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Marine recruits training at Parris Island experienced an unexpectedly high rate of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, despite preventive measures including a supervised, 2-week, pre-entry quarantine. We characterize SARS-CoV-2 transmission in this cohort.Between May and November 2020, we monitored 2,469 unvaccinated, mostly male, Marine recruits prospectively during basic training. If participants tested negative for SARS-CoV-2 by quantitative polymerase chain reaction (qPCR) at the end of quarantine, they were transferred to the training site in segregated companies and underwent biweekly testing for 6 weeks. We assessed the effects of coronavirus disease 2019 (COVID-19) prevention measures on other respiratory infections with passive surveillance data, performed phylogenetic analysis, and modeled transmission dynamics and testing regimens.Preventive measures were associated with drastically lower rates of other respiratory illnesses. However, among the trainees, 1,107 (44.8%) tested SARS-CoV-2-positive, with either mild or no symptoms. Phylogenetic analysis of viral genomes from 580 participants revealed that all cases but one were linked to five independent introductions, each characterized by accumulation of mutations across and within companies, and similar viral isolates in individuals from the same company. Variation in company transmission rates (mean reproduction number R 0 ; 5.5 [95% confidence interval [CI], 5.0, 6.1]) could be accounted for by multiple initial cases within a company and superspreader events. Simulations indicate that frequent rapid-report testing with case isolation may minimize outbreaks.Transmission of wild-type SARS-CoV-2 among Marine recruits was approximately twice that seen in the community. Insights from SARS-CoV-2 outbreak dynamics and mutations spread in a remote, congregate setting may inform effective mitigation strategies.

Rapid coronavirus disease 2019 (COVID-19) antigen tests can be used to aid in quickly identifying positive cases, which can help mitigate the spread of COVID-19 infection. Using previously characterized Omicron-positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), non-Omicron-positive SARS-CoV-2, and negative samples, we evaluated five brands of at-home rapid COVID-19 antigen tests (On/Go at-home COVID-19 rapid antigen self-test, iHealth COVID-19 antigen rapid test, QuickVue SARS antigen test, Abbott BinaxNOW COVID-19 card home test, and InBios SCoV-2 Ag detect rapid self-test). Our results showed that these rapid tests had similar levels of sensitivity to Omicron and non-Omicron variants (On/Go, 76.4% and 71.0%; iHealth, 73.0% and 71.0%; QuickVue, 84.3% and 74.3%; BinaxNOW, 69.7% and 71.0%; and InBios, 66.3% and 64.5%, respectively). In conclusion, rapid COVID-19 antigen tests can continue to be used as part of public health measures to combat the spread of the Omicron variant, as their sensitivity was not significantly affected.

We investigated the temporal profile of multiple components of the serological response after asymptomatic or mildly symptomatic SARS-CoV-2 infection, in a cohort of 67 previously SARS-CoV-2 naive young adults, up to 8.5 months after infection. We found a significant decrease of spike IgG and neutralization antibody titers from early (11 to 56 days) to late (4 to 8.5 months) time points postinfection. Over the study period, S1-specific IgG levels declined significantly faster than that of the S2-specific IgG. Further, serum antibodies from PCR-confirmed participants cross-recognized S2, but not S1, of the betacoronaviruses HKU1 and OC43, suggesting a greater degree of cross-reactivity of S2 among betacoronaviruses. Antibody-Dependent Natural Killer cell Activation (ADNKA) was detected at the early time point but significantly decreased at the late time point. Induction of serum Antibody-Dependent Monocyte Phagocytosis (ADMP) was detected in all the infected participants, and its levels remained stable over time. Additionally, a reduced percentage of participants had detectable neutralizing activity against the Beta (50%), Gamma (61 to 67%), and Delta (90 to 94%) variants, both early and late postinfection, compared to the ancestral strain (100%). Antibody binding to S1 and RBD of Beta, Gamma, Delta (1.7 to 2.3-fold decrease), and Omicron (10 to 16-fold decrease) variants was also significantly reduced compared to the ancestral SARS-CoV-2 strain. Overall, we found variable temporal profiles of specific components and functionality of the serological response to SARS-CoV-2 in young adults, which is characterized by lasting, but decreased, neutralizing activity and antibody binding to S1, stable ADMP activity, and relatively stable S2-specific IgG levels.

