is a ubiquitous organism that colonizes a variety of animal hosts and has the ability to persist within the environment. As such, it is not surprising that animals frequently share strains and contribute to environmental ecology. It has been well documented that poultry meat can serve as a reservoir of avian pathogenic (APEC) with the potential to cause human disease. However, the impact of backyard poultry rearing on household and community APEC sharing is less clear. In this study, we examined 1348 isolates from children, dogs, and chickens in 222 households in peri-urban communities of Quito, Ecuador, sampled across five timepoints. Extensive overlap between isolates from all three host sources were identified using Clermont phylotyping and multilocus sequence typing. Human and dog isolates also had a high rate of carriage (37% and 49%, respectively) of genes indicative of APEC. Phylogenetic analyses of dominant sequence types (ST10, ST155, ST117, ST2847, ST162, ST38, and ST354) provided examples of highly related clones found between host sources and households, and spanning timepoints. Overall, this study illustrates the apparent extensive sharing of that occurs across peri-urban communities. The high rates of carriage of APEC by humans and dogs in this study contrasts with previous work examining the carriage of APEC in mammalian hosts and suggests that widespread rearing of, and frequent contact with, backyard chickens may influence the dissemination of APEC within households and communities.

We examined the replication and adaptation of avian paramyxoviruses serotype 1 (APMV-1) using four isolates, Mallard/US(OH)/04-411/2004, Northern pintail/US(OH)/87-486/1987, Mottled duck/US(TX)/TX01-130/2001, and Mallard/US(MN)/MN00-39/2000, and assessed their potential as vaccine candidates for chickens. The adaptability of each virus was examined by serial passages in embryonated chicken eggs (ECE) and in Vero cells. All APMVs successfully replicated in ECE. In contrast, two isolates passaged in Vero cells showed successful replication and two showed a continuous decline in viral load during passages. Whole-genome sequencing analysis identified 14 genomic positions with significant variation in mean allele frequency. Changes of the predominant virus population were characterized by shifts of amino acid (aa) frequency at eight positions. Notably, four of these changes were located in the hemagglutinin-neuraminidase (HN) protein, one in matrix (M) protein, and two in the L-protein sequences. Remarkably, although the percentage of alternative amino acids in viral populations passaged in ECE showed limited variation-for example, at aa position 127 of HN, the frequency varied from 7.4% to 19.8% and at HN aa position 192 it varied from 5.1% to 43.5%-the variation of the viral populations passaged in Vero cells was significantly higher at the same positions (e.g., the frequency of the alternative amino acids at HN aa positions 127 and 192 changed from 20.8% to 95.2% and 7.2% to 91.2%, respectively). Isolate 2 passaged in Vero cells displayed a marked variation in alternative aa frequencies, specifically at positions 127 within the HN and 100 within the M proteins. Isolate 3, while showing no alterations at the same HN positions, showed a considerable change in alternative aa frequency in the L protein at position 1875, a change occurring only in the Vero cell environment. One-day-old specific-pathogen-free chickens inoculated with isolates passaged in ECE elicited serum antibody responses similar to those elicited by the LaSota reference strain. In contrast, APMVs passaged in Vero cells showed limited replication in chickens and reduced induction of systemic antibodies. Interestingly, one virus passaged in ECE and another in Vero cells elicited immunoglobulin A levels in lacrimal fluid comparable to the LaSota strain. We concluded that the four wild-bird APMV isolates tested demonstrated successful adaptation to ECE, with one isolate eliciting overall immune responses comparable to the LaSota virus, supporting their potential as vaccine candidates.

