The Spanish influenza pandemic of 1918 to 1919 swept the globe and resulted in the deaths of at least 20 million people. The basis of the pulmonary damage and high lethality caused by the 1918 H1N1 influenza virus remains largely unknown. Recombinant influenza viruses bearing the 1918 influenza virus hemagglutinin (HA) and neuraminidase (NA) glycoproteins were rescued in the genetic background of the human A/Texas/36/91 (H1N1) (1918 HA/NA:Tx/91) virus. Pathogenesis experiments revealed that the 1918 HA/NA:Tx/91 virus was lethal for BALB/c mice without the prior adaptation that is usually required for human influenza A H1N1 viruses. The increased mortality of 1918 HA/NA:Tx/91-infected mice was accompanied by (i) increased (>200-fold) viral replication, (ii) greater influx of neutrophils into the lung, (iii) increased numbers of alveolar macrophages (AMs), and (iv) increased protein expression of cytokines and chemokines in lung tissues compared with the levels seen for control Tx/91 virus-infected mice. Because pathological changes in AMs and neutrophil migration correlated with lung inflammation, we assessed the role of these cells in the pathogenesis associated with 1918 HA/NA:Tx/91 virus infection. Neutrophil and/or AM depletion initiated 3 or 5 days after infection did not have a significant effect on the disease outcome following a lethal 1918 HA/NA:Tx/91 virus infection. By contrast, depletion of these cells before a sublethal infection with 1918 HA/NA:Tx/91 virus resulted in uncontrolled virus growth and mortality in mice. In addition, neutrophil and/or AM depletion was associated with decreased expression of cytokines and chemokines. These results indicate that a human influenza H1N1 virus possessing the 1918 HA and NA glycoproteins can induce severe lung inflammation consisting of AMs and neutrophils, which play a role in controlling the replication and spread of 1918 HA/NA:Tx/91 virus after intranasal infection of mice.

Highly pathogenic avian influenza viruses (HPAIVs) of the A/goose/Guangdong/1/1996 lineage H5 clade 2.3.4.4b continue to have a devastating effect on domestic and wild birds. Full genome sequence analyses using 1369 H5N1 HPAIVs detected in the United States (U.S.) in wild birds, commercial poultry, and backyard flocks from December 2021 to April 2022, showed three phylogenetically distinct H5N1 virus introductions in the U.S. by wild birds. Unreassorted Eurasian genotypes A1 and A2 entered the Northeast Atlantic states, whereas a genetically distinct A3 genotype was detected in Alaska. The A1 genotype spread westward via wild bird migration and reassorted with North American wild bird avian influenza viruses. Reassortments of up to five internal genes generated a total of 21 distinct clusters; of these, six genotypes represented 92% of the HPAIVs examined. By phylodynamic analyses, most detections in domestic birds were shown to be point-source transmissions from wild birds, with limited farm-to-farm spread.

Avian influenza (AI) viruses vary in their ability to produce infection, disease and death in different bird species. Based on the pathobiological effect in chickens, AI viruses (AIV) are categorised as low pathogenic (LPAIV) or highly pathogenic (HPAIV). Typically, LPAIV cause asymptomatic infections in wild aquatic birds, but when introduced into domesticated poultry, infections may be asymptomatic or produce clinical signs and lesions reflecting pathophysiological damage to the respiratory, digestive and reproductive systems. The HPAIV have primarily been seen in gallinaceous poultry, producing high morbidity and mortality, and systemic disease with necrosis and inflammation in multiple visceral organs, nervous and cardiovascular systems, and the integument. Although HPAIV have rarely infected domestic waterfowl or wild birds, the Eurasian-African H5N1 HPAIV have evolved over the past decade with the unique capacity to infect and cause disease in domestic ducks and wild birds, producing a range of syndromes including asymptomatic respiratory and digestive tract infections; systemic disease limited to two or three critical organs, usually the brain, heart and pancreas; and severe disseminated infection and death as seen in gallinaceous poultry. Although experimental studies using intranasal inoculation have produced infection in a variety of wild bird species, the inefficiency of contact transmission in some of them, for example, passerines and Columbiformes, suggests they are unlikely to be a reservoir for the viruses, while others such as some wild Anseriformes, can be severely affected and could serve as a dissemination host over intermediate distances.

