scholarly journals Passaging impact of H9N2 avian influenza virus in hamsters on its pathogenicity and genetic variability

2014 ◽  
Vol 8 (05) ◽  
pp. 570-580 ◽  
Author(s):  
Houssam A. Shaib ◽  
Nelly Cochet ◽  
Thierry Ribeiro ◽  
Afif M Abdel Nour ◽  
Georges Nemer ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Avian Influenza ◽  
Mammalian Cells ◽  
Pcr Amplification ◽  
Amino Acid Sequences ◽  
Influenza Viruses ◽  
H9n2 Virus ◽  
World Health ◽  
The Impact

Introduction: Avian influenza viruses of the H9N2 subtype have been reported to cause human infections. This study demonstrates the impact of nasal viral passaging of avian H9N2 in hamsters on its cross species-pathogenic adaptability and variability of amino acid sequences of the hemagglutinin (HA) and neuraminidase (NA) stalk. Methodology: Three intranasal passagings of avian H9N2 in hamsters P1, P2, and P3 were accomplished. Morbidity signs and lesions were observed three days post viral inoculation. The HA test was used for presumptive detection of H9N2 virus in the trachea and lungs of the hamsters challenged with the differently passaged viruses. Different primers were used for PCR amplification of the HA1 and NA stalk regions of the differently passaged H9N2 viruses, followed by sequence alignment. Results: The morbidity signs indicated low pathogenicity of the differently passaged H9N2 viruses in hamsters. The frequency of gross and microscopic lesions in the tracheas and lungs were insignificantly different among hamsters challenged with the differently passaged H9N2 viruses (p > 0.05). There was 100% similarity in the amino acid sequence of the HA gene of most passaged viruses. The amino acid sequence of the neuraminidase in the third passaged H9N2 virus recovered from lungs showed a R46P mutation that might have a role in the pathogenic adaptability of P3 viruses in hamsters’ lungs. Conclusions: The apparent adaptation of avian H9N2 virus to mammalian cells is in agreement with the World Health Organization’s alertness for a possible public health threat by this adaptable virus.

scholarly journals Evolution and Adaptation of the Avian H7N9 Virus into the Human Host

2020 ◽  
Vol 8 (5) ◽  
pp. 778
Author(s):  
Andrew T. Bisset ◽  
Gerard F. Hoyne
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Influenza A ◽  
Amino Acid Sequences ◽  
Influenza Viruses ◽  
Amino Acid Substitutions ◽  
Upper Respiratory Tract ◽  
Viral Polymerase ◽  
Evolutionary Changes ◽  
Avian Origin

Influenza viruses arise from animal reservoirs, and have the potential to cause pandemics. In 2013, low pathogenic novel avian influenza A(H7N9) viruses emerged in China, resulting from the reassortment of avian-origin viruses. Following evolutionary changes, highly pathogenic strains of avian influenza A(H7N9) viruses emerged in late 2016. Changes in pathogenicity and virulence of H7N9 viruses have been linked to potential mutations in the viral glycoproteins hemagglutinin (HA) and neuraminidase (NA), as well as the viral polymerase basic protein 2 (PB2). Recognizing that effective viral transmission of the influenza A virus (IAV) between humans requires efficient attachment to the upper respiratory tract and replication through the viral polymerase complex, experimental evidence demonstrates the potential H7N9 has for increased binding affinity and replication, following specific amino acid substitutions in HA and PB2. Additionally, the deletion of extended amino acid sequences in the NA stalk length was shown to produce a significant increase in pathogenicity in mice. Research shows that significant changes in transmissibility, pathogenicity and virulence are possible after one or a few amino acid substitutions. This review aims to summarise key findings from that research. To date, all strains of H7N9 viruses remain restricted to avian reservoirs, with no evidence of sustained human-to-human transmission, although mutations in specific viral proteins reveal the efficacy with which these viruses could evolve into a highly virulent and infectious, human-to-human transmitted virus.

