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Abstract There have been several reports of skin manifestations in patients with coronavirus disease 2019 (COVID-19). However, it is unclear whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA can be detected on the skin surface, including the sebum, of these patients. In this study, SARS-CoV-2 RNA was detected using real-time reverse-transcription polymerase chain reaction (RT-PCR) assay of skin surface lipids (SSLs) collected using an oil-blotting film from the faces of hospitalized patients with COVID-19. Human transcriptome analysis was also performed using the same samples. In facial SSLs of patients with COVID-19, the RT-PCR positivity rate was 84.6% (11/13 samples) within 5 days and 30.4% (7/23 samples) by 6–10 days of symptom onset. In the transcriptome analysis, the most characteristic SSL-RNA profile was the upregulation of interferon-stimulated gene (ISG)-related genes, such as ISG15, IFITM1, and MX1. This study presents an alternative technique using SSLs for non-invasive SARS-CoV-2 RNA detection and simultaneous analysis of human molecular pathogenesis in patients with COVID-19.

Abstract Background The challenges in culturing Treponema pallidum have hindered molecular-biological analysis. This study aims to establish a molecular epidemiological analysis of syphilis among Japanese men who have sex with men (MSM) and to investigate the relationship between bacteremia and associated pathophysiology. Methods We used whole blood specimens from syphilis-diagnosed individuals in Tokyo, collected between February 2019 and June 2022. All individuals were MSM, and most were people with HIV (97.2%). We used a multi-locus sequence typing (MLST) scheme for epidemiological analysis. Sequences for MLST (TP0136, TP0548, and TP0705) were obtained. Results Out of 71 whole blood samples, 26 samples (36.6%) were positive for TP0136, and we sequenced three loci for MLST in 22 samples (31.0%). The most frequently detected sequence type (ST) was ST3 (n = 9), followed by ST6 (n = 6). Phylogenetic analysis revealed that 12 samples belonged to the SS14-like group (60%), and 8 samples belonged to the Nichols-like group (40%). Treponema pallidum subsp. endemicum (TEN), the cause of bejel was detected in three samples (12%). There was a significant association between TP0136 detection rate and C- reactive protein (CRP) (77.0% at a cut-off:0.5 mg/dL). Conclusion Both SS14-like and Nichols-like strains were circulating concurrently, and TEN could have been sexually transmitted among MSM with HIV. Elevated CRP may indicate the presence of the pathogen in the blood.

Abstract Background Antiretroviral therapy (ART) has significantly improved the prognosis for people living with human immunodeficiency virus (PLWH). However, alterations in the gut microbiota of PLWH affect their metabolic environment and may contribute to chronic inflammation and premature aging, despite effective ART. Given that inflammation in the body have been shown to be risk factors for age-related diseases, elucidating changes in the gut microbiota and metabolic levels over time in individuals would be for understanding the dynamics in the body that lead to disease development. Methods The study assessed changes in blood markers, gut microbiota, and metabolic environment of PLWH on ART for four years, with participants divided into two groups based on median CD4 cell count (580/µL). Results The high-CD4 group was more obese than the low-CD4 group, but there was no significant difference in comorbidities, such as diabetes, hypertension, and dyslipidemia. An investigation of the alterations in gut microbiota over a 4-year period revealed a reduction in specific groups of bacteria that produce short-chain fatty acids (SCFAs) and an increase in the occurrence of Enterobacteriaceae. In addition, a decline in bacterial thiamine synthesis was predicted, and a decreased abundance of the Ruminococcus gauvreauii group, which relies on supplementation with thiamine from symbiotic bacteria, was observed. The abundance of Collinsella and Catenibacterium at baseline was positively correlated with the predicted number of thiamine-synthesizing pathway. The presence of Catenibacterium at follow-up was inversely correlated with the rate of increase in plasma levels of lipopolysaccharide-binding protein, an indicator of a leaky gut. In terms of assessing the diversity of gut microbiota, a trend towards an increase in opportunistic bacteria with different genetic distances was observed in the high-CD4 group at follow-up. Conclusion These observations suggested that, despite effective ART, the thiamine-deficient environment in the gut continues to reduce thiamine-requiring bacteria, such as certain SCFA-producing bacteria. As a result, secondary effects, such as proliferation of opportunistic bacteria and chronic inflammation associated with increased intestinal permeability, proceed in PLWH. Thus, certain essential nutrient deficiencies for gut metabolic environment may disrupt the symbiotic relationship between gut bacteria and increase the risk of developing inflammatory diseases.

A patient in Japan with HIV began antiretroviral therapy because of acute hepatitis B virus (HBV) 15 years ago, with low hepatitis B surface antibody, and experienced breakthrough HBV reactivation 4 months after switching from bictegravir/emtricitabine/tenofovir alafenamide to cabotegravir/rilpivirine. An immune escape mutation, E164V, was identified in the isolated HBV DNA.

Abstract Background Person with human immunodeficiency virus type-1 (PWH) are prone to chronic inflammation due to residual viral production, even with antiretroviral therapy (ART), which increases the risk of age-related diseases. There is also limited information on changes in the intestinal environment of PWH during ART. In this longitudinal study, we investigated changes in the gut microbiota, persistence of chronic inflammation, interactions between the gut environment and inflammation, and metabolic changes in PWH using long-term ART. Results We analyzed changes in clinical parameters and gut microbiota in 46 PWH over a mean period of 4 years to understand the influence of gut dysbiosis on inflammation. Overall, changes in the gut microbiota included a decrease in some bacteria, mainly involved in short-chain fatty acid (SCFA) production, and an increase in certain opportunistic bacteria. Throughout the study period, an increase in bacterial-specific metabolic activity was observed in the intestinal environment. Continued decline in certain bacteria belonging to the Clostridia class and metabolic changes in gut bacteria involved in glucose metabolism. Additionally, patients with a low abundance of Parabacteroides exhibited low bacterial alpha diversity and a significant increase in body mass index (BMI) during the study period. Monocyte chemoattractant protein 1, a marker of macrophage activation in the plasma, continued to increase from baseline (first stool collection timepoint) to follow-up (second stool collection timepoint), demonstrating a mild correlation with BMI. Elevated BMI was mild to moderately correlated with elevated levels of plasma interleukin 16 and chemokine ligand 13, both of which may play a role in intestinal inflammation and bacterial translocation within the gut microbiota. The rate of BMI increase correlated with the rate of decrease in certain SCFA-producing bacteria, such as Anaerostipes and Coprococcus 3. Conclusion Our data suggest that despite effective ART, PWH with chronic inflammation exhibit persistent dysbiosis associated with gut inflammation, resulting in a transition to an intestinal environment with metabolic consequences. Moreover, the loss of certain bacteria such as Parabacteroides in PWH correlates with weight gain and may contribute to the development of metabolic diseases.

Abstract Escherichia coli O157 can cause foodborne outbreaks, with infection leading to severe disease such as hemolytic-uremic syndrome. Although phage-based detection methods for E. coli O157 are being explored, research on their specificity with clinical isolates is lacking. Here, we describe an in vitro assembly-based synthesis of vB_Eco4M-7, an O157 antigen-specific phage with a 68-kb genome, and its use as a proof of concept for E. coli O157 detection. Linking the detection tag to the C-terminus of the tail fiber protein, gp27 produces the greatest detection sensitivity of the 20 insertions sites tested. The constructed phage detects all 53 diverse clinical isolates of E. coli O157, clearly distinguishing them from 35 clinical isolates of non-O157 Shiga toxin-producing E. coli. Our efficient phage synthesis methods can be applied to other pathogenic bacteria for a variety of applications, including phage-based detection and phage therapy.