Avian reoviruses (ARVs) within genetic cluster (GC) 2 represent a large, heterogenous group of currently circulating ARVs that have been isolated from clinical cases of tenosynovitis and malabsorption. In this study, the pathogenesis of an ARV GC 2 isolate was investigated via quantitative reverse transcriptase PCR (RT-qPCR), hybridization (ISH), and histopathology, following oral or footpad inoculation. RT-qPCR detected ARV within the digital flexor tendon, heart, lung, liver, spleen, kidney, duodenum, cecum, cloacal bursa, and thymus. The highest viral RNA load was observed within the intestinal tract between 36 and 72 hr postinoculation (hpi). ISH demonstrated ARV within villous enterocytes throughout the intestines, follicle-associated epithelial (FAE) cells of the bursa, and the synovial membrane of the tendon. Histopathology within the intestine consisted of rare syncytia with negligible inflammation, whereas marked inflammation was present within the synovial tissues. The identity of infected enterocytes as avian "M cells" or infected synovial lining cells as macrophage-like synoviocytes (MLS) could not be histologically determined. However, the susceptibility of these varied cell types to infection with an ARV GC 2 virus demonstrates simultaneous potential enteric and arthritic roles in the pathogenesis of these reoviruses.

Detection of antibody response to Newcastle disease virus (NDV) or other avian orthoavulaviruses 1 (AOAVs-1) can serve as a useful tool for monitoring infection and response to vaccination. Two commonly used serologic tests for AOAV-1 are the hemagglutinin inhibition (HI) test and ELISA. Enzyme-linked lectin assay (ELLA) is based on the ability of the neuraminidase (NA), which is also part of the hemagglutinin-neuraminidase (HN) protein of AOAV-1, to cleave sialic acid residues from a substrate coated on the solid surface, and the assay is performed similarly to ELISA. The same ELLA can be applied to detect and quantify the neuraminidase inhibition (NI) antibody present in the sample. In this study, we first evaluated ELLA for measuring NA activity of different AOAV-1 strains on fetuin substrate. In contrast to influenza A virus, which shows optimal NA activity in neutral pH, all AOAV-1 strains tested showed higher NA activity under lower pH and the preference of specific pH varied by the strain. We established assay parameters, including the pH and temperature conditions, for optimal NA activity of two AOAV-1 strains and conducted ELLA to evaluate NI antibody (NI-ELLA). NI-ELLA was highly specific with antigen dose of 95% effective concentrations (EC), and showed less than 10% background nonspecific NI reactivity with all AOAV-1-negative sera collected from different poultry species. NI-ELLA was conducted to determine the endpoint titer of sera collected from birds vaccinated with Newcastle disease vaccine. The NI antibody titer determined by NI-ELLA correlated well with the HI antibody titer against the same antigen used for both tests, while showing a detection limit more than 20 times higher. Our study shows great potential of NI-ELLA as a functional antibody assay like the HI test to evaluate vaccine-induced antibody response against challenge virus, with the added advantages of being more sensitive and having a higher-throughput screening capability than the HI test.

The global outbreak of clade 2.3.4.4b H5 highly pathogenic avian influenza (HPAI) has caused tremendous losses in poultry worldwide. Although turkeys are a smaller sector in poultry production compared to chickens, they tend to be affected more severely by HPAI because they can usually be infected with a lower dose of virus (i.e., they are more susceptible). Vaccines can be effective tools to help control HPAI, but data on vaccine efficacy and antibody response in turkeys are somewhat limited. Here we evaluated an in-house-produced, reverse-genetics-generated H5N9 inactivated vaccine that has a clade 2.3.4.4b H5 HA from A/turkey/Indiana/22-003707-003/2022 (TK/IN/22) modified to be low pathogenic and a North American wild bird lineage N9 in a PR8 "backbone" for its efficacy in commercial broad-breasted white turkeys by homologous challenge. Turkeys were divided into three vaccination groups, where each group was vaccinated once at 3, 7, or 9 wk of age. Turkeys were challenged at 10 wk of age with TK/IN/22 HPAI virus. There was 100% survival in all vaccinated groups and 0% survival in the sham immunized group. A significant decrease in viral shedding was observed in all vaccinated groups compared to the sham immunized turkeys. Also, the 9 wk vaccination group shed significantly higher quantities by the cloacal route at 7 days postchallenge (DPC) compared to the 3 wk vaccination group, and two turkeys in the 9 wk vaccination group had mild clinical signs 6-7 DPC. The neuraminidase inhibition-enzyme-linked lectin assay (NI-ELLA) was used to evaluate antibodies to the vaccine and was more sensitive than the hemagglutinin inhibition assay. Also, when tested as a potential assay to differentiate vaccinated and infected animals, 50% to 90% of vaccinated turkeys (depending on the age at vaccination) were positive by NI-ELLA at 7 DPC for antibodies to the challenge virus, and 100% were positive at 14 DPC.

