Pathology and Tissue Distribution of an LPAI H5N8 of North American Lineage Isolated from an Outbreak in Commercial Japanese Quail (Coturnix c. japonica) in the Central Valley of California.

This report describes the pathology and tissue distribution of avian influenza (AI) antigens by immunohistochemistry (IHC) in the tissues of commercial layer quail from a natural outbreak of low pathogenic avian influenza (LPAI) H5N8. LPAI virus H5N8 of North American lineage was diagnosed in commercial Japanese quail hens ( Coturnix coturnix japonica) in California based on serology, reverse-transcriptase real-time polymerase chain reaction, virus isolation, and sequencing. The sudden increase in mortality in a flock of laying quail hens had prompted the submission of 15 live and 5 dead, 10- to 15-wk-old quail to the California Animal Health and Food Safety Laboratory System, Turlock branch in the beginning of April 2014. There was mild bilateral swelling of the eyelids and greenish diarrhea in 4/15 live quail submitted. On postmortem examination, there were severe, extensive hemorrhages and multifocal, confluent pale foci in the pancreas in 10/20 birds. Liver gross lesions in five birds ranged from a few pale areas to numerous disseminated foci. Histology revealed moderate to severe necrosis of acinar cells in the pancreas with little or no inflammation in most of the birds. Livers had acute multifocal coagulative necrosis of hepatocytes with fibrin exudation and infiltration of few to large numbers of heterophils and lymphocytes randomly scattered throughout. The AI virus was detected in the nucleus and cytoplasm of pancreatic acinar cells and hepatocytes by IHC targeting the nucleoprotein of the AI virus. A few birds had AI antigen in the reticuloendothelial cells of the spleen, endothelial cells of the lungs, epithelium of the respiratory mucosa, and lamina propria of the intestine. The severity of the lesions observed in this natural outbreak of LPAI in quail was higher than that expected for the pathotypic presentation in this species.

A Novel Mycoplasma sp. Associated with Phallus Disease in Goose Breeders: Pathological and Bacteriological Findings.

In April 2014, poor fertility in a major commercial goose breeder operation in California triggered the submission of six live affected Toulouse ganders ( Anser anser ) to the California Animal Health and Food Safety Laboratory, Turlock branch (University of California-Davis). Toulouse were principally affected among all breeds, and their egg fertility dropped from 65.7% to less than 33.9% in the first 40 days of the 2014 breeding season. The flock consisted of 410 adult birds, 90 males and 320 females, between 2 and 5 yr of age. Inspection of the flock revealed that 44.4% of the Toulouse ganders had severe phallic deformities that prevented them from mating. At postmortem examination, severe yellowish fibrocaseous exudate disrupted the architecture of the phallus and occasionally produced fistulating tracts through the wall of the organ. Microscopically, multifocal lymphoid nodules were noted in the mucosa and submucosa of the phallus and were associated with extensive granulomatous reaction, intralesional bacteria, and spermatozoa. Mycoplasma spp. were isolated from the phallus of affected and nonaffected birds, and PCR protocols targeting the 16S-23S ribosomal RNA intergenic spacer regions and the RNA polymerase beta subunit gene were performed to identify the isolates. Three distinct species were identified on sequencing and analysis using the National Center for Biotechnology Information basic local alignment search tool: Mycoplasma cloacale , Mycoplasma anseris , and an unknown novel Mycoplasma sp. Additionally, Pasteurella multocida , in combination with other bacteria, was also isolated from the phallic lesions and identified as serotype 3 with a DNA profile of 1511 (National Veterinary Service Laboratory). This is the first report of these Mycoplasma spp. and other bacteria associated with reproductive disease in ganders in the United States.

Characterization of Seven Outbreaks of Hemorrhagic Hepatopathy Syndrome in Commercial Pullets Following the Administration of a Salmonella Enteritidis Bacterin in California.

