Reporter : He Liping
Publisher : The Paper
Ref : http://news.sciencenet.cn/htmlnews/2020/5/439801.shtm
Translation, editing : Gan Yung Chyan
/ KUCINTA SETIA
Image courtesy : Arterra / UIG via Getty Images.
Since the outbreak, the ability of SARS-CoV-2 to spread through aerosols has been one of the academic focuses. The latest research by the team of the University of Hong Kong shows that in the animal model experiment of golden hamsters, the virus can be transmitted via aerosols.
The research was published online in the form of "Accelerated Article Preview" on Nature on 14 May 2020, entitled "Pathogenesis and transmission of SARS-CoV-2 in golden hamsters". The corresponding author of the paper is Hui-Ling Yen of the Li Ka-shing School of Medicine of the University of Hong Kong. The author of the paper also includes Professor Pan Liewen of the School of Public Health of the University of Hong Kong.
After the emergence of the SARS-CoV-2, researchers urgently need suitable small animal models to support the development of vaccines and treatments. The researchers reported the pathogenesis and infectivity of the SARS-CoV-2 (covi, in short) in golden hamsters (golden Syrian hamsters). Immunohistochemistry showed that there were viral antigens in the nasal mucosa, bronchial epithelial cells and lung consolidation area 2 and 5 days after the golden hamster was inoculated with the virus. Seven days after virus infection, the virus was quickly cleared and lung cells proliferated. Viral antigens were also found in duodenal epithelial cells of golden hamsters, and viral RNA was detected in feces.
It is worth noting that animal model tests have shown that covi can effectively spread from infected golden hamsters to naive golden hamsters through direct contact and aerosols. The efficiency of hamster cage transmission through the medium is low. Although viral RNA was detected in the nasal wash of vaccinated hamsters for 14 consecutive days, covi can spread for a short time. Inoculated and naturally infected hamsters showed significant weight loss, and all golden hamsters were able to detect neutralizing antibodies after recovery. This result indicates that the characteristics of golden hamster's covi infection are similar to those of mild human infection.
The paper points out that an appropriate animal model is essential for understanding the pathogenesis of covid and evaluating vaccines and therapeutic candidates. Previous animal studies of SARS-CoV have shown that the interaction between the viral spike protein (S protein) and the host ’s angiotensin converting enzyme 2 (ACE2) receptor, as well as the age and innate immune status of the infected subject It plays an important role in the pathogenesis. Like SARS-CoV, the S protein of SARS-CoV-2 also uses the ACE2 receptor (mainly distributed in the epithelial cells of the lung and small intestine) to enter the cell for viral replication. SARS-CoV-2 binds well to human ACE2, but has limited binding to murine ACE2, which limits the application of inbred mice in virus research. Rhesus monkeys and transgenic ICR mice expressing the human ACE2 receptor have been shown to be susceptible to SARS-CoV-2; however, these animal models are not so easily available. Crab-eating macaques and rhesus monkeys attacked by SARS-CoV-2 showed limited and moderate clinical symptoms, respectively. The infected transgenic mice showed moderate pneumonia, and no obvious histological changes in non-respiratory tissues. According to reports, the previously generated transgenic mice expressing human ACE2 receptors support the replication of SARS-CoV in respiratory epithelial cells, but because of the high expression of ACE2 in the brains of transgenic mice, the mortality of mice is also added to the nerve Variables related to systemic disease.
The golden hamster is a widely used experimental animal model. Previous studies have shown that SARS-CoV can replicate in its body, but MERS-CoV cannot. This is because MERS-CoV uses the DPP4 protein as a virus to enter cells. Body, not ACE2. Previously, the vaccination study of SARS-CoV (Urbani strain) of 5-week-old golden hamsters showed that the virus replicated strongly in the body, and the peak virus titer could be detected in the lung 2 days after the virus inoculation; Seven days after the virus, the golden hamster can quickly remove the virus. However, virus-infected golden hamsters did not lose weight or have obvious disease conditions. A follow-up study reported that different SARS-CoV strains were tested in golden hamsters and found to differ in virulence between SARS-CoV strains. According to reports, SARS-CoV (Frk-1 strain) is lethal to hamsters. The Frk-1 strain differs from the non-lethal Urbani strain in the L1148F mutation in the S2 domain. Hamsters can also be infected with other respiratory viruses, including human metapneumovirus, human parainfluenza virus, and influenza A virus, and may support the spread of influenza through exposure or air. The comparison of ACE2 proteins in humans, macaques, mice and hamsters indicates that hamster ACE2 may interact with the S protein of SARS-CoV-2 more effectively than murine ACE2. Here, the research team evaluated the pathogenesis and contact transmission ability of SARS-CoV-2 in 4-5 week old male golden hamsters.
