Serum Testosterone Is Associated With the Severity of COVID-19.

BACKGROUND/AIM: Coronavirus disease 2019 (COVID-19) is more likely to be severe in men than in women. Its association with sex hormones as an aggravating factor for male patients has been attracting attention. This study aimed to investigate whether serum testosterone is associated with the aggravation of COVID-19. PATIENTS AND METHODS: Serum testosterone concentrations in 116 male patients with COVID-19 and residual serum were measured and examined upon their admission to Sapporo Medical University Hospital between February 1, 2020 and March 31, 2021. RESULTS: Blood samples collected from these patients with COVID-19 were analyzed. The serum testosterone levels were 2.19±1.35, 1.29±0.88, and 0.75±0.58 ng/ml in mild, moderate, and severe groups, respectively. Patients with severe COVID-19 on admission had lower testosterone levels (p<0.001). At a cutoff level of 1.31 ng/ml, the area under the curve for the comparison of severe with non-severe cases was 0.825. Furthermore, serum testosterone levels negatively correlated with C-reactive protein and serum amyloid A levels but positively correlated with calcium, zinc, C3, and C4. CONCLUSION: In male patients with COVID-19, low serum testosterone levels correlated with disease severity, accompanied by a strong inflammatory reaction and proportion of complement consumption.

Factors affecting the sensitivity of quantitative severe acute respiratory syndrome coronavirus 2 antigen test.

INTRODUCTION: The accuracy of nucleic acid amplification tests (NAATs) is affected by various factors; however, studies examining the factors affecting the accuracy of quantitative severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigen test (QAT) are limited. METHODS: A total of 347 nasopharyngeal samples were collected from patients with coronavirus disease 2019 (COVID-19), and the date of onset was obtained from the electronic medical records. The SARS-CoV-2 antigen level was measured using Lumipulse Presto SARS-CoV-2 Ag (Presto), while NAAT was performed using the Ampdirect 2019-nCoV Detection Kit. RESULTS: Presto had a sensitivity rate of 95.1% (95% confidence interval: 92.8-97.4) in detecting the SARS-CoV-2 antigen in 347 samples. The number of days from symptom onset to sample collection was negatively correlated with the amount of antigen (r = -0.515) and sensitivity of Presto (r = -0.711). The patients' age was lower in the Presto-negative samples (median age, 39 years) compared with that in the Presto-positive samples (median age, 53 years; p < 0.01). A significant positive correlation was observed between age (excluding teenagers) and Presto sensitivity (r = 0.764). Meanwhile, no association was found between the mutant strain, sex, and Presto results. CONCLUSION: Presto is useful for the accurate diagnosis of COVID-19 owing to its high sensitivity when the number of days from symptom onset to sample collection is within 12 days. Furthermore, age may affect the results of Presto, and this tool has a relatively low sensitivity in younger patients.

Study of post-opening stability of active ingredients in hand sanitizers.

INTRODUCTION: Hand disinfection plays an important role in infection control. Currently, hand sanitizers containing ethanol and chlorhexidine gluconate as active ingredients are widely used. Most of hand sanitizers have a defined expiration date for use. However, there was no rule about the expiration date after opening defined with the evidence. Therefore, we examined the fluctuation of active ingredients and disinfection effect after opening the bottle. METHOD: Twelve hand sanitizers from 44 to 921 days after opening set in different places in the hospital were examined and unopened hand sanitizer used as a control. Chlorhexidine gluconate and ethanol of each samples were measured by high performance liquid chromatography and gas chromatography, respectively. The correlation between the concentration of each ingredient obtained and the number of days after opening, bottle weight, storage temperature and humidity was analyzed. A time-kill test based on ASTM E2315-03 was performed to confirm the actual disinfection effect. RESULTS: It was observed that active ingredients had not been decreased up to 921 days after opening and were not affected by storage conditions after opening. In addition, a decrease of disinfection effect was not observed in any sample. CONCLUSIONS: We found that hand sanitizers do not need to be discard after a number of days have passed because the active ingredients are retained even after opening in it.

Performance analysis among multiple fully automated anti-SARS-CoV-2 antibody measurement reagents: A potential indicator for the correlation of protection in the antibody titer.

