Congrong Miao

Therapeutic Monoclonal Antibody Treatment Targeting Respiratory Syncytial Virus (RSV) G Protein Mediates Viral Clearance and Reduces the Pathogenesis of RSV Infection in BALB/c Mice

Because the G protein of respiratory syncytial virus (RSV) has a CX3C chemokine motif that has been associated with the ability of RSV G protein to modulate the virus-induced host immune response, we examined whether therapeutic treatment with an anti-RSV G monoclonal antibody (mAb), 131-2G, that blocks the CX3C-associated activity of RSV G protein might decrease the pulmonary inflammation associated with infection in BALB/c mice. The results show that treatment with mAb 131-2G on day 3 after RSV infection reduces both inflammation and RSV titer in the lungs. Later administration of anti-RSV G mAb (day 5 after RSV infection) effectively reduced the viral titer but had a minimal effect on pulmonary inflammation. This study suggests that an anti-RSV G mAb might be an effective antiviral, either alone or in combination with anti-RSV F protein neutralizing antibodies, for decreasing the virus-induced host response to infection and improve treatment outcome.

Prophylactic Treatment with a G Glycoprotein Monoclonal Antibody Reduces Pulmonary Inflammation in Respiratory Syncytial Virus (RSV)-Challenged Naïve and Formalin-Inactivated RSV-Immunized BALB/c Mice

ABSTRACT We examined whether prophylactically administered anti-respiratory syncytial virus (anti-RSV) G monoclonal antibody (MAb) would decrease the pulmonary inflammation associated with primary RSV infection and formalin-inactivated RSV (FI-RSV)-enhanced disease in mice. MAb 131-2G administration 1 day prior to primary infection reduced the pulmonary inflammatory response and the level of RSV replication. Further, intact or F(ab′)2 forms of MAb 131-2G administered 1 day prior to infection in FI-RSV-vaccinated mice reduced enhanced inflammation and disease. This study shows that an anti-RSV G protein MAb might provide prophylaxis against both primary infection and FI-RSV-associated enhanced disease. It is possible that antibodies with similar reactivities might prevent enhanced disease and improve the safety of nonlive virus vaccines.

Family cluster of Middle East respiratory syndrome coronavirus infections, Tunisia, 2013.

In 2013 in Tunisia, 3 persons in 1 family were infected with Middle East respiratory syndrome coronavirus (MERS-CoV). The index case-patient’s respiratory tract samples were negative for MERS-CoV by reverse transcription PCR, but diagnosis was retrospectively confirmed by PCR of serum. Sequences clustered with those from Saudi Arabia and United Arab Emirates.

Monoclonal antibodies to SARS-associated coronavirus (SARS-CoV): Identification of neutralizing and antibodies reactive to S, N, M and E viral proteins

Abstract Monoclonal antibodies (Mabs) against the Urbani strain of the SARS-associated coronavirus (SARS-CoV) were developed and characterized for reactivity to SARS-CoV and SARS-CoV S, N, M, and E proteins using enzyme-linked immunoabsorbent (ELISA), radioimmunoprecipitation, immunofluorescence, Western Blot and microneutralization assays. Twenty-six mAbs were reactive to SARS-CoV by ELISA, and nine were chosen for detailed characterization. Five mAbs reacted against the S protein, two against the M protein, and one each against the N and E proteins. Two of five S protein mAbs neutralized SARS-CoV infection of Vero E6 cells and reacted to an epitope within amino acids 490–510 in the S protein. While two of the three non-neutralizing antibodies recognized at second epitope within amino acids 270–350. The mAbs characterized should prove useful for developing SARS-CoV diagnostic assays and for studying the biology of infection and pathogenesis of disease.

Health care worker contact with MERS patient, Saudi Arabia.

To investigate potential transmission of Middle East respiratory syndrome coronavirus (MERS-CoV) to health care workers in a hospital, we serologically tested hospital contacts of the index case-patient in Saudi Arabia, 4 months after his death. None of the 48 contacts showed evidence of MERS-CoV infection.

Multifacility Outbreak of Middle East Respiratory Syndrome in Taif, Saudi Arabia.

Enhanced surveillance and infection-control practices are needed to prevent outbreaks in healthcare settings.

Persistence of Antibodies against Middle East Respiratory Syndrome Coronavirus.

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.

Recombinant Protein-Based Assays for Detection of Antibodies to Severe Acute Respiratory Syndrome Coronavirus Spike and Nucleocapsid Proteins

ABSTRACT Recombinant severe acute respiratory syndrome (SARS) nucleocapsid and spike protein-based immunoglobulin G immunoassays were developed and evaluated. Our assays demonstrated high sensitivity and specificity to the SARS coronavirus in sera collected from patients as late as 2 years postonset of symptoms. These assays will be useful not only for routine SARS coronavirus diagnostics but also for epidemiological and antibody kinetic studies.

Coronavirus antibodies in bat biologists.

