In a recent study published in Nature, researchers investigated the cellular tropism, replication competence, persistence and evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in humans, and associated histopathological changes in infected tissues.
Coronavirus disease 2019 (COVID-19) has reportedly caused multiple organ derangements in the acute period, with a few infected individuals developing persistent symptoms, referred to as PASC (post-acute sequelae of COVID-19). However, the non-respiratory disease burden and the duration of SARS-CoV-2 clearance from non-respiratory tissues such as the brain has not been extensively investigated.
About the study
In the present study, researchers mapped and quantified the distribution, proliferation, and cellular tropism of SARS-CoV-2 in non-respiratory human body tissues such as the brain from the acute COVID-19 period to >7.0 months after the onset of symptoms.
Autopsied brain specimens of 44 unvaccinated and deceased COVID-19 patients were analyzed, and extensive CNS (central nervous system) sampling was performed for 11 individuals between April 26, 2020, and March 2, 2021. Polymerase chain reaction (PCR) analysis was performed to confirm SARS-CoV-2 positivity, and 38 samples were found to be SARS-CoV-2-positive. Three samples (P27, P36, and P37) were seronegative for SARS-CoV-2, and sera were not available for three cases (P3, P4, and P15).
Droplet digital PCR (ddPCR) analysis was performed for SARS-CoV-2 N (nucleocapsid) gene quantification, and ISH (in-situ hybridization) analysis was performed to validate the ddPCR results and to determine SARS-CoV-2 cell tropism. The IF (immunofluorescence) and IHC (immunohistochemistry) analyses were performed further to verify the viral presence within human brain tissues.
Further, subgenomic ribonucleic acid (RNA) was detected using real-time quantitative reverse transcription-PCR (RT– qPCR) analysis, and virus isolation experiments were performed using Vero E6 cells to demonstrate proliferation-capable SARS-CoV-2 among tissues of respiratory and another origin. SARS-CoV-2 S (spike) gene variant diversity and distribution were measured using HT-SGS (high-throughput, single-genome amplification and sequencing) analysis for six individuals.
Among the autopsied specimens, 17, 13, and 14 were categorized as early cases, mid-cases, and late cases based on the day of infection (d) at death within 14 days, between 15 days and 30 days, and beyond 31 days, respectively. Further, image analysis on interventricular septal tissues of 16 individuals was performed to assess the association between SARS-CoV-2 N RNA detected by ddPCR analysis and SARS-CoV-2 S RNA detected by ISH analysis. Furthermore, N protein-targeted ISH assays, IF analysis, and IHC-based analyses were performed to validate SARS-CoV-2 detection and distribution in the CNS.
Results
Among the study individuals, 30% were women, with a median age value of 63 years, and 61% of them suffered from at least three comorbid conditions. The median duration between the onset of symptoms to hospital admission and death was six days and 19 days, respectively, and the median post-mortem duration was 22 hours.
SARS-CoV-2 ribonucleic acid was present at 84 anatomical sites in significantly greater amounts among respiratory tissues than other tissues. SARS-CoV-2 RNA levels among early cases, mid-cases, and late-cases were 2.0 log10 nucleocapsid gene copies for every nanogram RNA, 1.4 log10 nucleocapsid gene copies for every nanogram RNA and 0.7 log10 nucleocapsid gene copies for every nanogram RNA, respectively. SARS-CoV-2 ribonucleic acid was detected in the perimortem sera of 11 and one early cases and mid-cases, respectively.
SARS-CoV-2 ribonucleic acid was persistently present in several tissues of late cases, despite being below detectable levels in sera of any case. SARS-CoV-2 ribonucleic acid was identified within the CNS among 91% (n= 10) cases, including across most brain areas evaluated in five (out of six) late cases. SARS-CoV-2 subgenomic ribonucleic acid was detected across all tissues and in several body fluids, including serum, vitreous humor, and pleural fluid.
The subgenomic ribonucleic acid RT-qPCR analysis and ddPCR analysis findings correlated closely for 1,025 specimens, particularly among 369 respiratory samples, 496 early cases, and 302 specimens showing SARS-CoV-2 positivity by RT-qPCR and ddPCR analyses. SARS-CoV-2 was isolated among Vero E6 cells from 45% (n=25) of specimens from the lymph nodes, heart, adrenal gland, gastrointestinal tissues, and ophthalmic tissues of early cases.
In addition, SARS-CoV-2 was isolated from the P38 thalamus among Vero E6-transmembrane serine protease 2 (TMPRSS2)-T2A-angiotensin-converting enzyme 2 (ACE2) cells. HT-SGS analysis of 46 specimens from six individuals did not show any non-synonymous SARS-CoV-2 genomic diversity in respiratory tissues and other tissues for P18, P19, and P27. Among P27 samples, two SARS-CoV-2 haplotypes, each comprising a synonymous mutation, were detected preferentially among non-respiratory tissues such as the mediastinal lymph nodes and the left and right ventricles.
