Showing papers by "Patrick C. Y. Woo published in 2009"

Coronavirus Diversity, Phylogeny and Interspecies Jumping:

Abstract: The SARS epidemic has boosted interest in research on coronavirus biodiversity and genomics. Before 2003, there were only 10 coronaviruses with complete genomes available. After the SARS epidemic, up to December 2008, there was an addition of 16 coronaviruses with complete genomes sequenced. These include two human coronaviruses (human coronavirus NL63 and human coronavirus HKU1), 10 other mammalian coronaviruses [bat SARS coronavirus, bat coronavirus (bat-CoV) HKU2, bat-CoV HKU4, bat-CoV HKU5, bat-CoV HKU8, bat-CoV HKU9, bat-CoV 512/2005, bat-CoV 1A, equine coronavirus, and beluga whale coronavirus] and four avian coronaviruses (turkey coronavirus, bulbul coronavirus HKU11, thrush coronavirus HKU12, and munia coronavirus HKU13). Two novel subgroups in group 2 coronavirus (groups 2c and 2d) and two novel subgroups in group 3 coronavirus (groups 3b and 3c) have been proposed. The diversity of coronaviruses is a result of the infidelity of RNA-dependent RNA polymerase, high frequency of homologous RNA recombination, and the large genomes of coronaviruses. Among all hosts, the diversity of coronaviruses is most evidenced in bats and birds, which may be a result of their species diversity, ability to fly, environmental pressures, and habits of roosting and flocking. The present evidence supports that bat coronaviruses are the gene pools of group 1 and 2 coronaviruses, whereas bird coronaviruses are the gene pools of group 3 coronaviruses. With the increasing number of coronaviruses, more and more closely related coronaviruses from distantly related animals have been observed, which were results of recent interspecies jumping and may be the cause of disastrous outbreaks of zoonotic diseases.

Comparative analysis of complete genome sequences of three avian coronaviruses reveals a novel group 3c coronavirus.

Abstract: In this territory-wide molecular epidemiology study of coronaviruses (CoVs) in Hong Kong involving 1,541 dead wild birds, three novel CoVs were identified in three different bird families (bulbul CoV HKU11 [BuCoV HKU11], thrush CoV HKU12 [ThCoV HKU12], and munia CoV HKU13 [MuCoV HKU13]). Four complete genomes of the three novel CoVs were sequenced. Their genomes (26,396 to 26,552 bases) represent the smallest known CoV genomes. In phylogenetic trees constructed using chymotrypsin-like protease (3CL pro ), RNA-dependent RNA polymerase (Pol), helicase, spike, and nucleocapsid proteins, BuCoV HKU11, ThCoV HKU12, and MuCoV HKU13 formed a cluster distantly related to infectious bronchitis virus and turkey CoV (group 3a CoVs). For helicase, spike, and nucleocapsid, they were also clustered with a CoV recently discovered in Asian leopard cats, for which the complete genome sequence was not available. The 3CL pro , Pol, helicase, and nucleocapsid of the three CoVs possessed higher amino acid identities to those of group 3a CoVs than to those of group 1 and group 2 CoVs. Unique genomic features distinguishing them from other group 3 CoVs include a distinct transcription regulatory sequence and coding potential for small open reading frames. Based on these results, we propose a novel CoV subgroup, group 3c, to describe this distinct subgroup of CoVs under the group 3 CoVs. Avian CoVs are genetically more diverse than previously thought and may be closely related to some newly identified mammalian CoVs. Further studies would be important to delineate whether the Asian leopard cat CoV was a result of interspecies jumping from birds, a situation analogous to that of bat and civet severe acute respiratory syndrome CoVs.

Viral loads in clinical specimens and SARS manifestations.