To the Editor: Severe acute respiratory syndrome–associated coronavirus (SARS-CoV) is a new coronavirus that caused an epidemic of 8,096 cases of SARS and 774 deaths during 2002–2003 (1). Attempts are ongoing to identify the natural reservoir of SARS-CoV. Several horseshoe bat species (Rhinolopus spp.) from Asia (2,3) and a sample of bats from Africa (4) have been found to be infected by and potential reservoirs for various SARS-like CoVs and various CoVs that are not SARS-like (2–4). However, transmission of bat SARS-CoV from bats to humans has not been reported. During October 2005, we looked for serologic evidence of infection among bat biologists attending an international meeting in the United States. After giving informed consent, volunteer biologists completed an anonymous survey and provided 10 mL of blood. Serum samples were tested at the Centers for Disease Control and Prevention (CDC) for antibodies against inactivated human SARS-CoV and against recombinant, expressed SARS-CoV nucleocapsid protein (SARS-CoV N) by enzyme immunoassays (EIAs) as described (5,6). This study was approved by the CDC Institutional Review Board. Of 350 registered biologists, 90 (26%) participated. Of participants, 89% had worked with or studied bats in North America, 21% in South America, 11% in Africa, 8% in Asia, 7% in Europe, and 6% in Australia. The primary genera studied by participants were Myotis (24%), Tadarida (13%), and Eptesicus (10%). A total of 20 (23%) participants had worked with or had contact with horseshoe bat species (Rhinolopus spp.). Because this genus has 69 species, distributed from Australia to Europe, some participants who indicated that they worked with the Rhinolopus spp. may likely have worked with species found outside of Asia. Involvement with bats most often consisted of capturing or handling them in the field (90%), followed by capturing or handling them in the laboratory (36%). Urine and feces were encountered most frequently (“always” or “most of the time” by 66%–68% of participants); contact with blood, saliva, or tissues and bites or scratches reportedly occurred less often (“always” or “most of the time” by 4%–28% of participants). The serum samples from all 90 participants were negative for antibodies against inactivated SARS-CoV, and samples from all but 1 were negative for SARS-CoV N protein. The 1 positive sample gave a strong signal (optical density 1.08 at 405 nm at a 1:400 dilution) by SARS-CoV N protein EIA and against SARS-CoV N by Western blot but gave no reactivity against recombinant SARS-CoV spike protein or inactivated SARS-CoV by either EIA or Western blot. Because the N protein has a region that is relatively conserved among all known coronaviruses (7), the antibodies against SARS-CoV N protein could have been induced by other CoVs. Previous studies have demonstrated that SARS-CoV N protein can cross-react with polyclonal antiserum induced by group 1 animal CoVs (8). To address the possibility that the antibodies from this serum sample were not specific to SARS-CoV, we tested it against recombinant N proteins of human CoVs, HCoV-229E, HCoV-OC43, NL63, and HKU-1. The serum reacted to all 4 N proteins, by EIA and Western blot, at titers of 400–1,600. We then tested the sample against 3 recombinant fragments of the N protein from each of 3 viruses: SARS-CoV, HCoV-229E, and HCoV-OC43. One of these fragments, N2, contains a highly conserved motif (FYYLGTGP) that should detect cross-reacting antibodies; the other 2 fragments should detect antibodies specific to the strain or group. The serum reacted to 2 of 3 fragments from HCoV-OC43 and -229E but to only the N2 fragment with the conserved motif from SARS-CoV (Figure), which suggests that the antibodies against SARS-CoV N were likely induced by a CoV that was not SARS-like. Figure Antibody reactivity to coronavirus (CoV) nucleocapsid (N) protein fragments by ELISA. A set of recombinant protein fragments covering the N protein sequence of human CoV (HCoV)–OC43, HCoV-229E, and severe acute respiratory syndrome (SARS)–CoV ... If the antibodies were induced by a SARS-like CoV infection, we would expect to have also detected antibodies against recombinant S protein (9) or recombinant fragments representing antigenically distinct regions of the N protein of SARS-CoV. We did not detect either; instead, we detected antibodies against the antigenically distinct N fragments from group 1 and 2 human CoVs. Thus, this survey of a sample of bat biologists, who were exposed primarily to North American bats but also to bats from Asia and Africa, showed no evidence of SARS-like CoV infection. Our survey found no evidence of SARS-CoV transmission from bats to humans. However, since the conclusion of this study, Dominguez et al. found coronavirus RNA in bats in North America, particularly Eptesicus fuscus and Myotis occultus (10), 2 species of the genera handled by 25% of the participants in our survey. Of interest is whether the bat biologists who worked with these bats might be at risk for infection with group 1 bat CoVs. Unfortunately, the high likelihood of infection with human group 1 CoVs will make it difficult to address this question. Additional studies of bat SARS-CoV infections in a larger number of persons who have been in contact with the species found to be positive for SARS-like CoV are needed before the risk for SARS-like CoV transmission from bats to humans can be clearly understood.

Decrease in Formalin-Inactivated Respiratory Syncytial Virus (FI-RSV) Enhanced Disease with RSV G Glycoprotein Peptide Immunization in BALB/c Mice

Respiratory syncytial virus (RSV) is a high priority target for vaccine development. One concern in RSV vaccine development is that a non-live virus vaccine would predispose for enhanced disease similar to that seen with the formalin inactivated RSV (FI-RSV) vaccine. Since a mAb specific to RSV G protein can reduce pulmonary inflammation and eosinophilia seen after RSV infection of FI-RSV vaccinated mice, we hypothesized that RSV G peptides that induce antibodies with similar reactivity may limit enhanced disease after subunit or other non-live RSV vaccines. In support of this hypothesis, we show that FI-RSV vaccinated mice administered RSV G peptide vaccines had a significant reduction in enhanced disease after RSV challenge. These data support the importance of RSV G during infection to RSV disease pathogenesis and suggest that use of appropriately designed G peptide vaccines to reduce the risk of enhanced disease with non-live RSV vaccines merits further study.