Alaska represents a globally important region for the ecology of avian-origin influenza A viruses (IAVs) given the expansive wetlands in this region, which serve as habitat for numerous hosts of IAVs that disperse among four continents during the annual cycle. Extensive sampling of wild birds for IAVs in Alaska since 1991 has greatly extended inference regarding intercontinental viral exchange between North America and East Asia and the importance of Beringian endemic species to IAV ecology within this region. Data on IAVs in aquatic birds inhabiting Alaska have also been useful for helping to establish global patterns of prevalence in wild birds and viral dispersal across the landscape. In this review, we summarize the main findings from investigations of IAVs in wild birds and wetlands of Alaska with the aim of providing readers with an understanding of viral ecology within this region. More specifically, we review viral detections, evidence of IAV exposure, and genetic characterization of isolates derived from wild bird samples collected in Alaska by host taxonomy. Additionally, we provide a short overview of wetland complexes within Alaska that may be important to IAV ecology at the continental scale.

Eimeria spp. are important intestinal pathogens of chickens (Gallus gallus domesticus). Anticoccidial feed additives, chemicals, and ionophores have traditionally been used to control Eimeria infections in broiler production. Thus, the trend toward antibiotic-free and organic production requires new approaches to coccidiosis prevention. Two not mutually exclusive methods are the use of plant extracts with antiparasitic activity and manipulation of the intestinal microbiota by pre- and probiotics. In the present study, birds were inoculated with a combination of Eimeria acervulina, Eimeria maxima, and Eimeria tenella. We profiled the jejunal microbiome at multiple time points postinfection to investigate the changes in jejunum microbiota and to identify the time point of the maximum difference between infected and noninfected birds. Additionally, we assessed the anticoccidial effects of two anecdotal treatment methods, green tea and apple cider vinegar, as well as amprolium. Green tea and apple cider vinegar had no effect on oocyst shedding, but green tea reduced the mild unspecific lesions in coccidia-infected birds; there was no influence on unspecific lesions in uninfected controls. Jejunal contents were collected on the day of the infection and 1, 2, 4, 6, 10, and 14 days postinfection (dpi) for investigation of the intestinal microbiota by 16S ribosomal (r)RNA gene sequencing. Comparison of the untreated-uninfected and the untreated-infected groups showed a maximum community dissimilarity of 10 dpi. From 4 days after infection, Clostridiales were significantly enriched at the expense of Lactobacillales in infected compared with uninfected birds. Interestingly, treatment with green tea prevented proliferation of Clostridiales induced by the coccidia and increased the relative abundance of Melainabacteria.

Trichomonas gallinae, a single-celled protozoan parasite, is a causative agent of the disease trichomonosis, which is distributed worldwide and has recently been highlighted as a pandemic threat to several wild bird species. The aim of this study was to determine the prevalence and genotypic diversity of T. gallinae in Riyadh, Saudi Arabia. For this purpose, 273 oral swab samples from different bird species (feral pigeon Columba livia, common mynah Acridotheres tristis, chicken Gallus gallus domesticus, turkey Meleagris gallopavo, and ducks Anatidae) were collected and tested for T. gallinae infection with InPouch™ TV culture kits. The results showed that the overall prevalence of T. gallinae in these samples was 26.4% (n = 72). The PCRs were used to detect the internal transcribed spacer (ITS) region of T. gallinae, and the results of the sequence analysis indicated genetic variation. Among 48 sequences, we found 15 different ribotypes, of which 12 were novel. Three had been previously described as ribotypes A, C, and II. To our knowledge, this study demonstrated the presence of T. gallinae strain diversity in Saudi Arabian birds for the first time and revealed that ribotypes A and C are predominant among Riyadh birds.

Aquatic wild birds, especially waterfowl, have been long considered the main reservoirs of the avian influenza A virus; however, recent surveys have found an important prevalence of these viruses among land birds as well. Migration has been suggested as an important factor in the avian influenza virus dissemination. We aimed to estimate the prevalence of influenza A viruses in wild birds (waterbirds and land birds; resident and migratory) in eastern Mexico, where the three main North American migration flyways converge and where there was no previous information on this subject. We detected influenza with reverse transcription coupled with a PCR approach. Of the 534 birds sampled between 2010 and 2012, we detected the influenza A virus in a high proportion of birds (39%). Prevalence was particularly high in land birds (49%) when compared to aquatic birds (26%); there was no difference in overall prevalence between resident (39%) and migratory birds (39%). The high prevalence of the avian influenza virus in land birds was noteworthy in the innermost sampling areas in northern Mexico (Coahuila [82%] and Nuevo Leon [43%]).

