Prevalence of yaws and syphilis in the Ashanti region of Ghana and occurrence of H. ducreyi, herpes simplex virus 1 and herpes simplex virus 2 in skin lesions associated with treponematoses.

Yaws affects children in tropical regions, while syphilis primarily affects sexually active adults worldwide. Despite various campaigns towards the eradication of yaws and elimination of syphilis, these two diseases are still present in Ghana. The aetiological agents of both diseases, two Treponema pallidum subspecies, are genetically similar. This study aimed to assess the prevalence of these treponematoses and the occurrence of pathogens causing similar skin lesions in the Ashanti region of Ghana. A point-of-care test was used to determine the seroprevalence of the treponematoses. Both yaws and syphilis were identified in the Ashanti region of Ghana. Multiplex PCR was used to identify treponemes and other pathogens that cause similar skin lesions. The results indicated that the seroprevalences of T. pallidum in individuals with yaws-like and syphilis-like lesions were 17.2% and 10.8%, respectively. Multiplex PCR results showed that 9.1%, 1.8% and 0.9% of yaws-like lesions were positive for Haemophilus ducreyi, herpes simplex virus-1 (HSV-1) and T. pallidum respectively. Among syphilis-like lesions, 28.3% were positive for herpes simplex virus -2 (HSV-2) by PCR. To our knowledge, this is the first time HSV-I and HSV-2 have been reported from yaws-like and syphilis-like lesions, respectively, in Ghana. The presence of other organisms apart from T. pallidum in yaws-like and syphilis-like lesions could impede the total healing of these lesions and the full recovery of patients. This may complicate efforts to achieve yaws eradication by 2030 and the elimination of syphilis and warrants updated empirical treatment guidelines for skin ulcer diseases.

Pro-angiogenic and antibacterial copper containing nanoparticles in PLGA/amorphous calcium phosphate bone nanocomposites.

Large bone defects after trauma demand for adequate bone substitutes. Bone void fillers should be antibacterial and pro-angiogenic. One viable option is the use of composite materials like the combination of PLGA and amorphous calcium phosphate (aCaP). Copper stimulates angiogenesis and has antibacterial qualities. Either copper oxide (CuO) nanoparticles (NPs) were therefore added to PLGA/aCaP/CuO in different concentrations (1, 5 and 10 w/w %) or copper-doped tricalcium phosphate NPs (TCP with 2% of copper) were electrospun into PLGA/CuTCP nanocomposites. Bi-layered nanocomposites of PLGA/aCaP with different copper NPs (CuO or TCP) and a second layer of pristine PLGA were fabricated. Two clinical bacterial isolates (Staphylococcus aureus and Staphylococcus epidermidis) were used to assess antibacterial properties of the copper-containing materials. For angiogenesis, the chorioallantoic membrane (CAM) assay of the chicken embryo was performed. The higher the CuO content, the higher were the antibacterial properties, with 10 % CuO reducing bacterial adhesion most effectively. Vessel and cell densities were highest in the 5 % CuO containing scaffolds, while tissue integration was more pronounced at lower CuO content. The PLGA/aCaP/CuO (1 % CuO) behaved similar like PLGA/CuTCP in all angiogenic and antibacterial readouts, based on the same copper fraction. We conclude that CuO NPs or CuTCP NPs are useful components to increase angiogenic properties of nanocomposites and at the same time exhibiting antibacterial characteristics.

Decreased skin colonization with Malassezia spp. and increased skin colonization with Candida spp. in patients with severe atopic dermatitis.

