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Human enteroviruses (HEVs) include many different types that cause a wide range of diseases, and an effective method of genus-level identification has therefore significant clinical implications. However, quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), the gold-standard method, still has shortfalls in diagnostic sensitivity and timeliness. Here we established a one-step real-time reverse-transcription recombinase-aided PCR assay (RT-RAP) to detect HEV fragment within an hour. The RT-RAP assay showed a detection limit of 5 copies/μL using recombinant plasmids and was extensively verified using 15 HEV strains. Among 15 types of HEV (species A-C), the sensitivity of RT-RAP was approximately 2–8 folds lower than that of the qRT-PCR in 9 types, and no-cross reaction with other viruses was observed. RT-RAP was further applied to analyze CSF and fecal specimens; the clinical performance demonstrated that the RT-RAP and the commercial qRT-PCR kit provided consistent results. These results indicated that RT-RAP assay may be a promising approach for rapid and sensitive detection of HEV.

Abstract Background The newly discovered severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and four seasonal human coronaviruses (HCoVs) (HCoV-229E, HCoV-OC43, HCoV-NL63 and HCoV-HKU1) still circulate worldwide. The early clinical symptoms of SARS-CoV-2 and seasonal HCoV infections are similar, so rapid and accurate identification of the subtypes of HCoVs is crucial for early diagnosis, early treatment, prevention and control of these infections. However, current multiplex molecular diagnostic techniques for HCoV subtypes including SARS-CoV-2 are limited. Methods We designed primers and probes specific for the S and N genes of SARS-CoV-2, the N gene of severe acute respiratory syndrome coronavirus (SARS-CoV), and the ORF1ab gene of four seasonal HCoVs, as well as the human B2M gene product. We developed and optimized a quadruple quantitative real-time PCR assay (qq-PCR) for simultaneous detection of SARS-CoV-2, SARS-CoV and four seasonal HCoVs. This assay was further tested for specificity and sensitivity, and validated using 184 clinical samples. Results The limit of detection of the qq-PCR assay was in the range 2.5 × 101 to 6.5 × 101 copies/μL for each gene and no cross-reactivity with other common respiratory viruses was observed. The intra-assay and inter-assay coefficients of variation were 0.5–2%. The qq-PCR assay had a 91.9% sensitivity and 100.0% specificity for SARS-CoV-2 and a 95.7% sensitivity and 100% specificity for seasonal HCoVs, using the approved commercial kits as the reference. Compared to the commercial kits, total detection consistency was 98.4% (181/184) for SARS-CoV-2 and 98.6% (142/144) for seasonal HCoVs. Conclusion With the advantages of sensitivity, specificity, rapid detection, cost-effectiveness, and convenience, this qq-PCR assay has potential for clinical use for rapid discrimination between SARS-CoV-2, SARS-CoV and seasonal HCoVs.

Human immunodeficiency virus 1 (HIV-1) attacks the immune system, making people susceptible to various diseases, thus increasing their risk of death. Comprehensive detection of major HIV-1 strains circulating in China is vital for effective HIV-1 infection prevention and treatment. HIV-1 nucleic acid detection is considered effective for HIV-1 diagnosis since traditional immunological testing may fail to detect HIV-1 infection during the window period. This work demonstrates a one-pot two-stage amplification assay (RT-RAP), a combination of reverse transcription recombinase (RT- RAA), and quantitative real-time polymerase chain reaction (qRT-PCR). The turn-around time of the assay is only 50 min and can be performed with commonly available laboratory equipment, the qPCR devices. The RT-RAP assay could detect approximately 5 and 14 copies/reaction of HIV-1 DNA and RNA using recombinant plasmids and standard reference strains, respectively. Additionally, we found that the clinical performance of RT-RAP (detected 169 samples out of 170 specimens) was consistent with that of qRT-PCR. The sensitivity and specificity of RT-RAP were 100.00% (99/99) and 98.59% (70/71), respectively, while its positive and negative predictive values were 99.00% (99/100) and 100.00% (70/70), respectively. The total coincidence rate of the RT-RAP was 99.41% (169/170), with a kappa value of 0.988 (p < 0.05). We demonstrated that RT-RAP could rapidly detect the common HIV-1 subtypes commonly circulating in China with comparable sensitivity and specificity to qRT-PCR.

Introduction Exposure to the fine particulate matter PM2.5 is strongly associated with atherosclerotic diseases, creating considerable public concern. Nevertheless, the mechanisms have not been fully elucidated. We exposed atherosclerosis-prone apoE-deficient mice to PM2.5 to begin investigating these mechanisms. Material and methods Thirty-two 8-week-old male apoE –/– mice were divided to two groups fed with high-fat diet: a control group instilled with 0.9% saline, and an experimental group instilled with PM2.5 (30 mg/kg/day) for 8 weeks. We measured PM2.5 in whole blood by the ICP-MS method, and lipids and inflammatory factors by standard methods. The whole descending arteries were stained with oil red O; Aortic roots were stained with Movat, Sirius Red and immunohistochemical stains for pathological analysis; Brachiocephalic arteries for scanning electron microscopy, the descending arteries for Q-PCR. Echocardiography was used to evaluate cardiac function. Results In PM2.5 group, we observed elevated heavy metal components, consistent with higher amounts of platelets in total blood. The PM2.5 group also had elevated serum inflammatory factor levels. Finally, the PM2.5 group showed larger atherosclerotic plaques (p = 0.0231), higher numbers of lesion macrophages (p = 0.0183), greater injury to endothelial layers with greater adherence of platelets and leukocytes, elevated inflammatory factor levels, the NAD(P)H oxidase subunits p22phox and p47phox (p = 0.0079 and p = 0.0294), the M1/M2 associated markers IL-6, TNF-α (p = 0.0291, p = 0.0286), iNOS, IL-12 (p = 0.0122 and p = 0.0280) and arginase-1, and CD206 (p = 0.0216 and p = 0.0317). Conclusions PM2.5 exposure activated circulating leukocytes, platelets and associated inflammatory factors, contributing to the progression of atherosclerosis in apoE –/– mice.

