Lymphomas in seabirds: case reports in a black skimmer (Rynchops niger) and a brown booby (Sula leucogaster).

A black skimmer (Rynchops niger) and a brown booby (Sula leucogaster) were rescued and gross, histopathological, immunohistochemical and polymerase chain reaction evaluations were conducted to investigate the cause of death. There were neoplastic infiltrations of CD3+ PAX5- lymphocytes in the black skimmer and CD3- PAX5+ lymphocytes in the brown booby. Molecular assays for viral agents were negative in both cases. This is the first report of disseminated lymphoma as the cause of stranding and death in these species in Brazil.

Molecular characterization of the meq oncogene of Marek's disease virus in vaccinated Brazilian poultry farms reveals selective pressure on prevalent strains.

Marek's disease virus (MDV) has become an increasingly virulent pathogen in the poultry industry despite vaccination efforts to control it. Brazil has experienced a significant rise of Marek's disease (MD) outbreaks in recent years. Our study aimed to analyze the complete meq gene sequences to understand the molecular epidemiological basis of MD outbreaks in Brazilian vaccinated layer farms. We detected a high incidence rate of visceral MD (67.74%) and multiple circulating MDV strains. The most prevalent and geographically widespread genotype presented several clinical and molecular characteristics of a highly virulent strain and evolving under positive selective pressure. Phylogenetic and phylogeographic analysis revealed a closer relationship with strains from the USA and Japan. This study sheds light on the circulation of MDV strains capable of infecting vaccinated birds. We emphasize the urgency of adopting preventive measures to manage MDV outbreaks threatening the poultry farming industry.

Comparative Epidemiological Assessment of Monkeypox Infections on a Global and Continental Scale Using Logistic and Gompertz Mathematical Models.

The monkeypox virus (MPXV) has caused an unusual epidemiological scenario-an epidemic within a pandemic (COVID-19). Despite the inherent evolutionary and adaptive capacity of poxviruses, one of the potential triggers for the emergence of this epidemic was the change in the status of orthopoxvirus vaccination and eradication programs. This epidemic outbreak of HMPX spread worldwide, with a notable frequency in Europe, North America, and South America. Due to these particularities, the objective of the present study was to assess and compare cases of HMPX in these geographical regions through logistic and Gompertz mathematical modeling over one year since its inception. We estimated the highest contagion rates (people per day) of 690, 230, 278, and 206 for the world, Europe, North America, and South America, respectively, in the logistic model. The equivalent values for the Gompertz model were 696, 268, 308, and 202 for the highest contagion rates. The Kruskal-Wallis Test indicated different means among the geographical regions affected by HMPX regarding case velocity, and the Wilcoxon pairwise test indicated the absence of significant differences between the case velocity means between Europe and South America. The coefficient of determination (R2) values in the logistic model varied from 0.8720 to 0.9023, and in the Gompertz model, they ranged from 0.9881 to 0.9988, indicating a better fit to the actual data when using the Gompertz model. The estimated basic reproduction numbers (R0) were more consistent in the logistic model, varying from 1.71 to 1.94 in the graphical method and from 1.75 to 1.95 in the analytical method. The comparative assessment of these mathematical modeling approaches permitted the establishment of the Gompertz model as the better-fitting model for the data and the logistic model for the R0. However, both models successfully represented the actual HMPX case data. The present study estimated relevant epidemiological data to understand better the geographic similarities and differences in the dynamics of HMPX.

Mathematical Modeling of COVID-19 Cases and Deaths and the Impact of Vaccinations during Three Years of the Pandemic in Peru.