Abstract: 1. A high viral load in nasopharyngeal aspirate (with or without a high viral load in serum) is a useful prognostic indicator of respiratory failure or mortality. The presence of viral RNA in multiple body sites is also indicative of poor prognosis. 2. Early treatment with an effective antiviral agent before day 10 may decrease the peak viral load, and thus ameliorate the clinical symptoms and mortality, and reduce viral shedding and the risk of transmission

Clinical and Molecular Epidemiology of Human Rhinovirus C in Children and Adults in Hong Kong Reveals a Possible Distinct Human Rhinovirus C Subgroup

Abstract: Background. A novel human rhinovirus (HRV) species, HRV-C, was recently discovered, but its clinical features and epidemiology, compared with HRV-A and HRV-B, remains poorly understood, especially in adults. Methods. One thousand two hundred nasopharyngeal aspirate samples obtained from hospitalized children and adults during a 1-year period were subject to reverse-transcriptase polymerase chain reaction to detect HRV. The clinical and molecular epidemiology of the 3 HRV species was analyzed. Results. HRVs were detected in 178 (29.7%) of 600 nasopharyngeal aspirate samples from children and 42 (7%) of 600 nasopharyngeal aspirate samples from adults. HRV-A was most prevalent (n = 111), followed by HRV-C (n = 91) and HRV-B (n = 18). Although upper respiratory tract infection was the most common presentation in children, 8 (62%) of the 13 adults with HRV-C infection had pneumonia, compared with 6 (27%) of the 22 adults with HRV-A infection (P<.05 Wheezing episodes were also more common among individuals with HRV-C (37%) and HRV-A (20%) infection than among those with HRV-B (0%) infection (P<.05). Clinical and molecular data analysis revealed HRV-C as a frequent cause of community and institutionalized outbreaks. A diverse set of HRV-C genotypes was circulating throughout the year, among which a potential distinct subgroup of strains was observed. Conclusion. HRV-C is associated with pneumonia in adults and outbreaks of respiratory infections requiring hospitalization. A potential novel HRV-C subgroup was identified.

Outbreak of Intestinal Infection Due to Rhizopus microsporus

Abstract: Sinopulmonary and rhinocerebral zygomycosis has been increasingly found in patients with hematological malignancies and bone marrow transplantation, but intestinal zygomycosis remains very rare in the literature. We investigated an outbreak of intestinal infection due to Rhizopus microsporus in 12 patients on treatment for hematological malignancies over a period of 6 months in a teaching hospital. The intake of allopurinol during hospitalization (P < 0.001) and that of commercially packaged ready-to-eat food items in the preceding 2 weeks (P < 0.001) were found to be independently significant risk factors for the development of intestinal zygomycosis. A total of 709 specimens, including 378 environmental and air samples, 181 food samples, and 150 drug samples, were taken for fungal culture. Among them, 16 samples of allopurinol tablets, 3 prepackaged ready-to-eat food items, and 1 pair of wooden chopsticks were positive for Rhizopus microsporus, which was confirmed by ITS1-5.8S-ITS2 rRNA gene cluster (internal transcribed spacer [ITS]) sequencing. The mean viable fungal counts of allopurinol obtained from wards and pharmacy were 4.22 x 10(3) CFU/g of tablet (range, 3.07 x 10(3) to 5.48 x 10(3)) and 3.24 x 10(3) CFU/g of tablet (range, 2.68 x 10(3) to 3.72 x 10(3)), respectively, which were much higher than the mean count of 2 x 10(2) CFU/g of food. Phylogenetic analysis by ITS sequencing showed multiple clones from isolates of contaminated allopurinol tablets and ready-to-eat food, of which some were identical to patients' isolates, and with one isolate in the cornstarch used as an excipient for manufacture of this drug. We attempted to type the isolates by random amplification of polymorphic DNA analysis, with limited evidence of clonal distribution. The primary source of the contaminating fungus was likely to be the cornstarch used in the manufacturing of allopurinol tablets or ready-to-eat food. Rhizopus microsporus is thermotolerant and can multiply even at 50 degrees C. The long holding time of the intermediates during the manufacturing process of allopurinol amplified the fungal load. Microbiological monitoring of drugs manufactured for highly immunosuppressed patients should be considered.