Background. Currently, there are few studies on the effects of hyperlipidemia on semen parameters and serum hormones in men. In this study, we divided the study subjects into two groups of normal and hyperlipidemia according to the criteria, and observed the differences between semen parameters and serum reproductive hormones in hyperlipidaemic men and normal men, to explore the potential associations between the indicators. Materials and Methods. Eight hundred eighty five men attending infertility clinics in six hospitals from September 2016 to June 2017 were selected. Their lipid levels, semen parameters, and serum reproductive hormone levels were tested, and a total of 480 men with normal lipids and 405 men with hyperlipidemia were selected according to the criteria, and the relationship between semen quality, serum reproductive hormones, lipids and semen parameters, and serum hormones was statistically analyzed. Results. There was no significant difference in semen parameters between hyperlipidaemic men and normal men (P >0:05), serum testosterone levels were significantly lower in hyperlipidaemic men (P <0:05), and there was a negative correlation between triglycerides (TG) and testosterone in the blood (P <0:05).Conclusion. Hyperlipidemia does not affect male semen parameters, and changes in testosterone in hyperlipidaemic men may be related to triglycerides. [ABSTRACT FROM AUTHOR]

Introduction: Exposure to the fine particulate matter PM2.5 is strongly associated with atherosclerotic diseases, creating considerable public concern. Nevertheless, the mechanisms have not been fully elucidated. We exposed atherosclerosis-prone apoE-deficient mice to PM2.5 to begin investigating these mechanisms.Material and Methods: Thirty-two 8-week-old male apoE-/- mice were divided to two groups fed with high-fat diet: a control group instilled with 0.9% saline, and an experimental group instilled with PM2.5 (30 mg/kg/day) for 8 weeks. We measured PM2.5 in whole blood by the ICP-MS method, and lipids and inflammatory factors by standard methods. The whole descending arteries were stained with oil red O; Aortic roots were stained with Movat, Sirius Red and immunohistochemical stains for pathological analysis; Brachiocephalic arteries for scanning electron microscopy, the descending arteries for Q-PCR. Echocardiography was used to evaluate cardiac function.Results: In PM2.5 group, we observed elevated heavy metal components, consistent with higher amounts of platelets in total blood. The PM2.5 group also had elevated serum inflammatory factor levels. Finally, the PM2.5 group showed larger atherosclerotic plaques (p = 0.0231), higher numbers of lesion macrophages (p = 0.0183), greater injury to endothelial layers with greater adherence of platelets and leukocytes, elevated inflammatory factor levels, the NAD(P)H oxidase subunits p22phox and p47phox (p = 0.0079 and p = 0.0294), the M1/M2 associated markers IL-6, TNF-α (p = 0.0291, p = 0.0286), iNOS, IL-12 (p = 0.0122 and p = 0.0280) and arginase-1, and CD206 (p = 0.0216 and p = 0.0317).Conclusions: PM2.5 exposure activated circulating leukocytes, platelets and associated inflammatory factors, contributing to the progression of atherosclerosis in apoE-/- mice. [ABSTRACT FROM AUTHOR]

Objective • To evaluate the performance of matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in the identification of clinical pathogenic microorganisms. Methods • Blood culture-positive specimens were collected from inpatients in our hospital from March to December 2022 and identified using VITEK 2XL (biochemical), VITEK MS (colony), VITEK MS (bacterial membrane) and VITEK MS (separating gel) methods, respectively, to compare the compliance rate and identification values of the four methods. Results • A total of 280 strains were included in the analysis, including 155 (55.36%) Gram-negative and 125 (44.64%) Gram-positive strains. 279 (99.64%) of the 280 strains were identified by VITEK 2XL (biochemical), including 154 (99.35%) Gram-negative and 125 (100%) Gram-positive strains. VITEK MS (colony) identified 278 (99.29%) strains, including 153 (98.71%) Gram-negative and 125 (100%) Gram-positive. 261 (93.21%) strains were identified in VITEK MS (bacterial membrane), including 148 (95.48%) Gram-negative and 113 (90.40%) Grampositive strains. VITEK MS (separating gel) identified 232 (82.86%) strains, including 136 (87.74%) Gram-negative and 96 (76.80%) Gram-positive strains. Conclusion • MALDI-TOF MS findings are highly consistent with traditional culture identification methods in terms of identification accuracy, and the VITEK MS (bacterial membrane) and VITEK MS (separating gel) identification methods significantly reduce the turnaround time for identification in the laboratory. [ABSTRACT FROM AUTHOR]