From 2015 to 2024, inclusion body hepatitis (IBH) was diagnosed in a total of 346 chicken necropsy cases submitted to the California Animal Health and Food Safety (CAHFS) laboratory system. The majority of cases (68%; 237/346) originated from a single commercial broiler company with a high density of poultry premises within the California Central Valley region. Reported mortality varied between 0.5% and 4.8% per day, and morbidity ranged between 5% and 10%. Clinical signs observed in affected flocks included depression, reluctance to move, ruffled feathers, greenish diarrhea, and, occasionally, icterus of skin and adipose tissue. Typical histopathological lesions included extensive hepatic necrosis, multifocal pancreatic necrosis, gizzard erosions, and glomerulonephropathy. Intranuclear inclusions were identified, mainly within degenerating hepatocytes, followed by pancreatic acinar cells, glandular epithelium of proventriculus and gizzard, and enterocytes of the small intestines and renal endothelium. Diagnosis was based on microscopic changes characteristic of IBH and a positive conventional PCR targeting the hexon gene of fowl adenovirus (FAdV) on liver tissue pools. Sequence analysis was performed on 41 selected FAdV-positive samples from diagnostic cases. The majority of FAdV PCR-positive samples aligned with FAdV-8b (16/41). However, FAdV-7 (14/41), FAdV-4 (7/41), and FAdV-11 (4/41) were also detected from IBH cases. Disease management was successfully achieved by the administration of an autogenous vaccine to breeders, at 11 and 18 wk of age, to confer maternal antibody protection to the broiler progeny.

Vaccination with a recombinant Newcastle disease virus (NDV) LaSota (LS) strain expressing Arkansas (Ark) -type infectious bronchitis virus (IBV) spike ectodomain (Se) protein and granulocyte-macrophage colony-stimulating factor (GMCSF) (rLS/ArkSe.GMCSF, where rLS stands for recombinant LS) was evaluated in chickens of commercial origin with NDV and IBV maternally derived antibodies (MDAs). Chickens were vaccinated ocularly with rLS/ArkSe.GMCSF at either 2, 8, 15, or 30 days of age (DOA). Control chickens were vaccinated with the rLS virus (not expressing IBV SE or GMCSF) on the same days. Specific-pathogen-free (SPF) chickens also were vaccinated with either virus at 2 days old. The results showed detection of NDV RNA in lacrimal fluids of vaccinated chickens, indicating successful replication of the recombinant virus at periocular mucosal sites. IBV IgA in lacrimal fluids and serum IBV antibodies were determined by ELISA using recombinant IBV Ark S1-protein-coated plates. Vaccination at 2 DOA with rLS/ArkSe.GMCSF in chickens with MDAs elicited an IBV IgA response in lacrimal fluids. Chickens with MDAs vaccinated with rLS/ArkSe.GMCSF at 8 days old showed IgA levels in lacrimal fluids not differing significantly from levels achieved upon vaccination at 2 DOA. Vaccination at 30 days old did not result in increased IBV IgA levels in tear fluids of birds with MDAs compared to unvaccinated birds with MDAs. Vaccination with rLS/ArkSe.GMCSF of chickens with MDAs resulted in limited IBV and NDV serum antibody responses. We conclude that vaccination with rLS/ArkSe.GMCSF induces IBV IgA at periocular mucosae but limited serum antibody responses in chickens with NDV MDAs.