Objective Patients with haemophilia and HIV who acquire hepatitis C virus (HCV) after receiving contaminated blood products can experience accelerated progression of liver fibrosis and a poor prognosis, making liver disease a prominent cause of mortality among these patients. In the current study, we aimed to evaluate the safety and tolerability of the potential antifibrotic agent OP-724—a CREB-binding protein/β-catenin inhibitor—in this patient subset.Design In this single-centre, open-label, non-randomised, phase I trial, we sequentially enrolled patients with cirrhosis following HIV/HCV coinfection classified as Child-Pugh (CP) class A or B. Five patients received an intravenous infusion of OP-724 at doses of 140 or 280 mg/m2 for 4 hours two times weekly over 12 weeks. The primary endpoint was the incidence of serious adverse events (SAEs). Secondary endpoints included the incidence of AEs and improved liver stiffness measure (LSM), as determined by vibration-controlled transient elastography. This study was registered at ClinicalTrials.gov (NCT04688034).Results Between 9 February 2021 and 5 July 2022, five patients (median age: 51 years) were enrolled. All five patients completed 12 cycles of treatment. SAEs were not observed. The most common AEs were fever (60%) and gastrointestinal symptoms (diarrhoea: 20%, enterocolitis: 20%). Improvements in LSM and serum albumin levels were also observed.Conclusion In this preliminary assessment, intravenous administration of 140 or 280 mg/m2/4 hours OP-724 over 12 weeks was well tolerated by patients with haemophilia combined with cirrhosis due to HIV/HCV coinfection. Hence, the antifibrotic effects of OP-724 warrant further assessment in patients with cirrhosis.Trial registration number NCT04688034.

Abstract Background People living with HIV (PLWH) with chronic inflammation may have an increasing risk for coronavirus disease 2019 (COVID-19) severity; however, the impact of their gut microbiota on COVID-19 is not fully elucidated. Here, we analyzed the temporal changes in the gut microbiota composition of hospitalized severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected PLWH (PLWH-CoV) and their correlation with COVID-19 severity. Result The 16S rRNA analysis results using stool samples (along the timeline from disease onset) from 12 hospitalized PLWH-CoV, whose median CD4 + T cell count was 671 cells/µl, were compared to those of 19 healthy people and 25 PLWH. Bacterial diversity in PLWH-CoV is not significantly different from that of healthy people and SARS-CoV-2 non-infected PLWH, but a significant difference in the microbiota diversity was observed in the classification according to the disease severity. Immediately after the disease onset, remarkable changes were observed in the gut microbiota of PLWH-CoV, and the changing with a decrease in some short-chain fatty acid-producing bacteria and an increase in colitis-related pathobiont. In the second week after disease onset, relative amounts of specific bacteria distinguished between disease severity. One month after the disease onset, dysbiosis of the gut microbiota persisted, and the number of Enterobacteriaceae, mainly Escherichia-Shigella, which is potentially pathogenic, increased and were enriched in patients who developed post-acute sequelae of COVID-19 (PASC). Conclusion The changes in the gut microbiota associated with SARS-CoV-2 infection observed in PLWH in this study indicated a persistent decrease in SCFA-producing bacteria and an intestinal environment with an increase in opportunistic pathogens associated with enteritis. This report demonstrates that the intestinal environment in PLWH tends to show delayed improvement even after COVID-19 recovery, and highlights the importance of the dysbiosis associated with SARS-CoV-2 infection as a potential factor in the COVID-19 severity and the PASC in PLWH.

Abstract We assessed whether the impact of cabotegravir plus rilpivirine on inflammation reduction differs from that of oral antiretrovirals, using real-world data. Inflammatory biomarkers and lipid profiles were followed from baseline to 8 months after switching. Seventy-eight participants were analyzed. The CD4/CD8 ratio and C-reactive protein did not change. There were transient decreases in CD8 and CD4 counts in the group that switched from the dolutegravir-based regimen, but not in the tenofovir alafenamide-based regimen group. High-density lipoprotein (HDL) cholesterol increased, resulting in a decrease in the total-cholesterol to HDL cholesterol ratio, whereas there was no significant change in low-density lipoprotein cholesterol.

This article - Recommendations and Guidelines of Integrative Medicine (IM) for COVID-19 Care - was one of the outcomes from an Asia-Pacific Economic Cooperation (APEC) Project (Integrative Medicine (IM) and COVID -19 Care) during the time between May 2022 and March 2023. With the efforts from care providers, researchers, health policy makers and healthcare administrative leaders among APEC economies, the purpose of this file was to provide comprehensive IM systems for COVID-19 care as recommendations and suggestive guidelines including care methods, tools, procedures, symptom conditions and targets selections, and points need to be considered during care applications. All cited COVID-19 care practices have confirmed their efficacy and usefulness either used alone or combined with conventional medicine. This article provides current useful medical information on IM for COVID-19 care which could benefit APEC economies and world health communities on their healthcare system.

Intestinal Behçet disease (BD), associated with myelodysplastic syndrome (MDS), is often refractory to treatment. An 80-year-old man with trisomy 8 MDS (refractory anemia) developed intermittent fever. Despite investigations to exclude infectious disease, autoimmune disease, and malignancy as the cause of the fever, the etiology could not be determined. A colonoscopy revealed several shallow round ulcers in the ileocecal region and ascending colon, and the biopsy specimens showed nonspecific inflammation. Thereafter, the patient experienced abdominal pain and diarrhea. Other than an oral aphthous ulcer, the patient did not show symptoms to meet the diagnostic criteria for BD. The patient was diagnosed with intestinal ulcers (intestinal BD-like disease) with MDS and trisomy 8. After treatment failure with 5-aminosalicylic acid, steroid, colchicine, and azacitidine, cerebral infarction occurred. Eating was difficult because of the patient’s impaired consciousness; hence, total parenteral nutrition (TPN) was commenced. The fever and abdominal symptoms improved with bowel rest over approximately 1 month. Small amounts of food were orally administered to the patient following recovery from the after-effects of the cerebral infarction, but diarrhea and fever repeatedly flared up. Therefore, TPN was continued at home. The patient has not experienced any further intestinal BD symptoms for approximately 1 year with bowel rest. Nutritional therapy, including bowel rest, may be an effective treatment option for intestinal BD with MDS, and might be used as an induction therapy of remission or a supportive therapy for other treatments.

Although antibiotics are most frequently prescribed for respiratory tract infections, effective interventions for their proper use by physicians have not been fully established. We assessed the impact of educational films on the rates of antibiotic prescriptions for respiratory tract infections using fictitious scenarios. In this nationwide web-based survey prospective study, a total of 1100 physicians were included. The physicians were required to view educational short films and determine the need for prescribing antibiotics in 10 fictitious scenarios involving adults diagnosed with different acute respiratory tract infectious diseases. The antibiotic prescription rates for each scenario were compared before and after viewing the educational short film. The rates of antibiotic prescription significantly decreased after viewing the educational film, especially in cases with a narrowly defined common cold (from 51% to 15%), mild pharyngolaryngitis (from 71% to 25%), and acute bronchitis without chronic respiratory underlying diseases (from 63% to 23%). Alternatively, a slight decrease in rates was observed in cases with moderate or severe rhinosinusitis (from 94% to 79%), moderate or severe acute pharyngitis (from 88% to 69%), and acute bronchitis with chronic lung disease (from 70% to 58%), for which antibiotics are recommended. Educational short films may encourage the proper use of antibiotics for respiratory tract infections; however, the possibility of undertreatment in patients requiring antibiotics must be considered.

