Verminous bronchitis and pneumonia by nasal trematodes in Greater Caribbean manatees from Puerto Rico.

Five adult Greater Caribbean manatees Trichechus manatus manatus were found stranded on various coasts of Puerto Rico; 2 stranded alive and 3 stranded dead. Clinical signs observed in live-stranded manatees included emaciation, weakness, bradypnea, arrhythmia, and nasal mucus discharge. Postmortem examinations revealed serosanguinous, mucohemorrhagic, or suppurative exudate in bronchi associated with luminal adult Pulmonicola cochleotrema (range: 18-182 trematodes), accompanied by pulmonary abscesses in 2 cases. Histologically, we observed eosinophilic bronchopneumonia of varying severity (n = 4) and chronic erosive to eosinophilic tracheobronchitis (n = 4) with squamous metaplasia (n = 3) and intralesional trematodes and eggs. The trematode identity was confirmed and compared through molecular analysis for the amplified 18S rDNA fragment. Comorbidities included enteric chiorchosis (n = 5), gastric heterocheilosis (n = 4), malnutrition (n = 4), trauma related to watercraft collision (n = 3), systemic toxoplasmosis (n = 1), acute bacterial peritonitis (n = 1), and interstitial nephritis (n = 1), suggesting that immunosuppression was a predisposing factor for lower respiratory tract pulmonicolosis. Based on lesion severity, clinical signs, and the presence and absence of other findings to explain death, this condition was considered the primary cause of death in 1 manatee, a contributory cause of death in 3 manatees, and an incidental finding in 1 individual. These clinicopathological descriptions will facilitate the diagnosis and clinical management of pulmonicolosis in T. manatus, a species endangered with extinction.

Outcome of patients with cystic fibrosis colonized by Scedosporium and Lomentospora species: A longitudinal cohort study.

Scedosporium and Lomentospora species rank second among the filamentous fungi colonizing the airways of cystic fibrosis (CF) patients. These fungi could be responsible for allergic bronchopulmonary mycosis (ABPM) and bronchitis before lung transplantation and invasive infections after. However, their role in CF lung disease is debated. This study aimed to identify clinical or environmental factors associated with an airway colonization by Scedosporium/Lomentospora species in patients with CF over a period of 7 years. A longitudinal cohort study was conducted from 2008 to 2014 in the CF reference centre in Lyon, France, to compare the characteristics of patients with Scedosporium/Lomentospora colonized and non-colonized patients. During the study period, 283 patients completed the clinical and microbiological follow-up. The analysis revealed that a higher number and duration of hospitalizations, an increased number of courses of parenteral antibiotic therapy, a history of ABPA, and treatment by itraconazole were significantly associated with an airway colonization by Scedosporium/Lomentospora species. The rate of decline of forced expiratory volume in the first second was not statistically different between colonized and non-colonized patients. This study provides evidence that patients colonized by Scedosporium/Lomentospora species require more medical care than non-colonized patients. Additional care could be in part explained by the management of Scedosporium/Lomentospora-related diseases such as ABPM or bronchitis. However, we did not demonstrate a faster rate of decline of respiratory function or body mass index in colonized patients, suggesting, as previously reported, that colonization of the airways by these fungi does not play a significant role in the progression of CF disease.

This prospective study did not demonstrate a faster rate of decline of respiratory function or body mass index in cystic fibrosis (CF) patients colonized by Scedosporium/Lomentospora species compared to non-colonized patients, suggesting that these fungi do not play a significant role in the progression of CF disease.

Pathogenesis of Bohle iridovirus infection in Krefft's freshwater turtle hatchlings (Emydura macquarii krefftii).