The Single E627K Amino Acid Substitution in PB2 Enhances the Pathogenicity of Wild-Bird-Origin H6N6 Subtype Avian Influenza Virus in Mice

2020 ◽  
Vol 7 (1) ◽  
Author(s):  
Miura H ◽  
◽  
Ozeki Y ◽  
Omatsu T ◽  
Katayama Y ◽  
...  
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Wild Bird ◽  
Virus Transmission ◽  
Influenza Viruses ◽  
Whole Genome Sequence ◽  
Species Barrier ◽  
The Impact ◽  
Original Virus ◽  
Wild Waterfowl

Avian Influenza Viruses (AIVs) are harbored by wild waterfowl as a natural host, and there is a species barrier restricting virus transmission from birds to mammals, including humans. However, it has been reported that, through genetic mutations, AIVs occasionally infect mammals and acquire high pathogenicity. The Amino Acid (aa) substitution of glutamic acid to lysine at position 627 (E627K) in polymerase basic protein 2 (PB2) is one of the wellknown factors underlying mammalian adaptation. Although this substitution was previously observed in mammalian-adapted H5, H7, and H9 AIV subtypes, the impact of this mutation on the mammalian adaptation of other AIV subtypes is not fully verified. Here, we isolated the low pathogenic AIV subtype H6N6 from a wild bird fecal sample in Tokachi Subprefecture, Hokkaido, Japan. We passaged this H6N6 subtype in BALB/c mice four times and acquired the mouse-adapted virus. Whole-genome sequence analysis showed that the adapted virus had only one aa substitution (E627K) in PB2. The adapted virus-inoculated mice tended to show increased weight loss and mortality compared with the original virus-inoculated mice. The viral titer in the lungs of the adapted virus-inoculated mice was significantly higher than that of the original virus-inoculated mice. Additionally, the virus isolated from the lung of the original virus-inoculated mice with serious symptoms harbored the E627K substitution. Our findings indicate the possibility that the PB2 E627K substitution in H6N6 subtype AIV rapidly appears in mammalian hosts and contributes to the enhanced pathogenicity of this virus.

scholarly journals Molecular determinants of pathogenicity and host specificity of highly pathogenic h5N1 BiH isolates

2019 ◽  
Vol 49 ◽  
Author(s):  
Teufik Goletić ◽  
Abdulah Gagić ◽  
Vladimir Savić ◽  
Emina Rešidbegović ◽  
Aida Kavazović ◽  
...  
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Host Specificity ◽  
Pcr Amplification ◽  
Influenza Viruses ◽  
Viral Rna ◽  
Amino Acid Residues ◽  
Avian Influenza Viruses ◽  
Highly Pathogenic ◽  
Cygnus Olor

ABSTRACT Background: Towards preparation for a possible influenza pandemic, investigation of the molecular characteristics of the circulating avian H5N1 influenza virus strains is of crucial importance. These H5N1 viruses continue to spread, to infect animals and humans and to evolve and diversify providing so an ever-looming pandemic threat.Aim: To identify genetic structure and molecular biological characteristics of BiH's isolates of H5N1 HPAI as well as to assess the level of pathogenicity, phylogenetic origin and host- specificity of the isolates.Material and Methods: SPF embryonated chicken eggs were used for virus isolation. Viral RNA extracted using QIAamp viral RNA kit and manufacturer’s protocol (QIAGEN®) was used for PCR amplification. cDNA synthesis and PCR amplification of the coding region, using gene specific primer sets (primer sequences available on request), were carried out for all eight viral RNA segments separately. The Prism Big Dye Terminator v1.1 cycle sequencing kit (Applied Biosystems) was used and products were analyzed on an automatic ABI PRISM 3130 genetic analyzer (Applied Biosystems). Nucleotide sequences were analyzed using Bioedit software (v. 7.0.9.0) with an engine based on the ClustalW 1.4 algorithm. MEGA software (v. 4,0), using the neighbor joining tree inference analysis with the Tamura-Nei γ-model, was used to estimate phylogenies and calculate bootstrap values from the nucleotide sequences.Results: Full-length nucleotide sequences of the A/Cygnus olor/BIH/1/2006 (H5N1) strain were deposited in EMBL Nucleotide Sequence Database under accession nos. FN186008 to FN186014 and FM20943. The pathogenicity and host specificity of this strain, as polygenic traits, are determined in silico by the structure of its proteins, especially surface glycoproteins, HA and NA. Multibasic amino acid stretch PQGERRRKKR/GLF, marker of strains highly pathogenic to poultry, was present at the HA cleavage site of BiH strain. The RBS was typical for avian influenza viruses and contained Gln and Gly at positions 238 and 240 (H5 numbering) that is,226 and 228 according to H3 numbering with seven potential glycosylated sites but with increased binding to alpha2-6 sialoglycans thanks to substitutions, as follows, 110N, 171N, 171N, 172A, 205R and 251P. NA structure assigned this strain to the Z genotype, characterized also by the deletion of the five amino acid residues of the NS1 protein (positions 80-84). Amino acid residues, typical for the avian influenza viruses, were revealed in 40 out of 43 positions of M1, M2, NP, PA, PB2 and HA, determining the host range specificity. Phylogenetic analysis of the HA gene revealed that BiH isolates belonged to genetic clade 2.2., and presence of aspartic acid at the position of 403 of HA locate BiH isolates in 2.2.2. sub-clade.Conclusions: The BiH’s isolates were determined as HPAI virus with genes sequences closely related to A/Cygnus olor/Astrakhan/Ast05-2-10/2005 (H5N1). Three residues (M2 - 28V and 78K, NP - 33I), typical of human influenza viruses, were found, indicating a certain degree of intercurrent evolutionary adaptive changes in BiH isolates. Sequence comparison of HA and NA segments with relevant sequences in GenBank revealed that the BiH isolates and the ones from the southern Russia (Astrakhan region) group together phylogenetically, forming a monophyleticcluster in both genes indicating that these isolates have evolved from the same origin. Sequence derived phenotype markers of NA protein (E99, V129, D131, R136, H255 and Y256) as well as of M2 protein (26L, 27V, 30A, S31 and G34) showed that the isolates have an oseltamivir and amantadine sensitive genotype. 