Avian reovirus (ARV) is considered an important pathogen affecting poultry and recently has gained more attention because of its genetic diversity, prevalence, and several disease conditions associated with its infection. In Egypt, ARV-associated diseases are frequently reported despite implementation of ARV vaccination programs. Therefore, continuous screening for new variants is essential to improve related control measures. During this study, samples were collected from three Egyptian governorates for determination of the circulating ARV strains. Phylogenetic analysis based on the amino acid sequences of the σC protein revealed the identification of two genotyping clusters (GCs) of ARV (GC 4.4 and 5.2). Amino acid identity within each cluster was more than 94.9%. Meanwhile, the amino acid identity between the two clusters was less than 37.8%. Low amino acid identity (less than 27.2%) was found among the identified isolates and vaccine strains. Cluster 4-related isolates ReoHS3 and ReoHS6 were clustered together within a distinct clade, sharing not more than 83.1% amino acid identity with the closest strains. In addition, unique nucleotide insertion and deletion were found at the heptad repeat region of the σC coding gene of these two isolates. Despite the nucleotide deletion and insertion and the hypervariability at the heptad repeat region, the same length and the hydrophobicity at position "a" (and to some extent position "d") were maintained among the identified isolates. These results can clarify the significance of both the length of the heptad repeat region and the hydrophobicity at position "a" in the maintenance of the function of this region.

To assess the efficacy of two inactivated vaccines against serovar B variant infection in laying hens, day-old female chicks were allocated into five experimental groups (G): G1 received a commercial vaccine V1, strains 17756, 0222, and Modesto of serotypes A, B, and C, respectively, at 8 and 12 wk of life; G2 received V1 through an early vaccination plan at 5 and 12 wk of life; G3 received another commercial vaccine V2, strains 083, Spross, 48, and H18 of serotypes A, B, Bvar, and C, respectively, against infectious coryza, which contained a serovar B variant strain in its formulation at 8 and 12 wk of life. Chickens in G4 were not vaccinated and challenged and were used as positive control, and G5 was not vaccinated and not challenged and served as negative control. At 25 wk of life, chickens from G1 to G4 were inoculated (by injection into the infraorbital sinus) with the serovar B variant of strain INTA H8, into the left infraorbital sinus of each animal. Clinical signs were monitored daily until Day 5 postchallenge. On that day, bacterial isolation was performed on both inoculated and noninoculated sinuses from each animal to determine the presence of . The interpretation of clinical signs involved grading on a scale from 0 to 4, depending on the severity of conjunctivitis and swelling of the periorbital area. On Day 2 postchallenge, the highest number of birds from all inoculated groups exhibiting clinical signs was recorded, gradually decreasing over the following days. The unvaccinated group (G4) had significantly (χ, < 0.05) more birds with clinical signs compared to vaccinated birds, regardless of the immunization plan or age. Overall, no significant differences were observed between G1 and G3 on any of the postchallenge observation days. However, G1 had significantly fewer birds exhibiting clinical signs compared to G2, the group that received the early vaccination plan. On the other hand, comparing the results of the noninoculated sinuses, no differences were found among vaccinated birds in G1, G2, and G3, whereas significantly more infected sinuses were found in the nonvaccinated birds. In conclusion, both V1 and V2 demonstrated efficacy in significantly reducing clinical signs, both with the conventional vaccination plan at 8 and 12 wk of life (V1 and V2) as well as with early vaccination starting at 5 wk of age (V1), compared to nonvaccinated birds. It also showed effectiveness in reducing the presence of in the infraorbital sinuses following the experimental challenge with the serovar B variant. Furthermore, V1, which does not have a Bvar strain, was proven to be effective against the serovar B variant of , specifically demonstrating its efficacy in protecting against clinical signs associated with this particular serovar.