Background: Atopic dermatitis (AD) is a chronic relapsing inflammatory skin disease in which patients are sensitized towards a plethora of allergens. The hosts fungal microbiota, the mycobiota, that is believed to be altered in patients suffering from AD acts as such an allergen. The correlation context of specific sensitization, changes in mycobiota and its impact on disease severity however remains poorly understood. Objectives: We aim to enhance the understanding of the specific sensitization towards the mycobiota in AD patients in relation to their fungal skin colonization. Methods: Sensitization pattern towards the Malassezia spp. and Candida albicans of 16 AD patients and 14 healthy controls (HC) were analyzed with the newly developed multiplex-assay ALEX2® and the established singleplex-assay ImmunoCAP®. We compared these findings with the fungal skin colonization analyzed by DNA sequencing of the internal transcribed spacer region 1 (ITS1). Results: Sensitization in general and towards Malassezia spp. and C. albicans is increased in AD patients compared to HC with a quantitative difference in severe AD when compared to mild to moderate AD. Further we saw an association between sensitization towards and skin colonization with Candida spp. yet a negative correlation between sensitization towards and skin colonization with Malassezia spp. Conclusion: We conclude that AD in general and severe AD in particular is associated with increased sensitization towards the hosts own mycobiota. There is positive correlation in Candida spp. skin colonization and negative in Malassezia spp. skin colonization when compared to AD, AD severity as well as to specific sensitization patterns.

Multicentric evaluation of a specific intrathecal anti-Treponema pallidum IgG index as a diagnostic biomarker of neurosyphilis: results from a retro-prospective case-control study.

BACKGROUND AND OBJECTIVES: The diagnosis of neurosyphilis (NS) lacks a true 'gold standard', making the diagnosis challenging while consequences of a misdiagnosis are potentially severe. The aim of this study was to evaluate the diagnostic performance of measuring an antibody index (AI) for the intrathecal synthesis of specific anti-Treponema pallidum (T. pallidum) IgG for the diagnosis of NS. METHODS: Specific anti-T. pallidum IgG were measured simultaneously in paired cerebrospinal fluid (CSF)-serum samples collected retrospectively and prospectively between 2007 and 2022, from patients suspected of NS, in Switzerland. An AI was calculated to account for blood-brain barrier integrity. Area under the receiver operating characteristic curve, sensitivity/specificity and positive/negative predictive values of AI test were estimated. Two NS definitions were used: NS1 included patients with NS suspicion presenting with neurological symptoms and/or acute neurosensory signs, and positive T. Pallidum Hemagglutinations Assay (TPHA)/T. pallidum particle agglutination assay (TPPA) serology and CSF-TPHA/TPPA ≥320, and either CSF-leucocytes >5 cells/mm3 and/or CSF-protein >0.45 g/L and/or a reactive CSF-venereal disease research laboratory (VDRL)/rapid plasma reagin (RPR) test. NS2 included patients with suspected NS presenting with acute ocular and/or otologic symptoms, and positive TPHA/TPPA serology, and a favourable response to NS treatment. Controls were patients diagnosed with any other central nervous system (CNS) pathologies and with positive TPHA/TPPA serology. RESULTS: The study included 71 NS (43 NS1 and 28 NS2) and 110 controls. With a threshold of ≥1.7, sensitivity and specificity of the specific AI test were 90.7% (CI 77.7 to 97.4) and 100% (CI 96.7 to 100.0), respectively, for NS1 and 14.3% (CI 4 to 32.7) and 100% (CI 96.7 to 100.0) for NS2. In patients suspected of NS with a CNS involvement (NS1 group), NS could be confirmed by the positivity of this specific AI. CONCLUSIONS: Measurement of an intrathecal synthesis index of specific anti-T. pallidum IgG in patients with CSF inflammatory signs appears to be a valuable diagnostic test. However, in otic or ocular syphilis, presenting few CSF abnormalities, AI is not sufficient alone to confirm NS diagnosis. TRIAL REGISTRATION: Swiss Association of Research Ethics Committees number 2019-00232.

Incidence of sexually transmitted infections and association with behavioural factors: Time-to-event analysis of a large pre-exposure prophylaxis (PrEP) cohort.

