Probable Animal-to-Human Transmission of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Delta Variant AY.127 Causing a Pet Shop-Related Coronavirus Disease 2019 (COVID-19) Outbreak in Hong Kong.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can infect human and other mammals, including hamsters. Syrian (Mesocricetus auratus) and dwarf (Phodopus sp.) hamsters are susceptible to SARS-CoV-2 infection in the laboratory setting. However, pet shop-related Coronavirus Disease 2019 (COVID-19) outbreaks have not been reported. METHODS: We conducted an investigation of a pet shop-related COVID-19 outbreak due to Delta variant AY.127 involving at least 3 patients in Hong Kong. We tested samples collected from the patients, environment, and hamsters linked to this outbreak and performed whole genome sequencing analysis of the reverse transcription polymerase chain reaction (RT-PCR)-positive samples. RESULTS: The patients included a pet shop keeper (Patient 1), a female customer of the pet shop (Patient 2), and the husband of Patient 2 (Patient 3). Investigation showed that 17.2% (5/29) and 25.5% (13/51) environmental specimens collected from the pet shop and its related warehouse, respectively, tested positive for SARS-CoV-2 RNA by RT-PCR. Among euthanized hamsters randomly collected from the storehouse, 3% (3/100) tested positive for SARS-CoV-2 RNA by RT-PCR and seropositive for anti-SARS-CoV-2 antibody by enzyme immunoassay. Whole genome analysis showed that although all genomes from the outbreak belonged to the Delta variant AY.127, there were at least 3 nucleotide differences among the genomes from different patients and the hamster cages. Genomic analysis suggests that multiple strains have emerged within the hamster population, and these different strains have likely transmitted to human either via direct contact or via the environment. CONCLUSIONS: Our study demonstrated probable hamster-to-human transmission of SARS-CoV-2. As pet trading is common around the world, this can represent a route of international spread of this pandemic virus.

Emergence of a Severe Acute Respiratory Syndrome Coronavirus 2 Virus Variant With Novel Genomic Architecture in Hong Kong.

Throughout the coronavirus disease 2019 (COVID-19) pandemic, divergent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lineages have emerged continuously, mostly through the genomic accumulation of substitutions. We report the discovery of a SARS-CoV-2 variant with a novel genomic architecture characterized by absent ORF7a, ORF7b, and ORF8, and a C-terminally modified ORF6 product resulting from partial 5'-untranslated region (UTR) duplication and transposition.

Nosocomial Outbreak of Coronavirus Disease 2019 by Possible Airborne Transmission Leading to a Superspreading Event.

BACKGROUND: Nosocomial outbreaks with superspreading of coronavirus disease 2019 due to a possible airborne transmission have not been reported. METHODS: Epidemiological analysis, environmental samplings, and whole-genome sequencing (WGS) were performed for a hospital outbreak. RESULTS: A superspreading event that involved 12 patients and 9 healthcare workers (HCWs) occurred within 9 days in 3 of 6 cubicles at an old-fashioned general ward with no air exhaust built within the cubicles. The environmental contamination by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA was significantly higher in air grilles (>2 m from patients' heads and not within reach) than on high-touch clinical surfaces (36.4%, 8 of 22 vs 3.4%, 1 of 29, P = .003). Six (66.7%) of 9 contaminated air exhaust grilles were located outside patient cubicles. The clinical attack rate of patients was significantly higher than of HCWs (15.4%, 12 of 78 exposed patients vs 4.6%, 9 of 195 exposed HCWs, P = .005). Moreover, the clinical attack rate of ward-based HCWs was significantly higher than of nonward-based HCWs (8.1%, 7 of 68 vs 1.8%, 2 of 109, P = .045). The episodes (mean ±â€…standard deviation) of patient-care duty assignment in the cubicles was significantly higher among infected ward-based HCWs than among noninfected ward-based HCWs (6.0 ±â€…2.4 vs 3.0 ±â€…2.9, P = .012) during the outbreak period. The outbreak strains belong to SARS-CoV-2 lineage B.1.36.27 (GISAID clade GH) with the unique S-T470N mutation on WGS. CONCLUSIONS: This nosocomial point source superspreading event due to possible airborne transmission demonstrates the need for stringent SARS-CoV-2 screening at admission to healthcare facilities and better architectural design of ventilation systems to prevent such outbreaks. Portable high-efficiency particulate filters were installed in each cubicle to improve ventilation before resumption of clinical service.

Unique Clusters of Severe Acute Respiratory Syndrome Coronavirus 2 Causing a Large Coronavirus Disease 2019 Outbreak in Hong Kong.