Prompt animal model experiment of aerosol transmission
In order to study the ability of SARS-CoV-2 to spread through aerosols in hamsters, the researchers placed the donor (receiving the covi inoculation) hamster and the infant hamster (healthy hamster) in two adjacent iron cages respectively One day after the body hamster was inoculated with the virus, the two cages were placed together for 8 hours.
The infectious virus in the nasal wash of the donor hamster can detoxify for 6 days, and the RNA of the virus can be continuously detected for 14 days, that is to say, in the later stage, the infected hamster does not have the ability to infect.
Viral RNA can be detected in donor stool samples at 2, 4, and 6 days after virus inoculation, but it is not an infectious virus.
The researchers found that the aerosol transmission of covi in hamsters was effective because infectious viruses were detected in all exposed hamster nasal washes 1 day after exposure and the viral load peaked 3 days after exposure.
Although infectious viruses were not isolated, viral RNA could be detected for 14 consecutive days from stool samples of infected aerosol contacts.
Hamsters exposed to aerosol showed the greatest weight loss 7 days after exposure (mean ± SD, -7.72 ± 5,42%, N = 3). Compared with donor hamsters, aerosol contact hamsters expel a considerable amount of virus in the nasal wash.
In order to evaluate the ability of covi to spread in hamsters, the researchers intranasally inoculated 3 donor hamsters with 8 × 10 minus 4 power of TCID50 virus. Twenty-four hours after inoculation, each donor was transferred to a new cage and raised with a juvenile hamster. The researchers monitored body weight changes and clinical signs daily, and collected nasal washes from donor hamsters and contact hamsters every other day for 14 days. In donor hamsters, although viral RNA can be detected for 14 consecutive days, the peak of infectious virus appears early after the virus inoculation, and then drops rapidly.
Six days after SARS-CoV-2 inoculation, hamsters showed the greatest average weight loss (mean ± SD, -11.97 ± 4.51%, N = 6).
The transmission from donor to contact is very efficient. SARS-CoV-2 can be detected in cohabitation hamsters 1 day after contact, and peak viral load is detected in cohabitation hamster nasal wash 3 days after contact.
Cohabitation hamsters experienced the largest average weight loss 6 days after exposure (mean ± SD, -10.68 ± 3.42%, N = 3), and all animals returned to their original body weight 11 days after exposure.
Using PRNT analysis, neutralizing antibodies were detected from donors 14 days after virus inoculation (all titers 1: 640) and 13 days after contact with cohabiting hamsters (at 1: 160, 1: 320, and 1: 160 titers) . The researchers detected viral RNA from the nasal washes of donor hamsters for 14 consecutive days, but the infectious virus titer dropped rapidly. The researchers repeated the experiment and lived with the child hamster 6 days after the virus inoculation. A small amount of viral RNA was detected in the nasal wash of cohabiting naive hamsters 3 days after contact, and no viral RNA was detected in the nasal wash 7 days after contact. Moreover, no infectious virus was detected in the nasal wash.
Cohabiting hamsters did not show weight loss
PRNT analysis did not detect neutralizing antibodies (<1:10) in cohabiting hamsters 12 days after exposure. The results showed that the virus inoculated with SARS-CoV-2 donors can be transmitted in less than 6 days. The subsequent transmission from the donor to the cohabitation contact is related to the detection of an infectious disease in the donor's nasal wash, and not to the detection of viral RNA.
Pathogenesis of SARS-CoV-2 in golden hamsters
In the experiment, the researchers infected the golden hamsters intranasally with an 8 x 10 negative 4th power TCID50 (50% tissue infection) of the new coronavirus (Beta-CoV / Hong Kong / VM20001061 / 2020 virus, GISAID # EPI_ISL_412028). The virus was isolated from Vero E6 cells of nasopharyngeal aspirates and throat swabs of a patient with new coronary pneumonia in Hong Kong. The researchers collected turbinate, brain, lung, heart, duodenum, liver, spleen, and kidney tissues at 2, 5, and 7 days after the golden hamsters were inoculated with the virus to monitor virus replication and histopathological changes. On the second day after the virus inoculation, the peak viral load of the golden hamster lungs can be detected, and the viral load began to decrease 5 days after the virus inoculation. Although high copies of viral RNA can continue to be detected, no infectious virus was detected 7 days after virus inoculation.
