2022年 · 第04卷第05期

As a foundation of life-science research and advancement, biobanking has played a critical role and made tremendous contributions to healthcare, biotechnology, disease control and prevention, as well as bio-conservation for the benefit of all humankind. This paper starts with a brief introduction of basic concepts, history, classification, and significance of biobanking, followed by a discussion on cryobiology fundamentals and key challenges faced by cryopreservation in biobanking. A special case discussion on the cryopreservation and biobanking of pathogenic microorganisms to meet both the unmet needs for biomedical research and the urgent demand for developing countermeasures against the ongoing coronavirus disease 2019 (COVID-19) pandemic is highlighted with insightful recommendations for future studies.

Biobanking facilities are well established in high-income settings, where substantial funding has been invested in infrastructure. In contrast, such facilities are much less developed in resource-restricted settings. However, low-and middle-income countries (LMICs) still face a disproportionately high infectious diseases burden. Thus, the further development of infrastructure facilities, including biobanks is warranted as an important component of this unfolding clinical research environment. This perspective manuscript summarises the challenges and enablers for biobanking in LMICs, with a particular focus on infectious diseases, incorporating some of the lessons learned from the recent coronavirus disease 2019 (COVID-19) pandemic.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused many deaths and contributed to a tremendous public health concern worldwide since 2020. Angiotensin-converting enzyme 2 (ACE2) binds to the SARS-CoV-2 virus as a receptor. The challenge of different nonhuman primate (NHP) species by SARS-CoV-2 virus demonstrated different effects on virus replication and disease pathology. This study characterizes differences between host ACE2 sequences of three NHP species: Macaca mulatta, Macaca fascicularis, and Chlorocebus sabaeus. In addition, the binding affinity between the ACE2 ectodomain and the SARS-CoV-2 S receptor-binding domain (RBD) was analyzed. Variation of ACE2 sequence among NHP species and the binding affinity may account for different susceptibility and responses to SARS-CoV-2 infection.

Viral isolation in cell cultures has been regarded for decades as the "gold standard" for the laboratory diagnosis of influenza viral infections. Not all viral strains could be isolated from clinical samples. This study aimed to quantify the viral load in the samples before isolation to save working time and improve working efficiency. Four hundred samples from patients with influenza-like cases were confirmed pdmH1N1 positive (200 cases) and B Victoria (BV) positive (200 cases) by whole-genome sequencing and analyzed by ddPCR for viral load in samples before isolation, and isolation results were verified by hemagglutination (HA) assay and hemagglutination-inhibition (HI) tests. Probit regression analysis was used to calculate the isolation viral load limit with a 95% probability level by SPSS 19.0 software. The results showed that the isolation limit of viral load was 4.9 × 10⁴ (95% CI: 2.5 × 10⁴-9.0 × 10⁴) copies/mL for pdmH1N1 and 1.9 × 10⁴ (95% CI: 7.8 × 10³-3.6 × 10⁴) copies/mL for BV. The isolation rate of clinical samples is positively correlated with the viral load in clinical samples, which can be used for viral culture, providing important guidance for daily work.

Reverse genetics via targeted modification of gene sequences to obtain a phenotype and the inference of a gene's function or regulatory mechanism is widely used as a potent tool in viral biology and application. However, while reverse genetics has contributed significantly to our understanding of molecular biology and the pathogenesis of viruses, its accessibility (operation) and openness (data) have raised many concerns regarding biosafety and biosecurity. In this review, we retrospectively examine the development of reverse genetics and its applications in virology, then emphasize global biosafety and biosecurity concerns regarding reverse genetics, and summarize global regulations, governance, and laws on reverse genetics. This review seeks to enhance our understanding and rational application of reverse genetics technology for the benefit of humankind.

The outbreak of coronavirus disease 2019 (COVID-19) has adversely affected the public domain causing unprecedented cases and high mortality across the globe. This has brought back the concept of biosafety into the spotlight to solve biosafety problems in developing diagnostics and therapeutics to treat COVID-19. The advances in nanotechnology and material science in combination with medicinal chemistry have provided a new perspective to overcome this crisis. Herein, we discuss the efforts of researchers in the field of material science in developing personal protective equipment (PPE), detection devices, vaccines, drug delivery systems, and medical equipment. Such a synergistic approach of disciplines can strengthen the research to develop biosafety products in solving biosafety problems.

Long-term and stable preservation of bacteriophages is of crucial importance. Although many efforts have been made in the past decades to explore the influence of external factors on bacteriophage preservation, there is still little understanding, and a systematic description is lacking. In this study, we explored the influence of different factors on the preservation of lytic bacteriophage VP3, one of the typing bacteriophages of Vibrio cholerae O1 biotype El Tor, and attempted to optimize its preservation. We examined external factors, including temperature, solution, and cryoprotectant, in stable cooling/freezing conditions or alternate cooling/freezing and thawing. We found that whether in Luria-Bertani (LB) medium or SM buffer, in terms of 20-week stable cooling or freezing, -20 °C was the most damaging while 4 °C, -80 °C, and -196 °C were protective. Thirteen cycles of alternate cooling/freezing and thawing caused a loss in the survival rates of bacteriophages. The addition of cryoprotectant, glycerol (30%, w/v) or dimethyl sulfoxide (DMSO, 10%, w/v) significantly improved the survival rates of bacteriophages preserved at -20 °C. However, at 4 °C, -80 °C, and -196 °C, the cryoprotectant effect was only slightly positive or even harmful. In summary, for bacteriophage VP3, the best preservation method is to directly preserve the bacteriophage stocks in LB medium at -80 °C or -196 °C instead of storing them in SM buffer or adding cryoprotectant. Our results provided insights into the external influencing factors on bacteriophage VP3 during preservation at low temperature and can be applied to the optimization of bacteriophage preservation in the future.