The COVID-19 pandemic has caused widespread infections, deaths, and substantial economic losses. Vaccine development efforts have led to authorized candidates reducing hospitalizations and mortality, although variant emergence remains a concern. Peru faced a significant impact due to healthcare deficiencies. This study employed logistic regression to mathematically model COVID-19's dynamics in Peru over three years and assessed the correlations between cases, deaths, and people vaccinated. We estimated the critical time (tc) for cases (627 days), deaths (389 days), and people vaccinated (268 days), which led to the maximum speed values on those days. Negative correlations were identified between people vaccinated and cases (-0.40) and between people vaccinated and deaths (-0.75), suggesting reciprocal relationships between those pairs of variables. In addition, Granger causality tests determined that the vaccinated population dynamics can be used to forecast the behavior of deaths (p-value < 0.05), evidencing the impact of vaccinations against COVID-19. Also, the coefficient of determination (R2) indicated a robust representation of the real data. Using the Peruvian context as an example case, the logistic model's projections of cases, deaths, and vaccinations provide crucial insights into the pandemic, guiding public health tactics and reaffirming the essential role of vaccinations and resource distribution for an effective fight against COVID-19.

Genomic characterization of Staphylococcus aureus from Canastra Minas Artisanal Cheeses.

Canastra Minas Artisanal Cheese is produced in the Brazilian State of Minas Gerais using raw milk, rennet, and pingo, a natural endogenous starter culture (fermented whey) collected from the previous day's production. Due to the use of raw milk, the product can carry microorganisms that may cause foodborne diseases (FBD), including Staphylococcus aureus. Genomic characterization of S. aureus is an important tool to assess diversity, virulence, antimicrobial resistance, and the potential for causing food poisoning due to enterotoxin production. This study is aimed at exploring the genomic features of S. aureus strains isolated from Canastra Minas Artisanal Cheeses. Multilocus sequence typing (MLST) classified these strains as ST1, ST5, and a new profile ST7849 (assigned to the clonal complex CC97). These strains belonged to four spa types: t008, t127, t359, and t992. We identified antimicrobial resistance genes with phenotypic correlation against methicillin (MRSA) and tetracycline. Virulome analysis revealed genes associated with iron uptake, immune evasion, and potential capacity for adherence and biofilm formation. The toxigenic potential included cyto- and exotoxins genes, and all strains presented the genes that encode for Panton-Valentine toxin and hemolysin, and two strains encoded 4 and 8 Staphylococcal enterotoxin (SE) genes. The results revealed the pathogenic potential of the evaluated S. aureus strains circulating in the Canastra region, representing a potential risk to public health. This study also provides useful information to monitor and guide the application of control measures to the artisanal dairy food production chain.

Antigenic and molecular characterization of isolates of the Brazilian genotype BR-I (GI-11) of infectious bronchitis virus supports its recognition as BR-I serotype.

The antigenic and molecular characteristics of BR-I infectious bronchitis viruses (IBVs) isolated from Brazil are reported. IBVs isolated from commercial flocks with different clinical manifestations between 2003 and 2019 were submitted to antigenic and molecular characterization. The complete S1 glycoprotein gene of 11 field isolates was amplified and sequenced. The virus neutralization (VN) test showed 94.75% neutralization with a BR-I isolate and 30% or less against other worldwide reference strains. The nucleotide and amino acid sequence analyses revealed 84.3-100% and 83.5-100% identity among them, respectively. The identity values ranged from 57.1 to 82.6% for nucleotides and from 46.6-84.4% for amino acids compared with those of other genotypes. By phylogenetic tree analysis, the Brazilian isolates were branched into the BR-I genotype (lineage GI-11), which was differentiated from foreign reference strains. Selective pressure analyses of BR-I IBVs revealed evolution under purifying selection (negative pressure) for the complete S1 gene but four specific sites (87, 121, 279, and 542) under diversifying selection (positive pressure). Profiles of cleavage sites and potential N-glycosylation sites differed from those of other genotypes. The low molecular relationship among the Brazilian viruses and foreign serotypes was concordant with the VN test results. The low antigenic relatedness (ranging from 5.3-30% between Brazilian genotype BR-I and reference IBV serotypes of North America, Europe, and Asia) indicates that the BR-I genotype is a different serotype, referred to for the first time and hereafter as serotype BR-I. RESEARCH HIGHLIGHTSStrains of the BR-I genotype presented robust antigenic and molecular similarity.BR-I strains evolved under purifying selection mode (negative pressure).The BR-I genotype originated in Brazil and dispersed to other countries.BR-I genotype viruses can be referred to as the BR-I serotype.

