Identification and evaluation in-vitro of conserved peptides with high affinity to MHC-I as potential protective epitopes for Newcastle disease virus vaccines.

BACKGROUND: Newcastle disease (ND) is a major threat to the poultry industry, leading to significant economic losses. The current ND vaccines, usually based on active or attenuated strains, are only partially effective and can cause adverse effects post-vaccination. Therefore, the development of safer and more efficient vaccines is necessary. Epitopes represent the antigenic portion of the pathogen and their identification and use for immunization could lead to safer and more effective vaccines. However, the prediction of protective epitopes for a pathogen is a major challenge, especially taking into account the immune system of the target species. RESULTS: In this study, we utilized an artificial intelligence algorithm to predict ND virus (NDV) peptides that exhibit high affinity to the chicken MHC-I complex. We selected the peptides that are conserved across different NDV genotypes and absent in the chicken proteome. From the filtered peptides, we synthesized the five peptides with the highest affinities for the L, HN, and F proteins of NDV. We evaluated these peptides in-vitro for their ability to elicit cell-mediated immunity, which was measured by the lymphocyte proliferation in spleen cells of chickens previously immunized with NDV. CONCLUSIONS: Our study identified five peptides with high affinity to MHC-I that have the potential to serve as protective epitopes and could be utilized for the development of multi-epitope NDV vaccines. This approach can provide a safer and more efficient method for NDV immunization.

Highly pathogenic avian influenza virus H5N1 clade 2.3.4.4b from Peru forms a monophyletic group with Chilean isolates in South America.

Highly pathogenic avian Influenza virus (HPAIV) has spread in an unprecedented extent globally in recent years. Despite the large reports of cases in Asia, Europe, and North America, little is known about its circulation in South America. Here, we describe the isolation, and whole genome characterization of HPAIV obtained from sampling 26 wild bird species in Peru, representing one of the largest studies in our region following the latest HPAIV introduction in South America. Out of 147 samples analyzed, 22 were positive for detection of avian influenza virus using a qRT-PCR-based assay. Following inoculation into embryonated chicken eggs, fourteen viral isolates were obtained from which nine isolates were selected for genome characterization, based on their host relevance. Our results identified the presence of HPAIV H5N1 subtype in a highly diverse wild bird species. Phylogenetic analysis revealed that these isolates correspond to the clade 2.3.4.4b, sharing a common ancestor with North American isolates and forming a monophyletic group along with isolates from Chile. Altogether, changes at the amino acid levels compared to their closest relatives indicates the virus is evolving locally, highlighting the need for constant genomic surveillance. This data evidence the chances for spillover events increases as the virus spreads into large populations of immunologically naïve avian species and adding conditions for cross species transmission.

Evaluation of commercial doses of a feed additive and silymarin on broiler performance with and without CCl4-induced liver damage.

Improving productive performance is a daily challenge in the poultry industry. Developing cost-effective additives and strategies that improve performance in antibiotic-free poultry production is critical to maintaining productivity and efficiency. This study evaluates the influence of a commercially available phytogenic feed additive (CA-PFA, that comprises silymarin, betaine and curcumin extracts as main ingredients) and silymarin on commercial broilers' productive performance and liver function with and without carbon tetrachloride (CCl4)-induced liver damage. The experiment was conducted in a completely randomized design, with six treatments, eight replicates, and eight birds per replicate in 18 one-day-old male broilers (Cobb Vantress 500) each; under a 3 × 2 factorial arrangement (3 diets x 2 levels of CCl4, 0 and 1 mL/kg body weight orally). The experimental treatments included 3 diets, commercially recommended doses of CA-PFA (500 mg/kg of feed; this dose provides 70 mg/kg of silymarin, besides the other active ingredients included in the formulation), silymarin (250 mg/kg of feed, containing 28% of active ingredient; this dose provides 70 mg/kg of silymarin as active ingredient) and an additive-free basal diet as a control. A standard commercial silymarin was used as a reference due to its well-known and extensively studied hepatoprotective properties that can mitigate the negative effects of CCl4 in the liver. The data were analyzed as a 2-way ANOVA, and the means showing significant (P ≤ 0.05) differences were then compared using the Post-Hoc Tukey HSD test. No interaction was detected between factors. Exposure to CCl4 had a noticeable detrimental effect on alertness, productive performance, and liver function of broilers without a significant increase in mortality. Including CA-PFA in the diet improved productive performance compared to the basal diet from day 21 to the end of the trial, on day 42. While no influence in feed intake was detected for any treatment, CA-PFA improved body weight gain (BWG) and feed conversion ratio (FCR) significantly (P < 0.05) from day 21 to the end of the trial in healthy and CCl4-exposed birds. The results show that CA-PFA supplementation improves performance parameters in broilers with and without CCl4-induced liver damage, when compared to a basal diet and the addition of a standard commercial silymarin product.

