Comparative study of COVID-19 infection in renal transplant recipients and non transplant recipients (preprint)

To analyse the difference in COVID-19 infection between kidney transplant patients and non-transplant patients. We included post-transplant patients with COVID-19 infection who attended Shenzhen No. 3 Hospital from December 2022 to February 2023, and enrolled the general population with COVID-19 infection who were hospitalized during the same period, matched by age and gender. They were divided into Kidney Transplant Recipients group (KTR) (n=194) and Non-Kidney Transplant Recipients Group（NKTR）(n=516) and the basic information, clinical symptoms, laboratory data, treatments and outcomes of these two groups were compared. The proportion of the renal transplant population classified as severe and critical was 15.5%, which was significantly higher than that in NKTR group (P < 0.05); the proportion of patients with pneumonia was also significantly higher than that in NKTR group.  The mean maximum fever temperature was slightly higher in the NKTR(P<0.001);Kidney transplant population having lower absolute lymphocyte counts on admission and 7 days after admission than the general population, with statistically significant differences（P<0.001, P＜0.001). The use of intravenous hormones was significantly higher (42.8% vs. 6.0%, p=0.000), as was the use of small molecules such as Azvudine and Paxlovid, compared to the general population. A total of 10 patients in the included population required ICU admission, all in the KTR group; six patients experienced death, also in the renal transplant group. Conclusion: Post-transplant COVID-19 infections are more severe and require hormonal and small molecule antiviral therapy, and the prognosis is worse than in the general population. Trial registration This study was approved and supervised by the ethics committee of the Third People's Hospital of Shenzhen (approval number 2023-036-02) and firstly registered in 03/07/2023, registration number was NCT05926076.

Characterization of Microbial Co-infections in the Respiratory Tract of hospitalized COVID-19 patients (preprint)

Summary Background Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global pandemic of Coronavirus disease 2019 (COVID-19). However, microbial composition of the respiratory tract and other infected tissues, as well as their possible pathogenic contributions to varying degrees of disease severity in COVID-19 patients remain unclear. Method Between January 27 and February 26, 2020, serial clinical specimens (sputum, nasal and throat swab, anal swab and feces) were collected from a cohort of hospitalized COVID-19 patients, including 8 mildly and 15 severely ill patients (requiring ICU admission and mechanical ventilation), in the Guangdong province, China. Total RNA was extracted and ultra-deep metatranscriptomic sequencing was performed in combination with laboratory diagnostic assays. Co-infection rates, the prevalence and abundance of microbial communities in these COVID-19 patients were determined. Findings Notably, respiratory microbial co-infections were exclusively found in 84.6% of severely ill patients (11/13), among which viral and bacterial co-infections were detected by sequencing in 30.8% (4/13) and 69.2% (9/13) of the patients, respectively. In addition, for 23.1% (3/13) of the patients, bacterial co-infections with Burkholderia cepacia complex (BCC) and Staphylococcus epidermidis were also confirmed by bacterial culture. Further, a time-dependent, secondary infection of B. cenocepacia with expressions of multiple virulence genes in one severely ill patient was demonstrated, which might be the primary cause of his disease deterioration and death one month after ICU admission. Interpretation Our findings identified distinct patterns of co-infections with SARS-CoV-2 and various respiratory pathogenic microbes in hospitalized COVID-19 patients in relation to disease severity. Detection and tracking of BCC-associated nosocomial infections are recommended to improve the pre-emptive treatment regimen and reduce fatal outcomes of hospitalized patients infected with SARS-CoV-2. Funding National Science and Technology Major Project of China, National Major Project for Control and Prevention of Infectious Disease in China, the emergency grants for prevention and control of SARS-CoV-2 of Ministry of Science and Technology and Guangdong province, Guangdong Provincial Key Laboratory of Genome Read and Write, Guangdong Provincial Academician Workstation of BGI Synthetic Genomics, and Shenzhen Engineering Laboratory for Innovative Molecular Diagnostics.

Population Bottlenecks and Intra-host Evolution during Human-to-Human Transmission of SARS-CoV-2Population Bottlenecks and Intra-host Evolution during Human-to-Human Transmission of SARS-CoV-2 (preprint)

The emergence of the novel human coronavirus, SARS-CoV-2, causes a global COVID-19 (coronavirus disease 2019) pandemic. Here, we have characterized and compared viral populations of SARS-CoV-2 among COVID-19 patients within and across households. Our work showed an active viral replication activity in the human respiratory tract and the co-existence of genetically distinct viruses within the same host. The inter-host comparison among viral populations further revealed a narrow transmission bottleneck between patients from the same households, suggesting a dominated role of stochastic dynamics in both inter-host and intra-host evolutions. Author summaryIn this study, we compared SARS-CoV-2 populations of 13 Chinese COVID-19 patients. Those viral populations contained a considerable proportion of viral sub-genomic messenger RNAs (sgmRNA), reflecting an active viral replication activity in the respiratory tract tissues. The comparison of 66 identified intra-host variants further showed a low viral genetic distance between intra-household patients and a narrow transmission bottleneck size. Despite the co-existence of genetically distinct viruses within the same host, most intra-host minor variants were not shared between transmission pairs, suggesting a dominated role of stochastic dynamics in both inter-host and intra-host evolutions. Furthermore, the narrow bottleneck and active viral activity in the respiratory tract show that the passage of a small number of virions can cause infection. Our data have therefore delivered a key genomic resource for the SARS-CoV-2 transmission research and enhanced our understanding of the evolutionary dynamics of SARS-CoV-2.

