Genetic characterization of a new candidate hemagglutinin subtype of influenza A viruses

Avian influenza viruses (AIV) have been classified on the basis of 16 subtypes of hemagglutinin (HA) and 9 subtypes of neuraminidase. Here we describe genomic evidence for a new candidate HA subtype, nominally H19, with a large genetic distance to all previously described AIV subtypes, derived from a cloacal swab sample of a Common Pochard (Aythya ferina) in Kazakhstan, in 2008. Avian influenza monitoring in wild birds especially in migratory hotspots such as central Asia is an important approach to gain information about the circulation of known and novel influenza viruses. Genetically, the novel HA coding sequence exhibits only 68.2% nucleotide and 68.5% amino acid identity with its nearest relation in the H9 (N2) subtype. The new HA sequence should be considered in current genomic diagnostic AI assays to facilitate its detection and eventual isolation enabling further study and antigenic classification

Exposure of wild Caspian seals (Pusa caspica) to parasites, bacterial and viral pathogens, evaluated via molecular and serological assays

Disease surveillance of marine mammal populations is essential to understand the causes of strandings, identify potential threats to animal health, and to support development of conservation strategies. Here we report the first large multi-pathogen screening of prevalence for viruses, bacteria and parasites in a sample of 177 live, healthy, wild Caspian seals (Pusa caspica), captured and released during satellite telemetry studies 2007-2017. Employing molecular and serological assays we assess prevalence of pathogens known to be of significance for marine mammal health worldwide, and evaluate the results in relation to Caspian seal health and conservation. RT-PCR, and PCR assays find evidence for infection by Canine Distemper Virus (CDV), Phocine herpes virus, phocine adenovirus and Influenza A at prevalences of 5%, 6.4%, 21.7%, and 4% respectively. The genomes of CDV isolates collected in 2008 showed 99.59% identity with the 2000 Caspian seal CDV epizootic strain. A partial coding sequence for the Us2 gene from the Caspian seal herpes virus was identical to PhHV-1 isolate PB84, previously reported from a harbor seal (Phoca vitulina), while amplicon sequences for the adenovirus polymerase gene indicated a novel strain. ELISA assays detected exposure to Influenza A (55% of tested samples), adenovirus (25%), coronavirus (6%), CDV (8%), herpes virus (94%), Toxoplasma gondii (2.6%) and heartworm (1%). Hemagglutination inhibition (HI) tests detected exposure to Influenza B at a prevalence of 20%, and Leptospira microscopic agglutination tests detected suspected exposure to Leptospira serovars in 9% of tested samples. Overall, the risks, profile and prevalence of pathogens in Caspian seals appear comparable to other wild phocid seal populations. Our results suggest Caspian seals have exposure pathways to pathogens with epizootic potential or ability to cause significant morbidity, and that disease impacts could reduce the resilience of the population to other conservation threats. Caspian seals are listed as Endangered by the International Union for Conservation of Nature (IUCN), and we recommend that resources are invested to support further surveillance programs and to understand how anthropogenic pressures may influence future disease risks. A translated version of this abstract is available in Russian and Kazakh in the Supplementary Material (Presentation 1 and Presentation 2

Novel avian paramyxovirus isolated from gulls in Caspian seashore in Kazakhstan.

Three isolates APMV/gull/Kazakhstan/5976/2014, APMV/gull/Kazakhstan/ 5977/2014 and APMV/gull/Kazakhstan/5979/2014, were obtained from independent samples during annual surveillance for avian influenza and paramyxoviruses in wild birds from the Caspian Sea coast in Western Kazakhstan, and were initially identified as putative paramyxoviruses on the basis of electron microscopy. Hemagglutination Inhibition Assays with antisera to nine known APMV serotypes (APMV1-9) indicated no relation to any of them. Next generation sequencing of whole genome sequences indicated the three isolates were genetically identical, and had a nucleotide structure typical for all APMVs, consisting of six genes 3'-NP-P-M-F-HN-L-5'. Phylogenetic analyses, and assessment of amino acid identities, suggested the most closely related lineages to be APMV-2, 8, 10 and 15, but the novel isolate had less than 64% identity to them and all other known avian paramyxoviruses. This value was above levels considered to generally define other APMV serotypes. Estimates of the evolutionary divergence of the nucleotide sequences of the genomes of APMVs have shown that novel Kazakhstan APMV strain was closest to APMV-2, APMV-8, APMV-10 and APMV-15, with calculated distance values of 2.057, 2.058, 2.026 and 2.286 respectively, which is above values considered to differentiate other serotypes (observed minimum was 1.108 between APMV-1 and recently isolated APMV/UPO216/Korea). Together, the data suggest that isolate APMV/gull/Kazakhstan/5976/2014 and other two should be considered as the first representative of a novel APMV-20 group, and is the first time that avian paramyxoviruses have been found infecting members of the gull family, extending the known taxonomic host range