White chick syndrome (WCS), a vertically transmitted disease caused by chicken astrovirus (CAstV), causes decreased hatchability of broiler breeder progeny and increased hatch of weak, white chicks. In the absence of a commercial vaccine, control measures for WCS have included serological surveillance of pullets, then movement of litter from farms with confirmed CAstV seroconversion to farms with seronegative birds. However, litter movement carries the risk of introducing other pathogens to farms, such as . CAstVs have been isolated from clinical cases of WCS and genetically characterized as chicken astrovirus, group Biv based on sequencing of the capsid protein. Autogenous vaccines (AVs) have included CAstVs, but no data exist to validate their ability to stimulate adequate immunity in breeders to prevent WCS. The objectives of this study were to 1) develop a CAstV Biv-inactivated oil emulsion vaccine using a CAstV isolate from a clinical case of WCS and 2) evaluate antibody production following injection into 3-wk-old specific-pathogen-free (SPF) leghorns. The CAstV isolate was propagated and titrated in leghorn male hepatoma cells, inactivated using beta-propriolactone, formulated into an oil emulsion with Montanide® ISA 70 VG adjuvant and then injected intramuscularly into 3-wk-old SPF leghorns. Birds received a second injection at approximately 9 wk of age. A negative control group received no vaccination at either time point. Serum was collected at 0, 2, 4, 8, 10, 12, and 14 wk post-initial vaccination (wpiv) for CAstV ELISA (BioChek®) and CAstV virus neutralization assay (VN). At 0, 2, and 4 wpiv no vaccinated birds had measurable antibody levels via enzyme-linked immunosorbent assay (ELISA) and VN tests. At 8, 10, 12, and 14 wpiv CAstV-specific antibodies were not detected by ELISA in either group; however, seroconversion was observed by VN in CAstV-vaccinated birds. Serum samples collected from breeder flocks with progeny that were clinically affected with WCS had consistently high ELISA titers, but the VN titers, while positive, were more variable. Control of WCS with a CAstV AV may be an option for use in broiler breeders. Progeny studies to detect maternal antibody transfer after vaccination would be useful in confirming this assertion. Based on this study, VN serology using a genetically similar CAstV as antigen is the preferred method of monitoring vaccinated flocks for seroconversion given the lack of antibody detection by ELISA. The ability to determine efficacy of any vaccine would rely solely on field observations as there is no established challenge model for WCS.

The unavailability of drugs and vaccines has resulted in a resurgence of histomoniasis outbreaks in recent years. Studies have been conducted to understand the infectivity of field isolates and develop early diagnostic tools for histomoniasis. To better understand the underlying physiological impact of , serum clinical chemistry analytes were evaluated in three independent studies. Poults were challenged with wild-type isolates by the cloacal route. In studies 1 and 2, blood samples were collected 9 days postchallenge (PC), while in study 3, blood samples were collected at 5 and 8 days PC for serum analysis. Among the several serum parameters evaluated, serum cholesterol and ALKP were consistently and significantly depleted in poults challenged with wild-type . Depletion of serum cholesterol and ALKP was documented as early as 5 days PC with .

We evaluated the infection and transmission potential of the highly pathogenic avian influenza (HPAI) H5N1 virus from infected crows () to healthy in-contact crows. Six crows were inoculated with 10 EID of H5N1 virus (A/crow/India/01CA249/2021), and 24 hr later six in-contact crows were introduced with them. All the birds were observed for signs of illness, and swabs were collected up to 14 days postinfection (dpi). One of the infected birds showed dullness, ruffled feathers, shivering, and reluctance to move at 7 dpi, while the other five infected crows showed no significant clinical signs except ruffled feathers and some signs of mental confusion. All the in-contact crows remained asymptomatic throughout the experiment. Virus could be isolated in swabs of infected birds from 1 to 7 dpi, and viral quantification revealed viral shedding up to 8 dpi. Although virus was not isolated from the swabs collected from in-contact birds, low numbers of viral RNA copies were detected in some samples. Virus was isolated from the brain and trachea of most of the infected birds along with other tissues. Virus was also isolated from different tissues collected from three of the in-contact birds, albeit with low numbers of viral copies. All six infected and five in-contact crows were found to be seroconverted against H5N1 virus when analyzed on 14 dpi. These findings suggest that the H5N1 virus was transmitted to in-contact birds, but the infectious doses were low, resulting in only a limited infection. Our study highlights that H5N1 viruses may be transmitted by direct contact within the house crow population, a feature that might play an important role in the epidemiology of avian influenza.