Genomic Characterization and Genetic Profiles of Salmonella Gallinarum Strains Isolated from Layers with Fowl Typhoid in Colombia.

Salmonella Gallinarum (SG) is the causative agent of fowl typhoid (FT), a disease that is harmful to the poultry industry. Despite sanitation and prophylactic measures, this pathogen is associated with frequent disease outbreaks in developing countries, causing high morbidity and mortality. We characterized the complete genome sequence of Colombian SG strains and then performed a comparative genome analysis with other SG strains found in different regions worldwide. Eight field strains of SG plus a 9R-derived vaccine were subjected to whole-genome sequencing (WGS) and bioinformatics analysis, and the results were used for subsequent molecular typing; virulome, resistome, and mobilome characterization; and a comparative genome study. We identified 26 chromosome-located resistance genes that mostly encode efflux pumps, and point mutations were found in gyrase genes (gyrA and gyrB), with the gyrB mutation S464T frequently found in the Colombian strains. Moreover, we detected 135 virulence genes, mainly in 15 different Salmonella pathogenicity islands (SPIs). We generated an SPI profile for SG, including C63PI, CS54, ssaD, SPI-1, SPI-2, SPI-3, SPI-4, SPI-5, SPI-6, SPI-9, SPI-10, SPI-11, SPI-12, SPI-13, and SPI-14. Regarding mobile genetic elements, we found the plasmids Col(pHAD28) and IncFII(S) in most of the strains and 13 different prophage sequences, indicating a frequently obtained profile that included the complete phage Gifsy_2 and incomplete phage sequences resembling Escher_500465_2, Shigel_SfIV, Entero_mEp237, and Salmon_SJ46. This study presents, for the first time, the genomic content of Colombian SG strains and a profile of the genetic elements frequently found in SG, which can be further studied to clarify the pathogenicity and evolutionary characteristics of this serotype.

Complete genome sequence data of two Salmonella enterica subsp. enterica serovar Gallinarum: A 9R vaccine strain and a virulent Brazilian field strain.

Salmonella Gallinarum (SG) is a host-restricted enterobacteria and the causative agent of fowl typhoid in poultry. Here, we report the complete genomes of two strains belonging to this serotype. SA68 is a field strain isolated from the livers of dead hen carcasses of a commercial layer farm presenting high mortality located in São Paulo city, Brazil, in 1990. Strain 9R corresponds to a live attenuated SG commercial vaccine. DNA was extracted from pure cultures and subjected to whole genome sequencing (WGS) using the Ion Torrent PGM System. The assemblies reached lengths of 4,657,435 (SA68) and 4,657,471 (9R) base pairs. Complete genomes were deposited in GenBank under the accession numbers CP110192 (SA68) and CP110508 (9R). Both genomes were analyzed and compared in terms of molecular typing, antibiotic resistance genes, virulence genes, Salmonella pathogenic islands (SPIs), insertion sequences and prophages. The data obtained show many similarities in the genetic content, with the exception of the SPI-12 and CS54 pathogenic islands, which are exclusive to the field strain. The information generated will help to understand the virulence differences of field and vaccinal SG strains and can be used to perform evolutionary and epidemiologic studies.

Evolutionary Analysis of a Parrot Bornavirus 2 Detected in a Sulphur-Crested Cockatoo (Cacatua galerita) Suggests a South American Ancestor.