After 2 months of relative quiescence, a large coronavirus disease 2019 outbreak occurred in Hong Kong in July 2020 after gradual relaxation of social distancing policy. Unique severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) phylogenetic clusters have been identified among locally acquired cases, with most genomes belonging to cluster HK1, which is phylogenetically related to SARS-CoV-2 reported overseas.

Molecular Characterization of Influenza C Viruses from Outbreaks in Hong Kong SAR, China.

In 2014, the Centre for Health Protection in Hong Kong introduced screening for influenza C virus (ICV) as part of its routine surveillance for infectious agents in specimens collected from patients presenting with symptoms of respiratory viral infection, including influenza-like illness (ILI). A retrospective analysis of ICV detections up to week 26 of 2019 revealed persistent low-level circulation, with two outbreaks having occurred in the winters of 2015 to 2016 and 2017 to 2018. These outbreaks occurred at the same time as, and were dwarfed by, seasonal epidemics of influenza types A and B. Gene sequencing studies on stored ICV-positive clinical specimens from the two outbreaks have shown that the hemagglutinin-esterase (HE) genes of the viruses fall into two of the six recognized genetic lineages (represented by C/Kanagawa/1/76 and C/São Paulo/378/82), with there being significant genetic drift compared to earlier circulating viruses within both lineages. The location of a number of encoded amino acid substitutions in hemagglutinin-esterase fusion (HEF) glycoproteins suggests that antigenic drift may also have occurred. Observations of ICV outbreaks in other countries, with some of the infections being associated with severe disease, indicates that ICV infection has the potential to have significant clinical and health care impacts in humans.IMPORTANCE Influenza C virus infection of humans is common, and reinfection can occur throughout life. While symptoms are generally mild, severe disease cases have been reported, but knowledge of the virus is limited, as little systematic surveillance for influenza C virus is conducted and the virus cannot be studied by classical virologic methods because it cannot be readily isolated in laboratories. A combination of systematic surveillance in Hong Kong SAR, China, and new gene sequencing methods has been used in this study to assess influenza C virus evolution and provides evidence for a 2-year cycle of disease outbreaks. The results of studies like that reported here are key to developing an understanding of the impact of influenza C virus infection in humans and how virus evolution might be associated with epidemics.

Posterior Oropharyngeal Saliva for the Detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has put tremendous pressure on the healthcare system worldwide. Diagnostic testing remained one of the limiting factors for early identification and isolation of infected patients. This study aimed to evaluate posterior oropharyngeal saliva (POPS) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection among patients with confirmed or suspected COVID-19. METHODS: The laboratory information system was searched retrospectively for all respiratory specimens and POPS requested for SARS-CoV-2 RNA detection between 1 February 2020 and 15 April 2020. The agreement and diagnostic performance of POPS against NPsp were evaluated. RESULTS: A total of 13772 specimens were identified during the study period, including 2130 POPS and 8438 nasopharyngeal specimens (NPsp). Two hundred and twenty-nine same-day POPS-NPsp paired were identified with POPS and NPsp positivity of 61.5% (95% confidence interval [CI] 55.1-67.6%) and 53.3% (95% CI 46.8-59.6%). The overall, negative and positive percent agreement were 76.0% (95% CI 70.2-80.9%), 65.4% (95% CI 55.5-74.2%), 85.2% (95% CI 77.4-90.8%). Better positive percent agreement was observed in POPS-NPsp obtained within 7 days (96.6%, 95% CI 87.3-99.4%) compared with after 7 days of symptom onset (75.0%, 95% CI 61.4-85.2%). Among the 104 positive pairs, the mean difference in Cp value was 0.26 (range: 12.63 to -14.74), with an overall higher Cp value in NPsp (Pearson coefficient 0.579). No significant temporal variation was noted between the 2 specimen types. CONCLUSIONS: POPS is an acceptable alternative specimen to nasopharyngeal specimen for the detection of SARS-CoV-2.

A tricyclic pyrrolobenzodiazepine produced by Klebsiella oxytoca is associated with cytotoxicity in antibiotic-associated hemorrhagic colitis.