Ranaviruses have been detected in over 12 families of reptiles including many genera of turtles, tortoises, and terrapins, but the pathogenesis of these infections is still poorly understood. Krefft's river turtle hatchlings (N = 36; Emydura macquarii krefftii) were inoculated intramuscularly with Bohle iridovirus (BIV, Ranavirus, isolate) or saline, and euthanized at 9 timepoints (3 infected and 1 control per timepoint) over a 24-day period. Samples of lung, liver, kidney, and spleen were collected for quantitative polymerase chain reaction (PCR); internal organs, skin, and oral cavity samples were fixed for histopathological examination. The earliest lesions, at 8 days postinoculation (dpi), were lymphocytic inflammation of the skin and fibrinoid necrosis of regional vessels at the site of inoculation, and mild ulcerative necrosis with lymphocytic and heterophilic inflammation in the oral, nasal, and tongue mucosae. Fibrinonecrotic foci with heterophilic inflammation were detected in spleen and gonads at 16 dpi. Multifocal hepatic necrosis, heterophilic inflammation, and occasional basophilic intracytoplasmic inclusion bodies were observed at 20 dpi, along with ulcerative lymphocytic and heterophilic tracheitis and bronchitis. Tracheitis, bronchitis, and rare bone marrow necrosis were present at 24 dpi. Of the viscera tested for ranaviral DNA by PCR, the liver and spleen had the highest viral loads throughout infection, and thus appeared to be major targets of viral replication. Testing of whole blood by qPCR was the most-effective ante-mortem method for detecting ranaviral infection compared with oral swabs. This study represents the first time-dependent pathogenesis study of a ranaviral infection in turtles.

Comparative trachea transcriptome analysis in SPF broiler chickens infected with avian infectious bronchitis and avian influenza viruses.

Infectious bronchitis virus (IBV) and avian influenza virus (AIV) are two major respiratory infections in chickens. The coinfection of these viruses can cause significant financial losses and severe complications in the poultry industry across the world. To examine transcriptome profile changes during the early stages of infection, differential transcriptional profiles in tracheal tissue of three infected groups (i.e., IBV, AIV, and coinfected) were compared with the control group. Specific-pathogen-free chickens were challenged with Iranian variant-2-like IBV (IS/1494), UT-Barin isolates of H9N2 (A/chicken/Mashhad/UT-Barin/2017), and IBV-AIV coinfection; then, RNA was extracted from tracheal tissue. The Illumina RNA-sequencing (RNA-seq) technique was employed to investigate changes in the Transcriptome. Up- and downregulated differentially expressed genes (DEGs) were detected in the trachea transcriptome of all groups. The Kyoto Encyclopedia of Genes and Genomes pathway and Gene Ontology databases were examined to identify possible relationships between DEGs. In the experimental groups, upregulated genes were higher compared to downregulated genes. A more severe immune response was observed in the coinfected group; further, cytokine-cytokine receptor interaction, RIG-I-like receptor signaling, Toll-like receptor signaling, NOD-like receptor signaling, Janus kinase/signal transducer, and activator of transcription, and apoptotic pathways were important upregulated genes in this group. The findings of this paper may give a better understanding of transcriptome changes in the trachea during the early stages of infection with these viruses.

Impact of feeding n-3 fatty acids to layer breeders and their offspring on concentration of antibody titres against infectious bronchitis, and Newcastle diseases and plasma fatty acids in the offspring.

1. The impact of feeding sources of n-3 fatty acids (FA) to ISA Brown and Shaver White breeders and their offspring on antibody titres and plasma FA profile was examined.2. Breeders were fed either a control diet (CON); CON + 1% microalgae (DMA: Aurantiochytrium limacinum) as a source of docosahexaenoic acid; or CON + 2.6% of a co-extruded mixture of full-fat flaxseed (FFF) as a source of α-linolenic acid. Day-old female offspring were assigned to diets (breeder-offspring): 1) CON-CON, 2) CON-DMA, 3) CON - FFF, 4) DMA - CON, 5) DMA - DMA, 6) FFF - CON or 7) FFF - FFF, followed by a standard layer diet through 18 weeks of age (WOA) to 42 WOA.3. Antibody titres against infectious bronchitis (IBV) and Newcastle disease (NDV) were measured at six days and six WOA, and plasma FA profile was measured at 18 and 42 WOA.4. Pullets from FFF-fed breeders had higher antibody titres against IBV and NDV than pullets fed DMA (P < 0.05). Feeding FFF to offspring increased plasma ∑n-3 FA at 18 and 42 WOA, whereas feeding DMA to offspring reduced ∑n-6 FA at 18 WOA.5. In conclusion, independent of breeder strain, alpha linoleic acid (ALA) and DHA sources showed varied responses. Feeding breeders FFF increased plasma concentration of antibody titres and n-3 FA whereas DMA reduced plasma concentration of ∑n-6 FA.

Development and immunogenic potentials of chitosan-saponin encapsulated DNA vaccine against avian infectious bronchitis coronavirus.

