A Case of Diarrhea Caused by Microsporidiosis in a 4-month-old Blue Fronted Amazon Parrot (Amazona aestiva)

Gerry M. Dorrestein and Marein van der Hage

Department of Pathology, Veterinary Faculty, Utrecht University, Utrecht, The Netherlands
Bilthoven Dierenkliniek, Schubertlaan 17, 3723 LM Bilthoven, The Netherlands

Abstract. A case is presented of intestinal microsporidiosis in a blue fronted Amazon parrot, including clinical presentation, diagnosis in vivo, pathology and ultrastructure.

Key Words: Microsporidium, Encephalitozoon, Parrot, Amazon, Amazona aestiva, Avian

Diarrhea has many causes in birds and it can take on a variety of appearances. The most common cause of diarrhea in pet avian species is bacterial infection. Most enteric pathogens belong to the gram-negative family Enterobacteriaceae. The diagnosis is based on culture and clinical evaluation, but a fecal quick-stain is useful for identifying bacterial populations and the presence of yeast.¹ Gastrointestinal protozoans are also common as a cause of diarrhea in many avian species. Giardia sp. is considered the most important flagellate in the lower gastrointestinal tract of psittacines. Although uncommonly identified in clinical avian practice, gastrointestinal cryptosporidiosis has been reported in many species, including psittacines. Diarrhea may be acute and severe. In this report we will document a case of severe acute diarrhea in a young blue fronted Amazon parrot that "started" as a bacterial/fungal problem and ended as a microsporidial infection.

A 4 month old blue fronted Amazon parrot (Amazona aestiva), called Edje, was presented to the clinic on September 28 with complaints of polydipsia and a severe watery diarrhea of 3 days duration. The owner acquired the bird from a pet shop two months before. The bird was still fed with a hand-raising formula, although it started to eat a seed mixture. In addition, the owner provided sprouted seeds and some fruits. The bird was accepting its food very well and had free access to drinking water.

At presentation the bird was active and friendly. The bodyweight was 351 grams and the body condition and feathers were fine.

Clinical examination gave no additional information. A fecal examination did not show any parasites. The quick stain smear of the feces revealed many yeast organisms.

Preliminary diagnosis of a yeast infection (presumably Candida albicans) was made. The bird was send home with an advisement for a diet change (a complete pelleted diet) and amphotheracin-B 0.1 mg/kcal PO (= 3.5 mg/bird = 10 mg/kg bw²) for one week. An appointment was made for one week later.

One week later the bird returned, but the problem had not changed.

On presentation on October 5, the bird was very ill (Fig 1). The diarrhea had not changed. The body weight had decreased to 281 grams.

Fig. 1.   Blue fronted amazon (A. aestiva) Edje, 4 months old on October 5, 1999, bodyweight 281 gram.

Clinical examination demonstrated loss of muscle mass and an empty abdomen. The bird was very weak. The feces were still watery (Fig 2).

Fig. 2.   Watery diarrhea on October 5

A heparin blood specimen and a fecal sample were taken. The fecal smear showed many bacteria and some yeasts (Fig 3). A sample was sent to the laboratory for culture.

Fig. 3.   Fecal smear stained with Hemacolor® on October 5.  (a) yeast, (b) bacteria (Hc, 100x Oil immersion)

The results of the blood examination were: Ht 51%, WBC 10x10⁹/L, WBC differential: lymphocytes 23%, heterophils 76%, TP 30 g/L, electrophoresis (see Fig 4).

Fig. 4.   Electrophoresis Dij5/1 October 5

Based on this information, the preliminary diagnosis was redefined as a severe bacterial enteritis with early dehydration and hypoproteinemia.

The bird was hospitalized and put on a force-feeding schedule. The bird was fed 3 times daily with Nutrilon-Soya® based on 3 times the BMR calculated using the bird’s normal bodyweight (350 gram = 35 kcal) and a water intake of 30 ml (BMR ~ 35 x 0.5 ml, plus extra water for the diarrhea). Based on the fecal smear, the medical treatment was adjusted to amphotericin-B 0.2 mg/kcal (= 7 mg/bird) and trimethoprim 0.47 mg/kcal (= 16.5 mg/bird) divided over the 3 feedings for 7 days.

The bird recovered quickly and started showing interest in food again. It even started "communicating". The feces were less watery, but remained slimy and brown.

The bacteriological culture isolated Pseudomonas fluorescens (++) and Streptococcus sp. (++). Both organisms were sensitive to amoxycillin, but were not very sensitive to TMP/S. The antibiotic was changed to amoxycillin/clavulanic acid (Synulox®, Pfizer Animal Health) at a dose of 4.5 mg amoxycillin/kcal (= 150 mg/bird). A follow-up culture of the feces the day after the start of the antibiotic treatment showed no bacterial growth.

Four days later (9 October) the bird collapsed. It was still eating the hand-feeding formula supplemented with Nutrilon-Soya® and antibiotics, but started regurgitating. The feces changed to a slimy diarrhea with dark black material indicating blood loss (Fig 5). A stained smear demonstrated that the bacteria had disappeared, but large numbers of small protozoa were now present (Fig 6). The size of these organisms was about 2 µm. Protozoa were often in clusters associated to epithelial cells or scattered within a background of denatured blood.