Cytotoxin-producing Klebsiella oxytoca is the causative agent of antibiotic-associated hemorrhagic colitis (AAHC). Recently, the cytotoxin associated with AAHC was identified as tilivalline, a known pentacyclic pyrrolobenzodiazepine (PBD) metabolite produced by K. oxytoca Although this assertion of tilivalline's role in AAHC is supported by evidence from animal experiments, some key aspects of this finding appear to be incompatible with toxicity mechanisms of known PBD toxins. We therefore hypothesized that K. oxytoca may produce some other uncharacterized cytotoxins. To address this question, we investigated whether tilivalline alone is indeed necessary and sufficient to induce cytotoxicity or whether K. oxytoca also produces other cytotoxins. LC-MS- and NMR-based metabolomic analyses revealed the presence of an abundant tricyclic PBD, provisionally designated kleboxymycin, in the supernatant of toxigenic K. oxytoca strains. Moreover, by generating multiple mutants with gene deletions affecting tilivalline biosynthesis, we show that a tryptophanase-deficient, tilivalline-negative K. oxytoca mutant induced cytotoxicity in vitro similar to tilivalline-positive K. oxytoca strains. Furthermore, synthetic kleboxymycin exhibited greater than 9-fold higher cytotoxicity than tilivalline in TC50 cell culture assays. We also found that the biosynthetic pathways for kleboxymycin and tilivalline appear to overlap, as tilivalline is an indole derivative of kleboxymycin. In summary, our results indicate that tilivalline is not essential for inducing cytotoxicity observed in K. oxytoca-associated AAHC and that kleboxymycin is a tilivalline-related bacterial metabolite with even higher cytotoxicity.

Two novel noroviruses and a novel norovirus genogroup in California sea lions.

In this study, two novel noroviruses (NoVs) were discovered from faecal samples from California sea lions from an oceanarium in Hong Kong, and named California sea lion NoV 1 (Csl/NoV1) and California sea lion NoV 2 (Csl/NoV2). Whole-genome sequencing showed that the genome organization and amino acid motifs of both Csl/NoV1 and Csl/NoV2 were typical of those of other NoVs in their open reading frames (ORFs). Csl/NoV1 possessed only 52.6-52.8 % amino acid identity in VP1 to the closest matches in genogroup GII. Therefore, Csl/NoV1 should constitute a novel genogroup of NoV. Shifting of the phylogenetic position of Csl/NoV1 in the RdRp, VP1 and VP2 trees was observed, which may have been due to recombination events and/or biased mutations. Csl/NoV2 possessed 55.4-56.2 % amino acid identity in VP1 to its closest relatives in genogroup GVI, which means that it represents a new genotype in genogroup GVI. Further studies will reveal what diseases these NoVs can cause in marine mammals.

Seasonal Outbreak of Bacillus Bacteremia Associated With Contaminated Linen in Hong Kong.

BACKGROUND: A high seasonal incidence of Bacillus bacteremia was associated with the use of contaminated hospital linens. METHODS: An outbreak investigation was conducted to study the incidence and source of Bacillus bacteremia during the baseline, outbreak, and postoutbreak period from 1 January 2012 through 31 July 2016 at a university-affiliated teaching hospital in Hong Kong. Replicate organism detection and counting plates were used for microbial screening of linen samples. The Bacillus species isolated from patient and linen samples were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and were phylogenetically analyzed. RESULTS: During the study period, a total of 113 207 blood cultures were collected from 43 271 patients, of which 978 (0.86%) specimens from 744 (1.72%) patients were identified as Bacillus species. The incidence of Bacillus bacteremia per 10 000 patient admissions and per 10 000 patient-days was significantly higher during the summer outbreak as compared with baseline and 1 year postoutbreak after cessation of the linen supply from the designated laundry and change of laundry protocol (39.97 vs 18.21 vs 2.27; 13.36 vs 5.61 vs 0.73; P < .001). The mean total aerobic bacterial count per 100 cm2 was significantly higher among the 99 linen samples screened during the outbreak period compared to the 100 screened in the postoutbreak period (916.0 ± 641.6 vs 0.6 ± 1.6; P < .001). Blood culture isolates of Bacillus cereus group in 14 of 87 (16.1%) patients were phylogenetically associated with 9 linen sample isolates. CONCLUSIONS: Suboptimal conditions of hospital laundry contributed to the seasonal outbreak of Bacillus bacteremia.

High prevalence of four novel astrovirus genotype species identified from rodents in China.

Astroviruses cause gastrointestinal and neurological infections in humans and animals. Since astrovirus is genetically diverse and different astrovirus genotypes can be found in the same animal species, astrovirus is a potential zoonotic threat to humans. In this study, we screened for astroviruses in rodents from Hong Kong, Hunan and Guangxi. Astrovirus was detected in 11.9 % (67/562) of rectal swab specimens. Phylogenetic analysis of the ORF1b region, which encodes the RdRp, showed that there were four distinct clusters (clusters A, B, C and D). Whole genome sequencing was performed for 11 representative strains from each of these four clusters. The mean amino acid genetic distances (p-dist) of full-length ORF2 were >0.634 between clusters A, B, C and other known astroviruses. The p-dist between clusters A and B, A and C, and B and C were 0.371-0.375, 0.517-0.549 and 0.524-0.555, respectively. Within cluster C, the p-dist between HN-014 and GX-006 was 0.372. Since strains with p-dist of ≥0.368 in ORF2 are now considered to be of separate genotypes species, cluster A, cluster B, cluster C-HN-014 and cluster C-GX-006 can be classified as novel genotype species. Cluster D was most closely related to the rodent astrovirus previously identified in Hong Kong. Since rodents live in close proximity to humans, interspecies jumping of these novel astroviruses may represent a threat to human health.