Genetic Analysis of Infectious Bronchitis Virus S1 Gene Reveals Novel Amino Acid Changes in the GI-16 Lineage in Peru.

Infectious bronchitis is a highly contagious viral disease that represents an economic threat for poultry despite the wide use of vaccination. To characterize the virus circulating in Peru, we analyzed 200 samples, including nasopharyngeal swabs and multiple tissues collected from animals suspected of being infected with infectious bronchitis virus (IBV) between January and August in 2015. All animals had at least one positive sample for IBV by RT-PCR. Out of these positive samples, eighteen (18) were selected for viral isolation and a partial S1 sequencing. Phylogenetic analysis showed that sixteen isolates clustered with members of GI-16 lineage, also known as Q1, with nucleotide homology ranging from 93% to 98%. The two remaining isolates grouped with members of the GI-1 lineage. Our study reveals circulation of GI-16 lineage during this period in poultry systems in Peru, along with GI-1 lineage (vaccine-derived). Moreover, those IBV GI-16 isolates showed unique nucleotide and amino acid changes compared to their closest relatives. Altogether, these findings reveal the circulation of GI-16 lineage while describing changes at key regions of the S protein that might be of relevance for vaccine evasion. These results highlight the importance of genetic surveillance for improving vaccination strategies against infectious bronchitis.

Intranasal vaccination of hamsters with a Newcastle disease virus vector expressing the S1 subunit protects animals against SARS-CoV-2 disease.

The coronavirus disease-19 (COVID-19) pandemic has already claimed millions of lives and remains one of the major catastrophes in the recorded history. While mitigation and control strategies provide short term solutions, vaccines play critical roles in long term control of the disease. Recent emergence of potentially vaccine-resistant and novel variants necessitated testing and deployment of novel technologies that are safe, effective, stable, easy to administer, and inexpensive to produce. Here we developed three recombinant Newcastle disease virus (rNDV) vectored vaccines and assessed their immunogenicity, safety, and protective efficacy against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in mice and hamsters. Intranasal administration of rNDV-based vaccine candidates elicited high levels of neutralizing antibodies. Importantly, the nasally administrated vaccine prevented lung damage, and significantly reduced viral load in the respiratory tract of vaccinated animal which was compounded by profound humoral immune responses. Taken together, the presented NDV-based vaccine candidates fully protected animals against SARS-CoV-2 challenge and warrants evaluation in a Phase I human clinical trial as a promising tool in the fight against COVID-19.

Avian influenza in wild birds, central coast of Peru.

To determine genotypes of avian influenza virus circulating among wild birds in South America, we collected and tested environmental fecal samples from birds along the coast of Peru, June 2006-December 2007. The 9 isolates recovered represented 4 low-pathogenicity avian influenza strains: subtypes H3N8, H4N5, H10N9, and H13N2.

Pathotyping and Phylogenetic Characterization of Newcastle Disease Viruses Isolated in Peru: Defining Two Novel Subgenotypes Within Genotype XII.