In P38, residue D80F was detected in all 31 respiratory, but none of the 490 cranial sequences and residue G1219V was limited to the cranial variants. Among intraventricular septum tissues, the mean SARS-CoV-2 N gene copies/nanogram RNA correlated significantly with the median SARS-CoV-2 S RNA-positive cells. In the second ISH assay, SARS-CoV-2 RNA and protein were observed in the cerebellum and hypothalamus of P38, basal ganglia of P40, and cervical spinal cord of P42.
The histopathological analysis findings indicated that 92% (n=35) of cases died due to diffuse alveolar injury or acute pneumonia, and the diffuse alveolar injury cases showed a temporal pattern of progression. Myocardial infiltrates, and paracortical and follicular hyperplasia was observed. However, despite widespread SARS-CoV-2 RNA distribution in the body, negligible evidence of direct SARS-CoV-2 cytopathology or inflammation was observed in non-respiratory tissues.
Overall, the study findings showed that SARS-CoV-2 could infect and replicate in non-respiratory tissues such as the brain early in infection and persist for months (up to 230 days) following symptom onset.
HALIFAX – The Nova Scotia government says it could be months before it reveals how many people are on the wait-list for a family doctor.
The head of the province’s health authority told reporters Wednesday that the government won’t release updated data until the 160,000 people who were on the wait-list in June are contacted to verify whether they still need primary care.
Karen Oldfield said Nova Scotia Health is working on validating the primary care wait-list data before posting new numbers, and that work may take a matter of months. The most recent public wait-list figures are from June 1, when 160,234 people, or about 16 per cent of the population, were on it.
“It’s going to take time to make 160,000 calls,” Oldfield said. “We are not talking weeks, we are talking months.”
The interim CEO and president of Nova Scotia Health said people on the list are being asked where they live, whether they still need a family doctor, and to give an update on their health.
A spokesperson with the province’s Health Department says the government and its health authority are “working hard” to turn the wait-list registry into a useful tool, adding that the data will be shared once it is validated.
Nova Scotia’s NDP are calling on Premier Tim Houston to immediately release statistics on how many people are looking for a family doctor. On Tuesday, the NDP introduced a bill that would require the health minister to make the number public every month.
“It is unacceptable for the list to be more than three months out of date,” NDP Leader Claudia Chender said Tuesday.
Chender said releasing this data regularly is vital so Nova Scotians can track the government’s progress on its main 2021 campaign promise: fixing health care.
The number of people in need of a family doctor has more than doubled between the 2021 summer election campaign and June 2024. Since September 2021 about 300 doctors have been added to the provincial health system, the Health Department said.
“We’ll know if Tim Houston is keeping his 2021 election promise to fix health care when Nova Scotians are attached to primary care,” Chender said.
This report by The Canadian Press was first published Sept. 11, 2024.
ST. JOHN’S, N.L. – Newfoundland and Labrador‘s chief medical officer is monitoring the rise of whooping cough infections across the province as cases of the highly contagious disease continue to grow across Canada.
Dr. Janice Fitzgerald says that so far this year, the province has recorded 230 confirmed cases of the vaccine-preventable respiratory tract infection, also known as pertussis.
Late last month, Quebec reported more than 11,000 cases during the same time period, while Ontario counted 470 cases, well above the five-year average of 98. In Quebec, the majority of patients are between the ages of 10 and 14.
Meanwhile, New Brunswick has declared a whooping cough outbreak across the province. A total of 141 cases were reported by last month, exceeding the five-year average of 34.
The disease can lead to severe complications among vulnerable populations including infants, who are at the highest risk of suffering from complications like pneumonia and seizures. Symptoms may start with a runny nose, mild fever and cough, then progress to severe coughing accompanied by a distinctive “whooping” sound during inhalation.
“The public, especially pregnant people and those in close contact with infants, are encouraged to be aware of symptoms related to pertussis and to ensure vaccinations are up to date,” Newfoundland and Labrador’s Health Department said in a statement.
Whooping cough can be treated with antibiotics, but vaccination is the most effective way to control the spread of the disease. As a result, the province has expanded immunization efforts this school year. While booster doses are already offered in Grade 9, the vaccine is now being offered to Grade 8 students as well.
Public health officials say whooping cough is a cyclical disease that increases every two to five or six years.
Meanwhile, New Brunswick’s acting chief medical officer of health expects the current case count to get worse before tapering off.
A rise in whooping cough cases has also been reported in the United States and elsewhere. The Pan American Health Organization issued an alert in July encouraging countries to ramp up their surveillance and vaccination coverage.
This report by The Canadian Press was first published Sept. 10, 2024.
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