Infectious Bronchitis (IB) is an economically important avian disease that considerably threatens the global poultry industry. This is partly, as a result of its negative consequences on egg production, weight gain as well as mortality rate.The disease is caused by a constantly evolving avian infectious bronchitis virus whose isolates are classified into several serotypes and genotypes that demonstrate little or no cross protection. In order to curb the menace of the disease therefore, broad based vaccines are urgently needed. The aim of this study was to develop a recombinant DNA vaccine candidate for improved protection of avian infectious bronchitis in poultry. Using bioinformatics and molecular cloning procedures, sets of monovalent and bivalent DNA vaccine constructs were developed based on the S1 glycoprotein from classical and variants IBV strains namely, M41 and CR88 respectively. The candidate vaccine was then encapsulated with a chitosan and saponin formulated nanoparticle for enhanced immunogenicity and protective capacity. RT-PCR assay and IFAT were used to confirm the transcriptional and translational expression of the encoded proteins respectively, while ELISA and Flow-cytometry were used to evaluate the immunogenicity of the candidate vaccine following immunization of various SPF chicken groups (A-F). Furthermore, histopathological changes and virus shedding were determined by quantitative realtime PCR assay and lesion scoring procedure respectively following challenge of various subgroups with respective wild-type IBV viruses. Results obtained from this study showed that, groups vaccinated with a bivalent DNA vaccine construct (pBudCR88-S1/M41-S1) had a significant increase in anti-IBV antibodies, CD3+ and CD8+ T-cells responses as compared to non-vaccinated groups. Likewise, the bivalent vaccine candidate significantly decreased the oropharyngeal and cloacal virus shedding (p < 0.05) compared to non-vaccinated control. Chickens immunized with the bivalent vaccine also exhibited milder clinical signs as well as low tracheal and kidney lesion scores following virus challenge when compared to control groups. Collectively, the present study demonstrated that bivalent DNA vaccine co-expressing dual S1 glycoprotein induced strong immune responses capable of protecting chickens against infection with both M41 and CR88 IBV strains. Moreso, it was evident that encapsulation of the vaccine with chitosan-saponin nanoparticle further enhanced immune responses and abrogates the need for multiple booster administration of vaccine. Therefore, the bivalent DNA vaccine could serve as efficient and effective alternative strategy for the control of IB in poultry.

Occlusive mycotic tracheobronchitis and systemic Alphaherpesvirus coinfection in a free-living striped dolphin Stenella coeruleoalba in Italy.

A juvenile female striped dolphin Stenella coeruleoalba live stranded on 4 March 2016 at Alassio, western Ligurian Sea coast, Italy. The dolphin died shortly after stranding, and a complete postmortem examination was performed. Necropsy revealed severe tracheal occlusion and unilateral bronchial stenosis with luminal accumulation of abundant green-yellow mucous-gelatinous material. Histological features suggestive of tracheobronchial aspergillosis were observed. Cultures of lung tissue and tracheo-bronchial exudate isolated Aspergillus fumigatus, identified by a Microseq D2 LSUrDNA fungal sequencing kit. A pan-Herpesvirus nested-PCR assay on frozen samples obtained from multiple organs was positive. Phylogenetic analysis on the partial DNA polymerase gene revealed that the striped dolphin isolate was closely related to known cetacean Alphaherpesvirus sequences from the same host species. Attempted virus isolation was unsuccessful. The tissue levels of different persistent organic pollutants and the toxicological stress, evaluated using a theoretical model, showed a severely impaired immune response. This study reports the first case of occlusive mycotic tracheobronchitis in a free-living cetacean and the first molecular identification of an Alphaherpesvirus in a free-ranging striped dolphin stranded on the coast of Italy.

Protection of chickens vaccinated with combinations of commercial live infectious bronchitis vaccines containing Massachusetts, Dutch and QX-like serotypes against challenge with virulent infectious bronchitis viruses 793B and IS/1494/06 Israel variant 2.

Infectious bronchitis virus (IBV) is a coronavirus which affects chickens of all ages. IBV mainly causes respiratory disease but can also result in reduced weight gain, reduced egg production, increased frequency of abnormal eggs and increased rates of mortality. Vaccination is the most important way to control the disease. Nevertheless, novel strains of infectious bronchitis (IB) continue to emerge in the field. In order to respond promptly, combinations of existing IB vaccines are frequently tested to see whether they can provide cross-protection. The efficacy of a combination of vaccines based on Massachusetts, Dutch and QX-like IB strains against emerging IB Israel variant 2 and IB 793B strains was assessed by means of four challenge studies. At least 80% of the birds vaccinated with IB H120 (Mass type) combined with IB D274 (Dutch type) followed by a QX-like IB vaccine booster or vaccinated with a combination of IB H120, IB D274 and QX-like IB were protected against a challenge with IB 793B. In addition, IB 1263 (Mass type) boosted by QX-like IB showed an 85% protection following challenge with IB 793B. A combination of IB H120 and IB D274 boosted by QX-like IB vaccine conferred 70% protection whilst H120 and IB D274 combination on its own showed 61.1% protection against Israel variant 2 challenge. IB 1263 boosted by a QX-like IB vaccine showed 50% protection against IB Israel variant 2. Therefore, it can be concluded that a combination of the IB H120, IB D274 and QX-like IB confers broad protection against different non-related virulent IB strains.