Guidelines for interpretation of 16S rRNA gene sequence-based results for identification of medically important aerobic Gram-positive bacteria

Abstract: This study is believed to be the first to provide guidelines for facilitating interpretation of results based on full and 527 bp 16S rRNA gene sequencing and MicroSeq databases used for identifying medically important aerobic Gram-positive bacteria. Overall, full and 527 bp 16S rRNA gene sequencing can identify 24 and 40 % of medically important Gram-positive cocci (GPC), and 21 and 34 % of medically important Gram-positive rods (GPR) confidently to the species level, whereas the full-MicroSeq and 500-MicroSeq databases can identify 15 and 34 % of medically important GPC and 14 and 25 % of medically important GPR confidently to the species level. Among staphylococci, streptococci, enterococci, mycobacteria, corynebacteria, nocardia and members of Bacillus and related taxa (Paenibacillus, Brevibacillus, Geobacillus and Virgibacillus), the methods and databases are least useful for identification of staphylococci and nocardia. Only 0–2 and 2–13 % of staphylococci, and 0 and 0–10 % of nocardia, can be confidently and doubtfully identified, respectively. However, these methods and databases are most useful for identification of Bacillus and related taxa, with 36–56 and 11–14 % of Bacillus and related taxa confidently and doubtfully identified, respectively. A total of 15 medically important GPC and 18 medically important GPR that should be confidently identified by full 16S rRNA gene sequencing are not included in the full-MicroSeq database. A total of 9 medically important GPC and 21 medically important GPR that should be confidently identified by 527 bp 16S rRNA gene sequencing are not included in the 500-MicroSeq database. 16S rRNA gene sequence results of Gram-positive bacteria should be interpreted with basic phenotypic tests results. Additional biochemical tests or sequencing of additional gene loci are often required for definitive identification. To improve the usefulness of the MicroSeq databases, bacterial species that can be confidently identified by 16S rRNA gene sequencing but are not found in the MicroSeq databases should be included.

Clinical and Molecular Epidemiology of Human Parainfluenza Virus 4 Infections in Hong Kong: Subtype 4B as Common as Subtype 4A

Abstract: In this 1-year study, 35 (1.2%) of 2,912 nasopharyngeal aspirates were positive for human parainfluenza virus 4 (HPIV4) by reverse transcription-PCR. Patients with HPIV4 infection were mainly young children and immunocompromised adults. In contrast to the reported predominance of HPIV4A infection, molecular subtyping revealed that 15 (44%) cases were caused by HPIV4B.

The Complete Genome and Proteome of Laribacter hongkongensis Reveal Potential Mechanisms for Adaptations to Different Temperatures and Habitats

Abstract: Laribacter hongkongensis is a newly discovered Gram-negative bacillus of the Neisseriaceae family associated with freshwater fish-borne gastroenteritis and traveler's diarrhea. The complete genome sequence of L. hongkongensis HLHK9, recovered from an immunocompetent patient with severe gastroenteritis, consists of a 3,169-kb chromosome with G+C content of 62.35%. Genome analysis reveals different mechanisms potentially important for its adaptation to diverse habitats of human and freshwater fish intestines and freshwater environments. The gene contents support its phenotypic properties and suggest that amino acids and fatty acids can be used as carbon sources. The extensive variety of transporters, including multidrug efflux and heavy metal transporters as well as genes involved in chemotaxis, may enable L. hongkongensis to survive in different environmental niches. Genes encoding urease, bile salts efflux pump, adhesin, catalase, superoxide dismutase, and other putative virulence factors-such as hemolysins, RTX toxins, patatin-like proteins, phospholipase A1, and collagenases-are present. Proteomes of L. hongkongensis HLHK9 cultured at 37 degrees C (human body temperature) and 20 degrees C (freshwater habitat temperature) showed differential gene expression, including two homologous copies of argB, argB-20, and argB-37, which encode two isoenzymes of N-acetyl-L-glutamate kinase (NAGK)-NAGK-20 and NAGK-37-in the arginine biosynthesis pathway. NAGK-20 showed higher expression at 20 degrees C, whereas NAGK-37 showed higher expression at 37 degrees C. NAGK-20 also had a lower optimal temperature for enzymatic activities and was inhibited by arginine probably as negative-feedback control. Similar duplicated copies of argB are also observed in bacteria from hot springs such as Thermus thermophilus, Deinococcus geothermalis, Deinococcus radiodurans, and Roseiflexus castenholzii, suggesting that similar mechanisms for temperature adaptation may be employed by other bacteria. Genome and proteome analysis of L. hongkongensis revealed novel mechanisms for adaptations to survival at different temperatures and habitats.