Parrot bornavirus (PaBV) is an RNA virus that causes Proventricular Dilatation Disease (PDD), neurological disorders, and death in Psittaciformes. Its diversity in South America is poorly known. We examined a Cacatua galerita presenting neuropathies, PDD, and oculopathies as the main signs. We detected PaBV through reverse transcription polymerase chain reaction (RT-PCR) and partial sequencing of the nucleoprotein (N) and matrix (M) genes. Maximum likelihood and Bayesian phylogenetic inferences classified it as PaBV-2. The nucleotide identity of the sequenced strain ranged from 88.3% to 90.3% against genotype PaBV-2 and from 80.2% to 84.4% against other genotypes. Selective pressure analysis detected signs of episodic diversifying selection in both the N and M genes. No recombination events were detected. Phylodynamic analysis estimated the time to the most recent common ancestor (TMRCA) as the year 1758 for genotype PaBV-2 and the year 1049 for the Orthobornavirus alphapsittaciforme species. Substitution rates were estimated at 2.73 × 10-4 and 4.08 × 10-4 substitutions per year per site for N and M, respectively. The analysis of population dynamics showed a progressive decline in the effective population size during the last century. Timescale phylogeographic analysis revealed a potential South American ancestor as the origin of genotypes 1, 2, and 8. These results contribute to our knowledge of the evolutionary origin, diversity, and dynamics of PaBVs in South America and the world. Additionally, it highlights the importance of further studies in captive Psittaciformes and the potential impact on endangered wild birds.

Molecular characterization of chicken astrovirus from high embryonic mortality, based on the capsid protein (ORF2) gene, in Brazil.

CAstV infections were found in farms and incubators with increased embryo mortality.Brazilian CAstV Biv strains were associated with white chick syndrome.Antigenic peptides were predicted on the surface of the capsid protein.

Detection of aves polyomavirus 1 (APyV) and beak and feather disease virus (BFDV) in exotic and native Brazilian Psittaciformes.

There are several viral diseases in captive birds. Aves polyomavirus 1 (APyV) and beak and feather disease virus (BFDV) are among the most important in Psittaciformes. The occurrence of these agents has been widely described in various parts of the world; however, little is known about these viruses in South America. APyV and BFDV can cause high morbidity with feather alterations and even mortality. Other variable symptoms could appear depending on the host's age and taxonomic group. The aim of this study was to detect APyV and BFDV in samples of captive exotic and native Psittaciformes in Brazil. Samples from 120 birds with clinical signs compatible with APyV and/or BFDV were examined. In total, 57 (47.5%) positive birds were found, of which 21 (17.5%) had APyV and 41 (34.17%) had BFDV. Five animals (4.17%) presented concurrent infection. Phylogenetic analysis showed a divergent APyV strain and a diversity of Brazilian BFDV strains. Our study shows that these viruses are present at a significant frequency in captive exotic and native Psittaciformes in Brazil. This study also highlights the need for constant epidemiologic surveillance to preserve bird biodiversity with a focus on endangered Psittaciformes species.

Outbreaks of Avipoxvirus Clade E in Vaccinated Broiler Breeders with Exacerbated Beak Injuries and Sex Differences in Severity.

Avipoxvirus affects chickens and wild birds, and it is characterized by lesions on the nonfeathered parts of the body (the cutaneous form), or necrotic lesions in the upper respiratory tract (the diphtheritic form). In poultry farming, avian pox is usually controlled by live attenuated vaccines. However, there have been many reports of outbreaks, even in flocks of vaccinated birds. In the present study, different outbreaks of the emerging clade E avipoxvirus were detected in commercial breeder flocks of chickens vaccinated against fowlpox virus in Southeast Brazil. Clinical manifestations of these outbreaks included a marked prevalence of moderate to severe progressive lesions in the beaks of affected birds, especially in roosters with increased mortality (up to 8.48%). Also, a reduced hatchability (up to 20.77% fewer hatching eggs) was observed in these flocks. Analysis of clinical samples through light and transmission electron microscopy revealed the presence of Bollinger bodies and poxvirus particles in epithelial cells and affecting chondrocytes. PCR, sequencing, and phylogenetic analysis of major core protein (P4b) and DNA polymerase (pol) genes identified this virus as clade E avipoxvirus. We also developed qPCR assays for open reading frames (ORFs) 49, 114, and 159 to detect and quantify this emergent virus. These results show the arrival and initial spread of this pathogen in the poultry industry, which was associated with harmful outbreaks and exacerbated clinical manifestations in vaccinated commercial breeder flocks. This study also highlights the relevance of permanent vigilance and the need to improve sanitary and vaccination programs.