Background Whether young adults who are infected with SARS-CoV-2 are at risk of subsequent infection is uncertain. We investigated the risk of subsequent SARS-CoV-2 infection among young adults seropositive for a previous infection. Methods This analysis was performed as part of the prospective COVID-19 Health Action Response for Marines study (CHARM). CHARM included predominantly male US Marine recruits, aged 18–20 years, following a 2-week unsupervised quarantine at home. After the home quarantine period, upon arrival at a Marine-supervised 2-week quarantine facility (college campus or hotel), participants were enrolled and were assessed for baseline SARS-CoV-2 IgG seropositivity, defined as a dilution of 1:150 or more on receptor-binding domain and full-length spike protein ELISA. Participants also completed a questionnaire consisting of demographic information, risk factors, reporting of 14 specific COVID-19-related symptoms or any other unspecified symptom, and brief medical history. SARS-CoV-2 infection was assessed by PCR at weeks 0, 1, and 2 of quarantine and participants completed a follow-up questionnaire, which included questions about the same COVID-19-related symptoms since the last study visit. Participants were excluded at this stage if they had a positive PCR test during quarantine. Participants who had three negative swab PCR results during quarantine and a baseline serum serology test at the beginning of the supervised quarantine that identified them as seronegative or seropositive for SARS-CoV-2 then went on to basic training at Marine Corps Recruit Depot—Parris Island. Three PCR tests were done at weeks 2, 4, and 6 in both seropositive and seronegative groups, along with the follow-up symptom questionnaire and baseline neutralising antibody titres on all subsequently infected seropositive and selected seropositive uninfected participants (prospective study period). Findings Between May 11, 2020, and Nov 2, 2020, we enrolled 3249 participants, of whom 3168 (98%) continued into the 2-week quarantine period. 3076 (95%) participants, 2825 (92%) of whom were men, were then followed up during the prospective study period after quarantine for 6 weeks. Among 189 seropositive participants, 19 (10%) had at least one positive PCR test for SARS-CoV-2 during the 6-week follow-up (1·1 cases per person-year). In contrast, 1079 (48%) of 2247 seronegative participants tested positive (6·2 cases per person-year). The incidence rate ratio was 0·18 (95% CI 0·11–0·28; p

Combining diagnostic specimens into pools has been considered as a strategy to augment throughput, decrease turnaround time, and leverage resources. This study utilized a multi-parametric approach to assess optimum pool size, impact of automation, and effect of nucleic acid amplification chemistries on the detection of SARS-CoV-2 RNA in pooled samples for surveillance testing on the Hologic Panther Fusion® System. Dorfman pooled testing was conducted with previously tested SARS-CoV-2 nasopharyngeal samples using Hologic’s Aptima® and Panther Fusion® SARS-CoV-2 Emergency Use Authorization assays. A manual workflow was used to generate pool sizes of 5:1 (five samples: one positive, four negative) and 10:1. An automated workflow was used to generate pool sizes of 3:1, 4:1, 5:1, 8:1 and 10:1. The impact of pool size, pooling method, and assay chemistry on sensitivity, specificity, and lower limit of detection (LLOD) was evaluated. Both the Hologic Aptima® and Panther Fusion® SARS-CoV-2 assays demonstrated >85% positive percent agreement between neat testing and pool sizes ≤5:1, satisfying FDA recommendation. Discordant results between neat and pooled testing were more frequent for positive samples with CT>35. Fusion® CT (cycle threshold) values for pooled samples increased as expected for pool sizes of 5:1 (CT increase of 1.92–2.41) and 10:1 (CT increase of 3.03–3.29). The Fusion® assay demonstrated lower LLOD than the Aptima® assay for pooled testing (956 vs 1503 cp/mL, pool size of 5:1). Lowering the cut-off threshold of the Aptima® assay from 560 kRLU (manufacturer’s setting) to 350 kRLU improved the assay sensitivity to that of the Fusion® assay for pooled testing. Both Hologic’s SARS-CoV-2 assays met the FDA recommended guidelines for percent positive agreement (>85%) for pool sizes ≤5:1. Automated pooling increased test throughput and enabled automated sample tracking while requiring less labor. The Fusion® SARS-CoV-2 assay, which demonstrated a lower LLOD, may be more appropriate for surveillance testing.