Both highly pathogenic (HP) and low pathogenic (LP) avian influenza virus (AIV) can cause decreases or even cessation of egg production in chickens and turkeys. Production of abnormal eggs (deformed, thin-shelled, soft-shelled) can also be caused by AIV infection. Additionally, egg surfaces and contents may also be contaminated with virus. Because data quantifying these effects are lacking, white Plymouth Rock hens were inoculated with HP or LP AIV while in production. No decreases in egg production or abnormal eggs were observed with LPAIV-infected hens. No lesions or viral antigen staining in ovary and oviduct were observed in LPAIV-infected hens 3 days postchallenge. LPAIV RNA was detected on eggs collected from 12 hr to 11 days postinoculation (PI) and was on or in 6.4% (15/234) of the eggs. Titer equivalents of LPAIV ranged from 1.3–2.5 log₁₀ 50% egg infectious doses (EID₅₀). No virus was detected in embryo tissue from eggs laid by LPAIV-infected hens. In contrast, egg production by HPAIV-inoculated hens decreased at 72 hr PI and 18.4% (16/87) of the eggs were abnormal. However, viability was similar to that of the sham inoculates. HPAIV RNA was detected in or on 11.1% (9/81) of the eggs from 36 hr through 96 hr PI, when the hens were euthanatized. HPAIV RNA was detected on 6.2% of eggshells, in 4.2% of albumin/yolk samples, and in 8.3% of embryo tissue. Forty percent of the abnormal eggs were positive for HPAIV RNA. Titer equivalents on or in HPAIV-contaminated eggs ranges from 1.0–4.0 log₁₀ EID₅₀. Lesions and viral antigen staining were present in the ovary and all sections of the oviduct of infected hens 3 days postchallenge. These data will inform models using production-based triggers for LPAIV monitoring and for risk assessments to determine the disposition of eggs from flocks infected with LPAIV or HPAIV.

Infection of the oviduct by an infectious bronchitis virus (IBV) in laying hens has been associated with the false layer syndrome. Because the diagnostic procedure for the detection of cystic oviducts by postmortem examinations in IBV-positive replacement pullet flocks could involve the unnecessary sacrifice of numerous healthy pullets without reproductive tract anomalies, the development of a noninvasive and nonlethal diagnostic procedure would be desirable. The first objective of the study was to evaluate the diagnostic accuracy of a transcutaneous ultrasonography method to predict the presence of cystic oviducts compared to postmortem examinations in a commercial pullet flock positive for an IBV genotype Delmarva (DMV) variant. The second objective was to evaluate the performance of the same ultrasonography method to later detect false layers in the same flock in sexually mature hens by identifying the presence of an egg in the oviduct due to the presence of atretic oviducts undetectable by ultrasonography and the absence of cystic oviducts at that age. In replacement pullets, the sensitivity (Se) and specificity (Sp) of the ultrasonography (index test) compared to the postmortem examination (reference standard test) were 73% and 91%, respectively. The positive predictive value (PPV) and negative predictive value (NPV) were 67% and 93%. The ultrasonography technique showed a positive likelihood ratio (LR+) of 7.82 and a negative likelihood ratio (LR–) of 0.30. In sexually mature hens, the Se, Sp, PPV, and NPV of the ultrasonography compared to the laying status were 98%. The LR+ was 49.00 and the LR– was 0.02 when compared to the laying status. In conclusion, the ultrasonography could replace postmortem examinations to detect cystic oviducts in commercial flocks of replacement pullets previously infected with an IBV-DMV 1639 variant. Although the test accuracy of ultrasonography was excellent for the hens at production peak to identify laying and nonlaying hens based on the presence of an egg in the reproductive tract, its practicality was limited due to atretic oviducts being not detectable.