Infectious viral load was significantly different between 2 and 7 days after inoculation (P = 0.019, Dunn's multiple comparison test), but there was no difference in RNA virus copy number (P = 0.076). Although low copies of viral RNA were detected at 2 and 5 days after virus inoculation, no infectious virus was detected in the kidney.
Histopathological examination found that 5-10% of the inflammatory cells in the lungs increased and consolidated 2 days after the virus inoculation.
Five days after the virus inoculation, 15-35% of the inflammatory cells in the lungs increased.
In areas where viral antigens were detected 2 and 5 days after virus inoculation, monocyte infiltration could be observed. The immunohistochemistry of the N protein of SARS-CoV-2 showed that viral antigens were present in bronchial epithelial cells of golden hamsters 2 days after virus inoculation.
Five days after virus inoculation, it developed into lung cells.
Seven days after virus inoculation, 30-60% of the lungs are consolidated. However, no viral antigens were detected, and type 2 lung cell proliferation was obvious.
CD3 positive T lymphocytes were detected in the area around the bronchus 5 days after virus inoculation, which may help to quickly clear infected cells.
There is infiltration of inflammatory cells in the turbinate. Viral antigens were detected in nasal epithelial cells and olfactory sensory neurons of the nasal mucosa.
Infection in olfactory neurons was further confirmed in cells expressing the N protein of SARS-CoV and neuron-specific β-III tubulin.
Compared with the control group, the number of olfactory neurons in the nasal mucosa of the golden hamster decreased 2 days after virus infection, as shown in the figure below
Seven days after virus inoculation, nasal epithelial cells were significantly attenuated. Tissue repair can then be observed 14 days after virus inoculation. Although there was no inflammation in the duodenal epithelial cells, viral antigens were detected in the duodenal epithelial cells 2 days after the virus inoculation.
Five days after virus inoculation, no obvious histopathological changes were observed from the brain, heart, liver, and kidneys.
Publisher : The Paper
Ref : http://news.sciencenet.cn/htmlnews/2020/5/439801.shtm
Translation, editing : Gan Yung Chyan
/ KUCINTA SETIA
Image courtesy : Arterra / UIG via Getty Images.
Since the outbreak, the ability of SARS-CoV-2 to spread through aerosols has been one of the academic focuses. The latest research by the team of the University of Hong Kong shows that in the animal model experiment of golden hamsters, the virus can be transmitted via aerosols.
The research was published online in the form of "Accelerated Article Preview" on Nature on 14 May 2020, entitled "Pathogenesis and transmission of SARS-CoV-2 in golden hamsters". The corresponding author of the paper is Hui-Ling Yen of the Li Ka-shing School of Medicine of the University of Hong Kong. The author of the paper also includes Professor Pan Liewen of the School of Public Health of the University of Hong Kong.
After the emergence of the SARS-CoV-2, researchers urgently need suitable small animal models to support the development of vaccines and treatments. The researchers reported the pathogenesis and infectivity of the SARS-CoV-2 (covi, in short) in golden hamsters (golden Syrian hamsters). Immunohistochemistry showed that there were viral antigens in the nasal mucosa, bronchial epithelial cells and lung consolidation area 2 and 5 days after the golden hamster was inoculated with the virus. Seven days after virus infection, the virus was quickly cleared and lung cells proliferated. Viral antigens were also found in duodenal epithelial cells of golden hamsters, and viral RNA was detected in feces.
It is worth noting that animal model tests have shown that covi can effectively spread from infected golden hamsters to naive golden hamsters through direct contact and aerosols. The efficiency of hamster cage transmission through the medium is low. Although viral RNA was detected in the nasal wash of vaccinated hamsters for 14 consecutive days, covi can spread for a short time. Inoculated and naturally infected hamsters showed significant weight loss, and all golden hamsters were able to detect neutralizing antibodies after recovery. This result indicates that the characteristics of golden hamster's covi infection are similar to those of mild human infection.