The ongoing evolution of SARS-CoV-2 requires monitoring the capability of immune responses to cross-recognize Variants of Concern (VOC). In this cross-sectional study, we examined serological and cell-mediated immune memory to SARS-CoV-2 variants, including Omicron, among a cohort of 18-21-year-old Marines with a history of either asymptomatic or mild SARS-CoV-2 infection 6 to 14 months earlier. Among the 210 participants in the study, 169 were unvaccinated while 41 received 2 doses of mRNA-based COVID-19 vaccines. Vaccination of previously infected participants strongly boosted neutralizing and binding activity and memory B and T cell responses including the recognition of Omicron, compared to infected but unvaccinated participants. Additionally, no measurable differences were observed in immune memory in healthy young adults with previous symptomatic or asymptomatic infections, for ancestral or variant strains. These results provide mechanistic immunological insights into population-based differences observed in immunity against Omicron and other variants among individuals with different clinical histories.

Background: The risk of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) subsequent infection among seropositive young adults was studied prospectively. Methods: The study population comprised 3,249 predominantly male, 18-20-year-old Marine recruits. Upon arrival at a Marine-supervised two-week quarantine, participants were assessed for baseline SARS-CoV-2 IgG seropositivity, defined as a 1:150 dilution or greater on receptor binding domain and full-length spike protein enzyme-linked immunosorbent (ELISA) assays. SARS-CoV-2 infection was assessed by PCR at initiation, middle and end of the quarantine. After appropriate exclusions, including participants with a positive PCR during quarantine, we performed three biweekly PCR tests in both seropositive and in seronegative groups once recruits left quarantine and entered basic training and baseline neutralizing antibody titers on all subsequently infected seropositive and selected seropositive uninfected participants. Findings: Among 189 seropositive participants, 19 (10.1%) had at least one positive PCR test for SARS-CoV-2 during the six-week follow-up (1.1 cases per person-year). In contrast, 1,079 (48.0%) of the 2,247 seronegative participants tested positive (6.2 cases per person-year). The incidence rate ratio was 0.18 (95% CI 0.11-0.28, p

The Naval Infectious Diseases Diagnostic Laboratory (NIDDL) serves as a reference clinical laboratory that supports Department of Defense (DoD) military treatment facilities worldwide in the detection and identification of high-risk and emerging infectious diseases. Since the emergence of Zika virus (ZIKV) in the Western Hemisphere in 2016, the NIDDL has been a central hub for ZIKV testing for DoD personnel and beneficiaries. Samples collected during patients' clinical evaluations were screened for evidence of possible exposure to ZIKV using molecular and serological methods. An in-house ZIKV plaque reduction neutralization test was used to confirm the presence of ZIKV immunoglobulin M antibody. Of 1,420 individuals tested, ZIKV infection was confirmed by quantitative real-time polymerase chain reaction (PCR) assay in 11 (0.8%); an additional 26 recent flaviviral infections (possibly ZIKV) were identified based on serology (1.8%). These findings contribute to the understanding of the burden of ZIKV infections among DoD personnel and beneficiaries and highlight the role of the NIDDL in clinical diagnosis during emerging infectious disease outbreaks.