Infectious bronchitis vaccine virus detection and part-S1 genetic variation following single or dual inoculation in broiler chicks.

An investigation was undertaken of the extent of genetic variation occurring within infectious bronchitis virus (IBV) vaccine strains following vaccination of day-old broiler chicks. Chicks were divided into seven groups, with two groups receiving single Massachusetts (Mass) vaccinations while the other four were inoculated with combinations of different IBV serotypes; Mass, 793B, D274 and Arkansas (Ark). The remaining group was maintained as an unvaccinated control. Following vaccination, swabs and tissues collected at intervals were pooled and RNA was extracted for detection of IBV by reverse transcription polymerase chain reaction. Positive amplicons were sequenced for the part-S1 gene and compared to the original vaccine strain sequences. Single nucleotide polymorphisms, amino acid variations and hydrophobicity changes were identified and recorded for each sampling point. A total of 106 single nucleotide polymorphisms were detected within 28 isolates. The average single nucleotide polymorphism counts of swab isolates were greater than those found in tissue samples. This translated into 64 amino acid changes; however only six resulted in a change to the hydrophobicity properties. All hydrophobic alterations occurred within swab isolates and the majority were recovered at 3 days post vaccination suggesting such changes to be detrimental to early virus survival. Nucleotide deletions were seen only in the group given the combination of Mass and Ark. Of the 16 sequenced samples in this group, 13 contained the same AAT deletion at position 1033 1035 in the Ark strains. Findings presented in this study demonstrate alteration in the S1 nucleotide sequence following co-administration of live IBV vaccines.

Comparative proteome analysis of tracheal tissues in response to infectious bronchitis coronavirus, Newcastle disease virus, and avian influenza virus H9 subtype virus infection.

Infectious bronchitis coronavirus (IBV), Newcastle disease virus (NDV), and avian influenza virus (AIV) H9 subtype are major pathogens of chickens causing serious respiratory tract disease and heavy economic losses. To better understand the replication features of these viruses in their target organs and molecular pathogenesis of these different viruses, comparative proteomic analysis was performed to investigate the proteome changes of primary target organ during IBV, NDV, and AIV H9 infections, using 2D-DIGE followed MALDI-TOF/TOF-MS. In total, 44, 39, 41, 48, and 38 proteins were identified in the tracheal tissues of the chickens inoculated with IBV (ck/CH/LDL/97I, H120), NDV (La Sota), and AIV H9, and between ck/CH/LDL/97I and H120, respectively. Bioinformatics analysis showed that IBV, NDV, and AIV H9 induced similar core host responses involved in biosynthetic, catabolic, metabolic, signal transduction, transport, cytoskeleton organization, macromolecular complex assembly, cell death, response to stress, and immune system process. Comparative analysis of host response induced by different viruses indicated differences in protein expression changes induced by IBV, NDV, and AIV H9 may be responsible for the specific pathogenesis of these different viruses. Our result reveals specific host response to IBV, NDV, and AIVH9 infections and provides insights into the distinct pathogenic mechanisms of these avian respiratory viruses.

Bovine Rhinitis B Virus Variant as the Putative Cause of Bronchitis in Goat Kids.

A diagnostic investigation into an outbreak of fatal respiratory disease among young goats in Iowa, USA revealed bronchitis lesions of unknown etiology and secondary bacterial bronchopneumonia. Hypothesis-free metagenomics identified a previously unreported picornavirus (USA/IA26017/2023), and further phylogenetic analysis classified USA/IA26017/2023 as an aphthovirus related to bovine rhinitis B virus. Viral nucleic acid was localized to lesions of bronchitis using in situ hybridization. This marks the first report of a picornavirus putatively causing respiratory disease in goats and highlights the potential for cross-species transmission of aphthoviruses.

