Replication of H9 influenza viruses in the human ex vivo respiratory tract, and the influence of neuraminidase on virus release

H9N2 viruses are the most widespread influenza viruses in poultry in Asia. We evaluated the infection and tropism of human and avian H9 influenza virus in the human respiratory tract using ex vivo respiratory organ culture. H9 viruses infected the upper and lower respiratory tract and the majority of H9 viruses had a decreased ability to release virus from the bronchus rather than the lung. This may be attributed to a weak neuraminidase (NA) cleavage of carbon-6-linked sialic acid (Sia) rather than carbon-3-linked Sia. The modified cleavage of N-acetlylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) by NA in H9 virus replication was observed by reverse genetics, and recombinant H9N2 viruses with amino acids (38KQ) deleted in the NA stalk, and changing the amino acid at position 431 from Proline-to-Lysine. Using recombinant H9 viruses previously evaluated in the ferret, we found that viruses which replicated well in the ferret did not replicate to the same extent in the human ex vivo cultures. The existing risk assessment models for H9N2 viruses in ferrets may not always have a strong correlation with the replication in the human upper respiratory tract. The inclusion of the human ex vivo cultures would further strengthen the future risk-assessment strategies.published_or_final_versio

Three Drosophila Hox Complex microRNAs Do Not Have Major Effects on Expression of Evolutionarily Conserved Hox Gene Targets during Embryogenesis

The discovery of microRNAs has resulted in a major expansion of the number of molecules known to be involved in gene regulation. Elucidating the functions of animal microRNAs has posed a significant challenge as their target interactions with messenger RNAs do not adhere to simple rules. Of the thousands of known animal microRNAs, relatively few microRNA:messenger RNA regulatory interactions have been biologically validated in an normal organismal context. Here we present evidence that three microRNAs from the Hox complex in Drosophila (miR-10-5p, miR-10-3p, miR-iab-4-5p) do not have significant effects during embryogenesis on the expression of Hox genes that contain high confidence microRNAs target sites in the 3′ untranslated regions of their messenger RNAs. This is significant, in that it suggests that many predicted microRNA-target interactions may not be biologically relevant, or that the outcomes of these interactions may be so subtle that mutants may only show phenotypes in specific contexts, such as in environmental stress conditions, or in combinations with other microRNA mutations

Spectroscopic properties of luminescent platinum(II) complexes containing 4,4′,4″-tri-tert-butyl-2,2′:6′,2″-terpyridine (tBu3tpy). Crystal structures of [Pt(tBu3tpy)Cl]ClO4 and [Pt(tBu3tpy){CH2C(O)Me}]ClO4

The 77 K emission spectrum of [Pt(tBu3tpy)Cl]ClO4, 1(ClO4) (tBu3tpy = 4,4′,4″-tri-tert-butyl-2,2′:6′,2″-terpyridine), in a 10-3 mol dm-3 MeOH/EtOH glassy solution exhibits a 625 nm band attributed to tBu3tpy π-π interactions, whereas a dilute solution (10-6 mol dm-3) shows a vibronic intraligand emission at λmax 467 nm. By comparing the UV-vis spectra of 1 with the 1IL-dominated spectrum of [Au(tBu3tpy)Cl]2+ (4) in CH3CN, the moderately intense absorption band at λmax 373-386 nm for the former is ascribed to a 1MLCT [(5d)Pt → π*(tBu3tpy)] transition. The absorption spectrum of 1 contains a weak shoulder at 465 nm (∈ = 57 dm3 mol-1 cm-1) which obeys Beer's law in the concentration range 5 × 10-4 to 1 × 10-2 mol dm-3, and a 3MLCT assignment is proposed. Ketonyl platinum(II) complexes [Pt(tBu3tpy){CH2C(O)R}]ClO4 (R = Me, 2 and R = Ph, 3) have been prepared from 1(ClO4) in the presence of aqueous NaOH and the corresponding methyl ketone. The molecular structures of 1(ClO4) and 2(ClO4)·C2H5OH reveal π-stacking orientations for the tBu3tpy ligands with interplanar separations of ca. 3.8 and 3.6 Å, respectively. Complexes 2 and 3 are luminescent in solution at room temperature, and 3MLCT excited states are assigned, while 1 is nonemissive. The superior photophysical properties of 2 and 3 are ascribed to the stronger σ-donating ability of the ketonyl ligands. Like 1, complexes 2 and 3 also display low-energy emission at ca. 620 nm in 10-3 mol dm-3 MeOH/EtOH glass at 77 K. Complex 1(ClO4) is emissive in crystalline form at 77 K only (λmax 560 nm). Solid-state emissions of 2(ClO4) and 3(ClO4) at room temperature (λmax 607 and 615 nm, respectively) are derived from weak stacking interactions between tBu3tpy groups, while their emission maxima are red-shifted at 77 K. © 1999 American Chemical Society.link_to_subscribed_fulltex