Increased fat content and inflammation are observed in woody breast (WB) muscles, suggesting dysregulation of fatty acids and inflammation in WB conditions. This study evaluated the gene expression related to fatty acid metabolism and inflammation responses in WB muscle. In total, 112 Ross × Ross 708 broiler chickens were raised following the commercial husbandry recommendations in a floor-pen chicken house. On day 41, the WB condition of each bird was assessed through palpation, and the birds were then categorized into normal and WB phenotypes. Next, 10 birds were euthanatized, five of which exhibited normal breast conditions (normal group) and five of which exhibited WB conditions (woody group), and breast muscle was sampled. RNA was extracted and reverse transcribed. The first-strand complementary DNA (cDNA) was analyzed using the real-time RT Profiler PCR Array, targeting 84 genes associated with chicken fatty acid metabolism and five housekeeping genes. Additionally, this cDNA was subjected to real-time quantitative PCR for 10 inflammatory genes, utilizing the chicken gene as housekeeping control. Fold changes were calculated using the difference in cycle threshold (ΔΔCt) method and were compared using the Student -test with a significance level set at ≤ 0.05. Eight of the 84 examined genes related to lipid metabolism showed differential expression ( ≤ 0.05) between normal breast and WB samples. Specifically, three genes were upregulated ( ≤ 0.05) in WB samples: acetyl-coenzyme A acetyltransferase 2, alcohol dehydrogenase 6, and glycerol kinase. In contrast, five genes were downregulated ( ≤ 0.05) in WB samples: acyl-coenzyme A (CoA) dehydrogenase, acyl-CoA synthetase medium-chain family member 4, glycerol-3-phosphate dehydrogenase 2, 5'-AMP-activated protein kinase subunit beta-2, and 5'-AMP-activated protein kinase subunit gamma-3, where AMP is adenosine monophosphate. Compared to the normal group, the expression levels of Toll-like receptor 2A, Toll-like receptor 4, interferon-γ, interleukin-1β, and interleukin-6 in the WB group were significantly increased ( ≤ 0.05). In conclusion, gene expression in WB muscle indicated upregulation of lipid biosynthesis, downregulation of fatty acid catabolism, and an activated inflammation response. This study revealed significant gene expression changes in fatty acid metabolism and inflammatory responses as key factors in the development of WB in broilers.

A breeder flock of 3200 Pekin ducks () experienced a pox virus infection, which affected 40% to 45% of the flock and lasted for a period of 6 wk. No biting insects or equipment that could have caused injury were found, although injury was considered to be the inciting factor. Feed and water consumption as well as egg production and hatchability were unaffected. Pox lesions occurred primarily on the bill, with some ducks having lesions on the eyelid. Infection was confirmed by gross and microscopic examination of affected hens as well as PCR and DNA sequencing. Genetic analysis revealed that the isolate causing the infection was related to pox viral infections in other wild waterfowl found in the United States. This is the first documented case of a naturally occurring pox virus infection in commercial Pekin ducks in the United States.

Bacterial infections such as and necrotic enteritis (NE) caused by (CP) are responsible for significant economic losses in the broiler chicken industry. Our previous studies have involved trying to develop alternatives to antimicrobials and immunoprotective agents to such pathogens. Previously, we demonstrated that delivery of a single dose of oligodeoxynucleotides containing unmethylated cytosine-phosphodiester-guanine motifs (CpG-ODN) can promote antimicrobial immunity against yolk sac infections caused by and by enriching immune compartments and activating immune cells. Recently, we have demonstrated delivery of CpG-ODN twice by the intramuscular (IM) route in neonatal broiler chickens at Days 1 and 4 of age to induce trained immunity and protect against lethal septicemia later in the grow-out period. The objectives of this study were to explore the ability of CpG-ODN to induce trained immunity in broiler chickens (1) by administering CpG-ODN by the route and intrapulmonary (IPL) route at hatch and (2) by administering CpG-ODN by the route and IPL delivery of a CP vaccine at hatch to protect against infections. Intramuscular (IM) delivery of CpG-ODN twice at Days 1 and 4 of age in neonatal broiler chickens induced trained immunity to protect against NE. Induction of trained immunity in broiler chickens led to a switch in cellular energy metabolism of immune cells from glycolysis to mitochondrial oxidative phosphorylation (OXPHOS) following two administrations of CpG-ODN. We have also demonstrated that delivery of CpG-ODN by the route followed by delivery of a live CP vaccine by the IPL route at hatch induced trained immunity and significantly ( < 0.0001) protected birds against septicemia at 27 days of age. Trained immunity was induced in broiler chickens only with administrations of CpG-ODN by the route followed by the IPL route at hatch or delivery of CpG-ODN followed by IPL delivery of a live CP vaccine at hatch. These birds were significantly ( < 0.0001) protected against lethal septicemia and NE later in the production cycle, demonstrating the utility of CpG-ODN for induction of trained immunity and broad-spectrum protection of broiler chickens against common lethal bacterial infections.

