-
Journal of Diabetes Investigation Jun 2024We investigated the relationship of circulating TSP-1 mRNA and miR-194 with diabetic kidney disease's degree.
Relationship between circulating thrombospondin-1 messenger ribonucleic acid and microribonucleic acid-194 levels in Chinese patients with type 2 diabetic kidney disease: The outcomes of a case-control study.
AIMS/INTRODUCTION
We investigated the relationship of circulating TSP-1 mRNA and miR-194 with diabetic kidney disease's degree.
MATERIALS AND METHODS
We enrolled 167 hospitalized type 2 diabetes patients in the endocrinology department. Patients were split into three groups according to urinary microalbumin: A, B and C. The control group comprised healthy outpatients (n = 163). The quantities of microribonucleic acid (miR)-194 and thrombospondin-1 (TSP-1) messenger ribonucleic acid (mRNA) in the participants' circulation were measured using a quantitative real-time polymerase chain reaction.
RESULTS
Circulating TSP-1 mRNA (P = 0.024) and miR-194 (P = 0.029) expressions significantly increased in type 2 diabetes patients. Circulating TSP-1 mRNA (P = 0.040) and miR-194 (P = 0.007) expression levels differed significantly among the three groups; circulating TSP-1 mRNA expression increased with urinary microalbumin. However, miR-194 declined in group B and increased in group C. Circulating TSP-1 mRNA was positively correlated with cystatin-c (r = 0.281; P = 0.021) and microalbumin/creatinine ratio (UmALB/Cr; r = 0.317; P = 0.009); miR-194 was positively correlated with UmALB/Cr (r = 0.405; P = 0.003). Stepwise multivariate linear regression analysis showed cystatin-c (β = 0.578; P = 0.021) and UmALB/Cr (β = 0.001; P = 0.009) as independent factors for TSP-1 mRNA; UmALB/Cr (β = 0.005; P = 0.028) as an independent factor for miR194. Areas under the curve for circulating TSP-1 mRNA and miR194 were 0.756 (95% confidence interval 0.620-0.893; sensitivity 0.69 and specificity 0.71, P < 0.01) and 0.584 (95% confidence interval 0.421-0.748; sensitivity 0.54 and specificity 0.52, P < 0.01), respectively.
CONCLUSIONS
Circulating TSP-1 mRNA and miR-194 expressions significantly increased in type 2 diabetes patients. The microalbumin group had lower levels of miR-194 (a risk factor that is valuable for type 2 diabetes kidney disease evaluation).
PubMed: 38932465
DOI: 10.1111/jdi.14252 -
Viruses Jun 2024Quantification of Torquetenovirus (TTV) viremia is becoming important for evaluating the status of the immune system in solid organ transplant recipients, monitoring the...
Quantification of Torquetenovirus (TTV) viremia is becoming important for evaluating the status of the immune system in solid organ transplant recipients, monitoring the appearance of post-transplant complications, and controlling the efficacy of maintenance immunosuppressive therapy. Thus, diagnostic approaches able to scale up TTV quantification are needed. Here, we report on the development and validation of a real-time PCR assay for TTV quantification on the Hologic Panther Fusion System by utilizing its open-access channel. The manual real-time PCR previously developed in our laboratories was optimized to detect TTV DNA on the Hologic Panther Fusion System. The assay was validated using clinical samples. The automated TTV assay has a limit of detection of 1.6 log copies per ml of serum. Using 112 samples previously tested via manual real-time PCR, the concordance in TTV detection was 93% between the assays. When the TTV levels were compared, the overall agreement between the methods, as assessed using Passing-Bablok linear regression and Bland-Altman analyses, was excellent. In summary, we validated a highly sensitive and accurate method for the diagnostic use of TTV quantification on a fully automated Hologic Panther Fusion System. This will greatly improve the turnaround time for TTV testing and better support the laboratory diagnosis of this new viral biomarker.
