Sequencing of pigeon circovirus and the first report of identification of beak and feather disease virus in clinical specimens of domestic pigeons.

An internationally recognized syndrome that leads to deaths among domestic and ornamental pigeons, particularly after racing, is young pigeon disease syndrome (YPDS). Pigeon circovirus (PiCV) is regarded as one of the potential factors contributing to the occurrence of YPDS. This survey was conducted to determine the prevalence of PiCV infection and molecularly characterize the PiCV in pigeons suspected of YPDS. Eighty fecal samples were collected from 80 diseased pigeons (exhibiting symptoms such as lethargy, weight loss, crop stasis, vomiting and diarrhea) from 20 lofts in different areas of Ahvaz, Iran. Also, 20 fecal samples were obtained from 20 clinically healthy pigeons. The nested broad spectrum polymerase chain reaction test was done to identify the circovirus, using primers targeting part of the replication-associated protein gene with 350 bp, and several positive samples were sequenced. This study showed that PiCV was detected in 86 out of the 100 samples (86.00%). Two types of circoviruses were determined in the samples. One type of the detected circoviruses was PiCV which based on phylogenetic analysis had high genetic similarity with A, B, G and H genotypes of PiCV. The other type of detected circoviruses was closely related to beak and feather disease virus (BFDV) which causes one of the most significant viral diseases in psittacine birds. This is the first report of BFDV identification in pigeons.

Phylogenetic analysis of pathogenic Candida spp. in domestic pigeons.

The current study was conducted to survey the prevalence of pigeon candidiasis in diseased pigeons suspected to candidiasis by isolation, microscopic examination, and polymerase chain reaction (PCR) method and to characterize Candida spp. phylogenetically. For this purpose, samples were obtained from 100 suspected pigeons from September 2018 to February 2019 in Ahvaz, Iran. Cloacal and oropharyngeal swab samples were collected from each diseased pigeon with diarrhea resistant to the antibiotics, crop stasis, white diphtheritic membrane in the mouth, regurgitation, and vomiting. Sabouraud dextrose agar was used as a culture medium. Selected colonies were stained with lactophenol cotton blue stain. In the culture and direct microscopic observation, 19.00% of birds were suspected to candidiasis. Twenty-two isolates were identified. All 22 isolates were confirmed as Candida spp. By PCR method. The PCR test confirmed the presence of Candida spp. in 19.00% of pigeons. Based on the sequencing results of some PCR products, the isolates belonged to Candida albicans and Candida glabrata. The results revealed a 99.78% accordance when compared with other sequences of C. albicans which were formerly deposited in GenBank® from Colombia, Indonesia, China, and Sudan. The results revealed a 99.54% accordance when compared with other sequences of C. glabrata which were formerly deposited in GenBank® from the Netherlands and Spain. The symptoms such as diarrhea resistant to antibiotics, crop stasis, white diphtheritic membrane in the mouth, regurgitation, and vomiting were the most prevalent clinical symptoms in positive pigeons.

Phylogenetic analysis of pigeon adenovirus 1 in clinical specimens of domestic pigeons (Columba livia domestica) in Iran.

An internationally identified syndrome that leads to deaths between domestic and ornamental pigeons, particularly after racing is young pigeon disease syndrome (YPDS). This study was conducted to determine the status of pigeon adenoviral infection and molecularly characterize the pigeon adenovirus in Ahvaz pigeons. Sixty stool samples of healthy pigeons (young pigeons and adult pigeons) and 60 stool samples of diseased pigeons (young and adults) with symptoms of lethargy, weight loss, crop stasis, vomiting and diarrhea were examined. Samples were screened for aviadenoviruses by polymerase chain reaction (PCR) assay and degenerated primers set to target the aviadenovirus polymerase (pol) gene were used which was designed in this study. Screening for pigeon adenovirus 1 (PiAdV-1) was performed using a primer pair that targeted the fiber gene of PiAdV-1. Out of 120 stool samples, six samples (5.00%) were positive for aviadenovirus. The results showed that independent from pigeons' age status, 5.00 and 3.33% of sick and of healthy pigeons were positive for PiAdV-1, respectively. Genomic sequencing revealed that the viruses detected in Ahvaz pigeons belonged to the PiAdV-1 genotype. The results in pigeons revealed a 98.10 - 99.53% nucleotide similarity when compared to other strains of PiAdV-1 (TR/SKPA20, P18-05523-6 and strain IDA4) formerly deposited in GenBank® in Türkiye, Australia and The Netherlands. As far as the authors know, this was the first record of phylogenetic analysis of PiAdV-1 in Iran.

