Advancing MicroRNA Detection: Enhanced Biotin-Streptavidin Dual-Mode Phase Imaging Surface Plasmon Resonance Aptasensor.

MicroRNAs (miRNAs) are novel tumor biomarkers owing to their important physiological functions in cell communication and the progression of multiple diseases. Due to the small molecular weight, short sequence length, and low concentration levels of miRNA, miRNA detection presents substantial challenges, requiring the advancement of more refined and sensitive techniques. There is an urgent demand for the development of a rapid, user-friendly, and sensitive miRNA analysis method. Here, we developed an enhanced biotin-streptavidin dual-mode phase imaging surface plasmon resonance (PI-SPR) aptasensor for sensitive and rapid detection of miRNA. Initially, we evaluated the linear sensing range for miRNA detection across two distinct sensing modalities and investigated the physical factors that influence the sensing signal in the aptamer-miRNA interaction within the PI-SPR aptasensor. Then, an enhanced biotin-streptavidin amplification strategy was introduced in the PI-SPR aptasensor, which effectively reduced the nonspecific adsorption by 20% and improved the limit of detection by 548 times. Furthermore, we have produced three types of tumor marker chips, which utilize the rapid sensing mode (less than 2 min) of PI-SPR aptasensor to achieve simultaneous detection of multiple miRNA markers in the serum from clinical cancer patients. This work not only developed a new approach to detect miRNA in different application scenarios but also provided a new reference for the application of the biotin-streptavidin amplification system in the detection of other small biomolecules.

Cytoplasm Hydrogelation-Mediated Cardiomyocyte Sponge Alleviated Coxsackievirus B3 Infection.

Viral myocarditis (VMC), commonly caused by coxsackievirus B3 (CVB3) infection, lacks specific treatments and leads to serious heart conditions. Current treatments, such as IFNα and ribavirin, show limited effectiveness. Herein, rather than inhibiting virus replication, this study introduces a novel cardiomyocyte sponge, intracellular gelated cardiomyocytes (GCs), to trap and neutralize CVB3 via a receptor-ligand interaction, such as CAR and CD55. By maintaining cellular morphology, GCs serve as sponges for CVB3, inhibiting infection. In vitro results revealed that GCs could inhibit CVB3 infection on HeLa cells. In vivo, GCs exhibited a strong immune escape ability and effectively inhibited CVB3-induced viral myocarditis with a high safety profile. The most significant implication of this study is to develop a universal antivirus infection strategy via intracellular gelation of the host cell, which can be employed not only for treating defined pathogenic viruses but also for a rapid response to infection outbreaks caused by mutable and unknown viruses.

Removing Negative Impacts from Inevitable Nonreproducible and Nonspecific Antibody-Probe Interactions in Viral Serology.

Serological assays are indispensable tools in public health. Presently deployed serological assays, however, largely overlook research progress made in the last two decades that jeopardizes the conceptual foundation of these assays, i.e., antibody (Ab) specificity. Challenges to traditional understanding of Ab specificity include Ab polyspecificity and most recently nonreproducible Ab-probe interactions (NRIs). Here, using SARS-CoV-2 and four common livestock viruses as a test bed, we developed a new serological platform that integrates recent understanding about Ab specificity. We first demonstrate that the response rate (RR) from a large-sized serum pool (∼100) is not affected by NRIs or by nonspecific Ab-probe interactions (NSIs), so RR can be incorporated into the diagnostic probe selection process. We subsequently used multiple probes (configured as a "protein peptide hybrid microarray", PPHM) to generate a digital microarray index (DMI) and finally demonstrated that DMI-based analysis yields an extremely robust probabilistic trend that enables accurate diagnosis of viral infection that overcomes multiple negative impacts exerted by NSI/NRI. Thus, our study with SARS-CoV-2 confirms that the PPHM-RR-DMI platform enables very rapid development of serological assays that outperform traditional assays (for both sensitivity and specificity) and supports that the platform is extendable to other viruses.

DDX17-regulated alternative splicing that produced an oncogenic isoform of PXN-AS1 to promote HCC metastasis.