Topics: Real-Time Polymerase Chain Reaction; Viremia; Humans; Viral Load; DNA Virus Infections; Sensitivity and Specificity; Torque teno virus; DNA, Viral; Limit of Detection; Reproducibility of Results; Automation, Laboratory
PubMed: 38932255
DOI: 10.3390/v16060963 -
Viruses Jun 2024Recently, a multiplex PCR-based titration (MPBT) assay was developed for simultaneous determination of infectious titers of all three Sabin strains of the oral...
Recently, a multiplex PCR-based titration (MPBT) assay was developed for simultaneous determination of infectious titers of all three Sabin strains of the oral poliovirus vaccine (OPV) to replace the conventional CCID assay, which is both time-consuming and laborious. The MPBT assay was shown to be reproducible, robust and sensitive. The conventional and MPBT assays showed similar results and sensitivity. The MPBT assay can be completed in two to three days, instead of ten days for the conventional assay. To prevent attenuated vaccine strains of poliovirus from reversion to virulence, a novel, genetically stable OPV (nOPV) was developed by modifying the genomes of conventional Sabin strains used in OPV. In this work, we evaluated the MPBT assay as a rapid screening tool to support trivalent nOPV (tnOPV) formulation development by simultaneous titration of the three nOPV strains to confirm stability as needed, for the selection of the lead tnOPV formulation candidate. We first assessed the ability of the MPBT assay to discriminate a 0.5 log titer difference by titrating the two tnOPV samples (undiluted and threefold-diluted) on the same plate. Once the assay was shown to be discriminating, we then tested different formulations of tnOPV drug products (DPs) that were subjected to different exposure times at 37 °C (untreated group and treated groups: 2 and 7 days at 37 °C), and to three freeze and thaw (FT) cycles. Final confirmation of the down selected formulation candidates was achieved by performing the conventional CCID assay, comparing the stability of untreated and treated groups and FT stability testing on the top three candidates. The results showed that the MPBT assay generates similar titers as the conventional assay. By testing two trivalent samples in the same plate, the assay can differentiate a 0.5 log difference between the titers of the tested nOPV samples. Also, the assay was able to detect the gradual degradation of nOPV viruses with different formulation compositions and under different time/temperature conditions and freeze/thaw cycles. We found that there were three tnOPV formulations which met the stability criteria of less than 0.5 log loss after 2 days' exposure to 37 ℃ and after three FT cycles, maintaining the potency of all three serotypes in these formulations. The ability of the MPBT assay to titrate two tnOPV lots (six viruses) in the same plate makes it cheaper and gives it a higher throughput for rapid screening. The assay detected the gradual degradation of the tnOPV and was successful in the selection of optimal formulations for the tnOPV. The results demonstrated that the MPBT method can be used as a stability indicating assay to assess the thermal stability of the nOPV. It can be used for rapid virus titer determination during the vaccine manufacturing process, and in clinical trials. The MPBT assay can be automated and applied for other viruses, including those with no cytopathic effect.
Topics: Poliovirus Vaccine, Oral; Poliovirus; Humans; Multiplex Polymerase Chain Reaction; Poliomyelitis; Vaccines, Attenuated; Reproducibility of Results; Sensitivity and Specificity
PubMed: 38932253
DOI: 10.3390/v16060961 -
Viruses Jun 2024This study aimed to determine the incidence and etiological, seasonal, and genetic characteristics of respiratory viral coinfections involving severe acute respiratory...