Abstract Background The mucosa serves as the first defence against pathogens and facilitates the surveillance and elimination of symbiotic bacteria by mucosal immunity. Recently, the mRNA vaccine against SARS-CoV-2 has been demonstrated to induce secretory antibodies in the oral and nasal cavities in addition to a systemic immune response. However, the mechanism of induced immune stimulation effect on mucosal immunity and commensal bacteria profile remains unclear. Methods Here, we longitudinally analysed the changing nasal microbiota and both systemic and nasal immune response upon SARS-CoV-2 mRNA vaccination, and evaluated how mRNA vaccination influenced nasal microbiota in 18 healthy participants who had received the third BNT162b. Results The nasal S-RBD IgG level correlated significantly with plasma IgG levels until 1 month and the levels were sustained for 3 months post-vaccination. In contrast, nasal S-RBD IgA induction peaked at 1 month, albeit slightly, and correlated only with plasma IgA, but the induction level decreased markedly at 3 months post-vaccination. 16 S rRNA sequencing of the nasal microbiota post-vaccination revealed not an overall change, but a decrease in certain opportunistic bacteria, mainly Fusobacterium. The decrease in these bacteria was more pronounced in those who exhibited nasal S-RBD IgA induction, and those with higher S-RBD IgA induction had lower relative amounts of potentially pathogenic bacteria such as Pseudomonas pre-vaccination. In addition, plasma and mucosal S-RBD IgG levels correlated with decreased commensal pathogens such as Finegoldia. Conclusions These findings suggest that the third dose of SARS-CoV-2 mRNA vaccination induced S-RBD antibodies in the nasal mucosa and may have stimulated mucosal immunity against opportunistic bacterial pathogens. This effect, albeit probably secondary, may be considered one of the benefits of mRNA vaccination. Furthermore, our data suggest that a cooperative function of mucosal and systemic immunity in the reduction of bacteria and provides a better understanding of the symbiotic relationship between the host and bacteria in the nasal mucosa.

Summary: EG.5.1 is a subvariant of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron XBB variant that is rapidly increasing in prevalence worldwide. However, the pathogenicity, transmissibility, and immune evasion properties of isolates of EG.5.1 are largely unknown. Here, we show that there are no obvious differences in growth ability and pathogenicity between EG.5.1 and XBB.1.5 in hamsters. We also demonstrate that, like XBB.1.5, EG.5.1 is transmitted more efficiently between hamsters compared to its predecessor, BA.2. In contrast, unlike XBB.1.5, we detect EG.5.1 in the lungs of four of six exposed hamsters, suggesting that the virus properties of EG.5.1 are different from those of XBB.1.5. Finally, we find that the neutralizing activity of plasma from convalescent individuals against EG.5.1 was slightly, but significantly, lower than that against XBB.1.5 or XBB.1.9.2. Our data suggest that the different virus properties after transmission and the altered antigenicity of EG.5.1 may be driving its increasing prevalence over XBB.1.5 in humans.

Abstract Background Rifaximin is a poorly absorbed broad-spectrum antibiotic used for hepatic encephalopathy. Although increased Lactobacillaceae and decreased Bacteroidetes abundance are characteristic of hepatic encephalopathy, rifaximin does not dramatically alter the stool microbiota. As the antimicrobial effect of rifaximin increases by micellization with bile acids, we hypothesized that rifaximin alters the microbiota in the duodenum and jejunum, where the levels of bile acids are abundant. Methods and results Eight-week-old BALB/c mice were injected with carbon tetrachloride (CCl4) intraperitoneally for 12 weeks to induce liver fibrosis. The mice were grouped into the control (n = 9), CCl4 (n = 13), and rifaximin group in which mice were treated with rifaximin for two weeks after CCl4 administration (n = 13). We analyzed the microbiota of the duodenum, jejunum, ileum, cecum, and stool using 16S ribosomal RNA gene analysis. The content of Lactobacillaceae, the most abundant bacterial family in the duodenum and small intestine, increased in the CCl4 group, especially in the jejunum (median 67.0% vs 87.8%, p = 0.03). Rifaximin significantly decreased Lactobacillaceae content in the duodenum (median 79.4% vs 19.0%, p = 0.006) and jejunum (median 87.8% vs 61.3%, p = 0.03), but not in the ileum, cecum, and stool. Bacteroidetes abundance tended to decrease on CCl4 administration and increased following rifaximin treatment in the duodenum and jejunum. S24_7, the most abundant family in Bacteroidetes, demonstrated a significant inverse correlation with Lactobacillaceae (duodenum, r = − 0.61, p

Although SARS-CoV-2 evolution seeds a continuous stream of antibody-evasive viral variants, COVID-19 mRNA vaccines provide robust protection against severe disease and hospitalization. Here, we asked whether mRNA vaccine–induced memory T cells limit lung SARS-CoV-2 replication and severe disease. We show that mice and humans receiving booster BioNTech mRNA vaccine developed potent CD8 T cell responses and showed similar kinetics of expansion and contraction of granzyme B/perforin-expressing effector CD8 T cells. Both monovalent and bivalent mRNA vaccines elicited strong expansion of a heterogeneous pool of terminal effectors and memory precursor effector CD8 T cells in spleen, inguinal and mediastinal lymph nodes, pulmonary vasculature, and most surprisingly in the airways, suggestive of systemic and regional surveillance. Furthermore, we document that: (a) CD8 T cell memory persists in multiple tissues for > 200 days; (b) following challenge with pathogenic SARS-CoV-2, circulating memory CD8 T cells rapidly extravasate to the lungs and promote expeditious viral clearance, by mechanisms that require CD4 T cell help; and (c) adoptively transferred splenic memory CD8 T cells traffic to the airways and promote lung SARS-CoV-2 clearance. These findings provide insights into the critical role of memory T cells in preventing severe lung disease following breakthrough infections with antibody-evasive SARS-CoV-2 variants.

Summary: The emergence and spread of new SARS-CoV-2 variants with mutations in the spike protein, such as the XBB.1.5 and XBB.1.9.1 sublineages, raise concerns about the efficacy of current COVID-19 vaccines and therapeutic monoclonal antibodies (mAbs). In this study, none of the mAbs we tested neutralized XBB.1.9.1 or XBB.1.5, even at the highest concentration used. We also found that the bivalent mRNA vaccine could enhance humoral immunity against XBB.1.9.1, but that XBB.1.9.1 and XBB.1.5 still evaded humoral immunity induced by vaccination or infection. Moreover, the susceptibility of XBB.1.9.1 to remdesivir, molnupiravir, nirmatrelvir, and ensitrelvir was similar to that of the ancestral strain and the XBB.1.5 isolate in vitro. Finally, we found the replicative fitness of XBB.1.9.1 to be similar to that of XBB.1.5 in hamsters. Our results suggest that XBB.1.9.1 and XBB.1.5 have similar antigenicity and replicative ability, and that the currently available COVID-19 antivirals remain effective against XBB.1.9.1.