BACKGROUND AND AIMS: The mechanism underlying HCC metastasis remains unclear, many oncogenes are known to regulate this process. However, the role of alternative splicing (AS) in pro-metastatic HCC is poorly understood. APPROACH AND RESULTS: By performing RNA sequencing on nine pairs of primary HCC tissues with extrahepatic metastasis (EHMH) and nine pairs of metastasis-free HCC (MFH) tissues, we depicted the AS landscape in HCC and found a higher frequency of AS events in EHMH compared with MFH. Moreover, 28 differentially expressed splicing regulators were identified in EHMH compared with MFH. Among these, DEAD-box RNA helicase 17 (DDX17) was significantly up-regulated in EHMH and was strongly associated with patient outcome. Functional studies indicated that DDX17 knockout inhibited the degradation of the extracellular matrix, and diminished the invasive ability of HCC cells. A significant reduction in lung metastasis induced by DDX17 deficiency was also demonstrated in a diethylnitrosamine-induced DDX17HKO mouse model. Mechanistically, high DDX17 induced intron 3 retention of PXN-AS1 and produced a transcript (termed PXN-AS1-IR3). The transcript PXN-AS1-IR3 acted as an important promoter of HCC metastasis by inducing MYC transcription activation via recruiting the complex of testis expressed 10 and p300 to the MYC enhancer region, which led to transcriptional activation of several metastasis-associated downstream genes. Finally, the PXN-AS1-IR3 level was significantly higher in serum and HCC tissues with extrahepatic metastasis. CONCLUSIONS: DDX17 and PXN-AS1-IR3 act as important metastatic promoters by modulating MYC signaling, suggesting that DDX17 and PXN-AS1-IR3 may be potential prognostic markers for metastatic HCC.

Peptide aptamer-based time-resolved fluoroimmunoassay for CHIKV diagnosis.

BACKGROUND: Chikungunya virus (CHIKV) and Dengue virus (DENV) have similar clinical symptoms, which often induce misdiagnoses. Therefore, an antigen detection diagnostic system that can clearly identify these two viruses is desirable. METHODS: In this study, we developed a novel peptide with high affinity and specificity to CHIKV, and further constructed peptide aptamer-based TRFIA assay to efficiently detect CHIKV. Peptide aptamer B2 (ITPQSSTTEAEL) and B3 (DTQGSNWI) were obtained through computer-aided design and selected as CHIKV-specific peptide aptamers based on their high binding affinity, strong hydrogen bonding, and RMSD of molecular docking. Then, a sandwich-Time-Resolved Fluoroimmunoassay (TRFIA) was successfully constructed for the detection of the interaction between peptide aptamers and viruses. RESULTS: When using B2 as the detection element, highly specific detection of CHIKV E2 was achieved with detection limits of 8.5 ng/ml in PBS solution. Variation coefficient between inter-assay showed the disturbances received from the detection of clinical fluid specimens (including serum and urine), were also within acceptable limits. The detection limits for 10-fold dilution serum and urine were 57.8 ng/mL and 147.3 ng/mL, respectively. The fluorescent signal intensity exhibited a good linear correlation with E2 protein concentration in the range of 0-1000 ng/mL, indicating the potential for quantitative detection of E2 protein. CONCLUSIONS: These results demonstrate that the construction of peptide aptamers with high affinity and specificity provides an excellent method for rapid diagnostic element screening, and the developed peptide aptamer B2 contributed to better detection of CHIKV viral particles compared to traditional antibodies.

Antibacterial Activity and Mechanism of Canagliflozin against Methicillin-Resistant Staphylococcus aureus.