This study aimed to determine the incidence and etiological, seasonal, and genetic characteristics of respiratory viral coinfections involving severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Between October 2020 and January 2024, nasopharyngeal samples were collected from 2277 SARS-CoV-2-positive patients. Two multiplex approaches were used to detect and sequence SARS-CoV-2, influenza A/B viruses, and other seasonal respiratory viruses: multiplex real-time polymerase chain reaction (PCR) and multiplex next-generation sequencing. Coinfections of SARS-CoV-2 with other respiratory viruses were detected in 164 (7.2%) patients. The most common co-infecting virus was respiratory syncytial virus (RSV) (38 cases, 1.7%), followed by bocavirus (BoV) (1.2%) and rhinovirus (RV) (1.1%). Patients ≤ 16 years of age had the highest rate (15%) of mixed infections. Whole-genome sequencing produced 19 complete genomes of seasonal respiratory viral co-pathogens, which were subjected to phylogenetic and amino acid analyses. The detected influenza viruses were classified into the genetic groups 6B.1A.5a.2a and 6B.1A.5a.2a.1 for A(H1N1)pdm09, 3C.2a1b.2a.2a.1 and 3C.2a.2b for A(H3N2), and V1A.3a.2 for the B/Victoria lineage. The RSV-B sequences belonged to the genetic group GB5.0.5a, with HAdV-C belonging to type 1, BoV to genotype VP1, and PIV3 to lineage 1a(i). Multiple amino acid substitutions were identified, including at the antibody-binding sites. This study provides insights into respiratory viral coinfections involving SARS-CoV-2 and reinforces the importance of genetic characterization of co-pathogens in the development of therapeutic and preventive strategies.
Topics: Humans; Coinfection; SARS-CoV-2; COVID-19; Middle Aged; Adult; Female; Male; Adolescent; Phylogeny; Child, Preschool; Child; Aged; Young Adult; Infant; Respiratory Tract Infections; Rhinovirus; Influenza A virus; Respiratory Syncytial Virus, Human; Nasopharynx; Whole Genome Sequencing; China; Seasons; Aged, 80 and over; Genome, Viral; Influenza B virus
PubMed: 38932250
DOI: 10.3390/v16060958 -
Viruses May 2024Congenital cytomegalovirus (cCMV) infection poses significant risks to fetal development, particularly affecting the nervous system. This study reports a fetal autopsy...
Congenital cytomegalovirus (cCMV) infection poses significant risks to fetal development, particularly affecting the nervous system. This study reports a fetal autopsy case, examining cCMV infection and focusing on CMV DNA measurements in various fetal organs before formalin fixation, a novel approach for comprehensive CMV DNA evaluations in fetal organs affected by cCMV. A 20-week-old male fetus was diagnosed with cCMV following the detection of CMV DNA in ascites obtained via abdominocentesis in utero. After the termination of pregnancy, multiple organs of the fetus, including the cerebrum, thyroid gland, heart, lungs, liver, spleen, kidneys, and adrenal glands, were extracted and examined for CMV DNA loads using a real-time polymerase chain reaction. Histopathological examination involved hematoxylin-eosin and CMV-specific immunostaining. A correlation was found between CMV DNA loads and pathology, with higher CMV-infected cell numbers observed in organs positively identified with both staining methods, exhibiting CMV DNA levels of ≥1.0 × 10 copies/mL, compared to those detected solely by CMV-specific immunostaining, where CMV DNA levels ranged from 1.0 × 10 to 1.0 × 10 copies/mL. These results highlight a quantifiable relationship between the organ infection extent and CMV DNA concentration, providing insights into cCMV pathogenesis and potentially informing future diagnostic and therapeutic strategies for cCMV infection.
Topics: Cytomegalovirus Infections; Humans; Cytomegalovirus; DNA, Viral; Viral Load; Male; Female; Fetus; Pregnancy; Adult; Autopsy; Pregnancy Complications, Infectious
PubMed: 38932183
DOI: 10.3390/v16060891 -
Viruses May 2024We have been encouraging practicing gynecologists to adopt molecular diagnostics tests, PCR, and cancer biomarkers, as alternatives enabled by these platforms, to...
We have been encouraging practicing gynecologists to adopt molecular diagnostics tests, PCR, and cancer biomarkers, as alternatives enabled by these platforms, to traditional Papanicolaou and colposcopy tests, respectively. An aliquot of liquid-based cytology was used for the molecular test [high-risk HPV types, (HR HPV)], another for the PAP test, and one more for p16/Ki67 dual-stain cytology. A total of 4499 laboratory samples were evaluated, and we found that 25.1% of low-grade samples and 47.9% of high-grade samples after PAP testing had a negative HR HPV-PCR result. In those cases, reported as Pap-negative, 22.1% had a positive HR HPV-PCR result. Dual staining with p16/Ki67 biomarkers in samples was positive for HR HPV, and 31.7% were also positive for these markers. Out of the PCR results that were positive for any of these HR HPV subtypes, n 68.3%, we did not find evidence for the presence of cancerous cells, highlighting the importance of performing dual staining with p16/Ki67 after PCR to avoid unnecessary colposcopies. The encountered challenges are a deep-rooted social reluctance in Mexico to abandon traditional Pap smears and the opinion of many specialists. Therefore, we still believe that colposcopy continues to be a preferred procedure over the dual-staining protocol.