Abstract Health state utilities are global measurements of quality of life and have been used to evaluate health outcomes for the cost-utility analysis. This study aimed to estimate the health state utilities of patients with hepatitis B (HB), hepatitis C (HC), and hepatitis-related diseases in Japan. We distributed a self-administered questionnaire, including the EuroQol 5-Dimension 5-Level (EQ-5D-5L), to 9,952 outpatients with several clinical conditions caused by HB or HC virus infection (such as asymptomatic chronic hepatitis, chronic hepatitis, compensated cirrhosis, and decompensated cirrhosis) and estimated the condition-specific utilities of patients with HB or HC. In patients with more severe conditions (patients with acute hepatitis, fulminant hepatitis, and hepatocellular carcinoma and patients undergoing post-liver transplantation), the utilities of these severe conditions were estimated by three hepatitis experts using the EQ-5D-5L. The means of the utilities for acute hepatitis, fulminant hepatitis, asymptomatic chronic hepatitis, chronic hepatitis, compensated cirrhosis, compensated cirrhosis, hepatocellular carcinoma stage I/II, hepatocellular carcinoma stage III/IV, and post-liver transplantation were 0.529, − 0.111, 0.904, 0.868, 0.845, 0.722, 0,675, 0,428, and 0.651 and 0.876, 0.821, 0.737, 0.671, 0.675, 0.428, and 0.651 in HB and HC, respectively. To the best of our knowledge, this is the first study that comprehensively assessed the health state utilities of patients with HB, HC and hepatitis-related conditions from a nationwide survey in Japan using the EQ-5D-5L.

The hepatitis B vaccine is recognised as the most effective approach in reducing hepatitis-B-related morbidity; vaccine-escape mutations are however capable of infecting vaccinated individuals. In this work, authors aim to establish a hepatitis B vaccine candidate, which they assess in rhesus macaques in terms of efficacy and safety.

Summary: SARS-CoV-2 has gradually acquired amino acid substitutions in its S protein that reduce the potency of neutralizing antibodies, leading to decreased vaccine efficacy. Here, we attempted to obtain mutant viruses by passaging SARS-CoV-2 in the presence of plasma samples from convalescent patients or vaccinees to determine which amino acid substitutions affect the antigenicity of SARS-CoV-2. Several amino acid substitutions in the S2 region, as well as the N-terminal domain (NTD) and receptor-binding domain (RBD), affected the neutralization potency of plasma samples collected from vaccinees, indicating that amino acid substitutions in the S2 region as well as those in the NTD and RBD affect neutralization by vaccine-induced antibodies. Furthermore, the neutralizing potency of vaccinee plasma samples against mutant viruses we obtained or circulating viruses differed among individuals. These findings suggest that genetic backgrounds of vaccinees influence the recognition of neutralizing epitopes.

ABSTRACT Nelfinavir, an orally administered inhibitor of human immunodeficiency virus protease, inhibits the replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro. We conducted a randomized controlled trial to evaluate the clinical efficacy and safety of nelfinavir in patients with SARS-CoV-2 infection. We included unvaccinated asymptomatic or mildly symptomatic adult patients who tested positive for SARS-CoV-2 infection within 3 days before enrollment. The patients were randomly assigned (1:1) to receive oral nelfinavir (750 mg; thrice daily for 14 days) combined with standard-of-care or standard-of-care alone. The primary endpoint was the time to viral clearance, confirmed using quantitative reverse-transcription PCR by assessors blinded to the assigned treatment. A total of 123 patients (63 in the nelfinavir group and 60 in the control group) were included. The median time to viral clearance was 8.0 (95% confidence interval [CI], 7.0 to 12.0) days in the nelfinavir group and 8.0 (95% CI, 7.0 to 10.0) days in the control group, with no significant difference between the treatment groups (hazard ratio, 0.815; 95% CI, 0.563 to 1.182; P = 0.1870). Adverse events were reported in 47 (74.6%) and 20 (33.3%) patients in the nelfinavir and control groups, respectively. The most common adverse event in the nelfinavir group was diarrhea (49.2%). Nelfinavir did not reduce the time to viral clearance in this setting. Our findings indicate that nelfinavir should not be recommended in asymptomatic or mildly symptomatic patients infected with SARS-CoV-2. The study is registered with the Japan Registry of Clinical Trials (jRCT2071200023). IMPORTANCE The anti-HIV drug nelfinavir suppresses the replication of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro. However, its efficacy in patients with COVID-19 has not been studied. We conducted a multicenter, randomized controlled trial to evaluate the efficacy and safety of orally administered nelfinavir in patients with asymptomatic or mildly symptomatic COVID-19. Compared to standard-of-care alone, nelfinavir (750 mg, thrice daily) did not reduce the time to viral clearance, viral load, or the time to resolution of symptoms. More patients had adverse events in the nelfinavir group than in the control group (74.6% [47/63 patients] versus 33.3% [20/60 patients]). Our clinical study provides evidence that nelfinavir, despite its antiviral effects on SARS-CoV-2 in vitro, should not be recommended for the treatment of patients with COVID-19 having no or mild symptoms.

The detailed mechanisms of COVID-19 infection pathology remain poorly understood. To improve our understanding of SARS-CoV-2 pathology, we performed a multi-omics and correlative analysis of an immunologically naïve SARS-CoV-2 clinical cohort from blood plasma of uninfected controls, mild, and severe infections. Consistent with previous observations, severe patient populations showed an elevation of pulmonary surfactant levels. Intriguingly, mild patients showed a statistically significant elevation in the carnosine dipeptidase modifying enzyme (CNDP1). Mild and severe patient populations showed a strong elevation in the metabolite L-cystine (oxidized form of the amino acid cysteine) and enzymes with roles in glutathione metabolism. Neutrophil extracellular traps (NETs) were observed in both mild and severe populations, and NET formation was higher in severe vs. mild samples. Our correlative analysis suggests a potential protective role for CNDP1 in suppressing PSPB release from the pulmonary space whereas NET formation correlates with increased PSPB levels and disease severity. In our discussion we put forward a possible model where NET formation drives pulmonary occlusions and CNDP1 promotes antioxidation, pleiotropic immune responses, and vasodilation by accelerating histamine synthesis.

Abstract Background Hepatitis C virus (HCV) has been mainly transmitted through injection drug use, but recently, sexual transmission among men who have sex with men (MSM), which is also a major route of HIV transmission, is increasing. However, the prevalence of HIV and the incidence of other sexually transmitted infections (STIs) among HCV patients have been rarely reported. Methods Using a healthcare insurance claim data of employees and their dependents covering seven-million people in Japan, we evaluated HIV prevalence among HCV patients aged 20–59 years. Hemophilia patients were excluded. HIV and HCV were defined by registered diagnoses and receiving viral RNA testing. The time course of HCV and HIV infections was analyzed. Incidences of syphilis, amebiasis, chlamydia, gonorrhea, hepatitis A, and hepatitis B were assessed. Results From April 2012 to August 2018, 6,422 HCV patients were identified. HIV prevalence was 0.48% (31/6422, 95% CI [confidence interval]: 0.33–0.68%). HIV was diagnosed after HCV in 3.2% (1/31), before HCV in 58.1% (18/31), and concurrently in 38.7% (12/31). Compared with HCV patients without HIV infection, HCV/HIV co-infected patients were younger (median age, 37 vs 51 years, p

Background and Aims: It remains unclear whether obesity increases the risk of hepatocellular carcinoma (HCC) in patients with chronic hepatitis C who achieved a sustained virological response (SVR) with antiviral therapy. Methods: In this multicenter cohort study, we enrolled patients with chronic hepatitis C who achieved SVR with interferon (IFN)-based therapy (IFN group) or direct-acting antiviral (DAA) therapy (DAA group) between January 1, 1990, and December 31, 2018. The patients underwent regular surveillance for HCC. Cumulative incidence of and the risk factors for HCC development after SVR were assessed using the Kaplan-Meier method and Cox proportional hazard regression analysis, respectively. Results: Among 2,055 patients (840 in the IFN group and 1,215 in the DAA group), 75 developed HCC (41 in the IFN group and 34 in the DAA group) during the mean observation period of 4.1 years. The incidence rates of HCC at 1, 2, and 3 years were 1.2, 1.9, and 3.0%, respectively. Multivariate analysis revealed that in addition to older age, lower albumin level, lower platelet count, higher alpha-fetoprotein level, and absence of dyslipidemia, obesity (body mass index ≥25 kg/m2) and heavy alcohol consumption (≥60 g/day) were independent risk factors for HCC development, with adjusted hazard ratio (HR) of 2.53 (95% confidence interval [CI]: 1.51–4.25) and 2.56 (95% CI: 1.14–5.75), respectively. The adjusted HR was not significant between the 2 groups (DAA vs. IFN; HR 1.19, 95% CI: 0.61–2.33). Conclusions: Obesity and heavy alcohol consumption increased the risk of HCC development after SVR.