BACKGROUND: To evaluate the performance of various reagents in automated analyzers for antibody detection against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). METHODS: Using 100 serum samples from 100 individual patients diagnosed with SARS-CoV-2 infection, the precision, linearity, determination agreement, and correlation of five qualitative reagents (Elecsys Anti-SARS-CoV-2, ARCHITECT SARS-CoV-2 IgG, ARCHITECT SARS-CoV-2 IgM, Access SARS-CoV-2 IgM, and SARS-CoV-2 IgM) and four quantitative reagents (Elecsys Anti-SARS-CoV-2 S, ARCHITECT SARS-CoV-2 IgG II, Access SARS-CoV-2 IgG 1st IS, and SARS-COV-2 IgG S) were analyzed. A surrogate virus-neutralizing test (sVNT) kit was used to evaluate the measurement value of each quantitative reagent corresponding to the amount of neutralizing antibody, similar to that of patients in the late stage of infection. RESULTS: Precision and linearity were found to be sufficient for clinical use. Five discrepant samples were observed in the positive and negative judgments of the qualitative reagents for IgG, and one discrepant sample was observed in the qualitative reagent for IgM. Although the measurement values of the quantitative reagents were different, they were correlated with each reagent. The reference values inferred from the sVNT were Elecsys Anti-SARS-CoV-2: 71.8 U/L, ARCHITECT SARS-CoV-2 IgGⅡ: 2976.3 AU/mL, Access SARS-CoV-2 IgG 1st IS: 689.6 IU/mL, and SARS-CoV-2 IgG S: 19.3 U/L. CONCLUSIONS: The performance observed for each anti-SARS-CoV-2 antibody detection reagent was sufficient. The reference values based on the inhibition rate of sVNT have potential as indicators of the correlation of protection and are expected to be leveraged in automated antibody tests.

Comparative study of rapid antigen testing and two nucleic acid amplification tests for influenza virus detection.

INTRODUCTION: This study aimed to evaluate rapid antigen detection (RAD) and rapid nucleic acid amplification tests (NAATs) to detect influenza virus (IV). METHODS: The conventional RAD test (Quick Chaser Flu A, B: QC), using silver amplified immunochromatography (Quick Chaser Auto Flu A, B: QCA), as well as two NAATs (Xpert Xpress Flu/RSV: Xpert, cobas Influenza A/B & RSV: cobas) were evaluated using nasopharyngeal swabs from suspected cases of influenza. A reference method was performed using real-time reverse transcription polymerase chain reaction according to the manual of the Japanese National Institute of Infectious Disease (NIID). RESULTS: From a total of 177 samples, 51 were positive according to the NIID assay. The kappa (κ) coefficient in Xpert and cobas for influenza A virus (IAV)/influenza B virus (IBV) was 1.00, which was the highest among the four detection assays. However, the κ coefficients in QC and QCA for IAV/IBV were 0.71-0.77 and 0.87-0.89, respectively. The sensitivities of the RAD tests were 41.7% in QC and 50.0% in QCA at < 6 h after onset, and 100.0% in both QC and QCA at 24-48 h after onset. The cycle threshold (Ct) values were significantly lower in the group in which all detection assays were positive for IAV. CONCLUSIONS: Xpert and cobas have comparable analytical performances and are highly useful as influenza virus detection assays. QC and QCA could show false negatives frequently in the early stage of infection and when viral load is low.

Does the timing of saliva collection affect the diagnosis of SARS-CoV-2 infection?

We evaluated the optimal timing of saliva sample collection to diagnose the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. We obtained 150 saliva samples at four specific time points from 13 patients with confirmed SARS-CoV-2 infection. The time points were (1) early morning (immediately after waking), (2) immediately after breakfast before tooth brushing, (3) 2 h after breakfast, and (4) before lunch. On the 2nd hospital day, patients collected saliva at the four time points by themselves. We collected samples at two time points, (1) and (3), from the 3rd hospital day to day 9 following symptom onset. In 52 samples collected at the four time points, there was no significant difference. Meanwhile, there was no significant difference in the positive proportion or the viral load between the two time points in both analyses by the day from symptom onset and by all samples. In this study, there was no difference in the positive proportions in saliva collected at various time points within 9 days after symptom onset. The timing of saliva collection was not affected by the diagnosis of SARS-CoV-2 infection.

Evaluation of false positives in the SARS-CoV-2 quantitative antigen test.