The paper points out that an appropriate animal model is essential for understanding the pathogenesis of covid and evaluating vaccines and therapeutic candidates. Previous animal studies of SARS-CoV have shown that the interaction between the viral spike protein (S protein) and the host ’s angiotensin converting enzyme 2 (ACE2) receptor, as well as the age and innate immune status of the infected subject It plays an important role in the pathogenesis. Like SARS-CoV, the S protein of SARS-CoV-2 also uses the ACE2 receptor (mainly distributed in the epithelial cells of the lung and small intestine) to enter the cell for viral replication. SARS-CoV-2 binds well to human ACE2, but has limited binding to murine ACE2, which limits the application of inbred mice in virus research. Rhesus monkeys and transgenic ICR mice expressing the human ACE2 receptor have been shown to be susceptible to SARS-CoV-2; however, these animal models are not so easily available. Crab-eating macaques and rhesus monkeys attacked by SARS-CoV-2 showed limited and moderate clinical symptoms, respectively. The infected transgenic mice showed moderate pneumonia, and no obvious histological changes in non-respiratory tissues. According to reports, the previously generated transgenic mice expressing human ACE2 receptors support the replication of SARS-CoV in respiratory epithelial cells, but because of the high expression of ACE2 in the brains of transgenic mice, the mortality of mice is also added to the nerve Variables related to systemic disease.
The golden hamster is a widely used experimental animal model. Previous studies have shown that SARS-CoV can replicate in its body, but MERS-CoV cannot. This is because MERS-CoV uses the DPP4 protein as a virus to enter cells. Body, not ACE2. Previously, the vaccination study of SARS-CoV (Urbani strain) of 5-week-old golden hamsters showed that the virus replicated strongly in the body, and the peak virus titer could be detected in the lung 2 days after the virus inoculation; Seven days after the virus, the golden hamster can quickly remove the virus. However, virus-infected golden hamsters did not lose weight or have obvious disease conditions. A follow-up study reported that different SARS-CoV strains were tested in golden hamsters and found to differ in virulence between SARS-CoV strains. According to reports, SARS-CoV (Frk-1 strain) is lethal to hamsters. The Frk-1 strain differs from the non-lethal Urbani strain in the L1148F mutation in the S2 domain. Hamsters can also be infected with other respiratory viruses, including human metapneumovirus, human parainfluenza virus, and influenza A virus, and may support the spread of influenza through exposure or air. The comparison of ACE2 proteins in humans, macaques, mice and hamsters indicates that hamster ACE2 may interact with the S protein of SARS-CoV-2 more effectively than murine ACE2. Here, the research team evaluated the pathogenesis and contact transmission ability of SARS-CoV-2 in 4-5 week old male golden hamsters.
Prompt animal model experiment of aerosol transmission
In order to study the ability of SARS-CoV-2 to spread through aerosols in hamsters, the researchers placed the donor (receiving the covi inoculation) hamster and the infant hamster (healthy hamster) in two adjacent iron cages respectively One day after the body hamster was inoculated with the virus, the two cages were placed together for 8 hours.
The infectious virus in the nasal wash of the donor hamster can detoxify for 6 days, and the RNA of the virus can be continuously detected for 14 days, that is to say, in the later stage, the infected hamster does not have the ability to infect.
Viral RNA can be detected in donor stool samples at 2, 4, and 6 days after virus inoculation, but it is not an infectious virus.
The researchers found that the aerosol transmission of covi in hamsters was effective because infectious viruses were detected in all exposed hamster nasal washes 1 day after exposure and the viral load peaked 3 days after exposure.
Although infectious viruses were not isolated, viral RNA could be detected for 14 consecutive days from stool samples of infected aerosol contacts.
Hamsters exposed to aerosol showed the greatest weight loss 7 days after exposure (mean ± SD, -7.72 ± 5,42%, N = 3). Compared with donor hamsters, aerosol contact hamsters expel a considerable amount of virus in the nasal wash.
In order to evaluate the ability of covi to spread in hamsters, the researchers intranasally inoculated 3 donor hamsters with 8 × 10 minus 4 power of TCID50 virus. Twenty-four hours after inoculation, each donor was transferred to a new cage and raised with a juvenile hamster. The researchers monitored body weight changes and clinical signs daily, and collected nasal washes from donor hamsters and contact hamsters every other day for 14 days. In donor hamsters, although viral RNA can be detected for 14 consecutive days, the peak of infectious virus appears early after the virus inoculation, and then drops rapidly.