Carbene and Isocyanide Ligation at Luminescent Cyclometalated 6-Phenyl-2,2′-bipyridyl Platinum(II) Complexes: Structural and Spectroscopic Studies

A series of luminescent cyclometalated platinum(II) diamino-carbene complexes, namely, [(CNN)Pt{C(NHR1)(NHR2)}]+ (HCNN = 6-phenyl-2,2′-bipyridine; for R1 = tBu, R2 = Me (2), NH2 (3), CH2Ph (4); for R1 = 2,6-Me2C6H3 (Ar′), R2 = Me (6), NH2 (7), CH2Ph (8)), are synthesized by nucleophilic attack of amines at the coordinated isocyanide ligands of [(CNN)-PtC≡NR1]+ (R1 = tBu (1), Ar′ (5)). The binuclear bridging bis(carbene) derivative [{(CNN)Pt}2μ-{C(NHtBu)(NH(CH2) 3)}2NMe]2+ (9) is prepared by treatment of 1 with excess 3,3′-diamino-N-methyldipropylamine. The molecular structures of 2(ClO4)•0.5H2O, 3(ClO4)• 0.5H2O, 4(ClO4), and 6(ClO4) reveal Pt-C(carbene) distances of 1.997(7), 1.992(4), 1.989(6), and 1.996(8) Å, respectively. Short N-C(carbene) bond lengths (mean 1.332 Å) imply substantial pπ-pπ interactions within the N-C(carbene)-N fragment, and minimal π bonding in the Pt-C(carbene) moiety is inferred. Weak π-π stacking interactions between the CNN ligands are observed in the crystal lattice of 2-4 and 6 (range 3.5-3.6 Å). Complexes 1-9 display structureless emissions, with λmax ranging from 528 to 558 nm in acetonitrile at 298 K, and they are assigned to 3MLCT excited states. The emissions of 1 and 5 exhibit noticeably higher quantum yields, which reflect the strong ligand field strength of the isocyanide auxiliaries. The 77 K emissions of 1-9 in frozen acetonitrile are blue-shifted and display well-resolved vibronic structures. At high concentrations (≥10-5 M), 1 and 5 show an additional low-energy band at 627 (in butyronitrile) and 730 nm which are ascribed to π-π excimeric and MMLCT emissions, respectively. The solid-state emissions of complexes 1(ClO4)-9(ClO4)2 (except 5(ClO4)) at 298 K are red-shifted from solution to λmax 553-579 nm with a shoulder at 595-612 nm. They are tentatively assigned to 3MLCT excited states with excimeric character due to weak CNN π-π interactions, as evident in the crystal structures. The remarkable low-energy luminescence of 5(ClO4) in the solid state (λmax 704 nm at 298 K; 781 nm at 77 K) is attributed to MMLCT transitions arising from solid-state intermolecular stacking interactions. Upon photolysis with UV-visible light, reaction between complex 2 and iodomethane in acetonitrile leads to decomposition of the diamino-carbene ligand and generation of the platinum(IV) diiodide cyanide derivative trans-[(CNN)-Pt(C≡N)I2] (10).link_to_subscribed_fulltex

Self-assembly of predesigned trimetallic macrocycles based on benzimidazole as nonlinear bridging motifs: Crystal structure of a luminescent platinum(II) cyclic trimer

A trimetallic macrocycle: Starting from nonlinear N-deprotonated benzimidazole and square-planar cyclometalated platinum(II) precursors, cyclic trimers with three metal vertices can be obtained by a general and rational strategy (shown schematically, M=Pt). A complex is formed with 2- (2-thienyl)pyridine as chelating ligand which photoluminesces in solution at room temperature.link_to_subscribed_fulltex

Probing d8-d8 interactions in luminescent mono- and binuclear cyclometalated platinum(II) complexes of 6-phenyl-2,2′-bipyridines