Diabetic foot infection (DFI) is a common complication in diabetes patients, with foot infections being the leading cause of amputations. Staphylococcus aureus is frequently found in diabetic foot infections, of which methicillin-resistant Staphylococcus aureus (MRSA) has become a major clinical and epidemiological challenge. Since MRSA strains are resistant to most ß-lactam antibiotics, and also partially resistant to other antibiotics, treatment is difficult and costly. The emergence of drug-resistant bacteria often arises from overuse or misuse of antibiotics. Clinically, canagliflozin is commonly used for the treatment of type 2 diabetes. On this basis, we investigated the antibacterial activity and mechanism of canagliflozin against MRSA, with the aim to discover novel functions of canagliflozin and provide new insights for the treatment of MRSA. Using the microbroth dilution method to determine the half maximal inhibitory concentration of drugs, we found that canagliflozin not only can inhibit the growth of methicillin-sensitive Staphylococcus aureus (MSSA) but also exhibits antibacterial activity against MRSA. The IC50 values, at approximately 56.01 µM and 57.60 µM, were almost the same. At 12 h, canagliflozin showed a significant antibacterial effect against MRSA at and above 30 µM. In addition, its combined use with penicillin achieved better antibacterial effects, which were increased by about three times. Additive antibacterial activity (FICI = 0.69) was found between penicillin and canagliflozin, which was better than that of doxycycline and canagliflozin (FICI = 0.95). Canagliflozin also affected bacterial metabolic markers, such as glucose, ATP, and lactic acid. The results of crystal violet staining indicate that canagliflozin disrupted the formation of bacterial biofilm. Our electron microscopy results showed that canagliflozin distorted the bacterial cell wall. The results of RT-PCR suggest that canagliflozin down-regulated the expressions of biofilm-related gene (clfA, cna, agrC, mgrA, hld) and methicillin-resistance gene (mecA), which was related to MRSA. Molecular docking also indicated that canagliflozin affected some interesting targets of MRSA, such as the sarA, crtM and fnbA proteins. In conclusion, canagliflozin exhibits antibacterial activity against MRSA by affecting bacterial metabolism, inhibiting its biofilm formation, distorting the bacterial cell wall, and altering the gene expression of biofilm formation and its virulence. Our study reveals the antibacterial activity of canagliflozin against MRSA, providing a new reference for treating diabetic foot infections.

Antibody responses to SARS-CoV-2 in healthy individuals returning to Shenzhen.

To verify reliability of antibody detection and investigate population immunity to severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) in the local Chinese population. A cross-sectional study was conducted in Shenzhen to detect anti-coronavirus antibodies including, immunoglobulin G (IgG), immunoglobulin M (IgM), and immunoglobulin A (IgA). In the COVID-19 group, nine patients were enrolled after diagnosis. In the control group, 1589 individuals without clinical symptoms (cough, fever, and fatigue) and returning from outside Shenzhen were enrolled. The first study enrollment occurred at the end of February 2020; the final study visit was 18 March 2020. In the COVID-19 group, the seven of nine patients were positive for IgM, IgG, and IgA. Meanwhile, six of the 1589 healthy individuals were found to be weakly positive for IgG. According to SARS-CoV-2 nucleic acid tests, the six individuals were all negative. Strong supplemental support for clinical information can be provided by antibody detection, especially for IgA. According to comparison with overseas reports, the infection rate of the Chinese population outside Shenzhen, China, is significantly low, so most of the population in China is still susceptible. Hence, social distancing measures are still inevitable until a vaccine is developed successfully.

Comparative analysis of carbapenemases, RND family efflux pumps and biofilm formation potential among Acinetobacter baumannii strains with different carbapenem susceptibility.

AIM: This study has conducted a comparative analysis of common carbapenemases harboring, the expression of resistance-nodulation-cell division (RND) family efflux pumps, and biofilm formation potential associated with carbapenem resistance among Acinetobacter baumannii (A. baumannii) strains with different carbapenem susceptibility. METHODS: A total of 90 isolates of A. baumannii from two tertiary hospitals of China were identified and grouped as carbapenem susceptible A. baumannii (CSAB) strains and carbapenem non-susceptible A. baumannii (CnSAB) strains based on the susceptibility to imipenem. Harboring of carbapenemase genes, relative expression of RND family efflux pumps and biofilm formation potential were compared between the two groups. RESULT: Among these strains, 12 (13.3 %) strains were divided into the CSAB group, and 78 (86.7 %) strains into the CnSAB group. Compared with CSAB strains, CnSAB strains increased distribution of blaOXA-23 (p < 0.001) and ISAba1/blaOXA-51-like (p = 0.034) carbapenemase genes, and a 6.1-fold relative expression of adeB (p = 0.002), while CSAB strains led to biofilm formation by 1.3-fold than CnSAB strains (p = 0.021). CONCLUSIONS: Clinically, harboring more blaOXA-23-like and ISAba1/blaOXA-51-like complex genes and overproduction of adeABC are relevant with carbapenem resistance, while carbapenem susceptible strains might survive the stress of antibiotic through their ability of higher biofilm formation.

Analysis of Differentially Expressed Novel MicroRNAs as Potential Biomarkers in Dengue Virus Type 1 Infection.