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to unprecedented social and economic disruption. Many nucleic acid testing (NAT) laboratories in China have been established to control the epidemic better. This proficiency testing (PT) aims to evaluate the participants’ performance in qualitative and quantitative SARS-CoV-2 NAT and to explore the factors that contribute to differences in detection capabilities. Two different concentrations of RNA samples (A, B) were used for quantitative PT. Pseudovirus samples D, E (different concentrations) and negative sample (F) were used for qualitative PT. 50 data sets were reported for qualitative PT, of which 74.00% were entirely correct for all samples. Forty-two laboratories participated in the quantitative PT. 37 submitted all gene results, of which only 56.76% were satisfactory. For qualitative detection, it is suggested that laboratories should strengthen personnel training, select qualified detection kits, and reduce cross-contamination to improve detection accuracy. For quantitative detection, the results of the reverse transcription digital PCR (RT-dPCR) method were more comparable and reliable than those of reverse transcription quantitative PCR (RT-qPCR). The copy number concentration of ORF1ab and N in samples A and B scattered in 85, 223, 50, and 106 folds, respectively. The differences in the quantitative result of RT-qPCR was mainly caused by the non-standard use of reference materials and the lack of personnel operating skills. Comparing the satisfaction of participants participating in both quantitative and qualitative proficiency testing, 95.65% of the laboratories with satisfactory quantitative results also judged the qualitative results correctly, while 85.71% of the laboratories with unsatisfactory quantitative results were also unsatisfied with their qualitative judgments. Therefore, the quantitative ability is the basis of qualitative judgment. Overall, participants from hospitals reported more satisfactory results than those from enterprises and universities. Therefore, surveillance, daily qualitiy control and standardized operating procedures should be strengthened to improve the capability of SARS-CoV-2 NAT.

Limited data is available on the coronavirus disease 2019 (COVID-19), critical illness rate, and in-hospital mortality in the African setting. This study investigates determinants of critical illness and in-hospital mortality among COVID-19 patients in Kenya. We conducted a retrospective cohort study at Kenyatta National Hospital (KNH) in Kenya. Multivariate logistic regression and Cox proportional hazard regression were employed to determine predictor factors for intensive care unit (ICU) admission and in-hospital mortality, respectively. In addition, the Kaplan-Meier model was used to compare the survival times using log-rank tests. As a result, 346 (19.3%) COVID-19 patients were admitted to ICU, and 271 (15.1%) died. The majority of those admitted to the hospital were male, 1,137 (63.4%) and asymptomatic, 1,357 (75.7%). The most prevalent clinical features were shortness of breath, fever, and dry cough. In addition, older age, male, health status, patient on oxygen (O₂), oxygen saturation levels (SPO₂), headache, dry cough, comorbidities, obesity, cardiovascular diseases (CVDs), diabetes, chronic lung disease (CLD), and malignancy/cancer can predicate the risk of ICU admission, with an area under the receiver operating characteristic curve (AUC-ROC) of 0.90 (95% confidence interval [CI]: 0.88-0.92). Survival analysis indicated 271 (15.1%) patients died and identified older age, male, headache, shortness of breath, health status, patient on oxygen, SPO₂, headache, comorbidity, CVDs, diabetes, CLD, malignancy/cancer, and smoking as risk factors for mortality (AUC-ROC: 0.90, 95% CI: 0.89-0.91). This is the first attempt to explore predictors for ICU admission and hospital mortality among COVID-19 patients in Kenya.

In order to effectively implement the Tianjin Biosecurity Guidelines for Codes of Conduct for Scientists, biosecurity awareness-raising and education are essential because if these are not in place scientists will not understand the need for biosecurity codes of conduct. In an effort to assist in the implementation of the guidelines, a small-scale survey was carried out in early 2022 of biosecurity awareness-raising and education projects that have been developed over the last two decades to discover what resources and experience have been accumulated. It is argued that the survey demonstrates that much of what is needed to implement the guidelines effectively has been developed, but that there are specific deficiencies that need to be remedied quickly. In particular, an updated teaching resource covering the core issues related to the Biological and Toxin Weapons Convention (BTWC) and the problem of dual use in scientific research needs to be made widely available and translated into at least the six official United Nations (UN) languages. Additionally, more specialists from the Humanities with expertise in ethics need to become involved in biosecurity awareness-raising and education activities. While advantage should be taken now of the available national, regional and international networks of people involved in related activities, it is suggested that in the longer term cooperation in biosecurity awareness-raising and education will benefit from the development of an equivalent organisation to the International Nuclear Security Education Network (INSEN) organised through the International Atomic Energy Agency (IAEA).