OBJECTIVES: Our objective was to obtain long-term data on the incidence of sexually transmitted infections (STIs) and their association with behavioural factors after widespread pre-exposure prophylaxis (PrEP) implementation. METHODS: This was a time-to-event analysis of a national PrEP cohort in Switzerland (SwissPrEPared study). Participants were people without HIV interested in taking PrEP with at least two STI screening visits. Primary outcomes were incidence rate of gonorrhoea, chlamydia, and syphilis. The association between behavioural factors and STI diagnosis was expressed using hazard ratios. We adjusted for testing frequency and calendar year. RESULTS: This analysis included 3907 participants enrolled between April 2019 and April 2022, yielding 3815.7 person-years of follow-up for gonorrhoea (15 134 screenings), 3802.5 for chlamydia (15 141 screenings), and 3858.6 for syphilis (15 001 screenings). The median age was 39 years (interquartile range [IQR] 32-47), 93.8% (n = 3664) identified as men who have sex with men (MSM). The incidence was 22.8 (95% confidence interval [CI] 21.3-24.4) per 100 person-years for gonorrhoea, 26.3 (95% CI 24.7-28.0) for chlamydia, and 4.4 (95% CI 3.8-5.1) for syphilis. Yearly incidence rates decreased between 2019 (all bacterial STIs: 81.6; 95% CI 59.1-109.9) and 2022 (all bacterial STIs: 49.8; 95% CI 44.6-55.3). Participants reporting chemsex substance use were at higher risk of incident STIs, as were those reporting multiple sexual partners. Younger age was associated with a higher risk of gonorrhoea and chlamydia. CONCLUSIONS: Incidence rates of bacterial STIs decreased over time. Young MSM, those with multiple partners, and those using chemsex substances were at increased risk of STIs.

Congenital syphilis in Switzerland: a retrospective cohort study, 2010 to 2019.

AIMS OF THE STUDY: We previously reported a re-emergence of syphilis from 2006 to 2009 with detection of congenital syphilis in Switzerland. This study aimed to reassess the incidence of children exposed to maternal syphilis during pregnancy and congenital syphilis in a following 10-year period in the canton of Zurich, the most populous canton in Switzerland with the highest incidences of syphilis. METHODS: Children were identified both by reviewing medical records at the four major neonatal and paediatric hospitals providing acute care in the canton of Zurich and by the serological database of the syphilis reference laboratory. Inclusion criteria for children were (a) date of birth in the period 2010-2019, (b) place of birth in the canton of Zurich, (c) evaluation for syphilis due to positive syphilis pregnancy screening and (d) age <1 year at diagnosis. Results were compared with epidemiological data provided by the Federal Office of Public Health (FOPH). RESULTS: We identified and evaluated 17 children after potential exposure to maternal syphilis. Residual antibodies of a past infection were found in 11 mothers. Six children were identified as having had real exposure to asymptomatic maternal syphilis. From an epidemiological perspective, the distribution of the cases followed a similar pattern as confirmed syphilis cases in women of childbearing age reported to the FOPH. No cases of congenital syphilis were observed. CONCLUSIONS: In contrast to the rise in syphilis infections, this study identified no cases of congenital syphilis in the canton of Zurich, Switzerland, in the period 2010-2019. Syphilis pregnancy screening may have prevented congenital syphilis by diagnosing and allowing adequate treatment of asymptomatic maternal syphilis.

Anti-Treponema pallidum IgA response as a potential diagnostic marker of syphilis.

