Jade L. L. Teng

The mitochondrial genome of the thermal dimorphic fungus Penicillium marneffei is more closely related to those of molds than yeasts.

We report the complete sequence of the mitochondrial genome of Penicillium marneffei, the first complete mitochondrial DNA sequence of a thermal dimorphic fungus. This 35 kb mitochondrial genome contains the genes encoding ATP synthase subunits 6, 8, and 9 (atp6, atp8, and atp9), cytochrome oxidase subunits I, II, and III (cox1, cox2, and cox3), apocytochrome b (cob), reduced nicotinamide adenine dinucleotide ubiquinone oxireductase subunits (nad1, nad2, nad3, nad4, nad4L, nad5, and nad6), ribosomal protein of the small ribosomal subunit (rps), 28 tRNAs, and small and large ribosomal RNAs. Analysis of gene contents, gene orders, and gene sequences revealed that the mitochondrial genome of P. marneffei is more closely related to those of molds than yeasts.

Discovery of seven novel Mammalian and avian coronaviruses in the genus deltacoronavirus supports bat coronaviruses as the gene source of alphacoronavirus and betacoronavirus and avian coronaviruses as the gene source of gammacoronavirus and deltacoronavirus.

ABSTRACT Recently, we reported the discovery of three novel coronaviruses, bulbul coronavirus HKU11, thrush coronavirus HKU12, and munia coronavirus HKU13, which were identified as representatives of a novel genus, Deltacoronavirus, in the subfamily Coronavirinae. In this territory-wide molecular epidemiology study involving 3,137 mammals and 3,298 birds, we discovered seven additional novel deltacoronaviruses in pigs and birds, which we named porcine coronavirus HKU15, white-eye coronavirus HKU16, sparrow coronavirus HKU17, magpie robin coronavirus HKU18, night heron coronavirus HKU19, wigeon coronavirus HKU20, and common moorhen coronavirus HKU21. Complete genome sequencing and comparative genome analysis showed that the avian and mammalian deltacoronaviruses have similar genome characteristics and structures. They all have relatively small genomes (25.421 to 26.674 kb), the smallest among all coronaviruses. They all have a single papain-like protease domain in the nsp3 gene; an accessory gene, NS6 open reading frame (ORF), located between the M and N genes; and a variable number of accessory genes (up to four) downstream of the N gene. Moreover, they all have the same putative transcription regulatory sequence of ACACCA. Molecular clock analysis showed that the most recent common ancestor of all coronaviruses was estimated at approximately 8100 BC, and those of Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus were at approximately 2400 BC, 3300 BC, 2800 BC, and 3000 BC, respectively. From our studies, it appears that bats and birds, the warm blooded flying vertebrates, are ideal hosts for the coronavirus gene source, bats for Alphacoronavirus and Betacoronavirus and birds for Gammacoronavirus and Deltacoronavirus, to fuel coronavirus evolution and dissemination.

Laribacter hongkongensis gen. nov., sp. nov., a Novel Gram-Negative Bacterium Isolated from a Cirrhotic Patient with Bacteremia and Empyema

ABSTRACT A bacterium was isolated from the blood and empyema of a cirrhotic patient. The cells were facultatively anaerobic, nonsporulating, gram-negative, seagull shaped or spiral rods. The bacterium grows on sheep blood agar as nonhemolytic, gray colonies 1 mm in diameter after 24 h of incubation at 37°C in ambient air. Growth also occurs on MacConkey agar and at 25 and 42°C but not at 4, 44, and 50°C. The bacterium can grow in 1 or 2% but not 3, 4, or 5% NaCl. No enhancement of growth is observed with 5% CO2. The organism is aflagellated and nonmotile at both 25 and 37°C. It is oxidase, catalase, urease, and arginine dihydrolase positive, and it reduces nitrate. It does not ferment, oxidize, or assimilate any sugar tested. 16S rRNA gene sequencing showed that there are 91 base differences (6.2%), 112 base differences (7.7%), and 116 base differences (8.2%) between the bacterium andMicrovirgulaaerodenitrificans,Vogesellaindigofera, andChromobacterium species, respectively. The G+C content (mean and standard deviation) is 68.0% ± 2.43%, and the genomic size is about 3 Mb. Based on phylogenetic affiliation, the bacterium belongs to the Neisseriaceae family of the β-subclass ofProteobacteria. For these reasons, a new genus and species, Laribacterhongkongensis gen. nov., sp. nov., is proposed, for which HKU1 is the type strain. Further studies should be performed to ascertain the potential of this bacterium to become an emerging pathogen.