BACKGROUND: Host-derived miRNAs are reported to play diverse roles in dengue virus (DENV) infection, but DENV-derived miRNAs have been rarely studied. METHODS: Here, we used serum samples from three patients infected with dengue virus type 1 (DENV1) and three healthy volunteers to detect and profile novel microRNAs in dengue virus-infected human serum. MicroRNAs in serum samples were sequenced using an Illumina HiSeq 2000 system, and clean reads were trimmed, aligned, normalized, and analyzed to identify differentially expressed and novel microRNAs. Four microRNAs were selected as DENV1 infection biomarkers and verified through 1:2 paired case-control study, using serum from 15 DENV1-infected patients and 30 healthy volunteers. RESULTS: We identified 182 potential novel miRNAs, with 20 novel miRNAs found to have miRDeep2 score ≥ 4.0. Fifty-eight known and 11 novel miRNAs were upregulated at least 2-fold in DENV1-infected serum. Twenty-two known and 4 novel miRNAs were downregulated at least 0.5-fold. The AUCs of four selected miRNAs, hsa-miR-106b-3p, hsa-miR-122-5p, hsa-miR-novel-chr17_35150, and hsa-miR-novel-chr10_24390, were respectively: 0.962 (95% CI: 0.913 - 1.000, p > 0.05), 0.924 (95% CI: 0.851 - 0.998, p < 0.05), 0.941 (95% CI:0.877 - 1.000, p > 0.05), and 0.991 (95% CI: 0.973 - 1.000, p > 0.05). CONCLUSIONS: In summary, our study demonstrates that serum miRNA levels are affected by DENV1 infection, identifies novel DENV1-associated miRNAs, and suggests that Hsa-miR-122-5p may be a potential biomarker for DENV1 infection.

Tumor-infiltrating plasma cells are the promising prognosis marker for esophageal squamous cell carcinoma.

BACKGROUND: There is an urgent need to improve the clinical and basic research of esophageal cancer. The purpose of this study was to explore the prognostic value of tumor-infiltrating plasma cells (TIP) on overall survival (OS) of patients with esophageal squamous cell carcinoma (ESCC). METHODS: Three independent cohorts, which include 116 consecutive cases who received radical resection of ESCC in our institution (set to be discovery set), 179 cases from public GEO database (validation GEO set) and 95 cases from TCGA (validation TCGA set), with a total of 390 cases were retrospectively enrolled in this study. RESULTS: TIP was detected by immunohistochemical staining of CD138 in the paraffin-embedded specimen after surgery in the discovery set and was validated by using an established computational algorithm in the GEO and TCGA sets. Kaplan-Meier survival analysis showed high TIP was coincidently and significantly associated with favorable OS of ESCC in discovery set (p = 0.004) and validation GEO set (p = 0.002), showed a trend of better survival in validation TCGA set (p = 0.256 for 5-year OS, p = 0.034 for 15-month OS). Univariate and multivariate Cox regression analysis, together with survival analysis of the interaction between TIP and other variables, confirmed TIP to be a significant and independent prognostic factor for OS of ESCC. The incorporation of TIP into the TNM staging system could improve the accuracy of prognosis prediction for ESCC. CONCLUSION: This study revealed that high TIP in ESCC was associated with positive regulation of adaptive immunity and anti-tumor activity.

CircMTO1 inhibits liver fibrosis via regulation of miR-17-5p and Smad7.

Circular RNAs (circRNAs), often dysregulated in a variety of human diseases, participate in the initiation and development of cancers. Recently, circMTO1 (a circRNA derived from MTO1 gene), identified as a tumor suppressor, has been shown to contribute to the suppression of hepatocellular carcinoma. The present study aimed to explore the clinical significance and roles of circMTO1 in liver fibrosis. Here, we found that serum circMTO1 was significantly down-regulated in chronic hepatitis B (CHB) patients. Interestingly, serum circMTO1 was negatively correlated with fibrosis stages as well as HAI scores. Receiver operating characteristic curve analysis revealed that serum circMTO1 may serve as a diagnostic biomarker for liver fibrosis in CHB patients. Notably, overexpression of circMTO1 led to the suppression of transforming growth factor-ß1-induced hepatic stellate cells (HSCs) activation. Bioinformatic analysis and luciferase activity assays indicated that circMTO1 was a target of mircoRNA-17-5p (miR-17-5p). Data from RNA pull-down assay further confirmed that circMTO1 interacted with miR-17-5p. The inhibitory effects of circMTO1 on HSC activation were suppressed by miR-17-5p mimics. Further studies showed that Smad7 was a target of miR-17-5p. Moreover, circMTO1-inhibited HSC activation was also blocked down by loss of Smad7. Taken together, we demonstrate that circMTO1 inhibits liver fibrosis via regulation of miR-17-5p and Smad7, and serum circMTO1 may be a novel promising biomarker of liver fibrosis.

