To explore the difference of peripheral immune indexes especially T subsets cell mitochondrial indexes in patients with chronic HBV infection (CHB), liver cirrhosis (LC), hepatocellular carcinoma (HCC) and healthy controls (HCs). From May 2023 to December 2023, HBV infected patients aged 40–59 years (HBV infection group, 236 cases) and healthy people of the same age group (control group, 112 cases) were selected as the study objects. More than 80 parameters related to lymphocyte including cell percentage, absolute count, mitochondrial mass and mitochondrial low membrane potential were obtained by flow cytometry. The mitochondrial mass of memory T cells was found to be sequentially raised in the disease progression of hepatitis B patients from CHB to LC and eventually to HCC, and mitochondrial low membrane potential ratio was sequentially decreased. Then we screened out 4 main indicators related to the stage of disease progression in HBV-infected patients and distinguished the differences among the control group, CHB, LC, and HCC groups through the calculation of MPOLS (mitochondrial parameters of lymphocyte subsets) value, which can be used as a novel biomarker for hepatitis B disease progression. Mitochondrial parameters of peripheral lymphocytes in HBV-infected patients are significant indicators of different stages of disease progression. The MPOLS provides rich clinical data support for clinical early warning of high-risk hepatitis B patients, and we believe that the MPOLS can be combined with a variety of clinical characteristics to achieve more accurate diagnosis and treatment.

Background: The rapid increase in the number of monkeypox cases poses a considerable threat to the international community, necessitating sensitive, fast, and available diagnostic methods. Therefore, the objective of this study was to develop a rapid, sensitive and simple method with high clinical applicability. Methods: We developed a simple, rapid point-of-care assay to detect monkeypox virus (MPXV) using multienzyme isothermal rapid amplification (MIRA) coupled with the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas13a system. The detection system was optimized by synthesizing plasmids, and the detection sensitivity was explored by the continuous dilution of the plasmid. We validated the accuracy of this assay on 202 clinical MPXV samples and 104 interference samples through the kappa test. The visual interpretation of the results was realized by combining the assay with lateral flow strips. In addition, we developed a PCR-based method to identify MPXV Clades I and II, and the accuracy was tested through a kappa test on 202 clinical monkeypox samples and 104 interference samples. Results: Our assay achieved an analytical sensitivity of 14.4 copies/ml and high selectivity, as it differentiated MPXV from three other Orthopoxvirus species. The clinical testing results for 202 monkeypox samples and 104 interference samples demonstrated 100% sensitivity and specificity. Compared with quantitative PCR (qPCR), three samples tested as positive using our assay, which showed that the performance of this assay was superior to that of the qPCR assay. Combined with lateral flow strips, its availability and simplicity provide an alternative point-of-care diagnostic method for MPXV testing in remote settings and resource-poor areas. The results of 32 clinical samples showed that lateral flow strips had a high detection sensitivity and could identify samples with Ct value of 39 as positive. The clade identification assay detected as few as 200 copies/ml within 40 min and no cross-reaction was observed between Clades I and II. The clinical samples tested were all Clade II, which was consistent with the circulating clade in the Chinese mainland. Conclusions: The MIRA-CRISPR-Cas13a-MPXV system offers a rapid, sensitive and specific approach for monkeypox diagnosis, with significance for monitoring monkeypox epidemics. The clade identification assay based on PCR could accurately distinguish Clade I from Clade II within 40 min and can be implemented for high-throughput operation. Graphical

Background: Nontuberculous mycobacteria (NTM) are easily misdiagnosed as multidrug-resistant tuberculosis (MDR-TB), and treatment drugs are very limited. The main objective of our study was to evaluate the minimal inhibitory concentration (MIC) in vitro of bedaquiline (BDQ), clofazimine (CFZ), linezolid (LZD), delamanid (DLM), and pretomanid (PA-824) for treatment of M. abscessus and M. intracellulare. Furthermore, we determined whether MAB_1448, MAB_4384, MAB_2299c, MAB_1483, MAB_0540, rplD, rplC, and rrl were related to drug resistance to provide an experimental basis for the use of these five drugs in the treatment of NTM. Methods: We identified sample characteristics of epidemics in 550 patients with suspected NTM infection in Nanjing from 2019 to 2021 using the PCR-reverse spot hybrid method. Furthermore, we evaluated the MIC of BDQ, CFZ, DLM, LZD, and PA-824 against 155 clinical isolates of NTM using the microbroth dilution method. The resistant isolates were sequenced using Sanger sequencing. Results: The top three dominant species of NTM distributed in Nanjing were M. intracellulare, M. avium, and M. abscessus. Notably, the proportion of M. abscessus infections increased. The proportion of M. abscessus increased from 12% in 2019 to 18% in 2021. Demographic analysis showed that female infection rates were substantialy greater than male for M. abscessus (P=0.017, < 0.05). Our results demonstrate that NTM are highly sensitive to bedaquiline and clofazimine in vitro. However, delamanid and pretomanid had little effect on M. abscessus and M. intracellulare. In addition, we found 30– 41 nucleotide deletion mutations and some novel point mutations in the MAB_0540 gene of M. abscessus that are resistant to clofazimine. Conclusion: Bedaquiline, clofazimine, and linezolid were more successful in vitro treatments against M. abscessus and M. intracellulare. The MAB_0540 mutation may be associated with resistance of M. abscessus to clofazimine.