Rapid and sensitive screening of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is essential to limit the spread of the global pandemic we are facing. Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) is currently used for the clinical diagnosis of SARS-CoV-2 infection using nasopharyngeal swabs, tracheal aspirates, or bronchoalveolar lavage (BAL) samples. Despite the high sensitivity of the qRT-PCR method, false negative outcomes might occur, especially in patients with a low viral load. Here, we developed a multiplex qRT-PCR methodology for the simultaneous detection of SARS-CoV-2 genome (N gene) and of the human RNAse P gene as internal control. We found that multiplex qRT-PCR was effective in detecting SARS-Cov-2 infection in human specimens with 100% sensitivity. Notably, patients with few copies of SARS-CoV-2 RNA (<5 copies/reaction) were successfully detected by the novel multiplex qRT-PCR method. Finally, we assessed the efficacy of multiplex qRT-PCR on human nasopharyngeal swabs without RNA extraction. Collectively, our results provide evidence of a novel and reliable tool for SARS-CoV-2 RNA detection in human specimens, which allows the testing capacity to be expanded and the RNA extraction step to be bypassed.

Purpose Does controlled ovarian stimulation (COS) and progesterone (P) luteal supplementation modify the vaginal and endometrial microbiota of women undergoing in vitro fertilization? Methods Fifteen women underwent microbiota analysis at two time points: during a mock transfer performed in the luteal phase of the cycle preceding COS, and at the time of fresh embryo transfer (ET). A vaginal swab and the distal extremity of the ET catheter tip were analyzed using next-generation 16SrRNA gene sequencing. Heterogeneity of the bacterial microbiota was assessed according to both the Bray-Curtis similarity index and the Shannon diversity index. Results Lactobacillus was the most prevalent genus in the vaginal samples, although its relative proportion was reduced by COS plus P supplementation (71.5 ± 40.6% vs. 61.1 ± 44.2%). In the vagina, an increase in pathogenic species was observed, involving Prevotella (3.5 ± 8.9% vs. 12.0 ± 19.4%), and Escherichia coli-Shigella spp. (1.4 ± 5.6% vs. 2.0 ± 7.8%). In the endometrium, the proportion of Lactobacilli slightly decreased (27.4 ± 34.5% vs. 25.0 ± 29.9%); differently, both Prevotella and Atopobium increased (3.4 ± 9.5% vs. 4.7 ± 7.4% and 0.7 ± 1.5% vs. 5.8 ± 12.0%). In both sites, biodiversity was greater after COS (p < 0.05), particularly in the endometrial microbiota, as confirmed by Bray-Curtis analysis of the phylogenetic distance among bacteria genera. Bray-Curtis analysis confirmed significant differences also for the paired endometrium-vagina samples at each time point. Conclusions Our findings suggest that COS and P supplementation significantly change the composition of vaginal and endometrial microbiota. The greater instability could affect both endometrial receptivity and placentation. If our findings are confirmed, they may provide a further reason to encourage the freeze-all strategy.

When the patients are correctly selected, endourological treatment plays an equally important role in the treatment of gynaecologically-related ureteric injuries when compared to open surgery. Special attention should be paid to the treatment of lesions caused by radical hysterectomy alone or associated with radiotherapy, as these may lead to poor results.

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