Optimizing heat inactivation for SARS-CoV-2 at 95 °C and its implications: A standardized approach.

Background: Standardized and validated heat inactivation procedure for Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are not available. For heat inactivation, various protocols were reported to prepare External Quality Assessment Programme (EQAP) samples without direct comparison between different durations. Objective: To assess the heat inactivation procedures against SARS-CoV-2. The efficacy of the optimized condition was reflected by the results from laboratories testing the EQAP samples. Study design: The SARS-CoV-2 strain was exposed to 95 °C in a water bath for three different time intervals, 5 min, 10 min and 15 min, respectively. The efficacy of inactivation was confirmed by the absence of cytopathic effects and decreasing viral load in 3 successive cell line passages. The viral stock inactivated by the optimal time interval was dispatched to EQAP participants and the result returned were analyzed. Results: All of the three conditions were capable of inactivating the SARS-CoV-2 of viral load at around cycle threshold value of 10. When the 95 °C 10 min condition was chosen to prepare SARS-CoV-2 EQAP samples, they showed sufficient homogeneity and stability. High degree of consensus was observed among EQAP participants in all samples dispatched. Conclusions: The conditions evaluated in the present study could be helpful for laboratories in preparing SARS-CoV-2 EQAP samples.

Non-specific signals in real-time RT-PCR for detecting respiratory viruses.

We describe our experiences in investigating the origin of non-specific signals during the development phase of a multiplex PCR assay for respiratory viruses. After ruling out various sources of error, eventually we discovered the non-specific signal was related to the particular lot of the PCR kit.

Lessons learned 1 year after SARS-CoV-2 emergence leading to COVID-19 pandemic.

Without modern medical management and vaccines, the severity of the Coronavirus Disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2) might approach the magnitude of 1894-plague (12 million deaths) and 1918-A(H1N1) influenza (50 million deaths) pandemics. The COVID-19 pandemic was heralded by the 2003 SARS epidemic which led to the discovery of human and civet SARS-CoV-1, bat SARS-related-CoVs, Middle East respiratory syndrome (MERS)-related bat CoV HKU4 and HKU5, and other novel animal coronaviruses. The suspected animal-to-human jumping of 4 betacoronaviruses including the human coronaviruses OC43(1890), SARS-CoV-1(2003), MERS-CoV(2012), and SARS-CoV-2(2019) indicates their significant pandemic potential. The presence of a large reservoir of coronaviruses in bats and other wild mammals, culture of mixing and selling them in urban markets with suboptimal hygiene, habit of eating exotic mammals in highly populated areas, and the rapid and frequent air travels from these areas are perfect ingredients for brewing rapidly exploding epidemics. The possibility of emergence of a hypothetical SARS-CoV-3 or other novel viruses from animals or laboratories, and therefore needs for global preparedness should not be ignored. We reviewed representative publications on the epidemiology, virology, clinical manifestations, pathology, laboratory diagnostics, treatment, vaccination, and infection control of COVID-19 as of 20 January 2021, which is 1 year after person-to-person transmission of SARS-CoV-2 was announced. The difficulties of mass testing, labour-intensive contact tracing, importance of compliance to universal masking, low efficacy of antiviral treatment for severe disease, possibilities of vaccine or antiviral-resistant virus variants and SARS-CoV-2 becoming another common cold coronavirus are discussed.

Detection of SARS-CoV-2 in conjunctival secretions from patients without ocular symptoms.

PURPOSE: To evaluate the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in conjunctival secretions from patients without ocular symptoms. METHODS: Conjunctival swabs were prospectively collected from laboratory-confirmed Coronavirus disease 2019 (COVID-19) patients without ocular symptoms for reverse transcription-polymerase chain reaction (RT-PCR) and viral culture. RESULTS: A total of 158 conjunctival swabs were obtained from 49 laboratory-confirmed COVID-19 patients. The median duration of illness when the first conjunctival swab was obtained was 10 days (range 2-27 days). Four conjunctival swabs from four different patients (4/49, 8.2%) were positive for SARS-CoV-2 RNA by RT-PCR. The Ct values ranged from 32.7 to 37.7 (mean 35.4). Viral cultures were negative for all four RT-PCR-positive conjunctival swabs. CONCLUSION: Conjunctival secretions of a minority of COVID-19 patients without ocular symptoms may contain relatively low levels of SARS-CoV-2 RNA, but their infectiousness remains undetermined. Appropriate infection control measures should be implemented during ophthalmological assessment of COVID-19 patients to prevent potential nosocomial transmission of SARS-CoV-2.

Polyclonal Burkholderia cepacia Complex Outbreak in Peritoneal Dialysis Patients Caused by Contaminated Aqueous Chlorhexidine.