scholarly journals Identification of cytokine-induced neutrophil chemoattractants (CINC), rat GRO/CINC-2α and CINC-2 β, produced by granulation tissue in culture: purification, complete amino acid sequences and characterization

1994 ◽  
Vol 301 (2) ◽  
pp. 545-550 ◽  
Author(s):  
H Nakagawa ◽  
N Komorita ◽  
F Shibata ◽  
A Ikesue ◽  
K Konishi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Granulation Tissue ◽  
Pcr Amplification ◽  
Neutrophil Infiltration ◽  
Amino Acid Sequences ◽  
Terminal Amino Acid ◽  
Reverse Transcription Pcr ◽  
Terminal Amino ◽  
The Difference

Four basic neutrophil chemotactic factors (chemokines) have been purified from conditioned medium of granulation tissue obtained from carrageenin-induced inflammation in the rat. On the basis of their N-terminal amino acid sequences, one of the chemokines was identical with rat GRO/cytokine-induced neutrophil chemoattractant (CINC) which we reported previously, and another was identical with rat macrophage inflammatory protein-2 (MIP-2). Two other chemokines were novel chemoattractants related to MIP-2. The novel chemokines are referred to as rat GRO/CINC-2 alpha and CINC-2 beta, and consequently CINC and rat MIP-2 are renamed rat GRO/CINC-1 and CINC-3 respectively. The complete amino acid sequences of purified CINC-2 alpha and CINC-3 were determined by analysis of the fragments isolated from proteinase V8-treated CINCs. The cDNA for CINC-2 beta was cloned by reverse transcription/PCR amplification using specific primers starting with total RNA extracted from lipopolysaccharide-stimulated rat macrophages. A comparison of the amino acid sequence encoded by the cDNA with the N-terminal amino acid sequence of purified CINC-2 beta revealed that mature CINC-2 beta is a 68-residue chemoattractant produced by cleavage of a 32-residue signal peptide. The difference in amino acid sequences between CINC-2 alpha and CINC-2 beta consisted of only three C-terminal residues. Rat GRO/CINC-2 alpha is a major chemokine, and the four purified chemokines have similar chemotactic activity, suggesting that they contribute to neutrophil infiltration into inflammatory sites in rats.

scholarly journals Multiple Natural Substitutions in Avian Influenza A Virus PB2 Facilitate Efficient Replication in Human Cells

2016 ◽  
Vol 90 (13) ◽  
pp. 5928-5938 ◽  
Author(s):  
Benjamin Mänz ◽  
Miranda de Graaf ◽  
Ramona Mögling ◽  
Mathilde Richard ◽  
Theo M. Bestebroer ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Avian Influenza ◽  
Mammalian Cells ◽  
Influenza A ◽  
Polymerase Activity ◽  
Host Adaptation ◽  
Influenza Viruses ◽  
Amino Acid Substitutions ◽  
Avian Influenza A Virus