Table_1_Has avian influenza virus H9 originated from a bat source?.docx

Influenza A viruses are important pathogens that can cause diseases with high mortality in humans, animals, and birds; and wild birds are considered the primary reservoir of all subtypes in nature. After discovering the H9 influenza A viruses in bats, questions arose about their potential to serve as an additional natural reservoir and about the priority of the viral origin: Did the virus initially circulate in bats and then transmit to birds or vice versa? Influenza A viruses of the H9 subtype are of particular interest because fatal infections of humans caused by H5, H7, and H10 influenza viruses contained RNA segments from H9 viruses. Recently, a novel subtype of influenza A virus (H19) was reported and it was closely related to the H9 bat influenza A virus by its hemagglutinin structure. The genome of novel H19 has revealed a mixed characteristic genomic signature of both avian and bat influenza viruses. The time to most recent common ancestor (TMRCA) estimates have shown that the divergence time between the bat and avian H9-similar influenza virus occurred approximately at the end of the XVIII century. This article discusses the evolution and possible origin of influenza viruses of the H9 subtype isolated from bats and birds. The obtained data, along with the known data, suggest that the primary reservoir of the H9 influenza virus is wild birds, from which the virus was transmitted to bats. We hypothesize that the novel H19 could be a descendant of an intermediate influenza virus that was in the transition stage of spillover from avian to bat hosts.</p

Table_2_Has avian influenza virus H9 originated from a bat source?.docx

Influenza A viruses are important pathogens that can cause diseases with high mortality in humans, animals, and birds; and wild birds are considered the primary reservoir of all subtypes in nature. After discovering the H9 influenza A viruses in bats, questions arose about their potential to serve as an additional natural reservoir and about the priority of the viral origin: Did the virus initially circulate in bats and then transmit to birds or vice versa? Influenza A viruses of the H9 subtype are of particular interest because fatal infections of humans caused by H5, H7, and H10 influenza viruses contained RNA segments from H9 viruses. Recently, a novel subtype of influenza A virus (H19) was reported and it was closely related to the H9 bat influenza A virus by its hemagglutinin structure. The genome of novel H19 has revealed a mixed characteristic genomic signature of both avian and bat influenza viruses. The time to most recent common ancestor (TMRCA) estimates have shown that the divergence time between the bat and avian H9-similar influenza virus occurred approximately at the end of the XVIII century. This article discusses the evolution and possible origin of influenza viruses of the H9 subtype isolated from bats and birds. The obtained data, along with the known data, suggest that the primary reservoir of the H9 influenza virus is wild birds, from which the virus was transmitted to bats. We hypothesize that the novel H19 could be a descendant of an intermediate influenza virus that was in the transition stage of spillover from avian to bat hosts.</p

Has avian influenza virus H9 originated from a bat source?