Topics: Humans; Female; Mexico; Uterine Cervical Neoplasms; Papillomavirus Infections; Molecular Diagnostic Techniques; Papanicolaou Test; Biomarkers, Tumor; Papillomaviridae; Cyclin-Dependent Kinase Inhibitor p16; Vaginal Smears; Colposcopy; Gynecology; Adult; Middle Aged; Ki-67 Antigen; Polymerase Chain Reaction; Early Detection of Cancer; Private Practice
PubMed: 38932179
DOI: 10.3390/v16060887 -
Viruses May 2024Alphabaculoviruses are lethal dsDNA viruses of Lepidoptera that have high genetic diversity and are transmitted in aggregates within proteinaceous occlusion bodies. This...
Alphabaculoviruses are lethal dsDNA viruses of Lepidoptera that have high genetic diversity and are transmitted in aggregates within proteinaceous occlusion bodies. This mode of transmission has implications for their efficacy as biological insecticides. A Nicaraguan isolate of Spodoptera frugiperda multiple nucleopolyhedrovirus (SfMNPV-NIC) comprising nine genotypic variants has been the subject of considerable study due to the influence of variant interactions on the insecticidal properties of mixed-variant occlusion bodies. As part of a systematic study on the replication and transmission of variant mixtures, a tool for the accurate quantification of a selection of genotypic variants was developed based on the quantitative PCR technique (qPCR). First, primer pairs were designed around a region of high variability in four variants named SfNic-A, SfNic-B, SfNic-C and SfNic-E to produce amplicons of 103-150 bp. Then, using cloned purified amplicons as standards, amplification was demonstrated over a dynamic range of 10-10 copies of each target. The assay was efficient (mean ± SD: 98.5 ± 0.8%), reproducible, as shown by low inter- and intra-assay coefficients of variation (<5%), and specific to the target variants (99.7-100% specificity across variants). The quantification method was validated on mixtures of genotype-specific amplicons and demonstrated accurate quantification. Finally, mixtures of the four variants were quantified based on mixtures of budded virions and mixtures of DNA extracted from occlusion-derived virions. In both cases, mixed-variant preparations compared favorably to total viral genome numbers by quantification of the () gene that is present in all variants. This technique should prove invaluable in elucidating the influence of variant diversity on the transmission and insecticidal characteristics of this pathogen.
Topics: Nucleopolyhedroviruses; Animals; Spodoptera; Genotype; Genetic Variation; Real-Time Polymerase Chain Reaction; DNA, Viral
PubMed: 38932173
DOI: 10.3390/v16060881 -
Viruses May 2024The presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater has been reported in several studies and similar research can be used as a...
Distribution of SARS-CoV-2 Genomes in Wastewaters and the Associated Potential Infection Risk for Plant Workers in Typical Urban and Peri-Urban Communities of the Buffalo City Region, South Africa.
The presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater has been reported in several studies and similar research can be used as a proxy for an early warning of potential Coronavirus disease 2019 (COVID-19) outbreaks. This study focused on profiling the incidence of SARS-CoV-2 genomes in wastewater samples obtained from facilities located in the Buffalo City Municipality. Raw samples were collected weekly using the grab technique for a period of 48 weeks. Ribonucleic acids were extracted from the samples, using the QIAGEN Powersoil Total RNA Extraction kit, and extracted RNA samples were further profiled for the presence of SARS-CoV-2 genomes using Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR) technique. Furthermore, various environmental matrices were utilized to estimate the potential health risk to plant operators associated with exposure to SARS-CoV-2 viral particles using the quantitative microbiological risk assessment (QMRA) model. Our findings revealed the prevalence of SARS-CoV-2 genomes with concentrations that ranged from 0.22 × 103 to 17.60 × 103 genome copies per milliliter (GC/mL). Different exposure scenarios were employed for the QMRA model, and the findings indicate a probability of infection (P(i)) ranging from 0.93% to 37.81% across the study sites. Similarly, the P(i) was highly significant ( < 0.001) for the 20 mL volumetric intake as compared to other volumetric intake scenarios, and high P(i) was also observed in spring, autumn, and winter for all WWTPs. The P(i) was significantly different ( < 0.05) with respect to the different seasons and with respect to different volume scenarios.
Topics: Wastewater; SARS-CoV-2; South Africa; COVID-19; Humans; Genome, Viral; Risk Assessment; RNA, Viral; Occupational Exposure; Cities
PubMed: 38932163
DOI: 10.3390/v16060871 -
Viruses May 2024In this study, we investigated the concentration of airborne influenza virus in daycare centers and influencing factors, such as common cold prevalence, air pollutants,...
In this study, we investigated the concentration of airborne influenza virus in daycare centers and influencing factors, such as common cold prevalence, air pollutants, and meteorological factors. A total of 209 air samples were collected from daycare centers in Kaohsiung and the influenza virus was analyzed using real-time quantitative polymerase chain reaction. Air pollutants and metrological factors were measured using real-time monitoring equipment. Winter had the highest positive rates of airborne influenza virus and the highest prevalence of the common cold, followed by summer and autumn. The concentration of CO was significantly positively correlated with airborne influenza virus. Daycare center A, with natural ventilation and air condition systems, had a higher concentration of airborne influenza A virus, airborne fungi, and airborne bacteria, as well as a higher prevalence of the common cold, than daycare center B, with a mechanical ventilation system and air purifiers, while the concentrations of CO, CO, and UFPs in daycare center A were lower than those in daycare center B. We successfully detected airborne influenza virus in daycare centers, demonstrating that aerosol sampling for influenza can provide novel epidemiological insights and inform the management of influenza in daycare centers.
Topics: Humans; Air Microbiology; Influenza, Human; Seasons; Child Day Care Centers; Influenza A virus; Orthomyxoviridae; Air Pollutants; Common Cold; Child, Preschool; Prevalence; Environmental Monitoring
PubMed: 38932115
DOI: 10.3390/v16060822 -
Viruses May 2024Climate change, unpredictable weather patterns, and droughts are depleting water resources in some parts of the globe, where recycling and reusing wastewater is a...
Climate change, unpredictable weather patterns, and droughts are depleting water resources in some parts of the globe, where recycling and reusing wastewater is a strategy for different purposes. To counteract this, the EU regulation for water reuse sets minimum requirements for the use of reclaimed water for agricultural irrigation, including a reduction in human enteric viruses. In the present study, the occurrence of several human enteric viruses, including the human norovirus genogroup I (HuNoV GI), HuNoV GII, and rotavirus (RV), along with viral fecal contamination indicator crAssphage was monitored by using (RT)-qPCR methods on influent wastewater and reclaimed water samples. Moreover, the level of somatic coliphages was also determined as a culturable viral indicator. To assess the potential viral infectivity, an optimization of a capsid integrity PMAxx-RT-qPCR method was performed on sewage samples. Somatic coliphages were present in 60% of the reclaimed water samples, indicating inefficient virus inactivation. Following PMAxx-RT-qPCR optimization, 66% of the samples tested positive for at least one of the analyzed enteric viruses, with concentrations ranging from 2.79 to 7.30 Log genome copies (gc)/L. Overall, most of the analyzed reclaimed water samples did not comply with current EU legislation and contained potential infectious viral particles.
Topics: Wastewater; Sewage; Humans; Capsid; Coliphages; Rotavirus; Norovirus; Water Microbiology; Real-Time Polymerase Chain Reaction; Feces; Enterovirus; Capsid Proteins
PubMed: 38932109
DOI: 10.3390/v16060816