ABSTRACTA strong restriction of the avian influenza A virus polymerase in mammalian cells generally limits viral host-range switching. Although substitutions like E627K in the PB2 polymerase subunit can facilitate polymerase activity to allow replication in mammals, many human H5N1 and H7N9 viruses lack this adaptive substitution. Here, several previously unknown, naturally occurring, adaptive substitutions in PB2 were identified by bioinformatics, and their enhancing activity was verified usingin vitroassays. Adaptive substitutions enhanced polymerase activity and virus replication in mammalian cells for avian H5N1 and H7N9 viruses but not for a partially human-adapted H5N1 virus. Adaptive substitutions toward basic amino acids were frequent and were mostly clustered in a putative RNA exit channel in a polymerase crystal structure. Phylogenetic analysis demonstrated divergent dependency of influenza viruses on adaptive substitutions. The novel adaptive substitutions found in this study increase basic understanding of influenza virus host adaptation and will help in surveillance efforts.IMPORTANCEInfluenza viruses from birds jump the species barrier into humans relatively frequently. Such influenza virus zoonoses may pose public health risks if the virus adapts to humans and becomes a pandemic threat. Relatively few amino acid substitutions—most notably in the receptor binding site of hemagglutinin and at positions 591 and 627 in the polymerase protein PB2—have been identified in pandemic influenza virus strains as determinants of host adaptation, to facilitate efficient virus replication and transmission in humans. Here, we show that substantial numbers of amino acid substitutions are functionally compensating for the lack of the above-mentioned mutations in PB2 and could facilitate influenza virus emergence in humans.

An Antimicrobial Peptide from the Australian Native Hardenbergia violacea Provides the First Functionally Characterised Member of a Subfamily of Plant Defensins

1997 ◽  
Vol 24 (5) ◽  
pp. 571 ◽  
Author(s):  
Stuart J. Harrison ◽  
John P. Marcus ◽  
Kenneth C. Goulter ◽  
Jodie L. Green ◽  
Donald J. Maclean ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Antimicrobial Peptide ◽  
Mammalian Cells ◽  
Amino Acid Sequences ◽  
Plant Defensin ◽  
Defensive Role ◽  
Plant Defensins

An antimicrobial peptide (HvAMP1) was isolated from seeds of the Australian native legume Hardenbergia violacea (Schneev.) Stearn. The peptide is 47 amino acid residues in length, contains 8 cysteines, and has a molecular weight of 5392 and a predicted pI of 10.41. HvAMP1 inhibited the growth of several plant pathogenic fungi at concentrations as low as 1 µM in vitro and produced distinct hyphal distortion and increased branching. This antimicrobial activity was greatly diminished in the presence of 1 mM CaCl2 and 50 mM KCl. The purified peptide at 40 µM did not inhibit three different a-amylase enzymes. Aeukaryotic cell-free translation system showed inhibition approaching 50% in the presence of ~100 µM of HvAMP1. The viability of plant and mammalian cells cultured in vitro was not adversely affected by concentrations of HvAMP1 as high as 40 mM. The amino acid sequence of HvAMP1 contained the consensus amino acids that define the plant defensin family of peptides. The HvAMP1 amino acid sequence showed 87% and 57% identity with the amino acid sequences deduced from cDNA sequences from defensins of Vigna unguiculata and Pisum sativum, respectively. Other plant defensin sequences showed less than 33% amino acid identity to the peptide. Therefore, HvAMP1 and the putative plant defensins of cowpea and pea define a distinct sequence subfamily of plant defensins which is at present limited to members of the Fabaceae. HvAMP1 is the first member of this subfamily to be purified and functionally characterised. The antimicrobial activity of HvAMP1 suggests a defensive role for this subfamily of peptides.

scholarly journals Genetic Characteristics of Avian Influenza Virus Isolated from Wild Birds in South Korea, 2019–2020

Viruses ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 381
Author(s):  
Eun-Jee Na ◽  
Young-Sik Kim ◽  
Sook-Young Lee ◽  
Yoon-Ji Kim ◽  
Jun-Soo Park ◽  
...  
Keyword(s):  
Avian Influenza ◽  
South Korea ◽  
Basic Amino Acid ◽  
Histopathological Change ◽  
Amino Acid Sequences ◽  
Wild Birds ◽  
Influenza Viruses ◽  
Economic Losses ◽  
Active Monitoring ◽  
Genetic Characteristics