Abstract Background As a blood-borne pathogen, hepatitis C virus (HCV) has long been a major threat associated with needle-stick injuries (NSIs) mainly because no vaccine is available for HCV. Following an NSI, we usually test the source patient for HCV antibody (HCV-Ab). Since HCV-Ab positivity does not necessarily indicate current infection, HCV RNA is further examined in patients positive for HCV-Ab. Direct-acting antivirals (DAAs) have enabled us to treat most HCV-infected patients; therefore, we speculate that the rate of HCV RNA positivity among HCV-Ab-positive patients decreased after the emergence of DAAs. This cross-sectional study was performed to investigate the change in the actual HCV RNA positivity rate in source patients before and after the interferon (IFN)-free DAA era. Methods This was a cross-sectional study of NSI source patients at a tertiary academic hospital in Japan from 2009 to 2019. IFN-free DAA regimens were first introduced in Japan in 2014. Accordingly, we compared HCV status of NSI source patients that occurred between 2009 and 2014 (the era before IFN-free DAAs) with those that occurred between 2015 and 2019 (the era of IFN-free DAAs) in a tertiary care hospital in Japan. Results In total, 1435 NSIs occurred, and 150 HCV-Ab-positive patients were analyzed. The proportion of HCV RNA-positive patients significantly changed from 2009 through 2019 (p = 0.005, Cochran–Armitage test). Between 2009 and 2014, 102 source patients were HCV-Ab-positive, 78 of whom were also positive for HCV RNA (76.5%; 95%CI, 67.4–83.6%). Between 2015 and 2019, 48 patients were HCV-Ab-positive, 23 of whom were also positive for HCV RNA (47.9%; 95%CI, 34.5–61.7%; p = 0.0007 compared with 2009–2014). In the era of IFN-free DAAs, 9 of 23 HCV RNA-negative patients (39.1%) and 2 of 22 HCV RNA-positive patients (9.1%) were treated with an IFN-free combination of DAAs (p = 0.0351). Regarding the departments where NSIs occurred, HCV RNA-negative patients were predominant in departments not related to liver diseases in the era of IFN-free DAAs (p = 0.0078, compared with 2009–2014). Conclusions Actual HCV RNA positivity in source patients of NSIs decreased after the emergence of IFN-free DAAs. IFN-free DAAs might have contributed to this reduction, and HCV RNA-negative patients were predominant in departments not related to liver diseases in the era of IFN-free DAAs.

ABSTRACT The role of the intestinal microbiota in coronavirus disease 2019 (COVID-19) is being elucidated. Here, we analyzed the temporal changes in microbiota composition and the correlation between inflammation biomarkers/cytokines and microbiota in hospitalized COVID-19 patients. We obtained stool specimens, blood samples, and patient records from 22 hospitalized COVID-19 patients and performed 16S rRNA metagenomic analysis of stool samples over the course of disease onset compared to 40 healthy individual stool samples. We analyzed the correlation between the changes in the gut microbiota and plasma proinflammatory cytokine levels. Immediately after admission, differences in the gut microbiota were observed between COVID-19 patients and healthy subjects, mainly including enrichment of the classes Bacilli and Coriobacteriia and decrease in abundance of the class Clostridia. The bacterial profile continued to change throughout the hospitalization, with a decrease in short-chain fatty acid-producing bacteria including Faecalibacterium and an increase in the facultatively anaerobic bacteria Escherichia-Shigella. A consistent increase in Eggerthella belonging to the class Coriobacteriia was observed. The abundance of the class Clostridia was inversely correlated with interferon-γ level and that of the phylum Actinobacteria, which was enriched in COVID-19, and was positively correlated with gp130/sIL-6Rb levels. Dysbiosis was continued even after 21 days from onset. The intestines tended to be an aerobic environment in hospitalized COVID-19 patients. Because the composition of the gut microbiota correlates with the levels of proinflammatory cytokines, this finding emphasizes the need to understand how pathology is related to the temporal changes in the specific gut microbiota observed in COVID-19 patients. IMPORTANCE There is growing evidence that the commensal microbiota of the gastrointestinal and respiratory tracts regulates local and systemic inflammation (gut-lung axis). COVID-19 is primarily a respiratory disease, but the involvement of microbiota changes in the pathogenesis of this disease remains unclear. The composition of the gut microbiota of patients with COVID-19 changed over time during hospitalization, and the intestines tended to be an aerobic environment in hospitalized COVID-19 patients. These changes in gut microbiota may induce increased intestinal permeability, called leaky gut, allowing bacteria and toxins to enter the circulatory system and further aggravate the systemic inflammatory response. Since gut microbiota composition correlates with levels of proinflammatory cytokines, this finding highlights the need to understand how pathology relates to the gut environment, including the temporal changes in specific gut microbiota observed in COVID-19 patients.

Embolization coil migration to the gastrointestinal tract is a rare complication. This report describes our experience of coil migration in the stomach and spontaneous excretion. A 77-year-old man, who was diagnosed with esophageal squamous cell carcinoma with multiple lymph node metastases, had a bleeding left gastric artery and splenic artery pseudoaneurysm associated with an abdominal lymph node mass, that was treated by coil embolization, after which the coil migrated into the stomach. Because there were no complications such as active bleeding or peritonitis, our patient was followed carefully, and excretion of the coil was documented. No standard management exists for migrated coils. Conservative treatment is an option, as in this case.

Summary: CRISPR-based diagnostics (CRISPR-dx), including the Cas12-based DETECTR and Cas13-based SHERLOCK Class 2 CRISPRs, have been used to detect the presence of DNA or RNA from pathogens, such as the 2009 pandemic influenza virus A (IAV) and the 2019 novel coronavirus SARS-CoV-2. Here, we describe the collateral single-stranded DNA cleavage with Class 1 type I CRISPR-Cas3 and highlight its potential for development as a Cas3-mediated rapid (within 40 min), low-cost, instrument-free detection method for SARS-CoV-2. This assay, which we call Cas3-Operated Nucleic Acid detectioN (CONAN), not only detects SARS-CoV-2 in clinical samples, but also offers specific detection of single-base-pair mutations in IAV variants. This tool allows rapid and accurate point-of-care testing for patients with suspected SARS-CoV-2 or drug-resistant IAV infections in hospitals.