Development of peptide-based chemiluminescence enzyme immunoassay (CLEIA) for diagnosis of dengue virus infection in human.

Dengue is the most prevalent mosquito-borne viral disease in tropical and subtropical regions worldwide. Since its clinical symptoms are non-specific and easily mistaken as other kinds of infection, laboratory diagnosis is required to confirm dengue infections. In this study, ten peptides (E1-E10) from the envelope protein of dengue virus (DENV) were first identified using bioinformatic tool. The screened peptides were then synthesized for the peptide-based chemiluminescence enzyme immunoassay (CLEIA). Two peptides, E1 and E7, were found as the best candidate antigen and therefore used as downstream application in the development of low-cost peptide-based anti-DENV immunoglobulin M antibodies (IgM) indirect CLEIA. 176 serum samples were used to study the presence of anti-DENV IgM antibodies to evaluate the diagnostic ability of IgM-CLEIA. Receiver operating characteristic curve (ROC) was used to estimate the diagnostic cut-off value. The sensitivity and the specificity reached 82.5% and 94.6% respectively when peptide E1 was used, but declined to 79.2% and 92.9% respectively when peptide E7 was used. Therefore, the combination of E1 and E7 was used to improve the sensitivity and the specificity to 85.0% and 96.4% respectively in 1.5 h assay time, providing a potentially practical use for the diagnosis of DENV infections in patients' serum.

Development of a Strategy Based on the Surface Plasmon Resonance Technology for Platelet Compatibility Testing.

BACKGROUND: This study was aimed to establish a novel strategy based on the surface plasmon resonance (SPR) technology for platelet compatibility testing. METHODS: A novel surface matrix was prepared based on poly (OEGMA-co-HEMA) via surface-initiated polymerization as a biosensor surface platform. Type O universal platelets and donor platelets were immobilized on these novel matrices via amine-coupling reaction and worked as a capturing ligand for binding the platelet antibody. Antibodies binding to platelets were monitored in real time by injecting the samples into a microfluidic channel. Clinical serum samples (n = 186) with multiple platelet transfusions were assayed for platelet antibodies using the SPR technology and monoclonal antibody-immobilized platelet antigen (MAIPA) assay. RESULTS: The novel biosensor surface achieved nonfouling background and high immobilization capacity and showed good repeatability and stability after regeneration. The limit of detection of the SPR biosensor for platelet antibody was estimated to be 50 ng/mL. The sensitivity and specificity were 92% and 98.7%. It could detect the platelet antibody directly in serum samples, and the results were similar to MAIPA assay. CONCLUSIONS: A novel strategy to facilitate the sensitive and reliable detection of platelet compatibility for developing an SPR-based biosensor was established in this study. The SPR-based biosensor combined with novel surface chemistry is a promising method for platelet compatibility testing.

Human Infection with Burkholderia thailandensis, China, 2013.

Burkholderia thailandensis infection in humans is uncommon. We describe a case of B. thailandensis infection in a person in China, a location heretofore unknown for B. thailandensis. We identified the specific virulence factors of B. thailandensis, which may indicate a transition to a new virulent form.

Prevalence and genetic diversity analysis of human coronaviruses among cross-border children.