OBJECTIVES: Serological tests for syphilis detect mainly total Ig, IgM or IgG antibodies. We aimed to evaluate the specific IgA response in syphilis patients according to disease stage. METHODS: A serum IgA-enzyme immunoassay was developed using commercially available microplates coated with recombinant treponemal antigens and an anti-IgA-conjugate. To define a cut-off, we used 91 syphilis positive and 136 negative sera previously defined by the rapid plasma reagin and the Treponema pallidum particle agglutination results. Then we determined the intra- and inter-assay precisions, diagnostic sensitivity according to the clinical stage (in 66, 55 and 42 sera from primary, secondary and latent syphilis patients, respectively) and specificity (in 211 sera from people with conditions different to syphilis). IgA values were further measured in 71 sera from patients with previously treated syphilis. RESULTS: The newly developed IgA-enzyme immunoassay showed a good discrimination between negative and positive samples with intra- and inter-assay variation coefficients <20%. The sensitivity was 80.3% (95% CI, 70.0-90.6), 100.0% (95% CI, 99.1-100.0) and 95.2% (95% CI, 87.6-100.0) in primary, secondary and latent syphilis, respectively, and the specificity was 98.1% (95% CI, 96.0-100.0). Further, IgA values were negative in 61.3% (38/62) of patients with previously treated syphilis. DISCUSSION: Our findings suggest serum IgA as a sensitive and specific marker of syphilis and its detection could be used as a screening assay for active infection. Further evaluation is needed in prospective longitudinal field studies.

S1 Guideline onychomycosis.

Onychomycosis is a fungal infection of the fingernails and toenails. In Europe, tinea unguium is mainly caused by dermatophytes. The diagnostic workup comprises microscopic examination, culture and/or molecular testing (nail scrapings). Local treatment with antifungal nail polish is recommended for mild or moderate nail infections. In case of moderate to severe onychomycosis, oral treatment is recommended (in the absence of contraindications). Treatment should consist of topical and systemic agents. The aim of this update of the German S1 guideline is to simplify the selection and implementation of appropriate diagnostics and treatment. The guideline was based on current international guidelines and the results of a literature review conducted by the experts of the guideline committee. This multidisciplinary committee consisted of representatives from the German Society of Dermatology (DDG), the German-Speaking Mycological Society (DMykG), the Association of German Dermatologists (BVDD), the German Society for Hygiene and Microbiology (DGHM), the German Society of Pediatric and Adolescent Medicine (DGKJ), the Working Group for Pediatric Dermatology (APD) and the German Society for Pediatric Infectious Diseases (DGPI). The Division of Evidence-based Medicine (dEBM) provided methodological assistance. The guideline was approved by the participating medical societies following a comprehensive internal and external review.

EAACI Molecular Allergology User's Guide 2.0.

Since the discovery of immunoglobulin E (IgE) as a mediator of allergic diseases in 1967, our knowledge about the immunological mechanisms of IgE-mediated allergies has remarkably increased. In addition to understanding the immune response and clinical symptoms, allergy diagnosis and management depend strongly on the precise identification of the elicitors of the IgE-mediated allergic reaction. In the past four decades, innovations in bioscience and technology have facilitated the identification and production of well-defined, highly pure molecules for component-resolved diagnosis (CRD), allowing a personalized diagnosis and management of the allergic disease for individual patients. The first edition of the "EAACI Molecular Allergology User's Guide" (MAUG) in 2016 rapidly became a key reference for clinicians, scientists, and interested readers with a background in allergology, immunology, biology, and medicine. Nevertheless, the field of molecular allergology is moving fast, and after 6 years, a new EAACI Taskforce was established to provide an updated document. The Molecular Allergology User's Guide 2.0 summarizes state-of-the-art information on allergen molecules, their clinical relevance, and their application in diagnostic algorithms for clinical practice. It is designed for both, clinicians and scientists, guiding health care professionals through the overwhelming list of different allergen molecules available for testing. Further, it provides diagnostic algorithms on the clinical relevance of allergenic molecules and gives an overview of their biology, the basic mechanisms of test formats, and the application of tests to measure allergen exposure.

Effective Biofilm Eradication on Orthopedic Implants with Methylene Blue Based Antimicrobial Photodynamic Therapy In Vitro.

