In this research letter, investigators report on the stability of SARS-CoV-2 and SARS-CoV-1 under experimental conditions. The viability of the two viruses was assessed in aerosols and on plastic, ...stainless steel, copper, and cardboard.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was identified as the etiologic agent associated with coronavirus disease, which emerged in late 2019. In response, we developed a ...diagnostic panel consisting of 3 real-time reverse transcription PCR assays targeting the nucleocapsid gene and evaluated use of these assays for detecting SARS-CoV-2 infection. All assays demonstrated a linear dynamic range of 8 orders of magnitude and an analytical limit of detection of 5 copies/reaction of quantified RNA transcripts and 1 x 10
50% tissue culture infectious dose/mL of cell-cultured SARS-CoV-2. All assays performed comparably with nasopharyngeal and oropharyngeal secretions, serum, and fecal specimens spiked with cultured virus. We obtained no false-positive amplifications with other human coronaviruses or common respiratory pathogens. Results from all 3 assays were highly correlated during clinical specimen testing. On February 4, 2020, the Food and Drug Administration issued an Emergency Use Authorization to enable emergency use of this panel.
To determine how long antibodies against Middle East respiratory syndrome coronavirus persist, we measured long-term antibody responses among persons serologically positive or indeterminate after a ...2012 outbreak in Jordan. Antibodies, including neutralizing antibodies, were detectable in 6 (86%) of 7 persons for at least 34 months after the outbreak.
Known human coronaviruses (hCoV) usually cause mild to moderate upper-respiratory tract illnesses, except SARS-CoV and MERS-CoV, which, in addition to mild illness can also be associated with severe ...respiratory diseases and high mortality rates. Well-characterized multiplexed serologic assays are needed to aid in rapid detection and surveillance of hCoVs. The present study describes development and evaluation of a multiplexed magnetic microsphere immunoassay (MMIA) to simultaneously detect immunoglobulin G (IgG) antibodies specific for recombinant nucleocapsid proteins (recN) from hCoVs 229E, NL63, OC43, HKU1, SARS-CoV, and MERS-CoV. We used paired human sera to screen for IgG with reactivity against six hCoVs to determine assay sensitivity, specificity and reproducibility. We found no signal interference between monoplex and multiplex assay formats (R
range = 0.87-0.97). Screening of paired human sera using MMIA, resulted in 92 of 106 (sensitivity: 86%) as positive and 68 of 80 (specificity: 84%) as negative. This study serves as a proof of concept that it is feasible to develop and use a multiplexed microsphere immunoassay as a next generation screening tool for use in large scale seroprevalence studies of hCoVs.
The need for high-affinity, SARS-CoV-2-specific monoclonal antibodies (mAbs) is critical in the face of the global COVID-19 pandemic, as such reagents can have important diagnostic, research, and ...therapeutic applications. Of greatest interest is the ~ 300 amino acid receptor binding domain (RBD) within the S1 subunit of the spike protein because of its key interaction with the human angiotensin converting enzyme 2 (hACE2) receptor present on many cell types, especially lung epithelial cells. We report here the development and functional characterization of 29 nM-affinity mouse SARS-CoV-2 mAbs created by an accelerated immunization and hybridoma screening process. Differing functions, including binding of diverse protein epitopes, viral neutralization, impact on RBD-hACE2 binding, and immunohistochemical staining of infected lung tissue, were correlated with variable gene usage and sequence.
We assessed the relationship between antigen and reverse transcription PCR (RT-PCR) test positivity and successful virus isolation. We found that antigen test results were more predictive of virus ...recovery than RT-PCR results. However, virus was isolated from some antigen-negative and RT-PCR‒positive paired specimens, providing support for the Centers for Disease Control and Prevention antigen testing algorithm.