New Hepatitis E Virus Genotype in Camels, the Middle East

In a molecular epidemiology study of hepatitis E virus (HEV) in dromedaries in Dubai, United Arab Emirates, HEV was detected in fecal samples from 3 camels. Complete genome sequencing of 2 strains showed >20% overall nucleotide difference to known HEVs. Comparative genomic and phylogenetic analyses revealed a previously unrecognized HEV genotype.

Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis

ABSTRACT A bacterium was isolated from the blood culture of a patient with infective endocarditis. The cells were facultative anaerobic, nonsporulating, gram-positive cocci arranged in chains. The bacterium grows on sheep blood agar as alpha-hemolytic, gray colonies of 0.5 to 1 mm in diameter after 24 h of incubation at 37°C in ambient air. Growth also occurs in 10 or 40% bile and on bile esculin agar but not in 6% NaCl. No enhancement of growth is observed in 5% CO2. It is nongroupable with Lancefield groups A, B, C, D, F, or G antisera and is resistant to optochin and bacitracin. The organism is aflagellated and is nonmotile at both 25 and 37°C. It is Voges-Proskauer test positive. It produces leucine arylamidase and β-glucosidase but not catalase, urease, lysine decarboxylase, or ornithine decarboxylase. It hydrolyzes esculin and arginine. It utilizes glucose, lactose, salicin, sucrose, pullulan, trehalose, cellobiose, hemicellulase, mannose, maltose, and starch. 16S rRNA gene sequencing showed that there were 3.6, 3.7, 4.3, 4.7, and 5.9% differences between the 16S rRNA gene sequence of the bacterium and those of Streptococcus gordonii, Streptococcus intermedius, Streptococcus constellatus, Streptococcus sanguis, and Streptococcus anginosus, respectively. The G+C content of it (mean ± standard deviation) was 53.0% ± 2.9%. Based on phylogenetic affiliation, it belongs to the mitis or anginosus group of Streptococcus. For these reasons a new species, Streptococcus sinensis sp. nov., is proposed, for which HKU4 is the type strain. Further studies should be performed to ascertain the potential of this bacterium to become an emerging cause of infective endocarditis.

Natural Occurrence and Characterization of Two Internal Ribosome Entry Site Elements in a Novel Virus, Canine Picodicistrovirus, in the Picornavirus-Like Superfamily

ABSTRACT Dicistroviridae and Picornaviridae are two phylogenetically related families of positive-sense single-stranded RNA viruses in the picornavirus-like superfamily with similar gene contents but different genome organizations and hosts. In a surveillance study involving 1,472 samples from 368 dogs over a 22-month period, we identified a novel picornavirus-like virus from 47 fecal and urine samples by the use of reverse transcription-PCR (RT-PCR). Sequencing and phylogenetic analysis of three complete genomes revealed that, although it seemed that the virus was most closely related to other picornaviruses, P1, P2, and P3 of the virus possessed very low amino acid identities of <30% to those of all other known picornaviruses and that the amino acid identities between the 3Dpol and 2C of the virus and the RNA-dependent RNA polymerases and helicases of all other picornaviruses were <35%. Distinct from other picornaviruses, the genomes of the virus contain two putative internal ribosome entry sites (IRESs) and two open reading frames, encoding two polyprotein precursors (844 and 1,406 amino acids), separated by an intergenic region (IGR) of 588 bases. A dual-luciferase activity assay using DNA and RNA transfection revealed that both IRESs were functional. Quantitative RT-PCR showed that numbers of viral RNAs ranged from 7.55 × 106 to 1.26 × 109 copies/ml of urine and 1.82 × 106 to 4.97 × 1010 copies/ml of fecal sample. This is the first report of the natural occurrence of two functional IRESs in nondicistroviruses. Based on our results, we have proposed a novel species, canine picodicistrovirus (CPDV), to describe this novel member of the picornavirus-like superfamily, which could represent a novel family of viruses.

The complete genome and proteome of Laribacter hongkongensis reveal potential mechanisms for adaptations to different temperatures and habitats.

Laribacter hongkongensis is a newly discovered Gram-negative bacillus of the Neisseriaceae family associated with freshwater fish–borne gastroenteritis and travelers 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°C (human body temperature) and 20°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°C, whereas NAGK-37 showed higher expression at 37°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.