Severe bronchiectasis resulting from chronic bacterial bronchitis and bronchopneumonia in a jungle cat.

Bronchiectasis is irreversible bronchial dilation that can be congenital or acquired secondary to chronic airway obstruction. Feline bronchiectasis is rare and, to our knowledge, has not been reported previously in a non-domestic felid. An ~10-y-old female jungle cat (Felis chaus) was presented for evaluation of an abdominal mass and suspected pulmonary metastasis. The animal died during exploratory laparotomy and was submitted for postmortem examination. Gross examination revealed consolidation of the left caudal lung lobe and hila of the cranial lung lobes. Elsewhere in the lungs were several pale-yellow pleural foci of endogenous lipid pneumonia. On cut section, there was severe distension of bronchi with abundant white mucoid fluid. The remaining lung lobes were multifocally expanded by marginal emphysema. Histologically, ectatic bronchi, bronchioles, and fewer alveoli contained degenerate neutrophils, fibrin, and mucin (suppurative bronchopneumonia) with rare gram-negative bacteria. Aerobic culture yielded low growth of Proteus mirabilis and Escherichia coli. There was chronic bronchitis, marked by moderate bronchial gland hyperplasia, lymphoplasmacytic inflammation, and lymphoid hyperplasia. The palpated abdominal mass was a uterine endometrial polyp, which was considered an incidental, but novel, finding. Chronic bronchitis and bronchopneumonia should be considered as a cause of bronchiectasis and a differential diagnosis for respiratory disease in non-domestic felids.

[Evaluierung der Langzeittherapie bei Katzen mit felinem Asthma und chronischer Bronchitis]. / Evaluation of long-term therapy in cats with feline asthma and chronic bronchitis.

GEGENSTAND UND ZIEL: Felines Asthma (FA) und chronische Bronchitis (CB) sind häufige entzündliche Erkrankungen der Atemwege der Katze. Obwohl beide Krankheitsbilder durch eine Infiltration mit unterschiedlichen Entzündungszelltypen gekennzeichnet sind, sind die therapeutischen Maßnahmen oft ähnlich. Über mögliche Unterschiede im therapeutischen Management dieser beiden Atemwegserkrankungen ist wenig bekannt. Ziel der Studie war es daher, bei Katzen mit FA und CB die Erst- und Langzeitbehandlung, Therapieerfolg, Nebenwirkungen und Besitzerzufriedenheit zu vergleichen. MATERIAL UND METHODEN: 35 Katzen mit FA und 11 Katzen mit CB wurden in die retrospektive Querschnittstudie eingeschlossen. Einschlusskriterien waren kompatible klinische und radiologische Befunde sowie der zytologische Nachweis einer eosinophilen Entzündung (FA) oder einer sterilen neutrophilen Entzündung (CB) in der bronchoalveolären Lavage-Flüssigkeit (BALF). Katzen mit CB wurden ausgeschlossen, wenn Hinweise auf pathologische Bakterien vorlagen. Besitzer wurden gebeten einen standardisierten Fragebogen zum therapeutischen Management und Ansprechen auf die Behandlung auszufüllen. ERGEBNISSE: Im Gruppenvergleich wurden keine statistisch signifikanten Unterschiede der Therapie festgestellt. Die meisten Katzen wurden anfänglich mit Kortikosteroiden mittels einer oralen (FA 63%/CB 64%, p=1), inhalativen (FA 34%/CB 55%, p=0,296) oder injizierbaren Applikationsform (FA 20%/CB 0%, p=0,171) behandelt. Zusätzlich wurden in einigen Fällen orale Bronchodilatatoren (FA 43%/CB 45%, p=1) und Antibiotika (FA 20%/CB 27%, p=0,682) verabreicht. In der Langzeittherapie erhielten 43% der Katzen mit FA und 36% der Katzen mit CB inhalative Kortikosteroide (p=1), orale Kortikosteroide (FA 17%/CB 36%, p=0,220) und orale Bronchodilatatoren (FA 6%/CB 27%, p=0,084) sowie phasenweise Antibiotika (FA 6%/CB 18%, p=0,238). Behandlungsbedingte Nebenwirkungen (Polyurie/Polydipsie, Pilzinfektion im Gesicht und Diabetes mellitus) wurden bei 4 Katzen mit FA und 2 Katzen mit CB registriert. Die Mehrheit der Besitzer gab an, mit dem Ansprechen auf die Behandlung äußerst oder sehr zufrieden zu sein (FA 57%/CB 64%, p=1). SCHLUSSFOLGERUNG: Signifikante Unterschiede hinsichtlich des Managements und des Therapieansprechens konnten bei beiden Erkrankungen laut Besitzerbefragung nicht festgestellt werden. KLINISCHE RELEVANZ: Laut Besitzerumfrage können chronische Bronchialerkrankungen der Katze wie Asthma und chronische Bronchitis können mit einer ähnlichen Behandlungsstrategie erfolgreich therapiert werden.