Whether Burkholderia cepacia complex should be an objectionable organism in antiseptic solutions with acceptable total bacterial counts is controversial. By using next-generation sequencing, we documented a polyclonal B. cepacia complex outbreak affecting peritoneal dialysis patients in Hong Kong that was caused by contaminated chlorhexidine solutions. Epidemiologic investigations at a manufacturing site identified a semiautomated packaging machine as the probable source of contamination in some of the brands. Use of whole-genome sequencing differentiated the isolates into 3 brand-specific clonal types. Changes in exit site care recommendations, rapid recall of affected products, and tightening of regulatory control for chlorhexidine-containing skin antiseptics could prevent future similar outbreaks. Environmental opportunistic pathogens, including B. cepacia complex, might be included in regular surveillance as indicator organisms for monitoring environmental contamination.

Injectable diblock copolypeptide hydrogel provides platform to deliver effective concentrations of paclitaxel to an intracranial xenograft model of glioblastoma.

INTRODUCTION: Surgical resection and systemic chemotherapy with temozolomide remain the mainstay for treatment of glioblastoma. However, many patients are not candidates for surgical resection given inaccessible tumor location or poor health status. Furthermore, despite being first line treatment, temozolomide has only limited efficacy. METHODS: The development of injectable hydrogel-based carrier systems allows for the delivery of a wide range of chemotherapeutics that can achieve high local concentrations, thus potentially avoiding systemic side effects and wide-spread neurotoxicity. To test this modality in a realistic environment, we developed a diblock copolypeptide hydrogel (DCH) capable of carrying and releasing paclitaxel, a compound that we found to be highly potent against primary gliomasphere cells. RESULTS: The DCH produced minimal tissue reactivity and was well tolerated in the immune-competent mouse brain. Paclitaxel-loaded hydrogel induced less tissue damage, cellular inflammation and reactive astrocytes than cremaphor-taxol (typical taxol-carrier) or hydrogel alone. In a deep subcortical xenograft model of glioblastoma in immunodeficient mice, injection of paclitaxel-loaded hydrogel led to local tumor control and improved survival. However, the tumor cells were highly migratory and were able to eventually escape the area of treatment. CONCLUSIONS: These findings suggest this technology may be ultimately applicable to patients with deep-seated inoperable tumors, but as currently formulated, complete tumor eradication would be highly unlikely. Future studies should focus on targeting the migratory potential of surviving cells.

Early-Morning vs Spot Posterior Oropharyngeal Saliva for Diagnosis of SARS-CoV-2 Infection: Implication of Timing of Specimen Collection for Community-Wide Screening.

BACKGROUND: Posterior oropharyngeal saliva is increasingly recognized as a valid respiratory specimen for SARS-CoV-2 diagnosis. It is easy to collect and suitable for community-wide screening. The optimal timing of collection is currently unknown, and we speculate that an early-morning specimen before oral hygiene and breakfast would increase the diagnostic yield. METHODS: Posterior oropharyngeal saliva was collected at 5 different time points within the same day from 18 patients with previously confirmed SARS-CoV-2 infection by molecular testing. Cycle threshold (Ct) values were compared. RESULTS: There was an overall trend of lower Ct values from specimens collected in the early morning, with a gradual decrease of viral load towards nighttime, but reaching statistical significance only when compared with the specimens collected at bedtime. Eight out of 13 subjects had a higher viral load in the early morning than the rest of the 4 time points (before lunch, before teatime at 3 pm, before dinner, before bedtime). CONCLUSIONS: The result suggests a diurnal variation of viral shedding from the upper respiratory tract with a trend showing higher viral load in the early morning. For community screening purposes, posterior oropharyngeal saliva could be taken throughout the day, but preferably in the early morning to maximize the yield.

SARS-CoV-2 shedding and seroconversion among passengers quarantined after disembarking a cruise ship: a case series.