More and more coronaviruses: human coronavirus HKU1.

Abstract: After human coronaviruses OC43, 229E and NL63, human coronavirus HKU1 (HCoV-HKU1) is the fourth human coronavirus discovered. HCoV-HKU1 is a group 2a coronavirus that is still not cultivable. The G + C contents of HCoV-HKU1 genomes are 32%, the lowest among all known coronaviruses with complete genome sequences available. Among all coronaviruses, HCoV-HKU1 shows the most extreme codon usage bias, attributed most importantly to severe cytosine deamination. All HCoV-HKU1 genomes contain unique tandem copies of a 30-base acidic tandem repeat of unknown function at the N-terminus of nsp3 inside the acidic domain upstream of papain-like protease 1. Three genotypes, A, B and C, of HCoV-HKU1 and homologous recombination among their genomes, are observed. The incidence of HCoV-HKU1 infections is the highest in winter. Similar to other human coronaviruses, HCoV-HKU1 infections have been reported globally, with a median (range) incidence of 0.9 (0 – 4.4) %. HCoV-HKU1 is associated with both upper and lower respiratory tract infections that are mostly self-limiting. The most common method for diagnosing HCoV-HKU1 infection is RT-PCR or real-time RT-PCR using RNA extracted from respiratory tract samples such as nasopharyngeal aspirates (NPA). Both the pol and nucleocapsid genes have been used as the targets for amplification. Monoclonal antibodies have been generated for direct antigen detection in NPA. For antibody detection, Escherichia coli BL21 and baculovirus-expressed recombinant nucleocapsid of HCoV-HKU1 have been used for IgG and IgM detection in sera of patients and normal individuals, using Western blot and enzyme-linked immunoassay.

Confirmation of the first Hong Kong case of human infection by novel swine origin influenza A (H1N1) virus diagnosed using ultrarapid, real-time reverse transcriptase PCR.

