BackgroundMetagenomic next-generation sequencing (mNGS) is increasingly being used to detect pathogens directly from clinical specimens. However, the optimal application of mNGS and subsequent result interpretation can be challenging. In addition, studies reporting the use of mNGS for the diagnosis of invasive fungal infections (IFIs) are rare.ObjectiveWe critically evaluated the performance of mNGS in the diagnosis of pulmonary IFIs, by conducting a multicenter retrospective analysis. The methodological strengths of mNGS were recognized, and diagnostic cutoffs were determined.MethodsA total of 310 patients with suspected pulmonary IFIs were included in this study. Conventional microbiological tests (CMTs) and mNGS were performed in parallel on the same set of samples. Receiver operating characteristic (ROC) curves were used to evaluate the performance of the logarithm of reads per kilobase per million mapped reads [lg(RPKM)], and read counts were used to predict true-positive pathogens.ResultThe majority of the selected patients (86.5%) were immunocompromised. Twenty species of fungi were detected by mNGS, which was more than was achieved with standard culture methods. Peripheral blood lymphocyte and monocyte counts, as well as serum albumin levels, were significantly negatively correlated with fungal infection. In contrast, C-reactive protein and procalcitonin levels showed a significant positive correlation with fungal infection. ROC curves showed that mNGS [and especially lg(RPKM)] was superior to CMTs in its diagnostic performance. The area under the ROC curve value obtained for lg(RPKM) in the bronchoalveolar lavage fluid of patients with suspected pulmonary IFIs, used to predict true-positive pathogens, was 0.967, and the cutoff value calculated from the Youden index was −5.44.ConclusionsIn this study, we have evaluated the performance of mNGS-specific indicators that can identify pathogens in patients with IFIs more accurately and rapidly than CMTs, which will have important clinical implications.

Circular RNAs (circRNAs) are highly enriched in the central nervous system and significantly involved in a range of brain-related physiological and pathological processes. Ischemic stroke is a complex disorder caused by multiple factors; however, whether brain-derived circRNAs participate in the complex regulatory networks involved in stroke pathogenesis remains unknown. Here, we successfully constructed a cerebral ischemia-injury model of middle cerebral artery occlusion (MCAO) in male Sprague-Dawley rats. Preliminary qualitative and quantitative analyses of poststroke cortical circRNAs were performed through deep sequencing, and RT-PCR and qRT-PCR were used for validation. Of the 24,858 circRNAs expressed in the rat cerebral cortex, 294 circRNAs were differentially expressed in the ipsilateral cerebral cortex between the MCAO and sham rat groups. Cluster, GO, and KEGG analyses showed enrichments of these circRNAs and their host genes in numerous biological processes and pathways closely related to stroke. We selected 106 of the 294 circRNAs and constructed a circRNA-miRNA-mRNA interaction network comprising 577 sponge miRNAs and 696 target mRNAs. In total, 15 key potential circRNAs were predicted to be involved in the posttranscriptional regulation of a series of downstream target genes, which are widely implicated in poststroke processes, such as oxidative stress, apoptosis, inflammatory response, and nerve regeneration, through the competing endogenous RNA mechanism. Thus, circRNAs appear to be involved in multilevel actions that regulate the vast network of multiple mechanisms and events that occur after a stroke. These results provide novel insights into the complex pathophysiological mechanisms of stroke.

Acute ischemia-reperfusion (IR)-induced brain injury is further exacerbated by a series of slower secondary pathogenic events, including delayed apoptosis due to neurotrophic factor deficiency. Neuritin, a neurotrophic factor regulating nervous system development and plasticity, is a potential therapeutic target for treatment of IR injury. In this study, Neuritin-overexpressing transgenic (Tg) mice were produced by pronuclear injection and offspring with high overexpression used to generate a line with stable inheritance for testing the neuroprotective capacity of Neuritin against transient global ischemia (TGI). Compared to wild-type mice, transgenic mice demonstrated reduced degradation of the DNA repair factor poly [ADP-ribose] polymerase 1 (PARP 1) in the hippocampus, indicating decreased hippocampal apoptosis rate, and a greater number of surviving hippocampal neurons during the first week post-TGI. In addition, Tg mice showed increased expression of the regeneration markers NF-200, synaptophysin, and GAP-43, and improved recovery of spatial learning and memory. Our findings exhibited that the window of opportunity of neural recovery in Neuritin transgenic mice group had a tendency to move ahead after TGI, which indicated that Neuritin can be used as a potential new therapeutic strategy for improving the outcome of cerebral ischemia injury.

Leishmania belongs to a genus of the protozoan parasites that causes leishmaniasis, and includes cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL). In this case, Leishmania amastigotes were found on cytomorphology examination of the bone marrow specimen, followed by 1,076 Leishmania donovani reads using metagenomic next generation sequencing (mNGS). Since being definitely diagnosed with VL/HIV coinfection, the patient was treated with liposomal amphotericin B as the parasite-resistant therapy and was discharged after clinical cure. But nearly a year later, on the mNGS follow-up, L. donovani was detected in the patient’s blood plasma specimen with 941 reads, suggesting that a relapse of leishmaniasis had occurred. These results indicate that leishmaniasis still exists in China and may represent a public health concern. This case could be helpful in the differential diagnosis of leishmaniasis, and for determining disease progression, prevention, and control of vectors and reservoir hosts.