Infections of poultry with virulent strains of avian paramyxovirus 1 (APMV-1), also known as Newcastle disease viruses (NDVs), cause Newcastle disease (ND). This highly contagious disease affects poultry and many other species of birds worldwide. In countries where the disease is prevalent, constant monitoring and characterization of isolates causing outbreaks are necessary. In this study, we report the results of pathogenicity testing and phylogenetic analyses of seven NDVs isolated from several regions of Peru between 2004 and 2015. Six viruses had intracerebral pathogenicity indices (ICPIs) of between 1.75 and 1.88, corresponding to a velogenic pathotype. The remaining virus had an ICPI of 0.00, corresponding to a lentogenic pathotype. These results were consistent with amino acid sequences at the fusion protein (F) cleavage site. All velogenic isolates had the polybasic amino acid sequence 112RRQKR↓F117 at the F cleavage site. Phylogenetic analyses of complete F gene sequences showed that all isolates are classified in class II of APMV-1. The velogenic viruses are classified in genotype XII, while the lentogenic virus is classified in genotype II, closely related to the LaSota vaccine strain. Moreover, tree topology, bootstrap values, and genetic distances observed within genotype XII resulted in the identification of novel subgenotypes XIIa (in South America) and XIIb (in China) and possibly two clades within genotype XIIa. All velogenic Peruvian viruses belonged to subgenotype XIIa. Overall, our results confirm the presence of genotype XII in Peru and suggest that it is the prevalent genotype currently circulating in our country. The phylogenetic characterization of these isolates helps to characterize the evolution of NDV and may help with the development of vaccines specific to our regional necessities.

Complete Genome Sequence of Fowl Aviadenovirus Serotype 8b Isolated in South America.

We present here the complete genome sequence of fowl aviadenovirus E (FAdV-E) serotype 8b strain FV211-16, isolated from chickens with inclusion body hepatitis in Peru. Genome comparisons with other FAdV-E strains revealed identities of 84.9 to 97.1% and the presence of 9 and 2 unique amino acid mutations in hexon and fiber proteins, respectively.

The Genetic Diversity of Influenza A Viruses in Wild Birds in Peru.

Our understanding of the global ecology of avian influenza A viruses (AIVs) is impeded by historically low levels of viral surveillance in Latin America. Through sampling and whole-genome sequencing of 31 AIVs from wild birds in Peru, we identified 10 HA subtypes (H1-H4, H6-H7, H10-H13) and 8 NA subtypes (N1-N3, N5-N9). The majority of Peruvian AIVs were closely related to AIVs found in North America. However, unusual reassortants, including a H13 virus containing a PA segment related to extremely divergent Argentinian viruses, suggest that substantial AIV diversity circulates undetected throughout South America.

Characterization and Sequencing of a Genotype XII Newcastle Disease Virus Isolated from a Peacock (Pavo cristatus) in Peru.

Here, we report the first complete sequence and biological characterization of a Newcastle disease virus (NDV) isolated from a peacock in South America (NDV/peacock/Peru/2011). This isolate, classified as genotype XII in class II, highlights the need for increased surveillance of noncommercial avian species.

Genome sequence and comparative analysis of Avibacterium paragallinarum.

BACKGROUND: Avibacterium paragallinarum, the causative agent of infectious coryza, is a highly contagious respiratory acute disease of poultry, which affects commercial chickens, laying hens and broilers worldwide. METHODOLOGY: In this study, we performed the whole genome sequencing, assembly and annotation of a Peruvian isolate of A. paragallinarum. Genome was sequenced in a 454 GS FLX Titanium system. De novo assembly was performed and annotation was completed with GS De Novo Assembler 2.6 using the H. influenzae str. F3031 gene model. Manual curation of the genome was performed with Artemis. Putative function of genes was predicted with Blast2GO. Virulence factors were identified by comparison with the Virulence Factor Database. RESULTS: The genome obtained has a length of 2.47 Mb with 40.66% of GC content. Seventy five large contigs (>500 nt) were obtained, which comprised 1,204 predicted genes. All the contigs are available in Genbank [GenBank: PRJNA64665]. A total of 103 virulence factors, reported in the Virulence Factor Database, were found in A. paragallinarum. Forty four of them are present in 7 species of Haemophilus, which are related with pathogenesis, virulence and host immune system evasion. A tetracycline-resistance associated transposon (Tn10), was found in A. paragallinarum, possibly acting as a defense mechanism. DISCUSSION AND CONCLUSION: The availability of A. paragallinarum genome represents an important source of information for the development of diagnostic tests, genotyping, and novel antigens for potential vaccines against infectious coryza. Identification of virulence factors contributes to better understanding the pathogenesis, and planning efforts for prevention and control of the disease.

Isolation of low-pathogenic H7N3 avian influenza from wild birds in Peru.

Wild birds serve as natural reservoirs and sometimes harbor low-pathogenic avian influenza viruses. However, mutation of the virus can result in highly pathogenic strains, often more common among H5 and H7 genotypes. We report the isolation of a low-pathogenic H7N3 avian influenza in a Peruvian wetland.