BACKGROUND: More than a decade after the outbreak of human coronaviruses (HCoVs) SARS in Guangdong province and Hong Kong SAR of China in 2002, there is still no reoccurrence, but the evolution and recombination of the coronaviruses in this region are still unknown. Therefore, surveillance on the prevalence and the virus variation of HCoVs circulation in this region is conducted. METHODS: A total of 3298 nasopharyngeal swabs samples were collected from cross-border children (<6 years, crossing border between Southern China and Hong Kong SAR) showing symptoms of respiratory tract infection, such as fever (body temperature > 37.5 °C), from 2014 May to 2015 Dec. Viral nucleic acids were analyzed and sequenced to study the prevalence and genetic diversity of the four human coronaviruses. The statistical significance of the data was evaluated with Fisher chi-square test. RESULTS: 78 (2.37%; 95%CI 1.8-2.8%) out of 3298 nasopharyngeal swabs specimens were found to be positive for OC43 (36;1.09%), HKU1 (34; 1.03%), NL63 (6; 0.18%) and 229E (2;0.01%). None of SARS or MERS was detected. The HCoVs predominant circulating season was in transition of winter to spring, especially January and February and NL63 detected only in summer and fall. Complex population with an abundant genetic diversity of coronaviruses was circulating and they shared homology with the published strains (99-100%). Besides, phylogenetic evolutionary analysis indicated that OC43 coronaviruses were clustered into three clades (B,D,E), HKU1 clustered into two clades(A,B) and NL63 clustered into two clades(A,B). Moreover, several novel mutations including nucleotides substitution and the insertion of spike of the glycoprotein on the viral surface were discovered. CONCLUSIONS: The detection rate and epidemic trend of coronaviruses were stable and no obvious fluctuations were found. The detected coronaviruses shared a conserved gene sequences in S and RdRp. However, mutants of the epidemic strains were detected, suggesting continuous monitoring of the human coronaviruses is in need among cross-border children, who are more likely to get infected and transmit the viruses across the border easily, in addition to the general public.

Wavelength-Scanning SPR Imaging Sensors Based on an Acousto-Optic Tunable Filter and a White Light Laser.

A fast surface plasmon resonance (SPR) imaging biosensor system based on wavelength interrogation using an acousto-optic tunable filter (AOTF) and a white light laser is presented. The system combines the merits of a wide-dynamic detection range and high sensitivity offered by the spectral approach with multiplexed high-throughput data collection and a two-dimensional (2D) biosensor array. The key feature is the use of AOTF to realize wavelength scan from a white laser source and thus to achieve fast tracking of the SPR dip movement caused by target molecules binding to the sensor surface. Experimental results show that the system is capable of completing a SPR dip measurement within 0.35 s. To the best of our knowledge, this is the fastest time ever reported in the literature for imaging spectral interrogation. Based on a spectral window with a width of approximately 100 nm, a dynamic detection range and resolution of 4.63 × 10-2 refractive index unit (RIU) and 1.27 × 10-6 RIU achieved in a 2D-array sensor is reported here. The spectral SPR imaging sensor scheme has the capability of performing fast high-throughput detection of biomolecular interactions from 2D sensor arrays. The design has no mechanical moving parts, thus making the scheme completely solid-state.

High-throughput imaging surface plasmon resonance biosensing based on an adaptive spectral-dip tracking scheme.

Imaging-based spectral surface plasmon resonance (λSPR) biosensing is predominantly limited by data throughput because of the multiplied data capacity emerging from 2-dimensional sensor array sites and the many data points required to produce an accurate measurement of the absorption dip. Here we present an adaptive feedback approach to address the data throughput issue in λSPR biosensing. A feedback loop constantly tracks the dip location while target-molecule binding occurs at the sensor surface. An adaptive window is then imposed to reduce the number of data points that each pixel has to capture without compromising measurement accuracy. Rapid wavelength scanning is performed with a liquid crystal tunable filter (LCTF). With the use of a feedback loop, our demonstration system can produce a dip measurement within 700ms, thus confirming that the reported λSPR approach is most suitable for real-time micro-array label-free biosensing applications.

Dysregulated Serum MiRNA Profile and Promising Biomarkers in Dengue-infected Patients.

OBJECTIVES: Pathological biomarkers and mechanisms of dengue infection are poorly understood. We investigated a new serum biomarker using miRNAs and performed further correlation analysis in dengue-infected patients. METHODS: Expression levels of broad-spectrum miRNAs in serum samples from three patients with dengue virus type 1 (DENV-1) and three healthy volunteers were separately analyzed using miRNA PCR arrays. The expressions of the five selected miRNAs were verified by qRT-PCR in the sera of 40 DENV-1 patients and compared with those from 32 healthy controls. Receiver operating characteristic (ROC) curve and correlation analyses were performed to evaluate the potential of these miRNAs for the diagnosis of dengue infection. RESULTS: MiRNA PCR arrays revealed that 41 miRNAs were upregulated, whereas 12 miRNAs were down-regulated in the sera of DENV-1 patients compared with those in healthy controls. Among these miRNAs, qRT-PCR validation showed that serum hsa-miR-21-5p, hsa-miR-590-5p, hsa-miR-188-5p, and hsa-miR-152-3p were upregulated, whereas hsa-miR-146a-5p was down-regulated in dengue-infected patients compared with healthy controls. ROC curves showed serum hsa-miR-21-5p and hsa-miR-146a-5p could distinguish dengue-infected patients with preferable sensitivity and specificity. Correlation analysis indicated that expression levels of serum hsa-miR-21-5p and hsa-miR-146a-5p were negative and positively correlated with the number of white blood cells and neutrophils, respectively. Functional analysis of target proteins of these miRNAs in silico indicated their involvement in inflammation and cell proliferation. CONCLUSION: Dengue-infected patients have a broad "fingerprint" profile with dysregulated serum miRNAs. Among these miRNAs, serum hsa-miR-21-5p, hsa-miR-146a-5p, hsa-miR-590-5p, hsa-miR-188-5p, and hsa-miR-152-3p were identified as promising serum indicators for dengue infection.