A path towards SARS-CoV-2 attenuation: metabolic pressure on CTP synthesis rules the virus evolution (preprint)

Fighting the COVID-19 epidemic summons deep understanding of the way SARS-CoV-2 taps into its host cell metabolic resources. We describe here the singular metabolic background that creates a bottleneck constraining coronaviruses to evolve towards likely attenuation in the long term. Cytidine triphosphate (CTP) is at the crossroad of the biosynthetic processes that allow the virus to multiply. This is because CTP is in demand for three essential steps. It is a building block of the virus genome, it is required for synthesis of the cytosine-based liponucleotide precursors of the viral envelope and, finally, it is a critical building block of the host transfer RNAs synthesis. The CCA 3-end of all the transfer RNAs required to translate the RNA genome and further transcripts into the proteins used to build active virus copies is not coded in the human genome. It must be synthesized de novo from CTP and ATP. Furthermore, intermediary metabolism is built on compulsory steps of synthesis and salvage of cytosine-based metabolites via uridine triphosphate (UTP) that keep limiting CTP availability. As a consequence, accidental replication errors tend to replace cytosine by uracil in the genome, unless recombination events allow the sequence to return to its ancestral sequences. We document some of the consequences of this situation in the function of viral proteins. We also highlight and provide a raison detre to viperin, an enzyme of innate antiviral immunity, which synthesizes 3-deoxy-3',4-didehydro-CTP (ddhCTP) as an extremely efficient antiviral nucleotide.

Intra-host Variation and Evolutionary Dynamics of SARS-CoV-2 Population in COVID-19 Patients (preprint)

As of middle May 2020, the causative agent of COVID-19, SARS-CoV-2, has infected over 4 million people with more than 300 thousand death as official reports1,2. The key to understanding the biology and virus-host interactions of SARS-CoV-2 requires the knowledge of mutation and evolution of this virus at both inter- and intra-host levels. However, despite quite a few polymorphic sites identified among SARS-CoV-2 populations, intra-host variant spectra and their evolutionary dynamics remain mostly unknown. Here, using deep sequencing data, we achieved and characterized consensus genomes and intra-host genomic variants from 32 serial samples collected from eight patients with COVID-19. The 32 consensus genomes revealed the coexistence of different genotypes within the same patient. We further identified 40 intra-host single nucleotide variants (iSNVs). Most (30/40) iSNVs presented in single patient, while ten iSNVs were found in at least two patients or identical to consensus variants. Comparison of allele frequencies of the iSNVs revealed genetic divergence between intra-host populations of the respiratory tract (RT) and gastrointestinal tract (GIT), mostly driven by bottleneck events among intra-host transmissions. Nonetheless, we observed a maintained viral genetic diversity within GIT, showing an increased population with accumulated mutations developed in the tissue-specific environments. The iSNVs identified here not only show spatial divergence of intra-host viral populations, but also provide new insights into the complex virus-host interactions.

Detection of a novel coronavirus (SARS-CoV-2) by real-time reverse transcription-polymerase chain reaction, China, 2020 (preprint)

Background: During the outbreak of unexplained pneumonia in the city of Wuhan in the late December, 2019, a novel coronavirus named SARS-CoV-2 was identified as the cause of this outbreak.Methods: A real-time polymerase chain reaction, which targets the orf1ab gene of viral genome, was established to detect and identify the SARS-CoV-2. We used this assay to screen 309 samples from persons with suspected SARS-CoV-2 infection in Wuhan. Then 6 close-phylogenic coronaviruses and 7 viruses which could cause pneumonia were detected. Moreover, 57 clinical samples infected with other viruses and 77 healthy samples were also tested.Results: The limit of detection of the assay was 6.25 copies per reaction in the detection of cRNA transcribed in vitro. The results of detection of throat and fecal swabs from persons with suspected SARS-CoV-2 infection showed throat swabs were more sensitive than fecal swabs during the first 15 days after onset of symptoms (throat: 56.80%, fecal: 30.43%), while the situation was reversed after 15 days (throat: 20.83%, fecal: 27.58%). And matched pair tests suggested the sputum samples had higher virus loads than throat swabs in the patients (P < 0.05). There was no cross-reaction when we detected the inactive culture of six other coronaviruses (human coronavirus 229E, NL63, OC43, HKU1, SARS-CoV, MERS-CoV) and seven other viruses (influenza virus A H1N1, influenza virus A H3N2, influenza virus B, parainfluenza viruses 1, 2, and 3; and respiratory syncytial virus). Besides, 27 BALF samples from pneumonia patients infected with human coronavirus 229E, OC43, HKU1 or human adenovirus 7, 30 throat swabs from patients infected with H1N1 and 77 throat swabs from healthy people tested negative by this assay.Conclusions: The results indicated that the assay specifically and sensitively detected the SARS-CoV-2.