Influenza A viruses are important pathogens that can cause diseases with high mortality in humans, animals, and birds; and wild birds are considered the primary reservoir of all subtypes in nature. After discovering the H9 influenza A viruses in bats, questions arose about their potential to serve as an additional natural reservoir and about the priority of the viral origin: Did the virus initially circulate in bats and then transmit to birds or vice versa? Influenza A viruses of the H9 subtype are of particular interest because fatal infections of humans caused by H5, H7, and H10 influenza viruses contained RNA segments from H9 viruses. Recently, a novel subtype of influenza A virus (H19) was reported and it was closely related to the H9 bat influenza A virus by its hemagglutinin structure. The genome of novel H19 has revealed a mixed characteristic genomic signature of both avian and bat influenza viruses. The time to most recent common ancestor (TMRCA) estimates have shown that the divergence time between the bat and avian H9-similar influenza virus occurred approximately at the end of the XVIII century. This article discusses the evolution and possible origin of influenza viruses of the H9 subtype isolated from bats and birds. The obtained data, along with the known data, suggest that the primary reservoir of the H9 influenza virus is wild birds, from which the virus was transmitted to bats. We hypothesize that the novel H19 could be a descendant of an intermediate influenza virus that was in the transition stage of spillover from avian to bat hosts

Retrospective Analysis of the Equine Influenza Virus A/Equine/Kirgizia/26/1974 (H7N7) Isolated in Central Asia

A retrospective phylogenetic characterization of the hemagglutinin, neuraminidase and nucleoprotein genes of equine influenza virus A/equine/Kirgizia/26/1974 (H7N7) which caused an outbreak in Kirgizia (a former Soviet Union republic, now Kyrgyzstan) in 1977 was conducted. It was defined that it was closely related to the strain London/1973 isolated in Europe and it shared a maximum nucleotide sequence identity at 99% with it. This Central Asian equine influenza virus isolate did not have any specific genetic signatures and can be considered as an epizootic strain of 1974 that spread in Europe. The absence of antibodies to this subtype EI virus (EIV) in recent research confirms its disappearance as of the 1990s when the antibodies were last found in unvaccinated horses

Assessment of Caspian Seal By-Catch in an Illegal Fishery Using an Interview-Based Approach.

The Caspian seal (Pusa caspica) has declined by more than 90% since 1900 and is listed as endangered by IUCN. We made the first quantitative assessment of Caspian seal by-catch mortality in fisheries in the north Caspian Sea by conducting semi-structured interviews in fishing communities along the coasts of Russia (Kalmykia, Dagestan), Kazakhstan and Turkmenistan. We recorded a documented minimum by-catch of 1,215 seals in the survey sample, for the 2008-2009 fishing season, 93% of which occurred in illegal sturgeon fisheries. Due to the illegal nature of the fishery, accurately quantifying total fishing effort is problematic and the survey sample could reflect less than 10% of poaching activity in the north Caspian Sea. Therefore total annual by-catch may be significantly greater than the minimum documented by the survey. The presence of high by-catch rates was supported independently by evidence of net entanglement from seal carcasses, during a mass stranding on the Kazakh coast in May 2009, where 30 of 312 carcasses were entangled in large mesh sturgeon net remnants. The documented minimum by-catch may account for 5 to 19% of annual pup production. Sturgeon poaching therefore not only represents a serious threat to Caspian sturgeon populations, but may also be having broader impacts on the Caspian Sea ecosystem by contributing to a decline in one of the ecosystem's key predators. This study demonstrates the utility of interview-based approaches in providing rapid assessments of by-catch in illegal small-scale fisheries, which are not amenable to study by other methods

Serological Screening for Middle East Respiratory Syndrome Coronavirus and Hepatitis E Virus in Camels in Kazakhstan

After the recent Middle East Respiratory Syndrome coronavirus (MERS&ndash;CoV) pandemic in 2013, more attention has been paid to the camel as an important source of zoonotic viral infections. Almost simultaneously, in 2013, new genotypes 7 and 8 of the hepatitis E virus (HEV) were discovered in dromedary and Bactrian camels, respectively. HEV 7 was further shown to be associated with chronic viral hepatitis in a transplant recipient. In this study, serological screening for antibodies to MERS-CoV and hepatitis E virus was carried out on large camel farms in the south and west of Kazakhstan. 6.42% of the tested camels were found to be positive for antibodies to the hepatitis E virus, which indicates its circulation in local camel population. For the first time, antibodies to the hepatitis E virus were found in Bactrians, which have been little studied to date. Antibodies to MERS-CoV were not found in the camel sera