Identification of microRNAs in Throat Swab as the Biomarkers for Diagnosis of Influenza.

BACKGROUND: Influenza is a serious worldwide disease that captures global attention in the past few years after outbreaks. The recent discoveries of microRNA (miRNA) and its unique expression profile in influenza patients have offered a new method for early influenza diagnosis. The aim of this study was to examine the utility of miRNAs for the diagnosis of influenza. METHODS: Thirteen selected miRNAs were investigated with the hosts' throat swabs (25 H1N1, 20 H3N2, 20 influenza B and 21 healthy controls) by real-time quantitative polymerase chain reaction (RT-qPCR) using U6 snRNA as endogenous control for normalization, and receiver operating characteristic (ROC) curve/Area under curve (AUC) for analysis. RESULTS: miR-29a-3p, miR-30c-5p, miR-34c-3p and miR-181a-5p are useful biomarkers for influenza A detection; and miR-30c-5p, miR-34b-5p, miR-205-5p and miR-449b-5p for influenza B detection. Also, use of both miR-30c-5p and miR-34c-3p (AUC=0.879); and miR-30c-5p and miR-449b-5p (AUC=0.901) are better than using one miRNA to confirm influenza A and influenza B infection, respectively. CONCLUSIONS: Given its simplicity, non-invasiveness and specificity, we found that the throat swab-derived miRNAs miR-29a-3p, miR-30c-5p, miR-34b-5p, miR-34c-3p, miR-181a-5p, miR-205-5p and miR-449b-5p are a useful tool for influenza diagnosis on influenza A and B.

Dysregulated Serum MicroRNA Expression Profile and Potential Biomarkers in Hepatitis C Virus-infected Patients.

OBJECTIVES: Circulating microRNAs (miRNAs) play critical roles in pathogen-host interactions. Aberrant miRNA expression profiles might have specific characteristics for virus strains, and could serve as noninvasive biomarkers for screening and diagnosing infectious diseases. In this study, we aimed to find new potential miRNA biomarkers of hepatitis C virus (HCV) infection. METHODS: Expression levels of broad-spectrum miRNAs in serum samples from 10 patients with HCV viremia and 10 healthy volunteers were analyzed using miRNA PCR arrays. Subsequently, the differential expression of four selected miRNAs (miR-122, miR-134, miR-424-3p, and miR-629-5p) was verified by qRT-PCR in the serum of 39 patients compared with that in 29 healthy controls. Receiver operating characteristic (ROC) curve analysis was performed to evaluate their potential for the diagnosis of HCV infection. RESULTS: miRNA PCR array assays revealed differential expression of 106 miRNAs in sera of HCV patients compared with that in healthy controls. Serum hsa-miR-122, miR-134, miR-424-3p, and miR-629-5p were well identified. The ROC curves showed that miR-122, miR-134, miR-424-3p, and miR-629-5p could distinguish HCV patients with preferable sensitivity and specificity. In addition, Correlation analysis indicated serum miR-122 expression was positive correlation with ALT/AST levels. Functional analysis of target proteins of these miRNAs indicated the involvement of viral replication, inflammation, and cell proliferation. CONCLUSION: HCV patients have a broad 'fingerprint' profile with dysregulated serum miRNAs compared with that in healthy controls. Among these, serum hsa-miR-122, miR-134, miR-424-3p, and miR-629-5p are identified as promising indication factors of the serum miRNA profile of HCV infection. Particularly, miR-122 could be one of serum biomarkers for early pathological process of HCV. However, more miRNA biomarkers and biological functions of these miRNAs require further investigation.