The reemergence of infectious coryza (IC) caused by Avibacterium paragallinarum (AP) as an acute and occasionally chronic respiratory disease in domestic poultry has caused severe losses in several U.S. states. The disease is also associated with decreased egg production in layers and increased condemnations from air sac infections in broilers. A series of applied experiments were performed to elucidate the persistence of AP in infected broiler flocks, to genotype AP strains isolated from field cases, and to evaluate commercial and autogenous vaccine protection in commercial and specific-pathogen-free (SPF) chickens. Experimental evaluation of environmental persistence suggests that AP did not persist more than 12 hr in a hypothetically contaminated environment. Additionally, other detected potential pathogens such as Gallibacterium anatis and infectious bronchitis virus caused mild respiratory signs in the exposed birds. The HMTp210 and HagA genes of four IC field strains were sequenced and compared with published sequences of HMTp210 and HagA. The HMTp210 phylogeny showed a marginally imperfect clustering of the sequences in genogroups A, B, and C. Although not definitive, this phylogeny provided evidence that the four field strains aligned with previously characterized serovar C strains. Moreover, the base pair homology of the four strains was 100% identical to serovar C reference strains (H-18 and Modesto). HagA phylogeny was unclear, but interestingly, the IC field strains were 100% homologous to C-1 strains reported from Mexico and Ecuador. Finally, vaccine protection studies in commercial hens indicate that clinical signs are induced by a combination of IC and other concomitant pathogens infecting commercial birds. Additionally, vaccine protection experiments performed in SPF hens indicated that protection provided by the two commercial vaccines tested provided a reduction in clinical signs and bacterial shedding after two applications.

Duck enteritis virus (DEV) causes an acute and contagious infection in duck. The present study was carried out to evaluate the pathogenicity and pathodynamics of DEV isolates from different natural outbreaks in the Assam Province of India. A total of six wild-type isolates of DEV were revived in ducklings to determine its biologic characterization. Postmortem examination of infected ducklings revealed DEV-specific gross lesions in different organs. The presence of DEV was confirmed by its genome amplification and the presence of viral antigens from collected tissue samples by indirect fluorescent antibody test. All the isolates revived in ducklings were further propagated in duck embryo fibroblast cells. Highly virulent and low virulent isolates of DEV were selected for further study based on median duck infectivity dose (DID₅₀) and median tissue culture infectivity dose (TCID₅₀). The highly virulent isolate of DEV had values of 10² DID₅₀/ml and 10^6.33 TCID₅₀/ml, whereas the low virulent strain had titers of 10 DID₅₀/ml and 10^4.83 TCID₅₀/ml in the cell culture. Our results showed replication of DEV in ducks with the highest and lowest viral titers in the thymus and bursa of Fabricius, respectively. In addition, microscopic analysis revealed necrosis and degeneration of submucosal esophageal glands and glandular epithelium. The study will be useful to understand the organ tropism and pathologic alteration among the virulent DEV isolates.

Marek's disease (MD) is a highly contagious lymphoproliferative disease of chickens caused by Gallid alphaherpesvirus type 2. Gallid alphaherpesvirus type 3 (GaHV-3) strain 301B/1 was previously shown to be an effective MD vaccine with synergistic efficacy when used as a bivalent vaccine with turkey herpesvirus. Since the nucleotide sequences of only two GaHV-3 strains have been determined, we sought to sequence the 301B/1 genome using Illumina MiSeq technology. Phylogenomic analysis indicated that 301B/1 is more closely related to other GaHV-3 strains (SB-1 and HPRS24) than to virulent or attenuated strains of GaHV-2. One hundred and twenty-six open reading frames (ORFs) have been identified within the 301B/1 genome with 108 ORFs showing a high degree of similarity to homologs found in the genomes of SB-1 and HPRS24; 14 ORFs are highly homologous (> 90% identity) with the corresponding ORFs within the SB-1 genome. The R-LORF8 and R-LORF9 genes are the most dissimilar to the collinear genes found in the SB-1 genome but are highly homologous (99%–100% identity) with those within the HPRS24 genome. Overall the 301B/1 genome is most similar to the SB-1 virus genome (99.1%) and to a lesser degree with the HPRS24 virus genome (97.7%). However, six 301B/1 ORFs (UL47, UL48, UL52, pp38, ICP4, and US10) have been identified that contain nonsynonymous substitutions relative to homologs found in the SB-1 genome. Notably, unlike the avian retrovirus long terminal repeat sequences found within the SB-1 genome, none were identified within the 301B/1 genome.