The purpose of this study was to evaluate a deep amplicon sequencing approach for estimating the relative abundances of different spp. oocysts in litter from commercial broiler farms that may or may not be experiencing necrotic enteritis (NE) infections. Oligonucleotide primers directed to the mitochondrial cytochrome oxidase I (COI) gene, a sequence that is conserved among all chicken spp., were first used to PCR amplify , , and oocyst DNA. COI amplification was applied to samples containing either a single species or an equal mixture of , , and oocysts. Amplicon sequencing and mapping to the relevant COI sequences in the GenBank database confirmed the expected ∼100% mapping to the appropriate sp. and in approximately equal percentages (∼33%) for mixtures of equal numbers of spp. oocysts. This approach was then applied to DNA derived from oocysts obtained at 0, 2, and 4 wk after chick placement (growout) from a total of 20 individual houses on six different commercial broiler farms. Of the seven spp. known to infect chickens, only five were consistently found in litter at each collection time point: , , , , and . The relative numbers of and non- () oocysts in all litter samples as estimated by COI deep amplicon sequencing showed a modest correlation with the respective or oocyst counts ( ∼ 0.30). The results revealed an interesting phenomenon that supports the role of in predisposing chickens to NE. In this study, the percentage of as estimated by deep amplicon sequencing at 0, 2, and 4 wk growout showed a strong positive correlation with NE incidence (0 wk, = 0.57; 2 wk, = 0.52; 4 wk, = 0.61). This study provides evidence for the usefulness of a deep amplicon sequencing approach to estimating the relative abundances of different oocysts infecting chickens because it allows reactions to take place in a single tube, thus avoiding the time-consuming, labor-intensive, species-specific internal transcribed spacer 1 (ITS1) PCR analyses. More importantly, it allows one to explore relationships between NE incidence and the abundance of minor species, which would have been missed by oocyst counting or ITS1 PCR because most species are not distinguishable by microscopy, and ITS1 PCR is not quantitative.

is an important pathogen that causes fowl cholera (FC) in turkeys (). The aim of this study was to isolate and characterize from outbreaks of FC in turkeys for the first time in Morocco, both molecularly and pathologically, and to develop and evaluate an inactivated vaccine based on the isolated strain. Two farms in central Morocco were suspected of having FC. From each of these farms, two isolates were characterized. Molecular analysis showed that both strains were identified as genotype A and serovars 3*4. Three oil-based vaccines were prepared using the virulent isolate strain after evaluating its pathogenicity in target species to determine the protective dose. The vaccines were evaluated through challenge tests and serologic responses. According to the results, a dose of 10 colony-forming units showed 100% protection after challenge, and all animals became seropositive. Additionally, two vaccination programs were proposed to evaluate the duration of immunity: a double vaccination schedule a triple vaccination schedule. The results indicated that the triple vaccination induced good protection after challenge and a high level of antibodies.

Numerous reports and epidemiologic investigations in recent years identify adenoviral infection as an ongoing to increasing, international disease in commercially raised chickens. Impression smear cytopathology was evaluated as a tool for the diagnosis of adenoviral inclusion body hepatitis (IBH). In this study, 92, paired, Romanowsky-stained cytopathologic preparations and hematoxylin-and-eosin-stained histopathologic liver sections were evaluated from 54 chickens with experimentally induced or naturally occurring IBH. Large intranuclear inclusion bodies typical of adenoviruses were visible within hepatocytes on both cytopathology and histopathology. Cytopathologic to histopathologic percent positive agreement and percent negative agreement were 94% and 90%, respectively, with Π= 0.81 (0.61-1.01, 95% confidence, < 0.001). A subset of 20 cytopathologic samples evaluated by eight veterinary professionals as consistent or inconsistent with IBH yielded an average of 66% positive agreement and 98% negative agreement to the histopathologic diagnosis, across all observers, with Π= 0.61 (0.53-0.68, 95% confidence, < 0.001), resulting in a positive predictive value of 99% and a negative predictive value of 67%. Interobserver agreement was slightly higher (76% positive agreement, 94% negative agreement, Π= 0.68 [0.65-0.71], 95% confidence, < 0.001) within the 12 of these samples that originated from natural disease, with stable positive and negative predictive values. A rapid, cost-effective, tentative diagnosis of IBH via impression smear cytopathology at the time of necropsy can streamline further confirmatory laboratory testing and facilitate timely communication in the interim to affected parties, especially in locations with delayed access to a diagnostic laboratory.