Periprosthetic joint infections (PJI) are difficult to treat due to biofilm formation on implant surfaces, often requiring removal or exchange of prostheses along with long-lasting antibiotic treatment. This in vitro study investigated the effect of methylene blue photodynamic therapy (MB-PDT) on PJI-causing biofilms on different implant materials. MB-PDT (664 nm LED, 15 J/cm2) was tested on different Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli and Cutibacterium acnes strains in both planktonic form and grown in early and mature biofilms on prosthetic materials (polyethylene, titanium alloys, cobalt-chrome-based alloys, and bone cement). The minimum bactericidal concentration with 100% killing (MBC100%) was determined. Chemical and topographical alterations were investigated on the prosthesis surfaces after MB-PDT. Results showed a MBC100% of 0.5-5 µg/mL for planktonic bacteria and 50-100 µg/mL for bacteria in biofilms-independent of the tested strain, the orthopedic material, or the maturity of the biofilm. Material testing showed no relevant surface modification. MB-PDT effectively eradicated common PJI pathogens on arthroplasty materials without damage to the materials, suggesting that MB-PDT could be used as a novel treatment method, replacing current, more invasive approaches and potentially shortening the antibiotic treatment in PJI. This would improve quality of life and reduce morbidity, mortality, and high health-care costs.

Swiss public health measures associated with reduced SARS-CoV-2 transmission using genome data.

Genome sequences from evolving infectious pathogens allow quantification of case introductions and local transmission dynamics. We sequenced 11,357 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes from Switzerland in 2020-the sixth largest effort globally. Using a representative subset of these data, we estimated viral introductions to Switzerland and their persistence over the course of 2020. We contrasted these estimates with simple null models representing the absence of certain public health measures. We show that Switzerland's border closures decoupled case introductions from incidence in neighboring countries. Under a simple model, we estimate an 86 to 98% reduction in introductions during Switzerland's strictest border closures. Furthermore, the Swiss 2020 partial lockdown roughly halved the time for sampled introductions to die out. Last, we quantified local transmission dynamics once introductions into Switzerland occurred using a phylodynamic model. We found that transmission slowed 35 to 63% upon outbreak detection in summer 2020 but not in fall. This finding may indicate successful contact tracing over summer before overburdening in fall. The study highlights the added value of genome sequencing data for understanding transmission dynamics.

Fast and Sensitive Multiplex Real-Time Quantitative PCR to Detect Cutibacterium Periprosthetic Joint Infections.

Diagnosis of Cutibacterium periprosthetic joint infections (PJIs) is challenging due to a long cultivation time of up to 14 days. Faster culture-independent diagnosis would improve patient care with early and accurate treatment. Specific primers and probes were designed for Cutibacterium acnes, Cutibacterium avidum, and Cutibacterium granulosum and evaluated in a multiplex TaqMan real-time quantitative PCR (qPCR) format on 57 skin swabs and 20 culture-negative cerebrospinal fluid samples. The multiplex qPCR was tested in a PJI cohort of 41 sonication fluid samples from removed implants infected with different pathogens. All five culture-positive Cutibacterium PJIs were detected with the corresponding Cutibacterium-specific probe (100% positive percent agreement). The multiplex qPCR additionally detected C. avidum in two PJI sonication fluid samples that were diagnosed as Staphylococcus species infections according to culture (95% negative percent agreement). The new multiplex qPCR can provide a Cutibacterium PJI diagnosis within 1 day, allowing early and accurate antibiotic treatment. A prospective diagnostic trial in PJI with a high number of Cutibacterium species infections (shoulder PJI) is needed for further evaluation.

Longitudinal Characterization of the Fungal Skin Microbiota in Healthy Subjects Over a Period of 1 Year.

Beneficial microorganisms on the skin contribute to the first line of defense against attacking pathogens. However, instability of the skin microbiota is associated with skin diseases. Hence, temporal analyses are crucial because they serve as a baseline to understand the development of dysbiosis in disease. In this study, we aim to improve the understanding of the fungal skin microbiota, the mycobiota, in healthy subjects. Skin swabs were taken monthly for a year from four different skin sites, that is, antecubital crease, dorsal neck, glabella, and vertex, and analyzed by DNA sequencing of the internal transcribed spacer 1 region. The mycobiota on the skin was dominated by the class Malasseziomycetes, and the core community was composed of Malassezia restricta, M. globosa, and M. sympodialis at all skin sites. Over the period of 1 year, the intrapersonal mycobiota remained largely stable, with some fluctuations of low abundant non-Malassezia fungi. We conclude that despite fluctuations of low abundant classes, fungal skin communities form a temporally robust and individual fingerprint in healthy subjects.