Abstract: The emergence of novel swine origin influenza A (H1N1) virus (S-OIV), first recognized in Mexico, has affected 4,379 patients in 29 countries, with 49 deaths as of 10 May 2009 (2-5, 7, 8). Here we describe the first case of S-OIV infection in Hong Kong, as confirmed by LightCycler and ultrarapid reverse transcriptase PCR (RT-PCR) analysis. A 25-year-old previously healthy and asymptomatic Mexican man arrived in Hong Kong from Mexico via Shanghai on 30 April 2009. He attended the hospital emergency room because of fever, productive cough, and myalgia on the same day. His temperature was 38°C. His total white cell count was 4.7 × 109/liter, with lymphopenia of 0.9 × 109/liter. Hemoglobin, platelet, liver, and renal function test results were normal. His chest radiograph was clear. The results of a test of his nasopharyngeal aspirate (NPA) for the presence of influenza A or B virus direct antigen were negative (9). The result from an RT-PCR test to detect the presence of the influenza A virus M gene was positive but that from an RT-PCR test to detect influenza A virus subtypes H1 (human) and H3 was negative. LightCycler and ultrarapid RT-PCR confirmed the presence of S-OIV. Oseltamivir therapy was started, and the symptoms subsided on the second day. A total of 12 specimens from close contacts, including passengers who had been in close proximity on the same flight and asymptomatic contacts at the same hotel, were negative upon testing. RNA extraction was performed using a QIAamp virus RNA minikit (Qiagen) (6). All procedures involving clinical specimens and influenza virus were performed in a biosafety level 2 laboratory with biosafety level 3 practices (1). By means of conventional RT-PCR, an 83-bp fragment of the hemagglutinin (HA) gene of swine influenza A H1 virus was amplified using 0.5 μM primers (LPW9948 [5′-GGTAAATGTAACATTGCT-3′], corresponding to nucleotides 208 to 225, and LPW9949 [5′-ACAATGTAGGACCATGAGCTT-3′], corresponding to nucleotides 270 to 290) designed by multiple alignment of the HA gene sequences of swine H1 virus and S-OIV strain A/California/04/2009 (available in GenBank). These primers were designed to detect swine H1 virus but not human seasonal H1N1 or H3N2 viruses. Positive-control experiments with a swine H1 virus (A/SW/HK/294/09) and S-OIV (A/California/04/2009) and negative-control experiments with human H1N1 and H3N2 viruses were performed. RT was performed using a SuperScript III kit (Invitrogen). Each PCR mixture (25 μl) contained cDNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2, 0.01% gelatin), 200 μM (each) deoxynucleoside triphosphates, and 1.0 U Taq polymerase (Boehringer). The mixtures were incubated at 95°C for 10 min and then amplified in 45 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 30 s, followed by a final extension at 72°C for 5 min in an automated thermal cycler (Applied Biosystems). Since early detection of this virus is crucial for timely implementation of infection control measures, we developed an ultrarapid RT-PCR assay which allows detection of S-OIV within 50 min by the use of the same PCR primers but with AmpliTaq Gold Fast PCR master mix, UP (2×) (Applied Biosystems). The mixtures were amplified using 45 cycles of 96°C for 3 s, 55°C for 3 s, and 68°C for 5 s and a final extension at 72°C for 10 s. Both conventional and ultrarapid RT-PCR analysis of RNA from the patient's NPA, S-OIV, and swine H1 virus positive controls, but not from seasonal H1N1 or H3N2 virus, showed bands of 83 bp. All of the positive-testing M and HA gene PCR products from the patient were confirmed by sequencing to be S-OIV. Real-time RT-PCR assays were performed, using the same PCR primers, as described previously (6). cDNA was amplified in a 2.0 LightCycler (Roche) using 20-μl reaction mixtures containing FastStart DNA master SYBR green I mix reagent (Roche), 2 μl of cDNA, 3.5 mM MgCl2, and 0.5 mM primers at 95°C for 10 min followed by 50 cycles of 95°C for 10 s, 62°C for 5 s, and 72°C for 5 s. The melting temperatures of S-OIV and swine H1 virus were 79.9°C and 83.0°C, respectively (Fig. ​(Fig.1).1). The viral load in the nasopharyngeal secretion dropped from 2.3 × 106 copies per ml on day 1 of illness to an undetectable level on day 6 (Table ​(Table1).1). A total of 150 specimens that tested positive for seasonal influenza A H3N2 or H1N1 virus tested negative in both LightCycler and conventional RT-PCR assays. A cytopathic effect was observed in an MDCK cell line at 72 h after inoculation of the first NPA. The culture supernatant was strongly positive for H1 S-OIV by LightCycler RT-PCR, while the cells were strongly positive for influenza A virus nucleoprotein by immunofluorescence (Dako) (9). Complete gene sequencing of the H1 virus (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"GQ168606","term_id":"237769862","term_text":"GQ168606"}}GQ168606) showed 99.9% identity to S-OIV (A/California/04/2009). FIG. 1. LightCycler real-time RT-PCR assays for S-OIV and swine H1 virus. (A) Standard curve for quantitative analysis of S-OIV H1 gene. (B) An amplification plot of fluorescence intensity against the PCR cycle. The amplification curves of patient NPA containing ... TABLE 1. Viral load of patient specimens collected at different days after onset of illness by LightCycler RT-PCR The ultrarapid and LightCycler assays described here can be used as rapid diagnostic tests to specifically detect S-OIV and other swine H1 viruses. Furthermore, the LightCycler assay may distinguish between novel S-OIV and other swine H1 viruses by determination of different melting temperatures. However, sequencing of longer or other gene segments should be required for more confident differentiation.