Fig. 5.   Aspect of feces on October 9. (a) slimy aspect, (b) dark material	Fig 6. Fecal smear stained with Hemacolor® on October 10. (a) protozoa, (b) epithelial cell nucleus (Hc, 100x)

A final diagnosis of intestinal microsporidiosis was made. Treatment with clazuril (Appertex®, Janssen-Cilag) 2.5 mg, in addition to the antibiotic/antimycotic treatment, was added to the tube-feeding regimen. Metoclopramide (Primperan® suspension, Lorex Synthelabo, 1mg/ml) was added at a dose of 0.5 mg/bird to suppress the vomiting. However, the feces became more hemorrhagic and Edje deteriorated very rapidly. He died on October 12.

A necropsy was performed within 1 hour after death, following a standard protocol, including macroscopy, histology (lung, liver, spleen, kidney and several parts of the intestinal tract), impression smears (liver, spleen, lung and intestine), parasitology, and aerobic microbiology of liver and intestine³. Impressions of liver, spleen, lung, and intestine were immunostained for Chlamydia (IFT). Intestinal material and a liver swab were taken for PCR analysis for circovirus and sent to VHL laboratories (Wageningen, NL). Fecal material and duodenum were fixed for transmission electron microscopy (TEM).

Grossly, the bird was in a poor condition; fat reserves were not seen (Fig 7). The body weight was 226 gram.

Fig. 7.   Macroscopy of the post mortem of the amazon with intestinal microsporidia (B98/2132).

Macroscopically the most prominent pathological change was found in the intestinal tract (Fig 8). The intestinal wall was hyperemic and the contents were clear and mucoid, mixed with some fresh blood. In mucosal scrapings from the duodenum, jejunum, and rectum, many microsporia were demonstrated after staining the smears with Hemacolor®.

Fig. 8.   Macroscopic changes to the intestinal tract of the amazon with intestinal microsporidia (B98/3132).

Histologically, the main pathological findings were restricted to the intestines. Specific alterations were not seen in the parenchymateous organs. Sections of duodenum lacked intact epithelium. In other parts of the intestine, the villi were atrophied and the epithelium was desquamated over large areas. In locations where the epithelium was still present, there was a multinuclear reaction of the epithelium with mitotic figures and anisokaryosis. "Cysts" with many microsporidia were seen in many epithelial cells. A inflammatory infiltrates were virtually absent. Many hemorrhages were present in the lamina propria and submucosa (Fig. 9 and 10).

Fig. 9.   Histology of the duodenum demonstrating the loss of epithelium. (HE obj 10X).	Fig 10. Discarded epithelial cells containing "cysts" filled with microsporidia (a) in the cytoplasm (HE obj 40X).

Bacteriology, IFT for Chlamydia, and PCR for circovirus nucleic acid were negative.

Parasites other than the microsporidia were not detected.

TEM demonstrated the typical structural features of Microsporidia (Fig 11 and 12).

Fig. 11.   TEM fecal material showing a typical microsporidium (the yellow bar is 200 nm).	Fig 12. TEM intestines showing a typical microsporidium (the bar is 200 nm).

Only a few reports of microsporidial infections in psittacine birds are included in scientific literature as single case reports or flock outbreaks. In most of these cases, organisms were identified histologically and sometimes ultrastructurally as microsporidia or as a member of the genus Encephalitozoon. Using molecular techniques, parasites in budgerigars, eclectus parrots, a lory, and lovebirds have been identified as Enc. hellum. For a recent overview see Snowden, et al.⁴

According to the same authors, a significant portion of asymptomatic birds appear to be carriers of microsporidial parasites. In our case, the parasite has not been characterized yet, but material is still available for doing so.

In man the parasite, Enc. hellum, has been diagnosed as a cause of respiratory, urogenital, and disseminated systemic disease in immunocompromised persons. An immunocompromised situation also was suspected in this parrot, but the specific cause of immunosuppression was not found.

This case is also interesting because it describes an antemortem diagnosis of microsporidiosis that subsequently was confirmed at necropsy. In the first fecal smears, the parasite was already present (Fig 3, c), but was missed because of the large number of bacteria and yeasts. After the successful elimination of the bacteria and yeast, the microsporidia became obvious, as well as the desquamation of the epithelial cells (Fig 6).

The attempted therapy was predominantly symptomatic, directed toward stabilization of the patient. A specific therapy for intestinal microsporidiosis in birds has not been documented. In our case, we attempted an anticoccidial approach, but without success. Based on in vitro studies, fumagillin, thiabendazole, albendazole, or oxibendazole may have been a better choice.⁵ Drugs that show no effect in vitro include itraconazole, toltrazuril, metronidazole, ronidazole, and ganciclovir.

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4. Snowden K, Phalen D, Barton C: Microsporidia: A common inapparent infection in pet birds. Proc. Association of Avian Veterinarians 1999. pp. 215-217.

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