Wild aquatic birds, a natural reservoir of avian influenza viruses (AIVs), transmit AIVs to poultry farms, causing huge economic losses. Therefore, the prevalence and genetic characteristics of AIVs isolated from wild birds in South Korea from October 2019 to March 2020 were investigated and analyzed. Fresh avian fecal samples (3256) were collected by active monitoring of 11 wild bird habitats. Twenty-eight AIVs were isolated. Seven HA and eight NA subtypes were identified. All AIV hosts were Anseriformes species. The HA cleavage site of 20 representative AIVs was encoded by non-multi-basic amino acid sequences. Phylogenetic analysis of the eight segment genes of the AIVs showed that most genes clustered within the Eurasian lineage. However, the HA gene of H10 viruses and NS gene of four viruses clustered within the American lineage, indicating intercontinental reassortment of AIVs. Representative viruses likely to infect mammals were selected and evaluated for pathogenicity in mice. JB21-58 (H5N3), JB42-93 (H9N2), and JB32-81 (H11N2) were isolated from the lungs, but JB31-69 (H11N9) was not isolated from the lungs until the end of the experiment at 14 dpi. None of infected mice showed clinical sign and histopathological change in the lung. In addition, viral antigens were not detected in lungs of all mice at 14 dpi. These data suggest that LPAIVs derived from wild birds are unlikely to be transmitted to mammals. However, because LPAIVs can reportedly infect mammals, including humans, continuous surveillance and monitoring of AIVs are necessary, despite their low pathogenicity.

scholarly journals Tools for the Recognition of Sorting Signals and the Prediction of Subcellular Localization of Proteins From Their Amino Acid Sequences

2020 ◽  
Vol 11 ◽  
Author(s):  
Kenichiro Imai ◽  
Kenta Nakai
Keyword(s):  
Amino Acid ◽  
Subcellular Localization ◽  
Amino Acid Sequences ◽  
Additional Information ◽  
Sorting Signals ◽  
Specific Alternative ◽  
Cell Type Specific ◽  
Future Direction ◽  
The Impact ◽  
New Algorithms

At the time of translation, nascent proteins are thought to be sorted into their final subcellular localization sites, based on the part of their amino acid sequences (i.e., sorting or targeting signals). Thus, it is interesting to computationally recognize these signals from the amino acid sequences of any given proteins and to predict their final subcellular localization with such information, supplemented with additional information (e.g., k-mer frequency). This field has a long history and many prediction tools have been released. Even in this era of proteomic atlas at the single-cell level, researchers continue to develop new algorithms, aiming at accessing the impact of disease-causing mutations/cell type-specific alternative splicing, for example. In this article, we overview the entire field and discuss its future direction.

scholarly journals H5N2 Avian Influenza Outbreak in Texas in 2004: the First Highly Pathogenic Strain in the United States in 20 Years?

2005 ◽  
Vol 79 (17) ◽  
pp. 11412-11421 ◽  
Author(s):  
Chang-Won Lee ◽  
David E. Swayne ◽  
Jose A. Linares ◽  
Dennis A. Senne ◽  
David L. Suarez
Keyword(s):  
United States ◽  
Amino Acid ◽  
Avian Influenza ◽  
Cleavage Site ◽  
Basic Amino Acid ◽  
The United States ◽  
Influenza Viruses ◽  
Single Point Mutation ◽  
Highly Pathogenic

ABSTRACT In early 2004, an H5N2 avian influenza virus (AIV) that met the molecular criteria for classification as a highly pathogenic AIV was isolated from chickens in the state of Texas in the United States. However, clinical manifestations in the affected flock were consistent with avian influenza caused by a low-pathogenicity AIV and the representative virus (A/chicken/Texas/298313/04 [TX/04]) was not virulent for experimentally inoculated chickens. The hemagglutinin (HA) gene of the TX/04 isolate was similar in sequence to A/chicken/Texas/167280-4/02 (TX/02), a low-pathogenicity AIV isolate recovered from chickens in Texas in 2002. However, the TX/04 isolate had one additional basic amino acid at the HA cleavage site, which could be attributed to a single point mutation. The TX/04 isolate was similar in sequence to TX/02 isolate in several internal genes (NP, M, and NS), but some genes (PA, PB1, and PB2) had sequence of a clearly different origin. The TX/04 isolate also had a stalk deletion in the NA gene, characteristic of a chicken-adapted AIV. By analyzing viruses constructed by in vitro mutagenesis followed by reverse genetics, we found that the pathogenicity of the TX/04 virus could be increased in vitro and in vivo by the insertion of an additional basic amino acid at the HA cleavage site and not by the loss of a glycosylation site near the cleavage site. Our study provides the genetic and biologic characteristics of the TX/04 isolate, which highlight the complexity of the polygenic nature of the virulence of influenza viruses.

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