Current technologies with next generation sequencing have revolutionized metagenomics analysis of clinical samples. To achieve the non-selective amplification and recovery of low abundance genetic sequences, a simplified Sequence-Independent, Single-Primer Amplification (SISPA) technique in combination with MiSeq platform was applied to target negative- and positive-sense single-stranded RNA viral sequences. This method allowed successful sequence assembly of full or near full length avian influenza virus (AIV), infectious bronchitis virus (IBV), and Newcastle disease virus (NDV) viral genome. Moreover, SISPA analysis applied to unknown clinical cases of mixed viral infections produced genome assemblies comprising 98% NDV and 99% of IBV genomes. Complete or near complete virus genome sequence was obtained with titers at or above 104.5 EID50/ml (50% embryo infectious dose), and virus identification could be detected with titers at or above 103 EID50/ml. Taken together, these studies demonstrate a simple template enrichment protocol for rapid detection and accurate characterization of avian RNA viruses.

Low pathogenicity avian influenza virus (LPAIV) and lentogenic Newcastle disease virus (lNDV) are commonly reported causes of respiratory disease in poultry worldwide with similar clinical and pathobiological presentation. Co-infections do occur but are not easily detected, and the impact of co-infections on pathobiology is unknown. In this study chickens and turkeys were infected with a lNDV vaccine strain (LaSota) and a H7N2 LPAIV (A/turkey/VA/SEP-67/2002) simultaneously or sequentially three days apart. No clinical signs were observed in chickens co-infected with the lNDV and LPAIV or in chickens infected with the viruses individually. However, the pattern of virus shed was different with co-infected chickens, which excreted lower titers of lNDV and LPAIV at 2 and 3 days post inoculation (dpi) and higher titers at subsequent time points. All turkeys inoculated with the LPAIV, whether or not they were exposed to lNDV, presented mild clinical signs. Co-infection effects were more pronounced in turkeys than in chickens with reduction in the number of birds shedding virus and in virus titers, especially when LPAIV was followed by lNDV. In conclusion, co-infection of chickens or turkeys with lNDV and LPAIV affected the replication dynamics of these viruses but did not affect clinical signs. The effect on virus replication was different depending on the species and on the time of infection. These results suggest that infection with a heterologous virus may result in temporary competition for cell receptors or competent cells for replication, most likely interferon-mediated, which decreases with time.

Intestinal samples collected from 43 commercial broiler and 33 commercial turkey flocks from all regions of the United States during 2005 and 2006 were examined for the presence of astrovirus, rotavirus, reovirus, and coronavirus by reverse transcription-polymerase chain reaction (PCR), and for the presence of groups 1 and 2 adenovirus by PCR. Phylogenetic analysis was performed to further characterize the viruses and to evaluate species association and geographic patterns. Astroviruses were identified in samples from 86% of the chicken flocks and from 100% of the turkey flocks. Both chicken astrovirus and avian nephritis virus (ANV) were identified in chicken samples, and often both viruses were detected in the same flock. Turkey astrovirus type-2 and turkey astrovirus type-1 were found in 100% and 15.4% of the turkey flocks, respectively. In addition, 12.5% of turkey flocks were positive for ANV. Rotaviruses were present in 46.5% of the chicken flocks tested and in 69.7% of the turkey flocks tested. Based upon the rotavirus NSP4 gene sequence, the chicken and turkey origin rotaviruses assorted in a species-specific manner. The turkey origin rotaviruses also assorted based upon geographical location. Reoviruses were identified in 62.8% and 45.5% of chicken and turkey flocks, respectively. Based on the reovirus S4 gene segment, the chicken and turkey origin viruses assorted separately, and they were distinct from all previously reported avian reoviruses. Coronaviruses were detected in the intestinal contents of chickens, but not turkeys. Adenoviruses were not detected in any chicken or turkeys flocks. Of the 76 total chicken and turkey flocks tested, only three chicken flocks were negative for all viruses targeted by this study. Most flocks were positive for two or more of the viruses, and overall no clear pattern of virus geographic distribution was evident. This study provides updated enteric virus prevalence data for the United States using molecular methods, and it reinforces that enteric viruses are widespread in poultry throughout the United States, although the clinical importance of most of these viruses remains unclear.