Disseminated Infections with Talaromyces marneffei in Non-AIDS Patients Given Monoclonal Antibodies against CD20 and Kinase Inhibitors.

Infections with the fungus Talaromyces (formerly Penicillium) marneffei are rare in patients who do not have AIDS. We report disseminated T. marneffei infection in 4 hematology patients without AIDS who received targeted therapy with monoclonal antibodies against CD20 or kinase inhibitors during the past 2 years. Clinicians should be aware of this emerging complication, especially in patients from disease-endemic regions.

Evidence for Elizabethkingia anophelis transmission from mother to infant, Hong Kong.

Elizabethkingia anophelis, recently discovered from mosquito gut, is an emerging bacterium associated with neonatal meningitis and nosocomial outbreaks. However, its transmission route remains unknown. We use rapid genome sequencing to investigate 3 cases of E. anophelis sepsis involving 2 neonates who had meningitis and 1 neonate's mother who had chorioamnionitis. Comparative genomics revealed evidence for perinatal vertical transmission from a mother to her neonate; the 2 isolates from these patients, HKU37 and HKU38, shared essentially identical genome sequences. In contrast, the strain from another neonate (HKU36) was genetically divergent, showing only 78.6% genome sequence identity to HKU37 and HKU38, thus excluding a clonal outbreak. Comparison to genomes from mosquito strains revealed potential metabolic adaptations in E. anophelis under different environments. Maternal infection, not mosquitoes, is most likely the source of neonatal E. anophelis infections. Our findings highlight the power of genome sequencing in gaining rapid insights on transmission and pathogenesis of emerging pathogens.

Middle east respiratory syndrome coronavirus 4a protein is a double-stranded RNA-binding protein that suppresses PACT-induced activation of RIG-I and MDA5 in the innate antiviral response.

UNLABELLED: Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging pathogen that causes severe disease in human. MERS-CoV is closely related to bat coronaviruses HKU4 and HKU5. Evasion of the innate antiviral response might contribute significantly to MERS-CoV pathogenesis, but the mechanism is poorly understood. In this study, we characterized MERS-CoV 4a protein as a novel immunosuppressive factor that antagonizes type I interferon production. MERS-CoV 4a protein contains a double-stranded RNA-binding domain capable of interacting with poly(I · C). Expression of MERS-CoV 4a protein suppressed the interferon production induced by poly(I · C) or Sendai virus. RNA binding of MERS-CoV 4a protein was required for IFN antagonism, a property shared by 4a protein of bat coronavirus HKU5 but not by the counterpart in bat coronavirus HKU4. MERS-CoV 4a protein interacted with PACT in an RNA-dependent manner but not with RIG-I or MDA5. It inhibited PACT-induced activation of RIG-I and MDA5 but did not affect the activity of downstream effectors such as RIG-I, MDA5, MAVS, TBK1, and IRF3. Taken together, our findings suggest a new mechanism through which MERS-CoV employs a viral double-stranded RNA-binding protein to circumvent the innate antiviral response by perturbing the function of cellular double-stranded RNA-binding protein PACT. PACT targeting might be a common strategy used by different viruses, including Ebola virus and herpes simplex virus 1, to counteract innate immunity. IMPORTANCE: Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging and highly lethal human pathogen. Why MERS-CoV causes severe disease in human is unclear, and one possibility is that MERS-CoV is particularly efficient in counteracting host immunity, including the sensing of virus invasion. It will therefore be critical to clarify how MERS-CoV cripples the host proteins that sense viruses and to compare MERS-CoV with its ancestral viruses in bats in the counteraction of virus sensing. This work not only provides a new understanding of the abilities of MERS-CoV and closely related bat viruses to subvert virus sensing but also might prove useful in revealing new strategies for the development of vaccines and antivirals.

Novel selective medium for isolation of Staphylococcus lugdunensis from wound specimens.