Identification of major histocompatibility complex class I C molecule as an attachment factor that facilitates coronavirus HKU1 spike-mediated infection.

Abstract: Human coronavirus HKU1 (HCoV-HKU1) is a recently discovered human coronavirus associated with respiratory tract infections worldwide. In this study, we have identified the major histocompatibility complex class I C molecule (HLA-C) as an attachment factor in facilitating HCoV-HKU1 spike (S)-mediated infection. HCoV-HKU1 S pseudotyped virus was assembled using a human immunodeficiency virus type 1-derived reporter virus harboring the human codon-optimized spike of HCoV-HKU1. We identified human alveolar epithelial A549 cells as the most susceptible cell line among those tested to infection by HCoV-HKU1 S pseudotypes. A549 cells were shown to bind purified soluble HCoV-HKU1 S(1-600) glycopeptide. To search for the functional receptor for HCoV-HKU1, an A549 cDNA expression library was constructed and transduced into the nonpermissive, baby hamster kidney cells line BHK-21. Transduced cells that bind soluble HCoV-HKU1 S(1-600) glycoprotein with C-terminal FLAG were sorted. Sequencing of two independent clones revealed cDNA inserts encoding HLA-C. Inhibition of HLA-C expression or function by RNAi silencing and anti-HLA-C antibody decreased HCoV-HKU1 S pseudotyped virus infection of A549 cells by 62 to 65%, whereas pretreatment of cells with neuraminidase decreased such infection by only 13%. When HLA-C was constitutively expressed in another nonpermissive cell line, NIH-3T3, quantitative PCR showed that the binding of HCoV-HKU1 S pseudotyped virus to cell surfaces was increased by 200-fold, but the cells remained nonsusceptible to HCoV-HKU1 S pseudotyped virus infection. Our data suggest that HLA-C is involved in the attachment of HCoV-HKU1 to A549 cells and is a potential candidate to facilitate cell entry. However, other unknown surface proteins on A549 cells may be concomitantly utilized by S glycoprotein of HCoV-HKU1 during viral entry. Further studies are required to elucidate other putative receptors or coreceptors for HCoV-HKU1 and the mechanism of HCoV-HKU1 S-mediated cell entry.

First Report of Tsukamurella Keratitis: Association between T. tyrosinosolvens and T. pulmonis and Ophthalmologic Infections

Abstract: We describe the first two cases of Tsukamurella keratitis, presented as eye pain with or without blurring of vision. One case was associated with trichiasis and the other with contact lens wear. The two isolates were identified as T. tyrosinosolvens and T. pulmonis, respectively, by phenotypic characterization and 16S rRNA sequencing.

Development of a multi-locus sequence typing scheme for Laribacter hongkongensis , a novel bacterium associated with freshwater fish-borne gastroenteritis and traveler's diarrhea

Abstract: Background Laribacter hongkongensis is a newly discovered, facultative anaerobic, Gram-negative, motile, sea gull-shaped rod associated with freshwater fish borne gastroenteritis and traveler's diarrhea. A highly reproducible and discriminative typing system is essential for better understanding of the epidemiology of L. hongkongensis. In this study, a multilocus sequence typing (MLST) system was developed for L. hongkongensis. The system was used to characterize 146 L. hongkongensis isolates, including 39 from humans and 107 from fish.

Clinical features and molecular epidemiology of coronavirus-HKU1-associated community-acquired pneumonia.

Abstract: 1. Coronavirus (CoV)-HKU1 accounts for 2.4% of community-acquired pneumonia. 2. Clinical features alone cannot differentiate this entity from other community-acquired pneumonia. 3. Further studies are needed to understand the significance of CoV-HKU1 in upper respiratory tract infection and its potential to cause outbreaks of acute viral respiratory illnesses.