The influenza A(H1N1)pdm09 virus that emerged in 2009 causes seasonal epidemic worldwide. The virus acquired several amino acid substitutions that were responsible for antigenic drift until the 2018–2019 influenza season. Viruses possessing mutations in the NA and PA proteins that cause reduced susceptibility to NA inhibitors and baloxavir marboxil, respectively, have been detected after antiviral treatment, albeit infrequently. Here, we analyzed HA, NA, and PA sequences derived from A(H1N1)pdm09 viruses that were isolated during the 2018–2019 and 2019–2020 influenza seasons in Japan. We found that A(H1N1)pdm09 viruses possessing the D187A and Q189E substitutions in HA emerged and dominated during the 2019–2020 season; these substitutions in the antigenic site Sb, a high potency neutralizing antibody-eliciting site for humans, changed the antigenicity of A(H1N1)pdm09 viruses. Furthermore, we found that isolates possessing the N156K substitution, which was predicted to affect the antigenicity of A(H1N1)pdm09 virus at the laboratory level, were detected at a frequency of 1.0% in the 2018–2019 season but 10.1% in the 2019–2020 season. These findings indicate that two kinds of antigenically drifted viruses—N156K and D187A/Q189E viruses—co-circulated during the 2019–2020 influenza season in Japan.

Malaria parasites cannot multiply in host erythrocytes without cholesterol because they lack complete sterol biosynthesis systems. This suggests parasitized red blood cells (pRBCs) need to capture host sterols, but its mechanism remains unknown. Here we identified a novel high-density lipoprotein (HDL)-delivery pathway operating in blood-stage Plasmodium. In parasitized mouse plasma, exosomes positive for scavenger receptor CD36 and platelet-specific CD41 increased. These CDs were detected in pRBCs and internal parasites. A low molecular antagonist for scavenger receptors, BLT-1, blocked HDL uptake to pRBCs and suppressed Plasmodium growth in vitro. Furthermore, platelet-derived exosomes were internalized in pRBCs. Thus, we presume CD36 is delivered to malaria parasites from platelets by exosomes, which enables parasites to steal HDL for cholesterol supply. Cholesterol needs to cross three membranes (RBC, parasitophorous vacuole and parasite’s plasma membranes) to reach parasite, but our findings can explain the first step of sterol uptake by intracellular parasites.

ABSTRACT Chronic inflammation is a hallmark of human immunodeficiency virus (HIV) infection and a risk factor for the development and progression of age-related comorbidities. Although HIV-associated gut dysbiosis has been suggested to be involved in sustained chronic inflammation, there remains a limited understanding of the association between gut dysbiosis and chronic inflammation during HIV infection. Here, we investigated compositional changes in the gut microbiome and its role in chronic inflammation in patients infected with HIV. We observed that the gut microbiomes of patients with low CD4 counts had reduced alpha diversity compared to those in uninfected controls. Following CD4 recovery, alpha diversity was restored, but intergroup dissimilarity of bacterial composition remained unchanged between patients and uninfected controls. Patients with HIV had higher abundance of the classes Negativicutes, Bacilli, and Coriobacteriia, as well as depletion of the class Clostridia. These relative abundances positively correlated with inflammatory cytokines and negatively correlated with anti-inflammatory cytokines. We found that gut dysbiosis accompanying HIV infection was characterized by a depletion of obligate anaerobic Clostridia and enrichment of facultative anaerobic bacteria, reflecting increased intestinal oxygen levels and intestinal permeability. Furthermore, it is likely that HIV-associated dysbiosis shifts the immunological balance toward inflammatory Th1 responses and encourages proinflammatory cytokine production. Our results suggest that gut dysbiosis contributes to sustaining chronic inflammation in patients with HIV infection despite effective antiretroviral therapy and that correcting gut dysbiosis will be effective in improving long-term outcomes in patients. IMPORTANCE Chronic inflammation is a hallmark of HIV infection and is associated with the development and progression of age-related comorbidities. Although the gastrointestinal tract is a major site of HIV replication and CD4+ T-cell depletion, the role of HIV-associated imbalance of gut microbiome in chronic inflammation is unclear. Here, we aimed to understand the causal relationship between abnormalities in the gut microbiome and chronic inflammation in patients with HIV. Our results suggest HIV-associated gut dysbiosis presents a more aerobic environment than that of healthy individuals, despite prolonged viral suppression. This dysbiosis likely results from a sustained increase in intestinal permeability, which supports sustained bacterial translocation in HIV patients, despite effective therapy. Additionally, we observed that several bacterial taxa enriched in HIV patients were associated with increased expression of inflammatory cytokines. Collectively, these results suggest that gut dysbiosis plays an important role in chronic inflammation in HIV patients.

Although there is strong evidence that SARS-CoV-2 infection is associated with adverse outcomes in certain ethnic groups, the association of disease severity and risk factors such as comorbidities and biomarkers with racial disparities remains undefined. This retrospective study between March 2020 and February 2021 explores COVID-19 risk factors as predictors for patients’ disease progression through country comparison. Disease severity predictors in Germany and Japan were cardiovascular-associated comorbidities, dementia, and age. We adjusted age, sex, body mass index, and history of cardiovascular disease comorbidity in the country cohorts using a propensity score matching (PSM) technique to reduce the influence of differences in sample size and the surprisingly young, lean Japanese cohort. Analysis of the 170 PSM pairs confirmed that 65.29% of German and 85.29% of Japanese patients were in the uncomplicated phase. More German than Japanese patients were admitted in the complicated and critical phase. Ethnic differences were identified in patients without cardiovascular comorbidities. Japanese patients in the uncomplicated phase presented a suppressed inflammatory response and coagulopathy with hypocoagulation. In contrast, German patients exhibited a hyperactive inflammatory response and coagulopathy with hypercoagulation. These differences were less pronounced in patients in the complicated phase or with cardiovascular diseases. Coagulation/fibrinolysis-associated biomarkers rather than inflammatory-related biomarkers predicted disease severity in patients with cardiovascular comorbidities: platelet counts were associated with severe illness in German patients. In contrast, high D-dimer and fibrinogen levels predicted disease severity in Japanese patients. Our comparative study indicates that ethnicity influences COVID-19-associated biomarker expression linked to the inflammatory and coagulation (thrombo-inflammatory) response. Future studies will be necessary to determine whether these differences contributed to the less severe disease progression observed in Japanese COVID-19 patients compared with those in Germany.

The case of a patient with left ventricular assist device (LVAD)-associated endocarditis involving multiple clones of Staphylococcus aureus is presented. Different clones with distinct colony morphology were identified from blood cultures collected on the same day and showed diverse antimicrobial resistance patterns. In addition, a difference in antimicrobial susceptibility was observed even within an identical clone recovered 400 days apart due to the loss of SCCmec for methicillin and modification of the 23S rRNA target site for linezolid during a long-term treatment course. Keywords: Staphylococcus aureus, Left ventricular assist device, Linezolid resistance, Endocarditis, Whole-genome sequencing

Background: To develop an effective vaccine against a novel viral pathogen, it is important to understand the longitudinal antibody responses against its first infection. Here we performed a longitudinal study of antibody responses against SARS-CoV-2 in symptomatic patients. Methods: Sequential blood samples were collected from 39 individuals at various timepoints between 0 and 154 days after onset. IgG or IgM titers to the receptor binding domain (RBD) of the S protein, the ectodomain of the S protein, and the N protein were determined by using an ELISA. Neutralizing antibody titers were measured by using a plaque reduction assay. Findings: The IgG titers to the RBD of the S protein, the ectodomain of the S protein, and the N protein peaked at about 20 days after onset, gradually decreased thereafter, and were maintained for several months after onset. Extrapolation modeling analysis suggested that the IgG antibodies were maintained for this amount of time because the rate of reduction slowed after 30 days post-onset. IgM titers to the RBD decreased rapidly and disappeared in some individuals after 90 days post-onset. All patients, except one, possessed neutralizing antibodies against authentic SARS-CoV-2, which they retained at 90 days after onset. The highest antibody titers in patients with severe infections were higher than those in patients with mild or moderate infections, but the decrease in antibody titer in the severe infection cohort was more remarkable than that in the mild or moderate infection cohort. Interpretation: Although the number of patients is limited, our results show that the antibody response against the first SARS-CoV-2 infection in symptomatic patients is typical of that observed in an acute viral infection. Funding: The Japan Agency for Medical Research and Development and the National Institutes of Allergy and Infectious Diseases.