Analysis of a Viridans Group Strain Reveals a Case of Bacteremia Due to Lancefield Group G Alpha-Hemolytic Streptococcus dysgalactiae subsp. equisimilis in a Patient with Pyomyositis and Reactive Arthritis

ABSTRACT Streptococcus dysgalactiae is classified by a combination of phenotypic and genotypic characteristics into Lancefield group C alpha-hemolytic Streptococcus dysgalactiae subsp. dysgalactiae and Lancefield group C, group G, and group L beta-hemolytic Streptococcus dysgalactiae subsp. equisimilis. In this study, we report the isolation of a catalase-negative, alpha-hemolytic, optochin- and bacitracin-resistant viridans group strain, which does not grow in 10 or 40% bile, on MacConkey agar or bile esculin agar, or in 6% NaCl, from the blood culture of a 73-year-old woman with pyomyositis and poststreptococcal reactive arthritis. Lancefield grouping revealed that the strain was a group G streptococcus. The Vitek system (GPI) showed that it was unidentified, and the API system (20 STREP) showed that it was 95.7% S. dysgalactiae subsp. dysgalactiae. 16S rRNA gene sequencing showed that it was a strain of S. dysgalactiae. Based on phylogenetic affiliation with 16S rRNA gene or GroEL amino acid (another bacterial gene, in addition to 16S rRNA gene, that is highly conserved) sequences, the strain is most closely related to Lancefield group C beta-hemolytic S. dysgalactiae subsp. equisimilis. PCR amplification and sequencing of the streptolysin S structural gene (sagA) and M protein gene (emm) hypervariable region showed the presence of these suspected primary virulence factors. Further studies would delineate whether the isolate is just a hemolysin-deficient variant of group G beta-hemolytic S. dysgalactiae subsp. equisimilis or a novel type of S. dysgalactiae. The present case showed that group G alpha-hemolytic S. dysgalactiae subsp. equisimilis can be associated with serious invasive infection and poststreptococcal sequelae.

Identification and characterization of bocaviruses in cats and dogs reveals a novel feline bocavirus and a novel genetic group of canine bocavirus.

We report the identification and genome characterization of a novel bocavirus, feline bocavirus (FBoV), and novel bocaviruses closely related to canine bocavirus (CBoV) strain Con-161 in stray cats and dogs in Hong Kong, respectively. FBoV was detected by PCR in 7.2, 0.3, 1.6, 2.0 and 0.8% of faecal, nasal, urine, kidney and blood samples, respectively, from 364 cats, while CBoV was detected in 4.6, 5.1, 6.3 and 0.3% of faecal, nasal, urine and blood samples, respectively, from 351 dogs. Three FBoV genomes sequenced revealed the presence of three ORFs characteristic of bocaviruses. Phylogenetic analysis showed that FBoVs were related only distantly to other bocaviruses, forming a distinct cluster within the genus, with ≤ 5.7% nucleotide identities to the genome of minute virus of canines. The four CBoV genomes sequenced shared 87.4-89.2% nucleotide identities with that of CBoV strain Con-161. In addition to the three bocavirus ORFs, they encoded an additional ORF, ORF4, immediately downstream of the ORF for non-structural protein 1 (NS1), which was not found in other bocaviruses including CBoV strain Con-161. They also possessed a putative second exon encoding the C-terminal region of NS1 and conserved RNA-splicing signals, previously described in human bocaviruses. Partial VP1/VP2 sequence analysis of 23 FBoV and 25 CBoV strains demonstrated inter-host genetic diversity, with two potential genetic groups of FBoV and a novel CBoV group, CBoV-HK, distinct from the three groups, CBoV-A to -C, found in the USA. Although the pathogenicity of FBoV and CBoV remains to be determined, their presence in different host tissues suggested wide tissue tropism.

Ecoepidemiology of Laribacter hongkongensis, a Novel Bacterium Associated with Gastroenteritis

ABSTRACT In a territory-wide surveillance study, Laribacter hongkongensis was isolated solely from freshwater fish (60% of grass carps, 53% of bighead carps, and 25% of mud carps). Comparing the pulsed-field gel electrophoresis patterns of fish and patient isolates revealed that most patient isolates were clustered together, suggesting that some clones could be more virulent.