A Sensitive and Specific Competitive Enzyme-Linked Immunosorbent Assay for Serodiagnosis of COVID-19 in Animals.

In addition to human cases, cases of COVID-19 in captive animals and pets are increasingly reported. This raises the concern for two-way COVID-19 transmission between humans and animals. Here, we developed a SARS-CoV-2 nucleocapsid protein-based competitive enzyme-linked immunosorbent assay (cELISA) for serodiagnosis of COVID-19 which can theoretically be used in virtually all kinds of animals. We used 187 serum samples from patients with/without COVID-19, laboratory animals immunized with inactive SARS-CoV-2 virions, COVID-19-negative animals, and animals seropositive to other betacoronaviruses. A cut-off percent inhibition value of 22.345% was determined and the analytical sensitivity and specificity were found to be 1:64-1:256 and 93.9%, respectively. Evaluation on its diagnostic performance using 155 serum samples from COVID-19-negative animals and COVID-19 human patients showed a diagnostic sensitivity and specificity of 80.8% and 100%, respectively. The cELISA can be incorporated into routine blood testing of farmed/captive animals for COVID-19 surveillance.

Molecular Evolution of Human Coronavirus 229E in Hong Kong and a Fatal COVID-19 Case Involving Coinfection with a Novel Human Coronavirus 229E Genogroup.

Compared to other human coronaviruses, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) are relatively understudied. We report a fatal case of COVID-19 pneumonia coinfected with HCoV-229E in Hong Kong. Genome sequencing of SARS-CoV-2 and HCoV-229E from a nasopharyngeal sample of the patient showed that the SARS-CoV-2 strain HK13 was most closely related to SARS-CoV-2 type strain Wuhan-Hu-1 (99.99% nucleotide identity), compatible with his recent history of travel to Wuhan. The HCoV-229E strain HK20-42 was most closely related to HCoV-229E strain SC0865 from the United States (99.86% nucleotide identity). To investigate if it may represent a newly emerged HCoV-229E genotype in Hong Kong, we retrieved 41 archived respiratory samples that tested positive for HCoV-229E from 2004 to 2019. Pneumonia and exacerbations of chronic airway diseases were common among infected patients. Complete RdRp, S, and N gene sequencing of the 41 HCoV-229E strains revealed that our contemporary HCoV-229E strains have undergone significant genetic drift with clustering of strains in chronological order. Two novel genogroups were identified, in addition to previously described genogroups 1 to 4, with recent circulating strains including strain HK20-42 belonging to novel genogroup 6. Positive selection was detected in the spike protein and receptor-binding domain, which may be important for viral evolution at the receptor-binding interphase. Molecular dating analysis showed that HCoV-229E shared the most recent common ancestor with bat and camel/alpaca 229E-related viruses at ∼1884, while camel/alpaca viruses had a relatively recent common ancestor at ∼1999. Further studies are required to ascertain the evolutionary origin and path of HCoV-229E.IMPORTANCE Since its first appearance in the 1960s, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) have been relatively understudied. In this study, we report a fatal case of COVID-19 coinfected with HCoV-229E in Hong Kong. Genome sequencing revealed that our SARS-CoV-2 strain is highly identical to the SARS-CoV-2 strain from Wuhan, compatible with the patient's recent travel history, whereas our HCoV-229E strain in this study is highly identical to a recent strain in the United States. We also retrieved 41 archived HCoV-229E strains from 2004 to 2019 in Hong Kong for sequence analysis. Pneumonia and exacerbations of chronic airway diseases were common diagnoses among the 41 patients. The results showed that HCoV-229E was evolving in chronological order. Two novel genogroups were identified in addition to the four preexisting HCoV-229E genogroups, with recent circulating strains belonging to novel genogroup 6. Molecular clock analysis dated bat-to-human and bat-to-camelid transmission to as early as 1884.

Multidisciplinary simulation training for endotracheal intubation during COVID-19 in one Hong Kong regional hospital: strengthening of existing procedures and preparedness.

Introduction: In early 2020, our hospital responded with high alertness when novel coronavirus SARS-CoV-2 appeared. A hospital-based training programme was rapidly arranged to prepare staff for the imminent threat. Objective: We developed a hospital-wide multidisciplinary infection control training programme on endotracheal intubation for healthcare workers to minimise nosocomial spread of COVID-19 during this high-stress and time-sensitive risky procedure. Methodology: Major stakeholders (Quality & Safety Department, Infection Control Team, Central Nursing Division, high-risk clinical departments and hospital training centre) formed a training programme task group. This group was tasked with developing high-fidelity scenario-based simulation training curriculum for COVID-19 endotracheal intubation with standard workflow and infection control practice. This group then implemented and evaluated the training programme for its effectiveness. Results: 101 training classes of 2-hour session were conducted from 5 February to 18 March 2020, involving 1415 hospital staff (~81% of target participants with training needs) either inside the hospital training centre or as in situ simulation training (intensive care unit or accident and emergency department). Learners' satisfaction was reflected by overall positive response percentage at 90%. Opinions of participating staff were incorporated into the standard airway management and infection control practice for endotracheal intubation of adult patients with COVID-19. Thirty-five patients with COVID-19 were intubated with the current workflow and guideline without any nosocomial transmission. Conclusion: An early planned and well-structured multidisciplinary hospital-wide simulation training programme was organised expeditiously to provide extensive staff coverage. The insight and experience gained from this project is valuable for future infectious disease challenges.