Complete Genome Characterization of Reticuloendotheliosis Virus Detected in Chickens with Multiple Viral Coinfections.

Reticuloendotheliosis virus (REV) is a retroviral pathogen capable of infecting several avian hosts and is associated with immunosuppression, anemia, proventriculitis, neoplasia, and runting-stunting syndrome. Its genome contains the three major genes, gag, pol, and env, and two flanking long terminal repeat (LTR) regions. Complete genome sequences of REV are limited in terms of geographical origin. The aim of this study was to characterize the complete genome of REV detected in Brazilian chickens with multiple viral coinfections and analyze the polymorphisms in the deduced amino acids sequences corresponding to its encoded proteins. We tested the presence and completeness of REV as well as other viral pathogens in samples from Brazilian poultry farms by qPCR. The complete genomes of two REV strains were sequenced by overlapping fragments through the dideoxy method. Phylogenetic analysis, pairwise identity matrix, polymorphism identification and protein modeling were performed along the entire genome. We detected REV in 65% (26/40) of the tested samples. Concomitant viral infections were detected in 82.5% (33/40) of the samples and in 90% (9/10) of the farms. Multiple infections included up to seven viruses. Phylogenetic analysis classified both Brazilian strains into REV subtype 3, and the pairwise comparison indicated that strains from the USA and fowlpox virus (FWPV)-related strains were the most identical. The subdomain p18 in gag, the reverse transcriptase/ribonuclease H in pol, and the surface (SU) in the env protein were the most polymorphic in genomic comparisons. The relevant motifs for each protein were highly conserved, with fewer polymorphisms in the fusion peptide, immunosuppression domain, and disulfide bonds on the surface (SU) and transmembrane (TM) of env. This is the first study to include complete genomes of REV in Brazil and South America detected in farms with multiple viral coinfections. Our findings suggest an involvement of REV as an immunosuppressor and active agent in the emergence and progression of multiple infectious diseases. We also found a possible etiological relationship between Brazilian strains and the USA and FWPV recombinant strains. This information highlights the need for epidemiological vigilance regarding REV in association with another pathogens.

Identification of Novel Candidate Epitopes on SARS-CoV-2 Proteins for South America: A Review of HLA Frequencies by Country.

Coronavirus disease (COVID-19), caused by the virus SARS-CoV-2, is already responsible for more than 4.3 million confirmed cases and 295,000 deaths worldwide as of May 15, 2020. Ongoing efforts to control the pandemic include the development of peptide-based vaccines and diagnostic tests. In these approaches, HLA allelic diversity plays a crucial role. Despite its importance, current knowledge of HLA allele frequencies in South America is very limited. In this study, we have performed a literature review of datasets reporting HLA frequencies of South American populations, available in scientific literature and/or in the Allele Frequency Net Database. This allowed us to enrich the current scenario with more than 12.8 million data points. As a result, we are presenting updated HLA allelic frequencies based on country, including 91 alleles that were previously thought to have frequencies either under 5% or of an unknown value. Using alleles with an updated frequency of at least ≥5% in any South American country, we predicted epitopes in SARS-CoV-2 proteins using NetMHCpan (I and II) and MHC flurry. Then, the best predicted epitopes (class-I and -II) were selected based on their binding to South American alleles (Coverage Score). Class II predicted epitopes were also filtered based on their three-dimensional exposure. We obtained 14 class-I and four class-II candidate epitopes with experimental evidence (reported in the Immune Epitope Database and Analysis Resource), having good coverage scores for South America. Additionally, we are presenting 13 HLA-I and 30 HLA-II novel candidate epitopes without experimental evidence, including 16 class-II candidates in highly exposed conserved areas of the NTD and RBD regions of the Spike protein. These novel candidates have even better coverage scores for South America than those with experimental evidence. Finally, we show that recent similar studies presenting candidate epitopes also predicted some of our candidates but discarded them in the selection process, resulting in candidates with suboptimal coverage for South America. In conclusion, the candidate epitopes presented provide valuable information for the development of epitope-based strategies against SARS-CoV-2, such as peptide vaccines and diagnostic tests. Additionally, the updated HLA allelic frequencies provide a better representation of South America and may impact different immunogenetic studies.