Between April 2013 and April 2015, seven flocks belonging to three different major commercial egg producers inCalifornia experienced a mild increase in mortality 2 to 3 wk after administration of Salmonella Enteritidis bacterins. Strains of chickens involved were H&N (flock A1, A2, B2, C1, C2, and C3) and Lohmann white (flock B1). Vaccination was administered individually through injection either in the breast muscles or subcutis in the legs between 11 and 18 wk of age in all flocks. Clinical signs ranged from inapparent to lameness, reluctance to walk, greenish diarrhea, and retching-like symptoms. The mortality ranged from 0.16% to 1.38% per week, with the highest peaks occurring usually 2 to 3 wk postvaccination, and then declined rapidly. Postmortem examinations revealed enlarged livers with disseminated hemorrhages and pale foci of necrosis. Also, severe extensive hemorrhages in the intestine, heart, and proventriculus were observed in a few birds. Various degrees of productive, exudative giant cell granulomatous myositis were observed invading deeply the muscles and subcutis at the site of vaccination. The myositis was always associated with optically empty vacuoles positive for neutral lipids by Oil Red O stain. Droplets of Oil Red O material were also noticed in the affected livers and intestines. Congo red stain highlighted the presence of amyloid in moderate to severe amounts in the breast muscles and moderate amounts in livers, spleens, and intestines. Salmonella antigens were detected in the injection sites and livers by immunohistochemical staining. No viruses or toxic substances were recovered from the liver, spleen, intestine, and pectoral muscles, and the few bacteria isolated were interpreted as secondary postmortem invaders. In addition, livers and bile tested for hepatitis E virus were negative by reverse-transcriptase polymerase chain reaction.

Trichodinosis associated with pathology of the reproductive tract in waterfowl.

Trichodinid ciliophorans are opportunistic parasites of many species of fish, amphibians, and molluscs, but yet never reported in association with lesions in birds. Postmortem and histopathological evaluation of a commercial adult Toulouse gander and female goose, and a wild Mallard drake revealed the presence of severe pathological parasitic colonization of their reproductive tracts. Histopathological findings included moderate to severe granulocytic inflammation, acanthosis, accentuation of the rete pegs, and proliferative hyperplastic squamous metaplasia of the mucosa of the ejaculatory ducts and groove, sulcus spermaticus, glandular part of the phallus (cavum penis), and oviduct in association with large numbers of ciliated protozoa anchored to the tissues or free in the lumen. These protozoa had characteristic morphological features analogous to the family of Trichodinidae. The source of this parasitism could not be determined. To our knowledge, this is the first report of trichodinosis associated with pathology in birds.

Atypical distribution of fowl pox lesions in broilers.

An unusual cutaneous fowl pox outbreak occurred in 8-wk-old broilers in California. Rounded and longitudinal, proliferative scratch-associated lesions were found only in feathered areas of the body. Both sides of the hip, the lower abdomen, pericloacal area, and lateral lower neck area were involved. The head, legs, feet, and toes did not have lesions. Birds in only one section of one of five houses were affected. Fifteen percent condemnations occurred in birds from the affected house due to the skin lesions. A diagnosis of fowl pox was achieved by histopathology, viral isolation, and direct electron microscopy. The unusual distribution of pox lesions was assumed to be associated with skin scratches. There was no evidence that mosquitoes or other types of insects were involved in this outbreak. To the knowledge of the authors, this is the first report of this kind of unusual fowl pox in the United States.

Respiratory tract trichomoniasis in breeder squab candidates in Northern California.

Breeder squab candidates between the ages of 6 and 16 wk were submitted to the California Animal Health and Food Safety Laboratory, Turlock branch, as a result of respiratory distress and increased mortality. These cases were submitted from one Northern California commercial squab operation on three separate occasions occurring between December 2007 and March 2008. Severe trichomoniasis was identified, primarily in the tracheal epithelium and lung of squabs, with few or no lesions in the oral cavity, crop, esophagus, and livers, where the organism commonly infiltrates. Infiltration of the trachea and lung sections with trichomonads was associated with a severe inflammatory response in the surrounding tissue. Diagnosis was confirmed with the use of histopathology and an immunoperoxidase special stain. Oxytetracycline supportive antibiotic therapy to prevent secondary bacterial infections was administered to remaining squabs on the farm, but no specific treatment regimen was instituted. This novel respiratory presentation of trichomoniasis continued over a period of 3 mo, until mortality gradually returned to normal.

Hexamita meleagridis (Spironucleus meleagridis) infection in chukar partridges associated with high mortality and intracellular trophozoites.