Avian gastric yeast (macrorhabdosis) in cockatiel, budgerigar and grey parrot: a focus on the clinical signs, molecular detection and phylogenetic evaluation.

Macrorhabdus ornithogaster is a microorganism that causes nonspecific and general clinical symptoms and to this day, diagnosis and also treatment have been yet hard. The present study was conducted to survey the prevalence of macrorhabdosis and to characterize M. ornithogaster phylogenetically in Psittaciformes suspected of macrorhabdosis from January 2018 to May 2019 in Ahvaz, Iran. For this purpose, fecal samples were collected from Psittaciformes with signs of the disease. Wet mounts were prepared from fecal samples and examined carefully using a light microscope. Samples from parrots with gastrointestinal symptoms of the disease were chosen for molecular diagnosis of the organism and DNA was extracted from these samples. For detection of M. ornithogaster, primer sets (BIG1, Sm4) and (AGY1, Sm4) which target the 18S rDNA gene were selected and Semi-nested polymerase chain reaction (Semi-nested PCR) was performed. The PCR method confirmed the presence of M. ornithogaster in 14.00% of the samples. Purified PCR products were sequenced for more accurate confirmation and according to the gene sequence all sequences were owned by M. ornithogaster. The results disclosed a 96.03 - 100% identity when compared to other sequences of M. ornithogaster which had previously been deposited in the GenBank® from Germany and the USA. The results of this study proved the circulation of M. ornithogaster between cockatiel, budgerigar and grey parrot. The prevalence of macrorhabdosis was higher in cockatiel compared to budgerigar and grey parrot. As far as the authors know, this was the first record of macrorhabdosis in African grey parrots.

Candidiasis in Birds (Galliformes, Anseriformes, Psittaciformes, Passeriformes, and Columbiformes): A Focus on Antifungal Susceptibility Pattern of Candida albicans and Non-albicans Isolates in Avian Clinical Specimens.

Candidiasis is a fungal infection caused by Candida species which has been reported in most domestic and wild birds and mammals. In this study, 196 samples from different species of birds with suspected symptoms of candidiasis were examined. Pharyngeal swabs, cloacal swabs, and fecal samples were taken from the birds. The samples were cultured in sabouraud dextrose agar (SDA) containing cycloheximide and chloramphenicol and incubated at 42°C. Suspected isolates of Candida were identified using PCR. To detect the candida genus, a primer set to target the candida rDNA (ITS1-ITS4) was selected. To detect Candida albicans (C albicans), a primer set to target cytochrome P-450 lanosterol-a-demethylase (P450-LIAl) gene (DH-1558) was selected. In direct microscopic observation and culture, 28.57% of the birds were suspected of candidiasis. In the molecular study, candidiasis was confirmed in 25% of the birds, and candidiasis caused by C albicans was confirmed in 14.28% of the birds. All isolates were subjected to antibiotic susceptibility by the disk diffusion method with glucose-enriched Mueller-Hinton Agar. 78.5% of the isolates were sensitive to nystatin and amphotericin B. None of the isolates were sensitive to itraconazole and more than 50% of the isolates were resistant to fluconazole, ketoconazole, and itraconazole. According to the results, it is suggested to use nystatin and amphotericin B in the treatment of avian candidiasis in the Ahvaz region. To the authors' knowledge, this is the first report of the molecular detection and antifungal susceptibility pattern of C albicans and non- albicans from Galliformes, Anseriformes, Psittaciformes, and Passeriformes in Iran.