Nine infectious bronchitis virus (IBV) strains belonging to the GI-7 lineage were isolated between 2009 and 2017 in China. Phylogenetic analysis and comparisons of full-length sequences of the S1 gene suggested that the GI-7 lineage should be further classified as Taiwan (TW)-I and TW-II sublineages, which correspond to the previous TW-I and TW-II genotypes. The nine IBV strains were clustered in the TW-II sublineage. Further investigation revealed that viruses in the TW-I and TW-II were not only genetically but also antigenically different. Moreover, the TW-II sublineage contained various clades and recombinants. A recombinant was found to originate from recombination events between field strains (TW-II ck/CH/LJL/090608- and GI-19 ck/ CH/LDL/091022-like viruses) in which the recombination in the S1 subunit coding sequences had led to changes in antigenicity of the viruses. A more in-depth investigation demonstrated that TW-II viruses appear to have undergone a significant evolution following introduction in mainland China, which resulted in the viruses diverging into different clades. The viruses between the different clades in TW-II sublineage exhibited a significant change in genetic and antigenic characteristics. In addition, the five TW-II viruses selected on the basis of the results of S1 nucleotide sequence phylogenetic trees showed different pathogenicity to specific-pathogen-free chickens, although they could induce nephritis in the infected chickens and thus were identified as nephropathogenic strains.

Despite routine vaccine use, sporadic outbreaks of infectious coryza in poultry continue to occur in Taiwan. This study was designed to determine the serotypes and the complete nucleotide sequences of a hemagglutinin gene (HMTp210) of Avibacterium paragallinarum isolated in Taiwan between 1994 and 2017. Hemagglutination inhibition tests showed that these isolates belong to serogroups B and C. Sequence analyses of the HMTp210 gene showed that Taiwanese serogroup B isolates are most similar (94.7%–98.2% identity) to strain FARPER-174 isolated in Peru in 2015. In contrast, Taiwanese serogroup C isolates are most similar (96.3%–99.8% identity) to strain H-18 isolated in Japan in 1976. This is the first report showing the presence of A. paragallinarum of serogroup B in Taiwan. In addition, one Taiwanese isolate showed cross-reactivity with serogroup B and C antisera. This isolate contains a chimeric HMTp210 gene that might result from recombination between serogroups B and C. These findings could be valuable for the epidemiologic study and molecular serotyping of A. paragallinarum.

This article outlines pathomorphologic findings of a study involving commercial mule ducks with confirmed influenza A H5N8 infections after a series of outbreaks in Bulgaria. Examinations were carried out after performing necropsy on dead birds from three different age groups (up to 15, 20 to 30, and 40+ days of age) fattened on different farms. Among birds of all ages, gross lesions were present as lesions affecting the heart. Histologically, the myocardium exhibited severe intermyofibrillar edema, moderate to massive hemorrhages, and degenerative changes. All lesions resulted in single or multiple and small to massive myocardial infarctions. Other affected organs included the brain, lungs, liver, spleen, and pancreas. Nonpurulent lymphocytic encephalitis was found postmortem in ducks that had shown prior clinical nervous signs. Among ducks of all ages, a viral antigen in the cardiomyocytes and the epithelium of air capillaries was found through immunohistochemical detection methods. The results of the present study allowed us to conclude that the highly pathogenic avian influenza A H5N8 viral infection may manifest itself as a systemic illness in commercial mule ducks with septicemic lesions, resulting in high morbidity and mortality rates of up to 100%. Pathomorphologic lesions were somewhat different from those previously reported in wild waterfowl.