An outbreak of infectious catarrhal enteritis, associated with the flagellated protozoan Spironucleus meleagridis (syn. Hexamita meleagridis), is reported from a commercial flock of chukar partridges in California. The disease affected birds between the ages of 4 and 6 wk and resulted in diarrhea, listlessness, depression, and high mortality. Concurrent infection with other intestinal pathogens, including Cryptosporidia, group E Salmonella, long-segmented filamentous microorganisms (LSFMOs), and Rotavirus-like virus particles, was found in some but not all affected birds. Dermatitis of the face, shanks, and feet, suggestive of B-complex vitamin deficiency, was present in most affected birds as well. Flagellated protozoan parasites could be found in the lumen of the duodenum and jejunum and in the intestinal crypts. In some cases the flagellates were wedged between epithelial cells or were located intracellularly within cells of the mucosal epithelium and the intestinal lamina propria.

Postural and neurological deficits in broiler chicks after cervical vaccination with live vaccine.

A disease characterized by paresis and paralysis was seen in 7-9-day-old broiler chicks after vaccination in the neck area at day-of-age with a live virus vaccine containing viruses of Marek's disease, fowl pox, and infectious bursal disease. Affected birds presented with variable signs of ataxia, lateral recumbency, leg paralysis, and twisting or S-shaped flexure of the neck. Gross lesions noted at necropsy included swelling and edema of the subcutaneous tissues and muscles of the neck at the injection site area. A heavy mononuclear inflammatory cell infiltration was seen in the subcutaneous tissues, connective tissues, and muscles of the neck at the injection site. In some cases, the inflammatory process extended along fascial planes to involve the epidural spaces surrounding the spinal cord. Fatty changes with possible demyelination of nerve fibers were noted in some sections of the spinal cord adjacent to the inflammatory lesions. Clusters of poxviruses were found within some inflammatory lesions on transmission electron photomicrographs.

Complementary randomly amplified polymorphic DNA (RAPD) analysis patterns and primer sets to differentiate Mycoplasma gallisepticum strains.

Randomly amplified polymorphic DNA (RAPD) analysis was used to differentiate 7 strains of Mycoplasma gallisepticum. Six commercially available primers or primer combinations were screened for their ability to differentiate vaccine and type strains. Although major and minor bands were produced with each primer, many of the primers were unsuitable for strain differentiation. The use of primer 6 and combined primers 3 and 4 resulted in complementary RAPD banding patterns for each M. gallisepticum strain. Eleven different isolates representing 7 different strains were segregated into 7 different patterns, corresponding to the 7 strains.

Randomly amplified polymorphic DNA (RAPD) analysis of Mycoplasma gallisepticum isolates from turkeys from the central valley of California.

Randomly amplified polymorphic DNA (RAPD) analysis was used to investigate the molecular epidemiology of 26 Mycoplasma gallisepticum (MG) isolates obtained from turkeys located in the central valley of California. The MG isolates were recovered from 5 different companies and 13 ranches. Each company had unique MG strains. No evidence of spread of MG between companies was detected. RAPD analysis of MG isolates within a ranch during an outbreak revealed only a single strain involved in each outbreak. RAPD analysis identified an isolate from 1 ranch with a banding pattern identical to that of the 6/85 vaccine strain, which had been used on that particular ranch. Similar RAPD banding patterns of isolates from different ranches within the same company suggested horizontal spread of MG between ranches. The use of 2 primer sets in RAPD analysis was critical to prevent misinterpretation of relationships between different isolates.

Pasteurella multocida infection involving cranial air spaces in White Leghorn chickens.

Seven 18-wk-old pullets from a commercial layer flock experiencing increased mortality associated with neurologic and respiratory symptoms were submitted to the California Veterinary Diagnostic Laboratory System at the Turlock Branch for necropsy. Clinical signs included depression, torticollis, swollen eyelids, conjunctivitis, and sinusitis. Meningoencephalitis and suppurative inflammation of the cranial air spaces were found on histopathology. The brain, sinuses, and air spaces of the cranium were infected with Pasteurella multocida. Complicating the condition was Mycoplasma gallisepticum infecting the sinus and paramyxovirus-I affecting the trachea.

Sticktight fleas associated with fowl pox in a backyard chicken flock in California.

A mixed breed rooster, from a backyard flock of 13 chickens, was received at California Veterinary Diagnostic Laboratory System-Turlock Branch for postmortem examination. The bird presented with thickened, featherless, scab-encrusted skin around the head region. Numerous sticktight fleas were found attached to the encrusted skin. Microscopic evaluation of the skin revealed a lymphoplasmacytic reaction in the dermis with visible embedded flea mouthparts. Also noted histologically in this region were epidermal hyperplasia and ballooned epidermal cells containing intracytoplasmic inclusions indicative of fowl poxvirus.