Isolation, molecular identification, and phylogenetic evaluation of Cryptococcus neoformans isolated from pigeon lofts, Psittaciformes, and Passeriformes in Ahvaz, Iran.

Cryptococcus neoformans, the main pathogen in immunocompromised patients, is a ubiquitous free-living fungus that can be isolated from avian excreta, soils, and plant material. This study was carried out to determine the infection rate of pigeon lofts, Passeriformes, and Psittaciformes in Ahvaz, the capital of Khuzestan province in Iran and to determine varieties of Cryptococcus neoformans (C. neoformans). The 80 samples were collected from pigeon lofts. Also, 163 feces of captive birds (Passeriformes and Psittaciformes) which kept in Ahvaz pet shops, and the 70 cloacal swabs of pet birds (Passeriformes and psittaciformes) referring to the department of avian medicine (the faculty of veterinary medicine of Shahid Chamran University of Ahvaz) were analyzed. The samples were directly inoculated on niger seed agar (NSA) and also enriched in brain heart infusion broth and then inoculated on NSA. Dark brown colonies suspected to C. neoformans subcultured on saborouds dextrose agar and pure cultures subjected to molecular (polymerase chain reaction (PCR)) diagnosis. For detection of C. neoformans, primer sets that targeting the CNLAC1 gene were selected and nested PCR was conducted. For identification of C. neoformans varieties, a primer set targeting the STR1 gene was selected. For more accurate confirmation, the purified PCR products of some isolates were also sequenced, and based on the gene sequences, all of the isolates belonged to C. neoformans variety grubii (var. grubii)(serotype A). Totally 16 out of 80 pigeon samples (20%) were contaminated with C. neoformans. The results in pigeons disclosed a 98.64% identity when compared with other strains of C. neoformans (CN1525, T4, and T1) which were previously deposited in GenBank from Italy and Thailand. Also, 21 out of 233 samples from Psittaciformes (9.01%) were contaminated with C. neoformans. The results in Psittaciformes disclosed a 99.7% identity when compared with other strains of C. neoformans (TIMM1313, IFM5882, CN1525, etc.) which were previously deposited in GenBank from Japan and Italy, etc. In the present study, the samples belonging to the passerine order were free of C. neoformans infection. According to the results, C. neoformans is prevalent in pigeon flocks and pet birds including Psittaciformes in the Ahvaz area, and should be considered by pigeon and captive bird breeders, veterinarians, and public health organizations in Ahvaz. The cryptococcus species isolated from captive birds and pigeons could be potential pathogens in humans.

Isolation, identification and antimicrobial sensitivity of Ornithobacterium rhinotracheale in broilers chicken flocks of Khuzestan, Iran.

Ornithobacterium rhinotracheale (ORT) is a bacterium associated with respiratory disease, growth retardation, decreased egg production and mortality in chickens and turkeys. The objective of this study was isolation, identification and evaluation of antimicrobial susceptibility of ORT bacterium in slaughtered broilers chicken flocks based on cultural and molecular tests in Khuzestan province, south-west of Iran. A total of 210 tracheal swab samples were collected from 21 broiler flocks slaughtered in abattoirs of the province. The results of cultural and biochemical tests showed that 23 (10.95%) isolates from tracheal swabs of 4 flocks (19.04%) were identified as ORT, but according to molecular characterization, 18 (8.57%) ORT isolates were positive in PCR assay and produced the predicted 784 bp amplification product. Finally, using the disk diffusion method, the drug resistance patterns of ORT isolates were determined against a panel of commonly used antimicrobial agents. Antimicrobial susceptibility test revealed that all isolates (100%) were sensitive to tetracycline, florfenicol and cephalexin. The highest antimicrobial resistance (89.00%) was seen for fosfomycin, sultrim and gentamicin. The results of present research showed that there was significant difference between the isolation rates of ORT from various areas of the province. As well, our findings indicated that the simultaneous use of both cultural and molecular techniques results in more comprehensive outcomes in the isolation and identification of the organismfrom understudy hosts.