BackgroundHuman microbiotas are communities of microorganisms living in symbiosis with humans. They play an important role in the host immune response to respiratory viral infection. However, evidence on the human microbiome and coronavirus disease (COVID-19) relationship is insufficient. The aim of this systematic literature review was to evaluate existing evidence on the association between the microbiome and COVID-19 in humans and summarize these data in the pandemic era.MethodsWe conducted a systematic literature review on the association between the microbiome and COVID-19 in humans by searching PubMed, Embase, and the Cochrane Library, CINAHL, and Web of Science databases for articles in English published up to October 31, 2020. The results were analyzed qualitatively. This study is registered with PROSPERO (CRD42020195982).ResultsOf the 543 articles identified by searching databases, 16 in line with the research objectives were eligible for qualitative review: eight sampled the microbiome using stool, four using nasopharyngeal or throat swab, three using bronchoalveolar lavage fluid, and one using lung tissue. Fecal microbiome dysbiosis and increased opportunistic pathogens were reported in COVID-19 patients. Several studies suggested the dysbiosis in the lung microbiome of COVID-19 patients with an abundance of opportunistic pathogens using lower respiratory tract samples. The association between COVID-19 severity and the human microbiome remains uncertain.ConclusionThe human fecal and respiratory tract microbiome changed in COVID-19 patients with opportunistic pathogen abundance. Further research to elucidate the effect of alternation of the human microbiome in disease pathogenesis is warranted.

Viral infections are influenced by various microorganisms in the environment surrounding the target tissue, and the correlation between the type and balance of commensal microbiota is the key to establishment of the infection and pathogenicity. Some commensal microorganisms are known to resist or promote viral infection, while others are involved in pathogenicity. It is also becoming evident that the profile of the commensal microbiota under normal conditions influences the progression of viral diseases. Thus, to understand the pathogenesis underlying viral infections, it is important to elucidate the interactions among viruses, target tissues, and the surrounding environment, including the commensal microbiota, which should have different relationships with each virus. In this review, we outline the role of microorganisms in viral infections. Particularly, we focus on gaining an in-depth understanding of the correlations among viral infections, target tissues, and the surrounding environment, including the commensal microbiota and the gut virome, and discussing the impact of changes in the microbiota (dysbiosis) on the pathological progression of viral infections.

The incidence of nonalcoholic steatohepatitis (NASH) is increasing worldwide, including in Asian countries. We reported that the hepatic expression of bile salt export pump (BSEP) was downregulated in patients with NASH, suggesting that BSEP is involved in the pathogenesis of NASH. To identify the underlying mechanism, we analyzed Bsep heterozygous knock-out (Bsep+/- mice) and wild-type (WT) C57BL/6J mice fed a high-fat diet (HFD) (32.0% animal fat) or normal diet. We examined histological changes, levels of hepatic lipids and hepatic bile acids, and expression of genes related to bile acid and cholesterol metabolism. HFD-fed Bsep+/- mice exhibited milder hepatic steatosis and less weight gain, compared to HFD-fed WT mice. The concentrations of total bile acid, triglycerides, and cholesterols were reduced in the liver of HFD-fed Bsep+/- mice. Regarding hepatic bile acid metabolism, the expression levels of Farnesoid X receptor (Fxr) and Multidrug resistance-associated protein 2 were significantly upregulated in HFD-fed Bsep+/- mice, compared to HFD-fed WT mice. Furthermore, several alterations were observed in upstream cholesterol metabolism in the liver. The expression levels of bile acid metabolism-related genes were also altered in the intestine of HFD-fed Bsep+/- mice. In conclusion, HFD-fed Bsep+/- mice exhibited significant alterations of the expression levels of genes related to bile acid and lipid metabolism in both the liver and ileum, resulting in alleviated steatosis and less weight gain. These results suggest the importance of BSEP for maintenance of bile acid and cholesterol metabolism. Further investigations of the involvement of BSEP in the pathogenesis of NASH will provide greater insight and facilitate the development of novel therapeutic modalities.

Abstract Background HIV-associated neurocognitive disorder (HAND) remains an important and yet potentially underdiagnosed manifestation despite the fact that the modern combination antiretroviral therapy (cART) has achieved effective viral suppression and greatly reduced the incidence of life-threatening events. Although HIV neurotoxicity is thought to play a central role, the potential of viral genetic signature as diagnostic and/or prognostic biomarker has yet to be fully explored. Results Using a manually curated sequence metadataset (80 specimens, 2349 sequences), we demonstrated that only three genetic features are sufficient to predict HAND status regardless of sampling tissues; the accuracy reached 100 and 94% in the hold-out testing subdataset and the entire dataset, respectively. The three genetic features stratified HAND into four distinct clusters. Extrapolating the classification to the 1619 specimens registered in the Los Alamos HIV Sequence Database, the global HAND prevalence was estimated to be 46%, with significant regional variations (30–71%). The R package HANDPrediction was implemented to ensure public availability of key codes. Conclusions Our analysis revealed three amino acid positions in gp120 glycoprotein, providing the basis of the development of novel cART regimens specifically optimized for HAND-associated quasispecies. Moreover, the classifier can readily be translated into a diagnostic biomarker, warranting prospective validation.

Hepatitis A virus (HAV) causes transient acute infection, and little is known of viral shedding via the duodenum and into the intestinal environment, including the gut microbiome, from the period of infection until after the recovery of symptoms. Therefore, in this study, we aimed to comprehensively observe the amount of virus excreted into the intestinal tract, the changes in the intestinal microbiome, and the level of inflammation during the healing process. We used blood and stool specimens from patients with human immunodeficiency virus who were infected with HAV during the HAV outbreak in Japan in 2018. Moreover, we observed changes in fecal HAV RNA and quantified the plasma cytokine level and gut microbiome by 16S rRNA analysis from clinical onset to at least 6 months after healing. HAV was detected from clinical onset up to a period of more than 150 days. Immediately after infection, many pro-inflammatory cytokines were elicited, and some cytokines showed different behaviors. The intestinal microbiome changed significantly after infection (dysbiosis), and the dysbiosis continued for a long time after healing. These observations suggest that the immunocompromised state is associated with prolonged viral shedding into the intestinal tract and delayed recovery of the intestinal environment.