Efficacy of a plant-produced infectious bronchitis virus-like particle vaccine in specific pathogen-free chickens.

Infectious bronchitis (IB) Gammacoronavirus causes a highly contagious respiratory disease in chickens that is listed by the World Organisation for Animal Health (WOAH). Its high mutation ability has resulted in numerous variants against which the commercially available live or recombinant vaccines singly offer limited protection. Agrobacterium-mediated transient expression in Nicotiana benthamiana (tobacco) plants was used here to produce a virus-like particle (VLP) vaccine expressing a modified full-length IBV spike (S) protein of a QX-like IB variant. In a challenge study with the homologous live IB QX-like virus, VLP-vaccinated birds produced S protein-specific antibodies comparable to those produced by live-vaccinated birds seroconverting with mean geometric titers of 6.8 and 7.2 log2, respectively. The VLP-vaccinated birds had reduced oropharyngeal and cloacal viral shedding compared to an unvaccinated challenged control and were more protected against tracheal ciliostasis than the live-vaccinated birds. While the results appeared similar, plant-produced IB VLPs are safer, more affordable, easier to produce and update to antigenically match any emerging IB variant, making them a more suitable alternative to IBV control than live-attenuated vaccines.

Correlation of clinical and radiographic variables in cats with lower airway disease.

BACKGROUND: Feline lower airway disease (FLAD) is frequently associated with radiographic abnormalities. OBJECTIVES: To evaluate whether radiographic changes in cats with naturally occurring FLAD improve with treatment and if radiographic changes correlate with clinical signs. ANIMALS: Twenty-four client-owned cats newly diagnosed with FLAD, based on medical history, typical clinical signs, radiographic findings, and examination of bronchoalveolar lavage fluid, were included in the prospective study. METHODS: At 2 examination time points (days 0 and 60), an owner questionnaire, clinical examination, and thoracic radiography were carried out. Information from the questionnaire and clinical examination were evaluated on the basis of a 12-point clinical score. Radiographs were assessed using a 10-point radiographic score. Individual treatment was given to all cats over the study period, based on severity of the disease and compliance of the cat. Clinical and radiographic scores were compared statistically for both examination time points and evaluated for correlation. RESULTS: All cats showed radiographic abnormalities at initial presentation. In addition to significant improvement in clinical variables, the total radiographic score improved significantly (P = .01) during the study period, with significant improvement in the severity of bronchial (P = .01) and interstitial lung pattern (P = .04). Improvement of the clinical and radiographic score was not correlated. CONCLUSION AND CLINICAL IMPORTANCE: In addition to clinical signs, repeated radiographic examination can be used as a diagnostic tool to evaluate treatment response in cats with FLAD.

The use of RT-PCR in the diagnosis and differentiation of vaccine strains of chicken infectious bronchitis and Newcastle disease.

Background: Infectious diseases of young and adult birds with respiratory syndrome are a significant deterrent to the development of industrial poultry farming due to decreased productivity and significant mortality. The only effective method of combating viral diseases is timely and targeted vaccination, which largely depends on laboratory diagnostic results. Aim: This article aims to study the real-time reverse transcription polymerase chain reaction, (RT-PCR) which has the prospect of more effective diagnosis of vaccine strains of chicken infectious bronchitis and Newcastle disease. Methods: The fastest and most accurate method for the differential diagnosis of pathogens in an associative viral infection is RT-PCR. The method proposed in the article for selecting primers for amplification made it possible to use this method for the simultaneous interspecies differential diagnosis of two or more viral agents, significantly accelerating their diagnosis. Results: The correlation of the nucleotide sequence obtained from sequencing to a specific virus strain is complicated by the lack of a single nomenclature mechanism for separating genetic groups. Conclusion: The results of this study will allow easy and fast typing of sequences into known and databased virus strains and avoid further confusion in the nomenclature of genetic groups in the future.