An atypical clinicopathological manifestation of fowlpox virus associated with reticuloendotheliosis virus in commercial laying hen flocks in Brazil.

Fowlpox (FP) is a common epitheliotropic disease in chickens that is usually controlled by live attenuated vaccines. However, there have been some reports of outbreaks of FP in recent years, even in vaccinated flocks, presenting as atypical lesions and feathering abnormalities in chickens. These findings can be associated with fowlpox virus (FPV) with the reticuloendotheliosis virus (REV) integrated into its genome. In the present study, outbreaks of atypical FP were explored in vaccinated commercial laying hen flocks to determine the nature of the causative agent by histopathologic and molecular approaches. FPV and REV were detected and classified into subclade A1 of the genus Avipoxvirus and subtype 3 of REV (REV3), respectively. Additionally, heterogeneous populations of FPV with partial (containing only a remnant long terminal repeat-LTR) or total (all functional genes) integration of REV were identified by heterologous PCRs and detected considering reference integration sites. These results indicate the mechanism of chimeric genome FPV-REV associated with outbreaks and atypical clinicopathological manifestations in commercial laying hens for the first time in Brazil and in South America. In addition, this study demonstrates the emergence of REV integrated in the FPV genome in Brazilian chicken flocks.

Detection and Molecular Characterization of a Natural Coinfection of Marek's Disease Virus and Reticuloendotheliosis Virus in Brazilian Backyard Chicken Flock.

Marek's disease virus (MDV) and the reticuloendotheliosis virus (REV) are two of the primary oncogenic viruses that significantly affect chickens. In Brazil, there have been no previous published reports on the presence of field REV alone or in coinfection. This retrospective study analyzes samples from a case of lymphoproliferative lesions from a backyard chicken flock. MDV and REV were detected by PCR and classified as MDV1 and REV3, respectively, through sequencing and phylogenetic analysis based on the glycoprotein B (gB) genes for MDV and the polymerase (pol) and envelope (env) genes for REV. Real-time PCR reactions were performed for MDV to rule out the presence of the Rispens vaccine strain. This is the first report of the presence of REV in coinfection with a MDV clinical case in Brazil and the first molecular characterization of REV in South America. This study highlights the importance of molecular diagnosis for REV and MDV in poultry. In addition, this study highlights the distribution of these two viruses worldwide and the latent risk of them solely or in coinfection to this part of the world.

A seminested RT-PCR for molecular genotyping of the Brazilian BR-I Infectious Bronchitis Virus Strain (GI-11).