First Report of a Fatal Case Associated with EV-D68 Infection in Hong Kong and Emergence of an Interclade Recombinant in China Revealed by Genome Analysis.

A fatal case associated with enterovirus D68 (EV-D68) infection affecting a 10-year-old boy was reported in Hong Kong in 2014. To examine if a new strain has emerged in Hong Kong, we sequenced the partial genome of the EV-D68 strain identified from the fatal case and the complete VP1, and partial 5'UTR and 2C sequences of nine additional EV-D68 strains isolated from patients in Hong Kong. Sequence analysis indicated that a cluster of strains including the previously recognized A2 strains should belong to a separate clade, clade D, which is further divided into subclades D1 and D2. Among the 10 EV-D68 strains, 7 (including the fatal case) belonged to the previously described, newly emerged subclade B3, 2 belonged to subclade B1, and 1 belonged to subclade D1. Three EV-D68 strains, each from subclades B1, B3, and D1, were selected for complete genome sequencing and recombination analysis. While no evidence of recombination was noted among local strains, interclade recombination was identified in subclade D2 strains detected in mainland China in 2008 with VP2 acquired from clade A. This study supports the reclassification of subclade A2 into clade D1, and demonstrates interclade recombination between clades A and D2 in EV-D68 strains from China.

Elizabethkingia anophelis bacteremia is associated with clinically significant infections and high mortality.

Unlike Elizabethkingia meningoseptica, the clinical importance of E. anophelis is poorly understood. We determined the clinical and molecular epidemiology of bacteremia caused by Elizabethkingia-like species from five regional hospitals in Hong Kong. Among 45 episodes of Elizabethkingia-like bacteremia, 21 were caused by Elizabethkingia, including 17 E. anophelis, three E. meningoseptica and one E. miricola; while 24 were caused by other diverse genera/species, as determined by 16S rRNA gene sequencing. Of the 17 cases of E. anophelis bacteremia, 15 (88%) were clinically significant. The most common diagnosis was pneumonia (n = 5), followed by catheter-related bacteremia (n = 4), neonatal meningitis (n = 3), nosocomial bacteremia (n = 2) and neutropenic fever (n = 1). E. anophelis bacteremia was commonly associated with complications and carried 23.5% mortality. In contrast, of the 24 episodes of bacteremia due to non-Elizabethkingia species, 16 (67%) were clinically insignificant. Compared to non-Elizabethkingia bacteremia, Elizabethkingia bacteremia was associated with more clinically significant infections (P < 0.01) and positive cultures from other sites (P < 0.01), less polymicrobial bacteremia (P < 0.01), and higher complication (P < 0.05) and mortality (P < 0.05) rates. Elizabethkingia bacteremia is predominantly caused by E. anophelis instead of E. meningoseptica. Elizabethkingia bacteremia, especially due to E. anophelis, carries significant morbidity and mortality, and should be considered clinically significant unless proven otherwise.

Hospital Outbreak of Pulmonary and Cutaneous Zygomycosis due to Contaminated Linen Items From Substandard Laundry.

BACKGROUND: Healthcare laundry-related infection is rare, and pulmonary zygomycosis due to contaminated hospital linens has never been reported. METHODS: We reported an outbreak investigation of zygomycosis in a university-affiliated teaching hospital. Air samplers, sponge swabs and Replicate Organism Detection and Counting (RODAC) contact plates were used for environmental sampling. The fungal isolates from clinical and environmental samples were identified by morphology, MALDI-TOF MS, and ITS1-5.8S-ITS2 rRNA gene cluster sequencing. RESULTS: From 2 June 2015 to 18 July 2015, 6 immunosuppressed patients developed pulmonary (n = 4) and/or cutaneous (n = 3) infection by a spore-forming mold, Rhizopus microsporus, through direct inhalation and skin contact of contaminated linen items supplied by a designated laundry. Seventy (27.8%) of 252 freshly laundered clothing and 15 (3.4%) of 443 nonclothing laundered linen items (pillow case, bed sheet, draw sheet) were contaminated by R. microsporus, which was significantly higher than those from other hospital laundries (0%, n = 451; P < .001) supplying linen to hospitals with no cases of zygomycosis reported during the same period. The fungal isolates from patients and linens were phylogenetically related. In sum, 61% of environmental samples and 100% of air samples at the designated laundry were also positive for zygomycetes, suggesting heavy environmental contamination. RODAC contact plates revealed mean total viable bacteria counts of freshly laundered items (1028 ± 611 CFU/100 cm(2)) far exceeded the "hygienically clean" standard of 20 CFU/100 cm(2) set by the US healthcare textile certification requirement. CONCLUSIONS: Suboptimal conditions of washing, drying, and storage contributed to the massive linen contamination and the outbreak of zygomycosis.