We compared a novel selective Staphylococcus lugdunensis (SSL) medium with routine media (blood and chocolate agars) for the detection of S. lugdunensis in 990 clinical specimens (from tissue, pus, or wound swabs). Significantly more S. lugdunensis isolates were detected on SSL medium (34/990) than on routine medium (7/990) (P = 0.001, McNemar's test).

Unraveling the molecular basis of temperature-dependent genetic regulation in Penicillium marneffei.

Penicillium marneffei is an opportunistic fungal pathogen endemic in Southeast Asia, causing lethal systemic infections in immunocompromised patients. P. marneffei grows in a mycelial form at the ambient temperature of 25°C and transitions to a yeast form at 37°C. The ability to alternate between the mycelial and yeast forms at different temperatures, namely, thermal dimorphism, has long been considered critical for the pathogenicity of P. marneffei, yet the underlying genetic mechanisms remain elusive. Here we employed high-throughput sequencing to unravel global transcriptional profiles of P. marneffei PM1 grown at 25 and 37°C. Among ∼11,000 protein-coding genes, 1,447 were overexpressed and 1,414 were underexpressed at 37°C. Counterintuitively, heat-responsive genes, predicted in P. marneffei through sequence comparison, did not tend to be overexpressed at 37°C. These results suggest that P. marneffei may take a distinct strategy of genetic regulation at the elevated temperature; the current knowledge concerning fungal heat response, based on studies of model fungal organisms, may not be applicable to P. marneffei. Our results further showed that the tandem repeat sequences (TRSs) are overrepresented in coding regions of P. marneffei genes, and TRS-containing genes tend to be overexpressed at 37°C. Furthermore, genomic sequences and expression data were integrated to characterize gene clusters, multigene families, and species-specific genes of P. marneffei. In sum, we present an integrated analysis and a comprehensive resource toward a better understanding of temperature-dependent genetic regulation in P. marneffei.

Two years after pandemic influenza A/2009/H1N1: what have we learned?

The world had been anticipating another influenza pandemic since the last one in 1968. The pandemic influenza A H1N1 2009 virus (A/2009/H1N1) finally arrived, causing the first pandemic influenza of the new millennium, which has affected over 214 countries and caused over 18,449 deaths. Because of the persistent threat from the A/H5N1 virus since 1997 and the outbreak of the severe acute respiratory syndrome (SARS) coronavirus in 2003, medical and scientific communities have been more prepared in mindset and infrastructure. This preparedness has allowed for rapid and effective research on the epidemiological, clinical, pathological, immunological, virological, and other basic scientific aspects of the disease, with impacts on its control. A PubMed search using the keywords "pandemic influenza virus H1N1 2009" yielded over 2,500 publications, which markedly exceeded the number published on previous pandemics. Only representative works with relevance to clinical microbiology and infectious diseases are reviewed in this article. A significant increase in the understanding of this virus and the disease within such a short amount of time has allowed for the timely development of diagnostic tests, treatments, and preventive measures. These findings could prove useful for future randomized controlled clinical trials and the epidemiological control of future pandemics.

Differential cell line susceptibility to the emerging novel human betacoronavirus 2c EMC/2012: implications for disease pathogenesis and clinical manifestation.

The emerging novel human betacoronavirus 2c EMC/2012 (HCoV-EMC) was recently isolated from patients with severe pneumonia and renal failure and was associated with an unexplained high crude fatality rate of 56%. We performed a cell line susceptibility study with 28 cell lines. HCoV-EMC was found to infect the human respiratory tract (polarized airway epithelium cell line Calu-3, embryonic fibroblast cell line HFL, and lung adenocarcinoma cell line A549), kidney (embryonic kidney cell line HEK), intestinal tract (colorectal adenocarcinoma cell line Caco-2), liver cells (hepatocellular carcinoma cell line Huh-7), and histiocytes (malignant histiocytoma cell line His-1), as evident by detection of high or increasing viral load in culture supernatants, detection of viral nucleoprotein expression by immunostaining, and/or detection of cytopathic effects. Although an infected human neuronal cell line (NT2) and infected monocyte and T lymphocyte cell lines (THP-1, U937, and H9) had increased viral loads, their relatively lower viral production corroborated with absent nucleoprotein expression and cytopathic effects. This range of human tissue tropism is broader than that for all other HCoVs, including SARS coronavirus, HCoV-OC43, HCoV-HKU1, HCoV-229E, and HCoV-NL63, which may explain the high mortality associated with this disease. A recent cell line susceptibility study showed that HCoV-EMC can infect primate, porcine, and bat cells and therefore may jump interspecies barriers. We found that HCoV-EMC can also infect civet lung fibroblast and rabbit kidney cell lines. These findings have important implications for the diagnosis, pathogenesis, and transmission of HCoV-EMC.