BACKGROUND: A cruise ship is a closed-off environment that simulates the basic functioning of a city in terms of living conditions and interpersonal interactions. Thus, the Diamond Princess cruise ship, which was quarantined because of an onboard outbreak of COVID-19 in February, 2020, provides an opportunity to define the shedding pattern of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and patient antibody responses before and after the onset of symptoms. METHODS: We recruited adult (≥18 years) passengers from Hong Kong who had been on board the Diamond Princess cruise ship docked in Yokohama, Japan in February, 2020. All participants had been found to be negative for SARS-CoV-2 by RT-PCR 4 days before disembarking and were transferred to further quarantine in a public estate in Hong Kong, where they were recruited. Participants were prospectively screened by quantitative RT-PCR (RT-qPCR) of nasopharyngeal and throat swabs, and serum IgG and IgM against internal nucleoprotein and the surface spike receptor-binding protein (RBD) of SARS-CoV-2 at baseline (upon entering quarantine) and on days 4, 8, and 12 of quarantine. FINDINGS: On Feb 22, 2020, 215 adults were recruited, of whom nine (4%; 95% CI 2-8) were positive for SARS-CoV-2 by RT-qPCR or serology and were hospitalised. Of these nine patients, nasopharyngeal swab RT-qPCR was positive in eight patients (89%; 57-99) at baseline. All nine patients were positive for anti-RBD IgG by day 8. Eight (89%; 57-99) were simultaneously positive for nasopharyngeal swab RT-PCR and anti-RBD IgG. One patient who was positive for anti-RBD IgG and had a negative viral load had multifocal peripheral ground-glass changes on high-resolution CT that were typical of COVID-19. Five patients (56%; 27-81) with ground-glass changes on high-resolution CT were found to have higher anti-nucleoprotein-IgG OD values on day 8 and 12 and anti-RBD IgG OD value on day 12 than patients without ground-glass changes. Six (67%; 35-88) patients remained asymptomatic throughout the 14-day quarantine period. INTERPRETATION: Patients with COVID-19 can develop asymptomatic lung infection with viral shedding and those with evidence of pneumonia on imaging tend to have an increased antibody response. Positive IgG or IgM confirmed infection of COVID-19 in both symptomatic and asymptomatic patients. A combination of RT-PCR and serology should be implemented for case finding and contact tracing to facilitate early diagnosis, prompt isolation, and treatment. FUNDING: Shaw Foundation Hong Kong; Sanming-Project of Medicine (Shenzhen); High Level-Hospital Program (Guangdong Health Commission).

Severe underlying liver diseases and high mortality associated with Laribacter hongkongensis bacteremia.

We characterized a strain of Laribacter hongkongensis isolated from the blood of a patient with fatal sepsis, who had alcoholic cirrhosis with ascites and portal hypertension. L. hongkongensis bacteremia is associated with underlying liver diseases (P < 0.001) and mortality (P < 0.05), whereas L. hongkongensis gastroenteritis is associated with recent travel history (P < 0.05).

Measles outbreak from Hong Kong International Airport to the hospital due to secondary vaccine failure in healthcare workers.

OBJECTIVE: To report an outbreak of measles with epidemiological link between Hong Kong International Airport (HKIA) and a hospital. METHODS: Epidemiological investigations, patients' measles serology, and phylogenetic analysis of the hemagglutinin (H) and nucleoprotein (N) genes of measles virus isolates were conducted. RESULTS: In total, 29 HKIA staff of diverse ranks and working locations were infected with measles within 1 month. Significantly fewer affected staff had history of travel than non-HKIA-related measles patients [10 of 29 (34.5%) vs 28 of 35 (80%); P < .01]. Of 9 airport staff who could recall detailed exposure history, 6 (66.7%) had visited self-service food premises at HKIA during the incubation period, where food trays, as observed during the epidemiological field investigation, were not washed after use. Furthermore, 1 airport baggage handler who was admitted to hospital A before rash onset infected 2 healthcare workers (HCWs) known to have 2 doses of MMR vaccination with positive measles IgG and lower viral loads in respiratory specimens. Infections in these 2 HCWs warranted contact tracing of another 168 persons (97 patients and 71 HCWs). Phylogenetic comparison of H and N gene sequences confirmed the clonality of outbreak strains. CONCLUSION: Despite good herd immunity with overall seroprevalence of >95% against measles, major outbreaks of measles occurred among HKIA staff having daily contact with many international pssengers. Lessons from severe acute respiratory syndrome (SARS) and measles outbreaks suggested that an airport can be a strategic epidemic center. Pre-exanthem transmission of measles from airport staff to HCWs with secondary vaccine failure poses a grave challenge to hospital infection control.

Fatal pancytopenia due to albendazole treatment for strongyloidiasis.

We report 7 cases of strongyloidiasis that had occurred from 2016 through 2017 in a tertiary hospital of southern China. Three of the 7 patients (age 66-77) with farming exposure many years ago developed symptomatic infection while receiving immunosuppressant for underlying medical conditions. The majority of them were treated with albendazole due to unavailability of ivermectin in mainland China. One of the 7 patients, with underlying IgG4 sclerosing cholangitis and secondary biliary cirrhosis was on immunosuppressives and developed severe pancytopenia 15 days after albendazole treatment. He ultimately died of polymicrobial sepsis. This was the second fatal case being reported in the literature as a consequence of albendazole-induced myelosuppression. We have undertaken a review of the literature regarding the use of albendazole for strongyloidiasis and its adverse effect with a focus on myelosuppression as a rare but potentially serious event.