Single-domain antibodies (sdAb), recombinantly produced variable heavy domains derived from the unconventional heavy chain antibodies found in camelids, provide stable, well-expressed binding elements with excellent affinity that can be tailored for specific applications through protein engineering. Complex matrices, such as plasma and serum, can dramatically reduce assay sensitivity. Thus, to achieve highly sensitive detection in complex matrices a highly efficient assay is essential. We produced sdAb as genetically linked dimers, and trimers, each including SpyTag at their C-terminus. The constructs were immobilized onto dyed magnetic microspheres to which SpyCatcher had been coupled and characterized in terms of their performance as capture reagents in sandwich assays. Initial tests on the ability of oriented monomer, dimer, and trimer captures to improve detection versus unoriented constructs in an assay for staphylococcal enterotoxin B spiked into buffer showed the oriented dimer format provided the best sensitivity while offering robust protein production. Thus, this format was utilized to improve a sdAb-based assay for the detection of dengue virus (DENV) nonstructural protein 1 (NS1) in serum. Detection of NS1 from each of the four DENV serotypes spiked into 50% normal human serum was increased by at least a factor of 5 when using the oriented dimer capture. We then demonstrated the potential of using the oriented dimer capture to improve detection of NS1 in clinical samples. This general method should enhance the utility of sdAb incorporated into any diagnostic assay, including those for high consequence pathogens.

Mosquito behavior is heavily influenced by the chemical molecules in the environment. This knowledge can be used to modify insect behaviors; particularly to reduce vector-host contact as a powerful method for disease prevention. N,N-Diethyl-meta-toluamide (DEET) is the most widely used insect repellent in the market and an excellent example of a chemical that has been used to modify insect behavior for disease prevention. However, genetic insensitivity and habituation in Aedes aegypti (L.) mosquitoes after preexposure to DEET have been reported. In this study, we investigated the effect of preexposure to DEET on the downstream blood-feeding behavior of Ae. aegypti mosquitoes and the duration of the effect. We exposed mosquitoes to four different DEET concentrations: 0.10, 0.12, 0.14, and 0.16% for 10 min then allowed the mosquitoes to blood-feed on an artificial blood-feeding system either immediately or after being held for 1, 3, 6, or 24 h following DEET exposure. We found that preexposing Ae. aegypti mosquitoes to 0.14 or 0.16% DEET lowered their blood engorgement level, but did not alter their landing and probing behavior when compared to the control test populations. The reduction in complete blood-feeding was observed at all time periods tested, but was only statistically significant at 3 and 6 h after the preexposure process. Because reduction in blood meal has been associated with increased refeeding, future studies analyzing the effect of this behavior using arbovirus-infected mosquitoes are needed to address the concern of potentially increased vectorial capacity.

No licensed vaccine or antiviral drug against dengue virus (DENV) is available; therefore, most of the effort to prevent this disease is focused on reducing vector-host interactions. One of the most widely accepted methods of blocking vector-human contact is to use insect repellents to interfere with mosquito host-seeking behavior. Some arboviruses can replicate in the nervous system of the vector, raising the concern that arboviral infection may alter the insect behavioral response toward chemical stimuli. Three different Aedes aegypti (L.) mosquito cohorts: DENV-1-injected, diluent-injected, and uninjected were subjected to behavioral tests using a high-throughput screening system with 2.5% DEET and 0.14% DEET on 1, 4, 7, 10, 14, and 17 d postinjection. All test cohorts exhibited significant contact irritant or escape responses when they were exposed to 2.5% or 0.14% DEET. However, we found no biologically relevant irritancy response change in DENV-1-infected Ae. aegypti mosquitoes when they were exposed to DEET. Further studies evaluating the effects of other arboviral infections on insect repellents activity are necessary in order to provide better recommendation on the prevention of vector-borne disease transmission.