Prevalence of Avian Gastric Yeast ( Macrorhabdus ornithogaster ) in Parrots and Parakeets: A Case Study

, green cheeks, and budgerigars with symptoms, such as lethargy, weight loss, regurgitation, and gasping referred to the Aria Veterinary Hospital, Mashhad, Iran, during 2021-2022. The samples were randomly evaluated for gastric yeast (Megabacter) using the gram-staining method of their feces. Conclusion: The results revealed that almost half of the symptomatic cases showed active or non-active yeast in their slides. A 14-day treatment of nystatin, apple vinegar, and metronidazole was applied for all cases, and then gram-staining slides were tested after three weeks showing that the treatment was successful. However, infection recurrence was detected in several birds, suggesting food hygiene control.


Introduction
Avian gastric yeast (Macroohabdus ornithogaster), which is more common in smaller companion birds like parrotlets, lovebirds, finches, cockatiels, and budgerigars, often colonizes the digestive tract of birds. A weakened immune system seems to be the most risk factor for the birds to be infected 1 .
The most common sign of infection is believed to be chronic weight loss, lethargy, regurgitation with or without undigested seeds or pellets in the droppings, and diarrhea. Therefore, the droppings can return to a proper environment for both relapses and potential shedding of the infection 2 .
The disease can be diagnosed by examining the droppings and the feces under the microscope. Both the gram-staining method and wet slides aid the diagnosis by positive identification of the organisms. As the organisms could be shed sporadically and in low numbers, repeated evaluation may be necessary 1 .
Controlling and treatment of avian gastric yeast are always difficult for clinicians, as subclinical infections can occur in many birds. Amphotericin B is the choice medication for treatment, as many antimicrobial and antifungal medications are ineffective against M. ornithogaster. It is recommended orally twice a day for 30 days at a dose of 100 mg/kg 3 . Other antibacterial and antifungal treatments, such as metronidazole and fluconazole are also suggested. As the organisms develop in an alkaline environment, oral administration of apple vinegar or grapefruit juice can be used to increase the proventricular activity. For prevention, good hygiene, like proper cleaning and disinfecting of the nest box, cages, and any feeding utensils, and providing highquality feed is crucial to minimize the infection and preserve the other birds 4 .

Materials and Methods
A total of 145 companion birds, including 50 cockatiels, 20 gray parrots, 20 Budgerigars, 5 parrots, 30 love birds, and 20 green cheeks, which had symptoms of lethargy, weight loss, regurgitation and gasping, were referred to Aria veterinary hospital, Mashhad, Iran. The birds were evaluated for avian gastric yeast (M. ornithogaster) by gram-staining of their feces during 2021-2022. After fluid therapy based on the dehydration and the weight of the birds, the fecal samples were collected and referred to the laboratory of Arya veterinary hospital clinical pathology, Mashhad, Iran. The samples were stained and checked with 100x lens (Olympious, Japan). A 14day medication, including nystatin (at a dose of 10 mg/kg q12h), apple vinegar (10 drops in 1.5 litter of drinking water), metronidazole (30 mg/kg q12h), and B-complex (10 drops in 1.5 litter of drinking water) was applied for the infected birds. After three weeks of treatment, all the infected birds were evaluated again by their clinical signs and gram-staining sampling.

Results
The laboratory revealed the following results. Of 50 cases of symptomatic cockatiels, 38 were positive for active, and 4 for non-active yeast in their fecal samples. The fecal samples of 10 gray parrots (out of 20) were positive for active, and 2 cases for non-active yeast. Of 20 Budgerigars, 12 and 2 cases were positive for active and non-active yeast, respectively. Regarding love birds (30 cases), 13 were infected by active and 1 case by non-active yeast. In green cheeks, 16 had active yeast in their fecal samples. Last but not least, only one parrot out of 5 was infected by active yeast in its fecal sample. Figure 1 and shows positive fecal samples for M. ornithogaster in a two years old female cockatiel and a three years old male African gray parrot.
Regarding the clinical signs after 14 days, all birds showed fewer symptoms, turning back to routine life. Another fecal sampling applied for all of them was gram-staining. Despite the treatment, nine cockatiels, two gray parrots, four Budgerigars, five love birds, and five green cheeks were still infected (no positive samples for parrots).

Discussion
Regarding the frequency of positive samples for M. ornithogaster, 6 reported 99 out of 150 birds (66%) were positive with the clinical signs of weight loss, regurgitation, gasping, and lethargy. In another study, 90 cases (90%) showed active yeast in their slides, but only 9 (0.09%) cases had non-active yeast 2 . Furthermore, the highest and lowest infection rates were for cockatiels (42/50) and parrots (1/5). This shows that avian gastric yeast is frequently observed by avian clinicians facing a companion bird with the mentioned clinical signs 5 . The infection rate of the present study was higher than one study in Peru, indicating an incidence rate of 15.9 ± 2.9% in companion birds 6 .
Regarding the treatment options and interruption of the infection cycle, control and treatment of M. ornothogaster are challenging for avian clinicians, as subclinical infections with any specific clinical sign can happen in many companion birds. It seems impossible to keep all the infected birds free from the disease. According to other studies, the only effective treatment for M. ornithogaster is amphotericin B at the oral dosage of 100/kg q12h for 30 days and apple vinegar or grapefruit juice in drinking water [5][6][7] . However, one study revealed that amphotericin B is not 100% effective for budgerigars, as some birds remained infected even after 30 days 8 . Furthermore, some reports showed that fluconazole (100mg/kg q12h) could be promising for the treatment of M. ornithogaster as the second choice. Despite that, mentioned dose of fluconazole can be highly toxic to budgerigars 7,8 . Although nystatin was effective as a treatment of M. ornithogaster in a small group of goldfinches, it was not effective enough for budgerigars in Australia 9 . To summarize the suggested treatments, iodine preparations, ketoconazole, terbinafine, lufenuron, and itraconazole were ineffective against M. ornithogaster in companion birds 7 . Although the information was completely against the present study where nystatin was found as a proper medication for the infected birds, it should be noticed that nystatin was not applied alone. As a proper treatment for avian gastric yeast, apple vinegar was added to the treatment regime 5 . Of note, none of the studies have mentioned the importance of non-infected feed, which could help return the disease. As mentioned in the present study, some birds (25/150) showed the infection after 2 months. This may be due to incomplete elimination of the microorganism from the gastric tract of the birds as nystatin is not considered the choice drug. Furthermore, there is a strong possibility for reinfection by contaminated feed used by the owners. Unfortunately, most bird keepers provide their feeds from invalid pet shops, which elevates the infection rate by contaminated feed.

Conclusion
Avian gastric yeast (M. ornithogaster) is one of the most important diseases that clinically or subclinically infects companion birds worldwide. Controlling and treating this disease has always been difficult for avian clinicians, as the bird could remain infected despite receiving the medications. Although amphotericin B is considered the best choice for M. ornithogaster, the current study claims that combining nystatin and apple vinegar can properly cure the infection. On the other hand, the importance of feed should be considered as the contaminated feed can cause reinfection. Every 3-month checkup by gram staining the fecal samples is recommended to all bird owners and clinicians.

Competing interests
There is no conflict of interest.

Authors' contribution
The final manuscript draft was reviewed by all authors, who also gave their approval.

Funding
No funding.

Ethical considerations
Ethical issues (including plagiarism, consent to publish, misconduct, data fabrication and/or falsification, double publication and/or submission, and redundancy) have been checked by all the authors.