Severe community-acquired pneumonia caused by macrolide-resistant Mycoplasma pneumoniae in a 6-year-old boy.

Mycoplasma pneumoniae is the commonest agent causing atypical pneumonia in children. Macrolides have long been used in the treatment of community-acquired pneumonia not responsive to beta-lactams alone. In this report, we describe the first locally acquired paediatric patient with severe community-acquired pneumonia caused by macrolide-resistant Mycoplasma pneumoniae, possessing an A-to-G transition at position 2063 of the 23s rRNA gene. In addition, we have detected two more strains of macrolide-resistant Mycoplasma pneumoniae out of a total of 10 cases with chest infection that were confirmed by polymerase chain reaction. Therefore macrolide-resistant Mycoplasma pneumoniae accounted for 33% (3 out of 10 patients) of the polymerase chain reaction-confirmed cases.

Oral bacterial flora of the Chinese cobra (Naja atra) and bamboo pit viper (Trimeresurus albolabris) in Hong Kong SAR, China.

OBJECTIVE: To determine the oral bacterial flora associated with two common local venomous snakes in Hong Kong, namely the Chinese cobra (Naja atra) and the bamboo pit viper (Trimeresurus albolabris). DESIGN: Cross-sectional study. SETTING: A non-government organisation and a regional hospital in Hong Kong. SUBJECTS: Thirty-two Chinese cobras and seven bamboo pit vipers. MAIN OUTCOME MEASURES: Species identification of bacteria in the oral cavity of both snakes and their antibiotic susceptibilities. RESULTS: The oral cavity of Chinese cobra harbour a wide range of pathogenic bacteria, including: Gram-negative bacterial species like Morganella morganii, Aeromonas hydrophila and Proteus, and Gram-positive bacteria like Enterococcus faecalis, coagulase-negative Staphylococcus as well as anaerobic species (clostridia). The oral cavity of the Chinese cobra is more likely than that of the bamboo pit viper to harbour pathogenic bacteria associated with snakebite infection (P<0.001). The median number of pathogenic bacteria per snake was significantly higher in the Chinese cobra (P<0.001). All pathogenic Gram-negative bacteria isolated were susceptible to levofloxacin. Amoxicillin/clavulanate provided good coverage against pathogenic Gram-positive bacteria (Enterococcus faecalis) and anaerobes. CONCLUSION: 'Prophylactic' antibiotic treatment for Chinese cobra bites may be beneficial, owing to the multiple pathogenic bacteria in its oral cavity and the higher risk of ensuing necrosis. The regimen of levofloxacin plus amoxicillin/clavulanate appears promising for this purpose, but further study is required to confirm its clinical utility in patients.

Computerized planning for multiprobe cryosurgery using a force-field analogy.

Cryosurgery is the destruction of undesired biological tissues by freezing. For internal organs, multiple cryoprobes are inserted into the tissue with the goal of maximizing cryoinjury within a predefined target region, while minimizing cryoinjury to the surrounding tissues. The objective of this study is to develop a computerized planning tool to determine the best locations to insert the cryoprobes, based on bioheat transfer simulations. This tool is general and suitable for all available cooling techniques and hardware. The planning procedure employs a novel iterative optimization technique based on a force-field analogy. In each iteration, a single transient bioheat transfer simulation of the cryoprocedure is computed. At the end of the simulation, regions of tissue that would have undesired temperatures apply "forces" to the cryoprobes directly moving them to better locations. This method is more efficient than traditional numerical optimization techniques, because it requires significantly fewer bioheat transfer simulations for each iteration of planning. For demonstration purposes, 2D examples on cross sections typical of prostate cryosurgery are given.

Computerized planning of cryosurgery using cryoprobes and cryoheaters.

In a typical minimally invasive cryoprocedure, multiple cryoprobes are inserted into the tissue with the goal of maximizing cryoinjury within a predefined target region, while minimizing cryoinjury to the surrounding tissues. A temperature-controlled electrical heater has been developed recently by this research team, in order to assist in limiting the cryoinjury to the target region. The new device has been termed a 'cryoheater,' and it can work with any cryosurgical cooling technique. A prototype computerized planning tool has been presented recently by this research team, which helps to determine the best locations in which to insert the cryoprobes. This prototype was designed for cryoprobes only. The planning procedure utilized a novel iterative optimization technique, based on a force-field analogy. The combination of cryoheaters with computerized planning is the subject matter of this report. The current report includes a review of cryoheater development, and presents an improved cryosurgery planning tool which incorporates cryoheaters.