Infectious bronchitis (IB) is one of the avian diseases with the greatest impact on poultry farming worldwide. In Brazil, strain BR-I (GI-11) is the most prevalent in poultry flocks. The present study aimed to develop a seminested RT-PCR assay specific for the diagnosis of BR-I IBV in Brazilian samples, targeting subunit 1 of the S gene. The detection limit of this assay was 10 copies of the IBV genome. In this study, 62.24% of 572 organ pools from the 5 regions of Brazil tested positive in a 3'UTR screening, and 84.83% were typed as BR-I IBV. BR-I was detected in the respiratory, digestive and urogenital tracts in pooled samples from all Brazilian geographical regions and in all the breeding systems analyzed. Specificity and sensitivity tests as well as phylogenetic analysis successfully confirmed the expected clustering of the sequences detected by this assay with the BR-I (GI-11) group. The nested PCR described in this study represents a suitable and valuable tool in the diagnosis, epidemiology, monitoring and vaccination decisions of IBV.

Draft Genome Sequences of Four Salmonella enterica subsp. enterica Serovar Gallinarum Strains Isolated from Layer Breeder Flocks in an Outbreak of Fowl Typhoid in Colombia.

This report describes the genome sequences of four Salmonella enterica subsp. enterica serovar Gallinarum (Salmonella Gallinarum) strains isolated in Colombia in 2017 from layer breeders of different ages. The layer breeder flocks were presenting with an elevated mortality with lesions typical of fowl typhoid (FT). These draft genome sequences revealed a highly conserved genome of Salmonella Gallinarum strains circulating in Colombia.

Sensitive SYBR Green-Real Time PCR for the Detection and Quantitation of Avian Rotavirus A.

Avian rotavirus A (ARtV-A) is a virus that affects young birds, causing acute diarrhea and economic losses in the poultry industry worldwide. The techniques used for the diagnosis of ARtV-A include electron microscopy, isolation in cell culture, and serology, as well as molecular techniques, such as the reverse transcription-polymerase chain reaction (RT-PCR). The objective of this work was to standardize a real-time RT-polymerase chain reaction (RT-qPCR) using SYBR Green chemistry for the rapid detection and quantification of ARtV-A from bird tissues and materials fixed on FTA cards on the basis of the nucleotide sequence of segment 6 (S6), which codes for the structural VP6 protein of ARtV-A. The results show the efficient amplification of the proposed target, with a limit of detection (LoD) of one copy gene (CG) per microliter of cDNA and a limit of quantification (LoQ) of 10 CGs per microliter. The efficiency of the primers was determined to be 95.66% using a standard curve, with an R² value of 0.999 and a slope of -3.43. The specificity was determined using samples coinfected with ARtV-A, the chicken parvovirus, the chicken astrovirus, and the avian nephritis virus as positive controls and commercially available vaccines of the infectious bronchitis virus, infectious bursa disease virus, avian reovirus and healthy organs as negative controls. This technique, which lacks nonspecific PCR products and dimers, demonstrated greater sensitivity and specificity than conventional RT-PCR, and it reduced the analysis time by more than 50%.

Genetic characterisation and analysis of infectious bronchitis virus isolated from Brazilian flocks between 2010 and 2015.

1. Infectious bronchitis virus (IBV) variants in Brazil were isolated during 2010-2015 for epidemiological and molecular analysis to characterise the different variants and perform a bioinformatic analysis to compare with sequences of variants collected over the previous 40 years. 2. Of the 453 samples examined, 61.4% were positive for IBV and 75.9% of these were considered to have the BR-I genotype and were detected in birds of all ages distributed in all five Brazilian regions. 3. The ratio of non-synonymous substitutions per non-synonymous site (dN) to synonymous substitutions per synonymous site (dS), i.e. dN/dS, revealed a predominance of codons with non-synonymous substitutions in the first third of the S1 gene and a dN/dS ratio of 0.67. Additionally, prediction of N-glycosylation sites showed that most of the BR-I variants (from 2003 to early 2014) had an extra site at amino acid position 20, whereas the newest variants lacked this extra site. 4. These results suggest that Brazilian IBV variants probably underwent drastic mutations at various points between 1983 and 2003 and that the selection processes became silent after achieving a sufficiently effective antigenic structure for invasion and replication in their hosts. Brazilian IBV genotype BR-I is currently the predominant genotype circulating in Brazil and South America.