INTRODUCTION: Highly sensitive reagents for detecting SARS-CoV-2 antigens have been developed for accurate and rapid diagnosis till date. In this study, we aim to clarify the frequency of false-positive reactions and reveal their details in SARS-CoV-2 quantitative antigen test using an automated laboratory device. METHODS: Nasopharyngeal swab samples (n = 4992) and saliva samples (n = 5430) were collected. We measured their SARS-CoV-2 antigen using Lumipulse® Presto SARS-CoV-2 Ag and performed a nucleic acid amplification test (NAAT) using the Ampdirect™ 2019 Novel Coronavirus Detection Kit as needed. The results obtained from each detection test were compared accordingly. RESULTS: There were 304 nasopharyngeal samples and 114 saliva samples were positive in the Lumipulse® Presto SARS-CoV-2 Ag test. All positive nasopharyngeal samples in the antigen test were also positive for NAAT. In contrast, only three (2.6%) of all the positive saliva samples in the antigen test were negative for NAAT. One showed no linearity with a dilute solution in the dilution test. Additionally, the quantitative antigen levels of all the three samples did not decrease after reaction with the anti-SARS-CoV-2 antibody. CONCLUSIONS: The judgment difference between the quantitative antigen test and NAAT seemed to be caused by non-specific reactions in the antigen test. Although the high positive and negative predictive value of this quantitative antigen test could be confirmed, we should consider the possibility of false-positives caused by non-specific reactions and understand the characteristics of antigen testing. We recommend that repeating centrifugation before measurement, especially in saliva samples, should be performed appropriately.

Luteinizing hormone (LH) formed a complex with an immunoglobulin G caused abnormally high levels of LH: A case report.

We encountered a 30-year-old woman with remarkably elevated luteinizing hormone (LH) levels, as measured by electrochemiluminescent immunoassay (ECLIA), and no specific symptoms. We performed the following investigations: dilution linearity test, polyethylene glycol (PEG) precipitation test, immunoprecipitation test, protein G addition test, and high-performance liquid chromatography (HPLC) analysis. The linearity of patient's serum was similar to that of a standard LH preparation, and non-specific reactions were not observed. The recovery rate of LH shown by the PEG precipitation test, immunoprecipitation test, and protein G addition test was low. Moreover, an abnormal peak in HPLC was located at a slightly larger molecular weight position than that of IgG. These results showed the presence of macro-LH, LH, and anti-LH-IgG autoantibody complex and suggested that the clearance of LH from the blood was delayed due to IgG binding, and therefore, the LH value was falsely high. We should keep the possibility of macro-LH in mind in cases of unexpectedly high LH values.

Evaluating a novel, highly sensitive, and quantitative reagent for detecting SARS-CoV-2 antigen.

INTRODUCTION: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is rapidly spreading all over the world. A new quantifying reagent for detecting SARS-CoV-2 antigen was developed for early and accurate detection. We evaluated the novel quantitative reagent for detecting SARS-CoV-2 antigen using an automated laboratory device. METHODS: One-hundred nasopharyngeal samples were collected from 47 SARS-CoV-2-infected patients, and 200 samples were collected from healthy donners. We measured the SARS-CoV-2 antigen and nucleic acid using Lumipulse Presto SARS-CoV-2 Ag and the 2019 Novel Coronavirus Detection Kit, respectively. RESULTS: The sensitivity and specificity of the SARS-CoV-2 antigen test were 75.7% (56/74) and 96.0% (192/200), respectively. The concordance rate in the positive group between the antigen and nucleic acid tests was 66% (66/100). In addition, the correlation coefficient between the concentration of SARS-CoV-2 antigen and the level of SARS-CoV-2 RNA was 0.74. There were 19 discrepant samples in which SARS-CoV-2 RNA was detected without SARS-CoV-2 antigen. There was significant difference between the discrepant and matched samples in terms of the time since symptom onset: the 19 discrepant samples were collected a median of 33 days after onset, while the 55 matched samples were collected a median of 19 days after onset. In addition, the 19 discrepant samples were collected from patients who were immune against SARS-CoV-2. CONCLUSIONS: This novel SARS-CoV-2 antigen detection assay is highly sensitive, rapid, accurate, easily diagnostic. It may be useful in both clinical diagnosis and in screening because it does not require special methods such as PCR.

Evaluation of new algorithm using TPLA as an initial syphilis screening test.