Susceptibility patterns of clinical and fish isolates of Laribacter hongkongensis: comparison of the Etest, disc diffusion and broth microdilution methods

Abstract: Results: All isolates were susceptible to imipenem and ciprofloxacin by all three methods, except for one strain which was resistant to ciprofloxacin by broth microdilution. All were susceptible to ampicillin/sulbactam by Etest and disc diffusion, but eight were resistant by broth microdilution. By broth microdilution, 90%, 100%, 46.7%, 100% and 8.3% of isolates were resistant to ampicillin, ceftriaxone, cefuroxime, erythromycin and tetracycline, respectively. Although broth microdilution generally yielded higher MICs of b-lactams, MICs obtained with Etest were in good correlation with broth microdilution for all drugs except ampicillin/sulbactam, with >90% agreement within 2 log2 dilutions for imipenem, ciprofloxacin, erythromycin and tetracycline. Comparison of susceptibilities between broth microdilution and the other two methods showed the highest (>95%) percentage agreement for imipenem, ciprofloxacin and tetracycline. The highest discrepancies were observed with erythromycin (58.3% agreement), with an apparent increase in susceptibility by disc diffusion. A higher proportion of human isolates than fish isolates were tetracycline-resistant by all three tests (P 50.022).

Environmental surveillance for Laribacter hongkongensis, a diarrhoeal pathogen discovered in Hong Kong.

Abstract: 1. Laribacter hongkongensis was isolated from the midguts and hindguts of 86 (24%) of 360 freshwater fish from retail markets, including grass carp (60%), bighead carp (53%), mud carp (25%), and large-mouth bass (5%). 2. This study is the first to demonstrate the presence of L hongkongensis in natural water environments, with the bacterium being isolated from the waters of six reservoirs, with higher recovery rates in summer and during days of higher water and ambient temperatures. 3. Molecular typing using pulsed field gel electrophoresis revealed a heterogeneous population of L hongkongensis in both the freshwater fish and drinking water reservoir isolates, suggesting that the bacterium is endemic in our freshwater environments. 4. Since freshwater fish are common food items for our population, the general public should be educated on the proper preparation and thorough cooking of freshwater fish before consumption to avoid L hongkongensis-associated gastroenteritis. 5. Although it is unlikely that treated drinking water is a significant source of L hongkongensis-associated gastroenteritis, it is important to be aware of the possibility of other contaminated water as a source of human infection.

Swine influenza: then and now.

Abstract: While everybody is worrying about when the avian influenza A (H5N1) virus will adapt well enough in human to cause a pandemic, the emergence of a novel swine-origin influenza A (H1N1) virus (S-OIV) has shocked the world. On 29 April 2009, the World Health Organization raised the pandemic alert level from phase 4 to 5. As of 15 May, there have been 7503 laboratory-confirmed cases of S-OIV infections and 65 confirmed deaths globally, with more than 95% of the confirmed cases and all the deaths from the American continent. The S-OIV genome is made up of a unique combination of gene segments that had not been observed in previously known influenza A viruses as a result of genetic reassortment.

An oral mucosal DNA vaccine for SARS coronavirus infections.

Abstract: 1. When different forms of SARS coronavirus (SARS-CoV) spike protein-based vaccines for generation of a neutralising antibody response to SARS-CoV were injected into a mouse model, all the mice immunised with intramuscular tPA-optimised 800 DNA vaccine boosted with intraperitoneal recombinant spike polypeptide generated by Escherichia coli and intramuscular CTLA4Hinge SARS800 DNA vaccine boosted with intraperitoneal S-peptide had neutralising antibody titres of>1:1280.2. This observation may have major practical value for field studies, such as the immunisation of civet cats, as the cost of recombinant proteins produced by E coli is much lower than those produced by eukaryotic systems.3. This study indicates that the type of vaccine used for priming is crucial for determining the type of immune response developed.Subsequent doses will boost the immune response generated by the first dose of vaccine.