Salmonella enteritidis, phase type 4 infection in a commercial layer flock in southern California: bacteriologic and epidemiologic findings.

Salmonella enteritidis, phage type 4 (SE PT4), was isolated from five of six 27-wk-old layer chickens submitted for necropsy from a flock of 43,000. Bacteriologic and epidemiologic investigations on the ranch revealed that five of the eight flocks (n = 176,000) were infected. The prevalence of SE PT4 in randomly selected healthy birds ranged from 1.7% (in caged birds) to 50% (in free-range birds) and prevalence in culled birds (kept on dirt floor houses) ranged from 14% to 42%. The estimated overall prevalence of group D Salmonella in eggs contaminated with group D Salmonella was 2.28 per 10,000. The estimated prevalence of group D Salmonella in eggs from caged birds in three infected houses ranged from 1.5 to 4.1 per 10,000, whereas in two houses of free-range birds, prevalence was 14.9 to 19.1 per 10,000. Three of the eight flocks on the ranch remained negative for Salmonella between May 1994 and December 1995 or until removed from the ranch. Salmonella enteritidis PT4 was also isolated from 12.5% (6 of 48) of mice; 57% (four of seven) of cats; and two of two skunks tested. Environmental drag swabs and well water samples yielded multiple serotypes of Salmonella (23/180 and 5/14, respectively) but not S. enteritidis.

Ascaridia dissimilis larval migration associated with enteritis and low market weights in meat turkeys.

A meat turkey ranch had a history of producing flocks with low market weights. Live bird samples and environmental samples from the ranch were collected over a period of eight months and analyzed at the California Veterinary Diagnostic Laboratory System-Turlock Branch. The disease condition consistently present was enteritis associated with Ascaridia dissimilis infection in birds from growout houses. After aggressive control measures for A. dissimilis were implemented, subsequent flocks reached expected market weights.

Esophageal trichomoniasis in chickens.

Esophageal trichomoniasis has been rarely reported in chickens. At the California Veterinary Diagnostic Laboratory System-Turlock Branch, this disease was recently diagnosed in two cases submitted from backyard chicken flocks. The esophageal lesions observed were similar to those seen in several other important diseases of chickens. The causative trichomonad organisms were readily demonstrated on wet smears and by histologic studies. In both cases, the investigated flocks were afflicted with several concurrent diseases. California has experienced an increase in the number of small nontraditional chicken production operations. These facilities are sometimes in close proximity to commercial poultry operations and biosecurity barriers occasionally fail. The poor husbandry practices often used in these small flocks make them a potential reservoir for rare diseases such as trichomoniasis and also for disease organisms that are devastating to commercial poultry.

Efficacy of miconazole nitrate against favus in oriental breed chickens.

Miconazole nitrate 2% was tested for its efficacy against Microsporum gallinae (the causative agent in favus) in a flock of various Oriental breed (Shamo and Aseel) and crossbreed chickens. Six adult males showing clinical signs of favus were randomized into control and experimental groups. The males were maintained on individual tiecords on the range, with no physical contact between birds. The experimental birds had the affected areas washed with soap and water and dried, and an ointment of miconazole nitrate 2% was applied. The experimental birds received the treatment twice a day for 34 days. Scrapings from the affected areas of all birds were cultured at the beginning and end of the test. At the end of the treatment period, the control birds were still positive for M. gallinae, but the organism could not be cultured from the treated birds.

Apoptosis as a cause of death in chicken embryos inoculated with Newcastle disease virus.

Eleven-day-old chicken embryos were inoculated by the allantoic route with the GB strain of Newcastle disease virus (NDV). At 0, 24, 36 and 42 h post-inoculation (p.i.), the brain and heart tissues were harvested for DNA extraction, and the thymus and the brain were fixed and processed for light and electron microscopy. At 42 h p.i., most of the embryos had died; however, no histopathological changes could be seen in the embryos at any stage of infection. DNA extracted from the brain cells started showing fragmentation at 24 h p.i., and from the heart muscle cells at 36 h p.i. Electron microscopy of the brain and thymus showed condensation of the nuclear chromatin, apoptotic bodies, various forms of crescent formation and some evidence of necrosis. Fragmentation of cellular DNA, crescent formation and apoptotic bodies are the typical signs of cells undergoing apoptosis. We suggest that apoptosis of the heart and brain is probably a cause of death of chicken embryos in acute Newcastle disease infection.