We retrospectively compared the performance of an existing syphilis diagnostic algorithm with a new algorithm that analyzes the results of Treponema pallidum latex agglutination (TPLA) tests. Of the 100 clinical blood samples, 51 were classified as positive through both Mediace TPLA and ESPLINE TP; 2/51 were classified as negative by Architect Syphilis TP, whereas 1/51 was negative as per LUMIPULSE Presto TP. The false positive rate when the results of Mediace TPLA and ESPLINE TP were combined was 1.96% versus 0% for both Architect Syphilis TP and LUMIPULSE Presto TP. The sensitivity of Mediace TPLA (98%) was comparable to that of Architect Syphilis TP (98%) but lower than that of LUMIPULSE Presto TP (100%). The specificity of Mediace TPLA was 98.0% versus 100% for Architect Syphilis TP, and versus 100% for LUMIPULSE Presto TP. We conclude that the performance of Mediace TPLA in combination with a reverse algorithm is nearly equal to that of enzyme immunoassay (EIA) or chemiluminescence immunoassay (CIA). Because TPLA is low cost, highly sensitive method for IgM detection, and is easy to operate, we have recommended its adoption for initial syphilis screening tests.

Stanniocalcin-1 promotes metastasis in a human breast cancer cell line through activation of PI3K.

Stanniocalcin-l (STC-1) is a secreted glycoprotein hormone that regulates calcium and phosphate homeostasis. STC-1 expression is upregulated in several cancers including breast cancer, and has been shown to be prognostic. Although these clinical observations implicate STC-1 as a potential tumor marker, it is still unclear whether STC-1 confers a malignant phenotype. In this study, this question was addressed by overexpressing STC-1 in the human breast cancer cell line MDA-MB-231 and examining the resultant phenotype in vitro and in vivo. Overexpression of STC-1 enhanced invasiveness of MDA-MB-231 cells in vitro and promoted their lung metastasis in vivo, while having no effect on proliferation, adhesion, or proteinase activity. The addition of soluble STC-1 to MDA-MB-231 cultures resulted in the activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway, suggesting a mechanistic basis for the observed increases in cell motility and metastasis. Taken together, it was indicated that secreted STC-1 promotes metastatic potential of breast cancer cells via activation of PI3K/AKT.

Prophylactic and therapeutic effects of Acanthopanax senticosus Harms extract on murine collagen-induced arthritis.

Evidences are accumulating that extract of Acanthopanax senticosus Harms (ASH; syn Eleutherococcus senticosus [Rupr. & Maxim.] Maxim), a shrub native to Northeastern Asia, has antiinflammatory effects. In this study, we examined prophylactic and therapeutic effects of ASH extract (ASHE) on rheumatoid arthritis using collagen-induced arthritis (CIA) mouse model. Acanthopanax senticosus Harms extract was administered before the onset of arthritis in the prophylaxis model. In the therapeutic model, ASHE was administered after the onset of arthritis with or without anti-TNF-α antibody. The ASHE treatment showed efficacy before onset of CIA but there was no effect after CIA was established. The ASHE treatment delayed the onset and decreased severity of CIA. In vitro examinations showed that ASHE is an antioxidant and that ASHE suppresses TNF-α and interleukin-6 production in human peripheral blood mononuclear cells. The combination therapy with ASHE and anti-TNF-α antibody reduced the severity of arthritis compared with anti-TNF-α antibody alone. The present study shows that ASHE has prophylactic effect against CIA and support therapeutic effect of anti-TNF-α antibody.

Reconstructing a 3-dimensional image of the results of antinuclear antibody testing by indirect immunofluorescence.

Indirect immunofluorescence anti-nuclear antibody testing (IIF-ANAT) is an essential screening tool in the diagnosis of various autoimmune disorders. ANA titer quantification and interpretation of immunofluorescence patterns are determined subjectively, which is problematic. First, we determined the examination conditions under which IIF-ANAT fluorescence intensities are quantified. Next, IIF-ANAT was performed using homogeneous, discrete speckled, and mixed serum samples. Images were obtained using Bio Zero BZ-8000, and 3-dimensional images were reconstructed using the BZ analyzer software. In the 2-dimensional analysis, homogeneous ANAs hid the discrete speckled pattern, resulting in a diagnosis of homogeneous immunofluorescence. However, 3-dimensional analysis of the same sample showed discrete speckled-type ANA in the homogeneous background. This study strengthened the current IIF-ANAT method by providing a new approach to quantify the fluorescence intensity and enhance the resolution of IIF-ANAT fluorescence patterns. Reconstructed 3-dimensional imaging of IIF-ANAT can be a powerful tool for routine laboratory examination.