Six days after SARS-CoV-2 inoculation, hamsters showed the greatest average weight loss (mean ± SD, -11.97 ± 4.51%, N = 6).
The transmission from donor to contact is very efficient. SARS-CoV-2 can be detected in cohabitation hamsters 1 day after contact, and peak viral load is detected in cohabitation hamster nasal wash 3 days after contact.
Cohabitation hamsters experienced the largest average weight loss 6 days after exposure (mean ± SD, -10.68 ± 3.42%, N = 3), and all animals returned to their original body weight 11 days after exposure.
Using PRNT analysis, neutralizing antibodies were detected from donors 14 days after virus inoculation (all titers 1: 640) and 13 days after contact with cohabiting hamsters (at 1: 160, 1: 320, and 1: 160 titers) . The researchers detected viral RNA from the nasal washes of donor hamsters for 14 consecutive days, but the infectious virus titer dropped rapidly. The researchers repeated the experiment and lived with the child hamster 6 days after the virus inoculation. A small amount of viral RNA was detected in the nasal wash of cohabiting naive hamsters 3 days after contact, and no viral RNA was detected in the nasal wash 7 days after contact. Moreover, no infectious virus was detected in the nasal wash.
Cohabiting hamsters did not show weight loss
PRNT analysis did not detect neutralizing antibodies (<1:10) in cohabiting hamsters 12 days after exposure. The results showed that the virus inoculated with SARS-CoV-2 donors can be transmitted in less than 6 days. The subsequent transmission from the donor to the cohabitation contact is related to the detection of an infectious disease in the donor's nasal wash, and not to the detection of viral RNA.
Pathogenesis of SARS-CoV-2 in golden hamsters
In the experiment, the researchers infected the golden hamsters intranasally with an 8 x 10 negative 4th power TCID50 (50% tissue infection) of the new coronavirus (Beta-CoV / Hong Kong / VM20001061 / 2020 virus, GISAID # EPI_ISL_412028). The virus was isolated from Vero E6 cells of nasopharyngeal aspirates and throat swabs of a patient with new coronary pneumonia in Hong Kong. The researchers collected turbinate, brain, lung, heart, duodenum, liver, spleen, and kidney tissues at 2, 5, and 7 days after the golden hamsters were inoculated with the virus to monitor virus replication and histopathological changes. On the second day after the virus inoculation, the peak viral load of the golden hamster lungs can be detected, and the viral load began to decrease 5 days after the virus inoculation. Although high copies of viral RNA can continue to be detected, no infectious virus was detected 7 days after virus inoculation.
Infectious viral load was significantly different between 2 and 7 days after inoculation (P = 0.019, Dunn's multiple comparison test), but there was no difference in RNA virus copy number (P = 0.076). Although low copies of viral RNA were detected at 2 and 5 days after virus inoculation, no infectious virus was detected in the kidney.
Histopathological examination found that 5-10% of the inflammatory cells in the lungs increased and consolidated 2 days after the virus inoculation.
Five days after the virus inoculation, 15-35% of the inflammatory cells in the lungs increased.
In areas where viral antigens were detected 2 and 5 days after virus inoculation, monocyte infiltration could be observed. The immunohistochemistry of the N protein of SARS-CoV-2 showed that viral antigens were present in bronchial epithelial cells of golden hamsters 2 days after virus inoculation.
Five days after virus inoculation, it developed into lung cells.
Seven days after virus inoculation, 30-60% of the lungs are consolidated. However, no viral antigens were detected, and type 2 lung cell proliferation was obvious.
CD3 positive T lymphocytes were detected in the area around the bronchus 5 days after virus inoculation, which may help to quickly clear infected cells.
There is infiltration of inflammatory cells in the turbinate. Viral antigens were detected in nasal epithelial cells and olfactory sensory neurons of the nasal mucosa.
Infection in olfactory neurons was further confirmed in cells expressing the N protein of SARS-CoV and neuron-specific β-III tubulin.
Compared with the control group, the number of olfactory neurons in the nasal mucosa of the golden hamster decreased 2 days after virus infection, as shown in the figure below
Seven days after virus inoculation, nasal epithelial cells were significantly attenuated. Tissue repair can then be observed 14 days after virus inoculation. Although there was no inflammation in the duodenal epithelial cells, viral antigens were detected in the duodenal epithelial cells 2 days after the virus inoculation.
Five days after virus inoculation, no obvious histopathological changes were observed from the brain, heart, liver, and kidneys.
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