Pathologic lesions caused by coinfection of Mycoplasma gallisepticum and H3N8 low pathogenic avian influenza virus in chickens.

Chickens were infected under experimental conditions with Mycoplasma gallisepticum and low pathogenic avian influenza (LPAI) strain A/mallard/Hungary/19616/07 (H3N8). Two groups of chickens were aerosol challenged with M. gallisepticum strain 1226. Seven days later, one of these groups and one mycoplasma-free group was challenged with LPAI H3N8 virus; one group without challenge remained as negative control. Eight days later, the birds were euthanized and examined for gross pathologic and histologic lesions. The body weight was measured, and the presence of antimycoplasma and antiviral antibodies was tested before the mycoplasma challenge, before the virus challenge, and at the end of the study to confirm both infections. Chickens in the mycoplasma-infected group developed antibodies against M. gallisepticum but not against the influenza virus. Chickens of the group infected with the influenza virus became serologically positive only against the virus, while the birds in the coinfected group developed antibodies against both agents. The LPAI H3N8 virus strain did not cause decrease in body weight and clinical signs, and macroscopic pathological lesions were not present in the chickens. The M. gallisepticum infection caused respiratory signs, airsacculitis, and peritonitis characteristic of mycoplasma infection. However, the clinical signs and pathologic lesions and the reduction in weight gain were much more significant in the group challenged with both M. gallisepticum and LPAI H3N8 virus than in the group challenged with M. gallisepticum alone.

Modulation of the innate immune system by lipopolysaccharide in the proventriculus of chicks inoculated with or without Newcastle disease and infectious bronchitis vaccine.

This study aimed to determine whether the innate immune system in the proventriculus of broiler chicks responds to lipopolysaccharide (LPS) and whether this response is affected by Newcastle disease and infectious bronchitis (ND/IB) vaccination. Chicks were divided into 4 groups: nonvaccinated and injected with PBS or LPS (V-L- and V-L+), and vaccinated and injected with PBS or LPS (V+L- and V+L+). Vaccination was performed on d 1, and LPS was intraperitoneally injected on d 11 of age. The gene expression and protein levels of immune molecules, including toll-like receptors (TLRs), antimicrobial peptides, interleukin-1ß (IL-1B), and immunoglobulin A (IgA) in the proventriculus and serum were analyzed. The results showed that the expression levels of TLR21 were higher in vaccinated (V+L-) group than in nonvaccinated (V-L-) group. Gene expression levels of avian ß-defensin (AvBDs) and cathelicidin1 (Cath1) were not different among the 4 groups. However, the results of LC/MS analysis showed that the levels of AvBD2, 6, and 7 significantly increased after the LPS challenge in nonvaccinated and vaccinated chicks; the levels were higher in V-L+ and V+L+ than in V-L- and V+L-, respectively. Immunohistochemistry analysis revealed the localization of AvBD1 protein in the epithelial cells of the surface glands and AvBD2 and CATH1 in the heterophil-like cells in the lamina propria of surface glands. Although IL-1B gene expression and protein concentration in the proventriculus tissues were not different among the 4 groups, serum IL-1B levels were upregulated by LPS in both the nonvaccinated and vaccinated groups (V-L- vs. V-L+, V+L- vs. V+L+). Moreover, IgA levels in the proventriculus and serum were not affected by vaccination or LPS challenge. Taken together, we conclude that LPS derived from gram-negative bacteria upregulates the innate immune system, including antimicrobial peptide synthesis in the proventriculus. ND/IB vaccination may not significantly affect antimicrobial peptide synthesis in response to LPS; however, TLR21 expression is upregulated by that vaccination. The antimicrobial peptides synthesized in the proventriculus probably prevent pathogenic microbes from entering the intestine.

Evaluation of Cholera Toxin Adjuvanticity Effect on the Production of Specific Antibodies Induced by Avian Infectious Bronchitis Vaccine in Chickens.