Implementation of radiotherapy in a tertiary hospital in Zhejiang province during the epidemic of COVID-19 / 中华放射医学与防护杂志

Objective@#To explore the clinical practice of delivering radiotherapy during the outbreak of 2019 novel coronavirus disease(COVID-19).@*Methods@#During this epidemic period, available methods including but not limited to: strict disinfection, body temperature monitoring, learning relevant knowledge by all staffs to ensure the safety of radiotherapy treatment. Relevant data including proportion of radiotherapy, time from scanning to the first time of radiation delivery and degree of satisfaction in the view of staffs and patients, respectively.@*Results@#A total of 60 patients received radiation therapy in the department of radiotherapy of Zhejiang Provincial People’s Hospital (2020-02-11). Compared with the same period in 2019 (after the Spring Festival), the total number of patients receiving radiotherapy was decreased from 72 to 60(83.3%). Among them, the number of patients receiving palliative radiation therapy decreased significantly, while the proportion of radical, preoperative and/or postoperative radiotherapy/radiochemotherapy did not significantly decrease. There was significant difference between different years (χ2=6.967, P<0.05). The median time for newly admitted patients to receive radiotherapy was two days, which was not significantly longer than the interval in 2019 (P>0.05). Staffs and patients were generally satisfied with the current prevention measures.@*Conclusions@#Using a variety of prevention and control methods, and taking full account of medical safety and patient benefits, radiation-related activities can be carried out during the epidemic.

Effect of large-scale testing platform in prevention and control of the COVID-19 pandemic: an empirical study with a novel numerical model (preprint)

Background: China adopted an unprecedented province-scale quarantine since January 23rd 2020, after the novel coronavirus (COVID-19) broke out in Wuhan in December 2019. Responding to the challenge of limited testing capacity, large-scale standardized and fully-automated laboratory (Huo-Yan) was built as an ad-hoc measure. There was so far no empirical data or mathematical model to reveal the impact of the testing capacity improvement since the quarantine. Methods: We integrated public data released by the Health Commission of Hubei Province and Huo-Yan Laboratory testing data into a novel differential model with non-linear transfer coefficients and competitive compartments, to evaluate the trends of suspected cases under different nucleic acid testing capacities. Results: Without the establishment of Huo-Yan, the suspected cases would increased by 47% to 33,700, the corresponding cost of the quarantine would be doubled, and the turning point of the increment of suspected cases and the achievement of "daily settlement" (all daily new discovered suspected cases were diagnosed according to the nucleic acid testing results) would be delayed for a whole week and 11 days. If the Huo-Yan Laboratory could ran at its full capacity, the number of suspected cases could started to decrease at least a week earlier, the peak of suspected cases would be reduced by at least 44% and the quarantine cost could be reduced by more than 72%. Ideally, if a daily testing capacity of 10,500 could achieved immediately after the Hubei lockdown, "daily settlement" for all suspected cases would be achieved immediately. Conclusions: Large-scale and standardized clinical testing platform with nucleic acid testing, high-throughput sequencing and immunoprotein assessment capabilities need to be implemented simultaneously in order to maximize the effect of quarantine and minimize the duration and cost. Such infrastructure like Huo-Yan, is of great significance for the early prevention and control of infectious diseases for both common times and emergencies.

Multiple approaches for massively parallel sequencing of HCoV-19 genomes directly from clinical samples (preprint)

COVID-19 has caused a major epidemic worldwide, however, much is yet to be known about the epidemiology and evolution of the virus. One reason is that the challenges underneath sequencing HCoV-19 directly from clinical samples have not been completely tackled. Here we illustrate the application of amplicon and hybrid capture (capture)-based sequencing, as well as ultra-high-throughput metatranscriptomic (meta) sequencing in retrieving complete genomes, inter-individual and intra-individual variations of HCoV-19 from clinical samples covering a range of sample types and viral load. We also examine and compare the bias, sensitivity, accuracy, and other characteristics of these approaches in a comprehensive manner. This is, to date, the first work systematically implements amplicon and capture approaches in sequencing HCoV-19, as well as the first comparative study across methods. Our work offers practical solutions for genome sequencing and analyses of HCoV-19 and other emerging viruses.