A series of luminescent mono- and binuclear cyclometalated platinum(II) complexes, namely [Pt(L1-6)Cl] (1a-6a; HL1-6 = 4-(aryl)-6-phenyl-2,2′-bipyridine; aryl = H (1), phenyl (2), 4-chlorophenyl (3), 4-tolyl (4), 4-methoxyphenyl (5), 3,4,5-trimethoxyphenyl (6)), [Pt(L1)E]+ (E = py (7), PPh3 (8)), [Pt2(L1-6)2(μ-dppm)]2+ (1b-6b, dppm = bis(diphenylphosphino)methane), [Pt2(L1)2(μ-pz)]+ (9, Hpz = pyrazole), and [Pt2(L1)2(μ-dppCn)]2+ (dppCn = bis(diphenylphosphino)propane (10, n = 3) and -pentane (11, n = 5)), were synthesized in order to examine fluid- and solid-state oligomeric d8-d8 and ligand-ligand interactions. The molecular structures of 4b(ClO4)2 and 9(PF6) reveal intramolecular Pt-Pt distances of 3.245(1) and 3.612(2) Å, respectively. While minimal metal-metal communication is expected for 9, weak π-π interactions are possible. All complexes described in this work are emissive in fluid solution at room temperature. Negligible changes in emission energy are detected by incorporating different aryl substituents into the 4-position of 6-phenyl-2,2′-bipyridine, and this indicates little electronic delocalization between them. Self-quenching of the 3MLCT emission by the mononuclear derivatives are observed in CH2Cl2 at 298 K, and a red shift in the emission energy is exhibited by complex 7 in acetonitrile at 77 K. The fluid emissions of the μ-dppm species 1b-6b at λmax 652-662 nm appear at substantially lower energies than their mononuclear counterparts and show dramatic solvatochromic effects. These emissions are ascribed to 3[dσ*, π*] excited states. In contrast, the emission of 10 and 11, bearing long bridging diphosphine ligands, are attributed to 3MLCT states of non-interacting [Pt(L1)] moieties. Significantly, the luminescence of the μ-pyrazolate complex 9 displays transitional features which are reminiscent of both 3[dσ*, π*} and 3MLCT excited states. Hence a relationship is observed between emission energy, the nature of the lowest energy excited state, and metal-metal interactions. The excited-state redox potential [E(*Pt2 2+/Pt2 +)] of 1b has been estimated by electrochemical studies (1.61 V vs NHE) and by quenching experiments with aromatic hydrocarbons (1.63 V vs NHE). © 1999 American Chemical Society.link_to_subscribed_fulltex

Synthesis of organoplatinum oligomers by employing N-donor bridges with predesigned geometry: Structural and photophysical properties of luminescent cyclometalated platinum(II) macrocycles

A series of luminescent di- and trimeric cyclometalated platinum(II) macrocycles, namely, [Pt(C-N)(N-N)]2 (N-N = pyrazolate, pz (2); 7-azaindolate, aza (3); C-N = 2-(2′-thienyl)-pyridyl, Thpy (a); 7,8-benzoquinolate, Bzqn (b); 2-phenylpyridyl, Phpy (c)) and [Pt(C-N)-(N-N)]3 (N-N = benzimidazolate, bzim (4); C-N = Thpy (a) and Bzqn (b)), are synthesized in high yields (70-90%). The rigid, predefined coordination geometry of the pz, aza, and bzim bridging ligands ensure the efficient and selective assembly of the respective cyclic oligomers. The stacking arrangements in the crystal structures of 1a and 4a reveal π-π interactions between 2-(2′-thienyl)pyridyl moieties. The lowest energy absorption band in the UV-vis spectra is assigned to metal-to-ligand charge transfer (MLCT) transitions. The 298 K fluid emissions of complexes 2-4a bearing Thpy ligands are assigned to transitions with mixed MLCT and IL (intraligand) character. In crystalline form, the observed red-shift is attributed to excimeric emission arising from π-stacking in the solid state. In contrast, a number of Bzqn derivatives are nonemissive in CH2Cl2 at 298 K. The energy of the lowest energy UV-vis and emissive bands for Thpy oligomers are red-shifted relative to the Bzqn and Phpy congeners. Significantly, Thpy complexes and especially the μ-pyrazolate species 2a (τo = 15.5 μs, Φo = 0.18) emit with longer lifetimes and higher quantum yields.link_to_subscribed_fulltex

Sapovirus detection by quantitative real-time RT-PCR in clinical stool specimens

Sapovirus (SV) is one of the major causative agents of viral gastroenteritis affecting all age groups worldwide. A new method for the quantitative detection of SV from clinical stool specimens by real-time reverse transcription-polymerase chain reaction (RT-PCR) based on TaqMan ® MGB technology was described. Primers and probe were designed to target the RNA-dependent RNA polymerase/capsid genes junction. Performance of the newly developed assay was validated against a panel of 244 clinical stool specimens collected for patients with gastroenteritis. SV was detected in eight (3.3%) specimens. Phylogenetic analysis of the positive isolates suggested that the assay could detect at least SV genogroups I, II and IV. In addition, the assay had an increased detection rate compared with a widely used conventional RT-PCR assay. Quantitative analysis showed that the assay could detect as low as 10 copies of viral cDNA per reaction. No cross-reactivity with norovirus and rotavirus was observed. In conclusion, the assay is a sensitive and specific method for the detection of SV from clinical stool specimens. © 2005 Elsevier B.V. All rights reserved.link_to_subscribed_fulltex

Luminescent metal complexes derived from carbene and related ligands: Tuning excited-state properties with metal-carbon multiple bonds

This account begins with a report on the photophysical responses to pH exhibited by the luminescent chelating bis(carbene) platinum(II) complexes [(C10H21N4)Pt(CN)(CNtBu)] (1) and [(C10H21N4)Pt(CNtBu) 2]+ (2), and concludes with an overview of recent work by our group regarding luminescent platinum and rhenium derivatives bearing metal-carbon multiple bonds. © 2001 Elsevier Science B.V.link_to_subscribed_fulltex