33% of blood, and 50% of SSS samples of untreated Smear-Negative leprosy patients. The statistical significant difference (p = 0.004) was observed between BI Negative (Slit Skin Smear test) and RLEP PCR positivity in urine samples of untreated leprosy group. CONCLUSION: The PCR positivity using Rlep gene target (129 bp) was highest in all clinical samples among the study groups, across all types of leprosy. Untreated tuberculoid and PNL leprosy patients showed the highest PCR positivity in urine samples, indicating its potential as a non-invasive diagnostic tool for leprosy and even for contact screening.

Microbial Dynamics of a Specialty Italian Raw Ewe's Milk Cheese Curdled with Extracts from Spontaneous and Cultivated Onopordum tauricum Willd.

Milk coagulants prepared by maceration of flowers harvested from both spontaneous and cultivated Onopordum tauricum Willd. and a commercially available coagulant obtained from Cynara cardunculus L. (control) were assayed for small-scale manufacturing of Caciofiore, an Italian specialty raw ewe's milk cheese produced in a family run dairy farm located in the Marche region (Central Italy). The microbiota of the three thistle-based milk coagulants and their effect on the microbial dynamics of raw milk cheeses during fermentation and maturation (from day 0 up until day 60) were investigated through a combined approach based on viable counting and Illumina DNA sequencing. In both the control and experimental cheeses, despite the slight differences emerged depending on the coagulant used, Lactococcus lactis and Debaryomyces hansenii were the prevalent species among bacteria and fungi, respectively. Moreover, raw ewe's milk was the main factor affecting the evolution of both the bacterial and fungal microbiota in all cheeses. The overall similarities between control and experimental cheeses herein analyzed supports the exploitation of Onopordum tauricum Willd. as an alternative milk coagulating agent for production of Caciofiore and, more in general, raw ewe's milk cheeses.

Nannizzia incurvata as a rare cause of favus and tinea corporis in Cambodia and Vietnam.

Nannizzia (N.) incurvata (formerly Microsporum incurvatum) represents a geophilic dermatophyte which has been previously classified as belonging to the species complex of N. gypsea (formerly Microsporum gypseum). A 42-year-old Vietnamese female from Saxony, Germany, suffered from tinea corporis of the right buttock after she returned from a 2-week-visit to her homeland Vietnam. From skin scrapings of lesions, N. incurvata grew on Sabouraud's dextrose agar. Treatment by ciclopirox olamine cream twice daily for 4 weeks was successful. A 6-year-old Cambodian boy living near river Mekong with contact history to chicken, dogs and cattle suffered from tinea faciei and capitis. Symptoms of the favus-like tinea capitis and tinea faciei were erythema and scaly patches with areas of alopecia. N. incurvata grew on Sabouraud's dextrose agar. The boy was treated with oral terbinafine 125 mg daily, topical miconazole cream and ketoconazole shampoo. The symptoms healed within 4 weeks of treatment. Cultivation of the samples revealed growth of N. incurvata. For confirmation of species identification, the isolates were subject to sequencing of ITS (internal transcribed spacer) region of the rDNA, and addition of the "translation elongation factor 1 α" (TEF 1 α) gene. Sequencing of the ITS region showed 100% accordance with the sequence of N. incurvata deposited at the NCBI database under the accession number MF415405. N. incurvata is a rare, or might be underdiagnosed geophilic dermatophyte described in Sri Lanka and Vietnam until now. This is the first isolation of N. incurvata in Cambodia, and the first description of favus in a child due to this dermatophyte.

Four novel mutations of ADAR1 in Chinese patients with dyschromatosis symmetrica hereditaria.

BACKGROUND: Novel mutations in adenosine deaminase acting on RNA 1 gene (ADAR1) are responsible for dyschromatosis symmetrica hereditaria (DSH). DSH patients display a mixture of hyperpigmented and hypopigmented macules on the dorsal aspects of the extremities, and freckle-like macules on the face. AIMS: To provide new evidence for further study of the etiopathogenisis of DSH. METHODS: Genomic DNA was extracted and used as a template for the polymerase chain reaction (PCR) amplification of all 15 coding exons as well as intron-exon boundaries of ADAR1. The PCR products were sequenced directly. RESULTS: We identified eight mutations of ADAR1 in four Chinese pedigrees and four individual patients, which were c.2722G>T, p.(Asp908Tyr), c.1657delA, p.(Ser553fs), c.2563_2564delCT, p.(Leu855fs), c.526T>G, p.(Leu176Val) as well as four previously reported mutations c. 3363_3364insT, p.(Lys1122fs), c. 2865_2866delGT, p.(Val955fs), c.1630C>T, p.(Arg544X), and c.2894C>T, p.(Pro965Leu). In silico analysis predicted that all the mutations reported were pathogenic. LIMITATIONS: We did not study how ADAR1 played its role in DSH. So, the exact pathogenic mechanism of ADAR1 in DSH patients wasn't clarified in this study. CONCLUSION: We found four novel ADAR1 mutations in this study. Our results enlarge the database on ADAR1 mutations associated with DSH.