Clinical presentation and serological diagnosis of syphilis reinfection in men living with HIV: a retrospective cohort study.

OBJECTIVES: Studies on the characteristics of syphilis reinfection are scarce despite increasing numbers and proportions of cases. We aimed to gain insights into the clinical and serological presentation of reinfected men living with HIV and to evaluate diagnostic criteria for syphilis reinfection. METHODS: We conducted a retrospective cohort study of 259 HIV-positive men diagnosed with syphilis between January 1999 and September 2015 at the University Hospital Zurich. We compared patients with a single syphilis infection (n=109) to patients with reinfections (n=150). RESULTS: The two groups matched in age, sexual orientation and numbers of other STIs. Reinfected patients more often presented with latent syphilis than patients with a single syphilis episode (41.9% vs 8.9%; p<0.001). Although generally high venereal diseases research laboratory (VDRL) or rapid plasma reagin (RPR) titres (median 1:32) were seen in reinfected patients, 19.4% had titres ≤1:8. Treponema pallidum passive particle agglutination (TPPA) titres were significantly higher (1:81 840 vs 1:10 240; p<0.001), while IgM values were significantly lower (1.27 vs 3.5; p<0.001) in syphilis reinfections than in first infections. The TPPA increased ≥fourfold in >92.3% of reinfected patients. CONCLUSIONS: Our data highlight the paramount importance of regularly screening patients at risk as syphilis reinfections in men living with HIV are more likely to be latent infections, that is, without symptoms. As non-treponemal tests might be biologically false-positive (up to a titre of 1:8) due to various conditions, a ≥fourfold increase of the TPPA might be considered as optional criterion for the diagnosis of syphilis reinfections. This could be especially valuable for diagnosing reinfected latent stage patients.

Evolutionary Processes in the Emergence and Recent Spread of the Syphilis Agent, Treponema pallidum.

The incidence of syphilis has risen worldwide in the last decade in spite of being an easily treated infection. The causative agent of this sexually transmitted disease is the bacterium Treponema pallidum subspecies pallidum (TPA), very closely related to subsp. pertenue (TPE) and endemicum (TEN), responsible for the human treponematoses yaws and bejel, respectively. Although much focus has been placed on the question of the spatial and temporary origins of TPA, the processes driving the evolution and epidemiological spread of TPA since its divergence from TPE and TEN are not well understood. Here, we investigate the effects of recombination and selection as forces of genetic diversity and differentiation acting during the evolution of T. pallidum subspecies. Using a custom-tailored procedure, named phylogenetic incongruence method, with 75 complete genome sequences, we found strong evidence for recombination among the T. pallidum subspecies, involving 12 genes and 21 events. In most cases, only one recombination event per gene was detected and all but one event corresponded to intersubspecies transfers, from TPE/TEN to TPA. We found a clear signal of natural selection acting on the recombinant genes, which is more intense in their recombinant regions. The phylogenetic location of the recombination events detected and the functional role of the genes with signals of positive selection suggest that these evolutionary processes had a key role in the evolution and recent expansion of the syphilis bacteria and significant implications for the selection of vaccine candidates and the design of a broadly protective syphilis vaccine.

A Comparative Study of Real-Time RT-PCR-Based SARS-CoV-2 Detection Methods and Its Application to Human-Derived and Surface Swabbed Material.