Histomoniasis is a significant disease of gallinaceous birds caused by Histomonas meleagridis. Transmission of this parasite is dependent on use of the cecal nematode Heterakis gallinarum. To define the host range of this nematode, cecal contents from 399 game birds and poultry, representing eight species, were examined for Heterakis spp. The majority of these species (five of eight) were infected with Heterakis nematodes. Heterakis gallinarum was detected in free-ranging wild turkeys (Meleagridis gallopovo), captive-raised ring-necked pheasants (Phasianus colchicus), chukars (Alectoris chukar), and domestic chickens (Gallus gallus domesticus), whereas H. isolonche was found in ruffed grouse (Bonasa umbellus). No Heterakis species were identified in the domestic turkey (Meleagridis gallopovo), American woodcock (Scolopax minor), and dabbling duck (Anas spp.) samples. Genetic characterization indicated that nematodes identified as H. gallinarum were present in two distinct clades. One clade of H. gallinarum sequenced from this study grouped with chicken-derived sequences from other countries. The other group of sequences consisted of a sister clade to a group of parasites morphologically identified as H. isolonche. Currently it is unknown if this group represents a genetic variant of H. gallinarum, a variant of H. isolonche, or a novel species. These results indicate Heterakis infection varies among poultry and game bird species but is common among select gallinaceous species in Pennsylvania.

Avian chlamydiosis is an infection caused by obligate intracellular and Gram-negative bacteria belonging to the family Chlamydiaceae and has been reported in more than 450 avian species distributed in 30 orders. In particular, a high prevalence of infection has been demonstrated in wild passerine populations, including both asymptomatic and clinically ill individuals, suggesting a role of these avian species as important carriers. In May 2018, avian chlamydiosis was diagnosed in a 1-year-old male Gouldian finch (Erythrura gouldiae) at the Turlock Branch of the California Animal Health and Food Safety Laboratory System. The bird belonged to an outdoor aviary with mixed avian species, including Gouldian finches, doves (Geopelia cuneata and Spilopelia chinensis), and psittacines (Aratinga, Psittacula, Pyrrhura, and Trichoglossus sp.). Severe respiratory distress and mortality were noted among the finches. Gross and histopathologic lesions were concentrated in the liver and spleen, with a mild involvement of the upper respiratory tract. Chlamydia spp. were detected in the spleen and kidney by real-time PCR and were further confirmed by immunohistochemistry. Subsequently, Chlamydia psittaci was isolated from the liver and spleen and characterized as a CP3-like strain (genotype B). In addition, viral particles compatible with circovirus were identified in the liver by direct electron microscopy. To the authors' knowledge, this is the first report of avian chlamydiosis with hepatic viral particles consistent with circovirus infection in a Gouldian finch.

Avian poxvirus (APV) is an enveloped double-stranded DNA virus that affects many domestic and wild birds worldwide. APVs are classified into three clades (A to C), represented by fowlpox (FP) virus (clade A), canarypox virus (clade B), and psittacinepox virus (clade C), although two additional clades (D and E) have been proposed. In this study, a tumorlike skin lesion found in a domestic fowl was submitted for molecular diagnosis of Avipoxvirus by PCR and sequencing. The phylogenetic analysis revealed that the amplified segment of the corelike 4b protein and polymerase genes clustered in clade E. The APVs in clade E were previously reported from outbreaks in Hungary (flock of turkeys) and in Mozambique (layer chickens), associated with a possible vaccine failure to protect against clade E viruses. To our knowledge, this report is the first identification of clade E in this country, providing new information about host range and genetic diversity of APVs in Brazil, and may represent a potential risk of FP disease outbreaks in commercial poultry.

A grey-hooded parakeet (Psilopsiagon aymara) and two budgerigars (Melopsittacus undulatus) from different owners presented with decreased activity, vomitus, and diarrhea. A microscopic examination of feces showed trophozoites of the protozoan flagellate Giardia. A commercial immunochromatographic dipstick test for Giardia sp. antigens confirmed the infection. These findings were assured by PCR of the small subunit ribosomal RNA (SSU rRNA) gene and coproantigen ELISA. Sequencing of PCR products of the SSU rRNA (292 bp) and β-giardin genes (511 bp) identified Giardia psittaci as the species involved. Therefore, our results show that a GSA 65–based coproantigen ELISA, which was established for diagnosis of Giardia duodenalis is applicable for the detection of G. psittaci. A treatment with ronidazole was started. Additionally, fecal examination and dissection of the dead birds revealed coinfection with the fungal pathogen Macrorhabdus ornithogaster. One budgerigar survived and repeatedly tested negative after treatment with ronidazole. The described cases indicate that a single infection with G. psittaci has a good prognosis, whereas the prognosis is poor when coinfections occur, especially with M. ornithogaster.