Co-infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other viruses has been reported. We evaluated cell lines commonly used to isolate viruses and diagnose related ...diseases for their susceptibility to SARS-CoV-2. Although multiple kidney cell lines from monkeys were susceptible to SARS-CoV-2, we found many cell types derived from humans, dogs, minks, cats, mice, and chicken were not. We analyzed MDCK cells, which are most commonly used for surveillance and study of influenza viruses, and found that they were not susceptible to SARS-CoV-2. The low expression level of the angiotensin converting enzyme 2 receptor and lower receptor affinity to SARS-CoV-2 spike, which could be overcome by overexpression of canine angiotensin converting enzyme 2 in trans, strengthened the cellular barrier to productive infection. Moreover, a D614G mutation in the spike protein did not appear to affect SARS-CoV-2 cell tropism. Our findings should help avert inadvertent propagation of SARS-CoV-2 from diagnostic cell lines.
To assess transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a detention facility experiencing a coronavirus disease outbreak and evaluate testing strategies, we ...conducted a prospective cohort investigation in a facility in Louisiana, USA. We conducted SARS-CoV-2 testing for detained persons in 6 quarantined dormitories at various time points. Of 143 persons, 53 were positive at the initial test, and an additional 58 persons were positive at later time points (cumulative incidence 78%). In 1 dormitory, all 45 detained persons initially were negative; 18 days later, 40 (89%) were positive. Among persons who were SARS-CoV-2 positive, 47% (52/111) were asymptomatic at the time of specimen collection; 14 had replication-competent virus isolated. Serial SARS-CoV-2 testing might help interrupt transmission through medical isolation and quarantine. Testing in correctional and detention facilities will be most effective when initiated early in an outbreak, inclusive of all exposed persons, and paired with infection prevention and control.
Middle East respiratory syndrome coronavirus, MERS‐CoV, was identified in Saudi Arabia in 2012, and as of January 29, 2018, there were 2,123 laboratory‐confirmed MERS‐CoV cases reported to WHO (WHO, ...2018, https://www.who.int/emergencies/mers-cov/en/). Multiple studies suggest that dromedary camels are a source for human MERS‐CoV infection. MERS‐CoV‐specific antibodies have been detected in the serum of dromedary camels across Northern Africa and across the Arabian Peninsula. Israel's geographic location places Israel at risk for MERS‐CoV infection. To date, MERS‐CoV‐related illness has not been reported and the burden of MERS‐CoV infection in the Israeli population is unknown. The seroprevalence of MERS‐CoV‐specific antibodies in Israeli dromedary camels is unknown. The objective of this study was to determine the prevalence of MERS‐CoV seropositivity in dromedary camels in Israel. The prevalence of MERS‐CoV antibodies in Israeli camels was examined in 71 camel sera collected from four farms across Israel by MERS‐CoV‐specific microneutralization (Mnt) assay and confirmed by MERS‐CoV‐specific immunofluorescence assay (IFA). Although this study cannot rule out potential antibody cross‐reactivity by IFA, the presence of bovine coronavirus‐specific antibodies do not appear to impact detection of MERS‐CoV antibodies by Mnt. MERS‐CoV neutralizing antibodies were detectable in 51 (71.8%) camel sera, and no association was observed between the presence of neutralizing antibodies and camel age or gender. These findings extend the known range of MERS‐CoV circulation in Middle Eastern camels. The high rate of MERS‐CoV‐specific antibody seropositivity in dromedary camels in the absence of any reported human MERS cases suggests that there is still much to be learned about the dynamics of camel‐to‐human transmission of MERS‐CoV.
Virus shedding in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can occur before onset of symptoms; less is known about symptom progression or infectiousness associated with initiation ...of viral shedding. We investigated household transmission in 5 households with daily specimen collection for 5 consecutive days starting a median of 4 days after symptom onset in index patients. Seven contacts across 2 households implementing no precautionary measures were infected. Of these 7, 2 tested positive for SARS-CoV-2 by reverse transcription PCR on day 3 of 5. Both had mild, nonspecific symptoms for 1-3 days preceding the first positive test. SARS-CoV-2 was cultured from the fourth-day specimen in 1 patient and from the fourth- and fifth-day specimens in the other. We also describe infection control measures taken in the households that had no transmission. Persons exposed to SARS-CoV-2 should self-isolate, including from household contacts, wear a mask, practice hand hygiene, and seek testing promptly.
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