Cholera toxin (CT) is one of the most well-known immunostimulants. Mammalian studies have shown that CT can generate immune responses against antigen. However, it has not exhibited a definite effect on poultry yet. In this study, focusing on a cost-effective method, the effect of different concentrations of CT obtained from Vibrio cholerae biotype El Tor and serotype Inaba was investigated on the immunogenicity of infectious bronchitis vaccine. After culturing and concentrating CT, different concentrations of CT (0.1, 1, 2, and 5 micrograms) were combined with avian infectious bronchitis vaccine strain H120 produced by Razi Vaccine and Serum Research Institute (RVSRI) and, at 7 days of age, inoculated via the eye drop administration in 42 specific-pathogen-free chickens (seven groups of six chicks that included four experimental groups, two negative control groups (PBS and toxin), and one positive control group). Blood samples were taken weekly from the wing veins of the chickens, and the immunoglobulin G (IgG) titer was checked by enzyme-linked immunosorbent assay. The results showed that 2 µg of CT in comparison with other concentrations caused a significant increase in the antibody titer against avian infectious bronchitis in the blood serums of the chickens. One-way ANOVA test showed that all the results of this study were significant at P<0.05 level. Our data show that CT has the potential to further stimulate the immune system of chickens and may increase the immunogenicity of the infectious bronchitis vaccine. However, more research is needed to examine all aspects of the use of this toxin in animal vaccines.

The Pathology of Infectious Bronchitis.

Infectious bronchitis (IB) is an acute disease of chickens caused by a gammacoronavirus, infectious bronchitis virus (IBV). Infection of the nasal and tracheal mucosa causes a rapid loss of ciliated epithelium and impaired mucociliary clearance that predispose chickens to secondary bacterial infections. In poultry production, disease progression and severity are influenced by other live virus vaccines, immunosuppression, and coexisting pathogens. The digestive tract supports viral replication in the proventriculus, intestines, cloaca, and the bursa of Fabricius. Acute enteritis and stunted growth in young chickens are caused by an enterotropic IBV. IBV spreads systemically by infection of tracheal macrophages and blood monocytes, deep respiratory infections, and potentially ascending viral infection from the cloaca. Nephrotropic IBV causes severe disease in the kidney with necrosis of tubular epithelial cells, inflammation, and renal failure. Viral infection of the female reproductive tract in the first 2 weeks of life causes necrosis and scarring of the oviduct mucosa, resulting in a chronic cystic oviduct that precludes egg formation when the hen matures. Virus infection of mature hens causes necrosis and inflammation of the oviduct mucosa, leading to the deterioration of egg quality and transient interruption of egg production. In males, IBV infection of seminiferous tubules in the testicle and efferent ductules in the epididymis results in epididymitis and epididymal lithiasis, decreases in sperm production and fertility, and viral shed to semen, leading to venereal transmission. The role IBV in gastrointestinal and urogenital disease merits further study.

Estudio recapitulativo- La patología de la bronquitis infecciosa: una revisión La bronquitis infecciosa es una enfermedad aguda de los pollos causada por un gammacoronavirus, el virus de la bronquitis infecciosa (IBV). La infección de la mucosa nasal y traqueal provoca una pérdida rápida del epitelio ciliado y la alteración de la función mucociliar que predispone a los pollos a infecciones bacterianas secundarias. En la producción avícola, el desarrollo y la severidad de la enfermedad están influenciadas por otras vacunas con virus vivos, inmunosupresión y patógenos coexistentes. En el tracto digestivo se lleva a cabo la replicación viral en el proventrículo, los intestinos, la cloaca y en la bolsa de Fabricio. La enteritis aguda y el retraso del crecimiento en pollos jóvenes son causados por virus de bronquitis infecciosa enterotrópicos. El virus de la bronquitis infecciosa se propaga sistémicamente por infección de macrófagos traqueales y monocitos sanguíneos, infecciones respiratorias profundas e infección viral potencialmente ascendente desde la cloaca. Los virus de la bronquitis infecciosa nefrotrópicos causan una enfermedad severa en el riñón con necrosis de las células epiteliales tubulares, inflamación e insuficiencia renal. La infección viral del aparato reproductor femenino en las primeras dos semanas de vida causa necrosis y cicatrización de la mucosa del oviducto, lo que resulta en un oviducto quístico crónico que impide la formación de huevos cuando la gallina llega a la madurez. La infección por el virus en gallinas maduras causa necrosis e inflamación de la mucosa del oviducto, lo que conduce al deterioro de la calidad del huevo y la interrupción transitoria de la producción de huevos. En los machos, la infección por bronquitis infecciosa de los túbulos seminíferos en el testículo y los conductos eferentes en el epidídimo da como resultado epididimitis y litiasis epididimaria, disminución de la producción y fertilidad de espermatozoides, y la diseminación viral al semen, lo que lleva a la transmisión venérea. El papel del virus de la bronquitis en las enfermedades gastrointestinales y urogenitales merece un estudio más a fondo.