Real-time RT-PCR remains a gold standard in the detection of various viral diseases. In the coronavirus 2019 pandemic, multiple RT-PCR-based tests were developed to screen for viral infection. As an emergency response to increasing testing demand, we established a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) PCR diagnostics platform for which we compared different commercial and in-house RT-PCR protocols. Four commercial, one customized, and one in-house RT-PCR protocols were evaluated with 92 SARS-CoV-2-positive and 92 SARS-CoV-2-negative samples. Furthermore, economical and practical characteristics of these protocols were compared. In addition, a highly sensitive digital droplet PCR (ddPCR) method was developed, and application of RT-PCR and ddPCR methods on SARS-CoV-2 environmental samples was examined. Very low limits of detection (1 or 2 viral copies/µL), high sensitivities (93.6% to 97.8%), and high specificities (98.7% to 100%) for the tested RT-PCR protocols were found. Furthermore, the feasibility of downscaling two of the commercial protocols, which could optimize testing capacity, was demonstrated. Tested commercial and customized RT-PCR detection kits show very good and comparable sensitivity and specificity, and the kits could be further optimized for use on SARS-CoV-2 viral samples derived from human and surface swabbed samples.

Prolonged Unfrozen Storage and Repeated Freeze-Thawing of SARS-CoV-2 Patient Samples Have Minor Effects on SARS-CoV-2 Detectability by RT-PCR.

Reliable transportation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) patient samples from a swabbing station to a diagnostics facility is essential for accurate results. Therefore, cooling or freezing the samples is recommended in case of longer transportation times. In this study, SARS-CoV-2 detectability by RT-PCR was assessed after prolonged unfrozen storage or repetitive freeze-thawing of SARS-CoV-2 samples. SARS-CoV-2-positive patient swabs stored in viral transport medium were exposed to different temperatures (4°C, 25°C, and 35°C) and to repetitive freeze-thawing, to assess the effect of storage conditions on RT-PCR detection. SARS-CoV-2 RNA was still reliably detected by RT-PCR after 21 days of storage in viral transport medium, even when the samples had been stored at 35°C. The maximum observed change in cycle threshold value per day was 0.046 (±0.019) at 35°C, and the maximum observed change in cycle threshold value per freeze-thaw cycle per day was 0.197 (±0.06). Compared with storage at 4°C, viral RNA levels deviated little but significantly when stored at 25°C or 35°C, or after repeated freeze-thawing. The results of this study indicate that viral RNA levels are relatively stable at higher temperatures and repetitive freeze-thawing.

Systemic and mucosal antibody responses specific to SARS-CoV-2 during mild versus severe COVID-19.

BACKGROUND: Whereas severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific antibody tests are increasingly being used to estimate the prevalence of SARS-CoV-2 infection, the determinants of these antibody responses remain unclear. OBJECTIVES: Our aim was to evaluate systemic and mucosal antibody responses toward SARS-CoV-2 in mild versus severe coronavirus disease 2019 (COVID-19) cases. METHODS: Using immunoassays specific for SARS-CoV-2 spike proteins, we determined SARS-CoV-2-specific IgA and IgG in sera and mucosal fluids of 2 cohorts, including SARS-CoV-2 PCR-positive patients (n = 64) and PCR-positive and PCR-negtive health care workers (n = 109). RESULTS: SARS-CoV-2-specific serum IgA titers in patients with mild COVID-19 were often transiently positive, whereas serum IgG titers remained negative or became positive 12 to 14 days after symptom onset. Conversely, patients with severe COVID-19 showed a highly significant increase of SARS-CoV-2-specific serum IgA and IgG titers after symptom onset. Very high titers of SARS-CoV-2-specific serum IgA were correlated with severe acute respiratory distress syndrome. Interestingly, some health care workers with negative SARS-CoV-2-specific serum antibody titers showed SARS-CoV-2-specific IgA in mucosal fluids with virus-neutralizing capacity in some cases. SARS-CoV-2-specific IgA titers in nasal fluids were inversely correlated with age. CONCLUSIONS: Systemic antibody production against SARS-CoV-2 develops mainly in patients with severe COVID-19, with very high IgA titers seen in patients with severe acute respiratory distress syndrome, whereas mild disease may be associated with transient production of SARS-CoV-2-specific antibodies but may stimulate mucosal SARS-CoV-2-specific IgA secretion.