Abstract Background Daptomycin has been reported to cause artificial prolongation of prothrombin time (PT) by interacting with some test reagents of PT. This prolongation was particularly prominent with high concentrations of daptomycin in vitro. However, whether this prolongation is important in clinical settings and the optimal timing to assess PT remain unclear. Methods A prospective clinical study was conducted with patients who received daptomycin for confirmed or suspected drug-resistant, gram-positive bacterial infection at a university hospital in Japan. PT at the peak and trough of daptomycin was tested using nine PT reagents. Linear regression analyses were used to examine the difference in daptomycin concentration and the relative change of PT-international normalized ratios (PT-INR). Results Thirty-five patients received daptomycin (6 mg/kg). The mean ± standard deviation of the trough and peak concentrations of daptomycin were 13.5 ± 6.3 and 55.1 ± 16.9 μg/mL, respectively. Twelve patients (34%) received warfarin. With five PT reagents, a significant proportion of participants experienced prolongation of PT-INR at the daptomycin peak concentration compared to the PT-INR at the trough, although the mean relative change was less than 10%. None of the participants clinically showed any signs of bleeding. A linear, dose-dependent prolongation of PT was observed for one reagent [unadjusted coefficient β 3.1 × 10−3/μg/mL; 95% confidence interval (CI) 2.3 × 10−5–6.3 × 10−3; p = 0.048]. When patients were stratified based on warfarin use, this significant linear relationship was observed in warfarin users for two PT reagents (adjusted coefficient β, 6.4 × 10−3/μg/mL; 95% CI 3.5 × 10−3–9.3 × 10−3; p

Immunodominant T cell epitopes preferentially targeted in multiple individuals are the critical element of successful vaccines and targeted immunotherapies. However, the underlying principles of this “convergence” of adaptive immunity among different individuals remain poorly understood. To quantitatively describe epitope immunogenicity, here we propose a supervised machine learning framework generating probabilistic estimates of immunogenicity, termed “immunogenicity scores,” based on the numerical features computed through sequence-based simulation approximating the molecular scanning process of peptides presented onto major histocompatibility complex (MHC) by the human T cell receptor (TCR) repertoire. Notably, overlapping sets of intermolecular interaction parameters were commonly utilized in MHC-I and MHC-II prediction. Moreover, a similar simulation of individual TCR-peptide interaction using the same set of interaction parameters yielded correlates of TCR affinity. Pathogen-derived epitopes and tumor-associated epitopes with positive T cell reactivity generally had higher immunogenicity scores than non-immunogenic counterparts, whereas thymically expressed self-epitopes were assigned relatively low scores regardless of their immunogenicity annotation. Immunogenicity score dynamics among single amino acid mutants delineated the landscape of position- and residue-specific mutational impacts. Simulation of position-specific immunogenicity score dynamics detected residues with high escape potential in multiple epitopes, consistent with known escape mutations in the literature. This study indicates that targeting of epitopes by human adaptive immunity is to some extent directed by defined thermodynamic principles. The proposed framework also has a practical implication in that it may enable to more efficiently prioritize epitope candidates highly prone to T cell recognition in multiple individuals, warranting prospective validation across different cohorts.

Reverse transcription-quantitative PCR (RT-qPCR)-based tests are widely used to diagnose coronavirus disease 2019 (COVID-19). As a result that these tests cannot be done in local clinics where RT-qPCR testing capability is lacking, rapid antigen tests (RATs) for COVID-19 based on lateral flow immunoassays are used for rapid diagnosis. However, their sensitivity compared with each other and with RT-qPCR and infectious virus isolation has not been examined. Here, we compared the sensitivity among four RATs by using severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) isolates and several types of COVID-19 patient specimens and compared their sensitivity with that of RT-qPCR and infectious virus isolation. Although the RATs read the samples containing large amounts of virus as positive, even the most sensitive RAT read the samples containing small amounts of virus as negative. Moreover, all RATs tested failed to detect viral antigens in several specimens from which the virus was isolated. The current RATs will likely miss some COVID-19 patients who are shedding infectious SARS-CoV-2.

The hepatitis C virus nonstructural protein NS5A is involved in resistance to the host immune response, as well as the viral lifecycle such as replication and maturation. Here, we established transgenic mice expressing NS5A protein in the liver and examined innate immune responses against lipopolysaccharide (LPS) in vivo. Intrahepatic gene expression levels of cytokines such as interleukin-6, tumor necrosis factor-α, and interferon-γ were significantly suppressed after LPS injection in the transgenic mouse liver. Induction of the C-C motif chemokine ligand 2, 4, and 5 was also suppressed. Phosphorylation of the signal transducer and activator of transcription 3, which is activated by cytokines, was also reduced, and expression levels of interferon-stimulated genes, 2'-5' oligoadenylate synthase, interferon-inducible double-stranded RNA-activated protein kinase, and myxovirus resistance 1 were similarly suppressed. Since LPS binds to toll-like receptor 4 and stimulates the downstream pathway leading to induction of these genes, we examined the extracellular signal-regulated kinase and IκB-α. The phosphorylation levels of these molecules were reduced in transgenic mouse liver, indicating that the pathway upstream of the molecules was disrupted by NS5A. Further analyses revealed that the interaction between interleukin-1 receptor-associated kinase-1 and tumor necrosis factor receptor associated factor-6 was dispersed in transgenic mice, suggesting that NS5A may interfere with this interaction via myeloid differentiation primary response gene 88, which was shown to interact with NS5A. Since the gut microbiota, a source of LPS, is known to be associated with pathological conditions in liver diseases, our results suggest the involvement of NS5A in the pathogenesis of HCV infected-liver via the suppression of innate immunity.

Pre-existing low-frequency resistance-associated variants (RAVs) may jeopardize successful sustained virological responses (SVR) to HCV treatment with direct-acting antivirals (DAAs). However, the potential impact of low-frequency (∼0.1%) mutations, concatenated mutations (haplotypes), and their association with genotypes (Gts) on the treatment outcome has not yet been elucidated, most probably owing to the difficulty in detecting pre-existing minor haplotypes with sufficient length and accuracy. Herein, we characterize a methodological framework based on Illumina MiSeq next-generation sequencing (NGS) coupled with bioinformatics of quasispecies reconstruction (QSR) to realize highly accurate variant calling and genotype-haplotype detection. The core-to-NS3 protease coding sequences in 10 HCV monoinfected patients, 5 of whom had a history of blood transfusion, and 11 HCV/HIV coinfected patients with hemophilia, were studied. Simulation experiments showed that, for minor variants constituting more than 1%, our framework achieved a positive predictive value (PPV) of 100% and sensitivities of 91.7-100% for genotyping and 80.6% for RAV screening. Genotyping analysis indicated the prevalence of dominant Gt1a infection in coinfected patients (6/11 vs 0/10, p = 0.01). For clinical samples, minor genotype overlapping infection was prevalent in HCV/HIV coinfected hemophiliacs (10/11) and patients who experienced whole-blood transfusion (4/5) but none in patients without exposure to blood (0/5). As for RAV screening, the Q80K/R and S122K/R variants were particularly prevalent among minor RAVs observed, detected in 12/21 and 6/21 cases, respectively. Q80K was detected only in coinfected patients, whereas Q80R was predominantly detected in monoinfected patients (1/11 vs 7/10, p < 0.01). Multivariate interdependence analysis revealed the previously unrecognized prevalence of Gt1b-Q80K, in HCV/HIV coinfected hemophiliacs [Odds ratio = 13.4 (3.48-51.9), p < 0.01]. Our study revealed the distinct characteristics of viral quasispecies between the subgroups specified above and the feasibility of NGS and QSR-based genetic deconvolution of pre-existing minor Gts, RAVs, and their interrelationships.

We report a patient in Japan infected with Cryptococcus gattii genotype VGIIa who had no recent history of travel to disease-endemic areas. This strain was identical to the Vancouver Island outbreak strain R265. Our results suggest that this virulent strain has spread to regions outside North America.