Avian reovirus (ARV) infections pose a significant economic threat to poultry production despite biocontainment and routine vaccination efforts. Quantifying ARV load is essential for understanding infection dynamics. There is a widespread misperception that ARVs in cultures do not produce countable plaques, leading most in the field to use the less useful 50% tissue culture infectious dose quantifications. Here, we compare the suitability of two well-known avian cell lines, quail myoblast clone 5 (QM5) and leghorn male hepatocellular carcinoma (LMH), for use in plaque assays for the quantification of ARV. LMH cells, which exhibit syncytia formation postinfection, proved unsuitable for plaque assays due to poor substrate adherence, the tendency to form syncytia-like conglomerations of cells even when uninfected, and the tendency for areas of the substrate cleared by viral cytopathic effect (CPE) to quickly fill in with new cell growth. In contrast, QM5 cells demonstrated clear contact inhibition and well-defined CPE, enabling distinct, countable plaques even at high virus titers. Therefore, although LMH cells are advantageous for viral propagation, QM5 cells are better suited for plaque assays to assess ARV infectivity, with QM5 enabling more precise viral growth tracking and quantification.

This case report details a severe outbreak of acute fowl cholera (FC) in commercial, slow-growing broiler chickens. Three necropsy cases of slow-growing broilers were submitted for postmortem examination at the California Animal Health Food Safety (CAHFS) Laboratory System at three different time points in their production cycle following an acute elevation in mortality on the farm. Broilers were submitted at 5 wk, 11 wk, and 14 wk of age. The birds were submitted from a flock of 25,000 broilers on a 262,500-broiler ranch that experienced 100% mortality by the time of processing at 14 wk of age. Gross findings included petechia, mottled livers and spleens, and congested and edematous lungs. On histology, major findings included hepatic necrosis, splenic necrosis, pulmonary edema, and bacterial colonies proliferation in multiple organs. A definitive diagnosis of FC was made with qPCR detection and aerobic culture and isolation of . The isolate was classified as serovar 1, fingerprint profile HI 0001 with serotyping and restriction enzyme analysis, respectively. A review of all FC cases detected in avian species submitted to the CAHFS laboratory in the last 11 yr revealed 10 cases of FC in slow-growing broilers and no cases in traditional broilers. This case underscores the need for enhanced biosecurity and potential vaccination strategies in slow-growing poultry systems to mitigate future risks of FC outbreaks.

Between May 2021 and June 2024, a series of slaughterhouse condemnations was documented in the male population of a commercial turkey farm in Lower Austria, Austria. The primary reasons for condemnation were severe airsacculitis, often accompanied by ascites and pleural effusion. Across various production cycles, the flocks either remained asymptomatic with lesions only being detected during carcass inspection at the slaughterhouse or showed a slight increase in mortality accompanied by clinical signs such as dyspnea, open-mouth breathing, and head shaking in some birds during the final weeks of the fattening period, at 16-22 weeks of age. Gross examination prior to slaughter typically revealed thoracic air sacs filled with large, firm, yellowish-white plaques, occupying a significant proportion. During the last two occurrences, microbiological and pathological investigations were conducted to identify the causative pathogens of this recurrent condition. Various infectious agents, including , , and , were identified. The findings revealed the multifactorial nature of the condition, raising concerns about the impact on the health and welfare of turkeys, as well as the economic implications of such condemnations for the turkey industry.