Amoeba protozoa

In microbiological quality control of laboratory animals, parasite infection control is also a critical component. A wide variety of internal and external parasites can cause infections in animals, among which the amoeba protozoan stands out as a significant one.

Amoebas parasitize a variety of organs in humans and animals, including the intestines, skin, and internal organs. They are a globally distributed zoonotic intestinal protozoan disease. There are many species of amoebas, some of which exhibit pathogenic potential. Among experimental animals, the most commonly encountered species is Entamoeba histolytica, which primarily inhabits the intestines and liver of both humans and animals, often leading to amebic dysentery or hepatic abscesses. Additionally, occasional cases of infection with Entamoeba muris have been reported in rodent-based laboratory animals.

● Category: Amoebas belong to the order Amoebida, the class Lobosasida, and the phylum Sarcomastiugophora. Among them, the parasitic species *Entamoeba*—including other closely related parasites such as those in the genera *Iodamoeba*, *Dientamoeba*, and *Endolimax*—are collectively referred to as "endamoebae." In contrast, free-living amoebas are predominantly found in soil and water, occasionally entering animal bodies but typically not causing any disease symptoms.

● Form: Amoebas are widely distributed in the environment, and their life cycle features distinct morphological forms at different developmental stages, primarily including the trophozoite and cyst phases. Moreover, the sizes of the trophozoites and cysts vary significantly among different species of amoebas. Notably, the most pathogenic species, Entamoeba histolytica, has two types of trophozoites: large and small.

The trophozoite is the pathogenic form, while the cyst is the harmless parasitic stage. The trophozoite measures 10–60 μm in size, primarily found in the intestines and fresh feces; it exhibits high motility and a highly variable morphology. After iron hematoxylin staining, its distinct vesicular nucleus becomes clearly visible.

The small trophozoite, also known as the luminal trophozoite, measures 7–20 μm in diameter, moves slowly, and is commonly found in carriers who exhibit no pathological symptoms.

The cysts typically measure between 8.5 and 19 μm in length. During excystation, the tetranuclear form becomes highly active, detaching itself from the cyst wall before escaping through a small pore, with the nuclei clustering together afterward. Outside the cyst, nucleic acids begin to separate from the surrounding cytoplasm, simultaneously dividing into an octonuclear, cyst-like trophozoite form. At this stage, the morphology of the Entamoeba histolytica parasite is slightly smaller than the typical trophozoites usually observed in patients (about 8 μm). Subsequently, the organism continues to absorb nutrients and grow steadily, eventually reaching the characteristic size of a mature trophozoite.

● Life History: The life cycle of Entamoeba histolytica essentially follows this sequence: cyst → trophozoite → cyst. During the cyst stage, the parasite spreads via the fecal-oral route or through direct human-to-human contact, entering an animal host and initiating infection. Once inside the intestine, the cyst ruptures in the small intestine after passing through the mouth and esophagus, releasing the mature trophozoites that then migrate to the large intestine. In the trophozoite stage, Entamoeba histolytica multiplies rapidly in the colon through binary fission. Finally, the parasites form cysts in the colon before being excreted in the feces, thus completing the entire life cycle.

This disease is mostly transmitted via the fecal-oral route and can cause human amebic dysentery. It can also infect dogs, cats, pigs, primates, and house mice; additionally, laboratory animals such as rats, mice, guinea pigs, gerbils, hamsters, and rabbits can all serve as hosts.

The primary focus of domestic research has consistently been on investigating infection cases in both humans and non-human primates. Additionally, there have been reports of infections in pet dogs and cats as well. The main sources of amoebiasis transmission are carriers—individuals or animals that continuously shed cysts in their feces. Infection occurs primarily through the oral route: humans become infected by consuming food or drinking water contaminated with mature cysts, while dogs and pigs are often infected after ingesting cysts found in feces or feed and water tainted by fecal matter. Amoebic trophozoites exhibit extremely weak resistance—they perish within hours at room temperature and are rapidly killed by gastric acid upon entering the digestive tract, rendering them incapable of spreading the disease. In contrast, cysts demonstrate remarkable resilience in external environments, surviving for more than two weeks in moist, cool conditions or within fecal matter, and remaining viable in water for 9 to 30 days. However, they are highly vulnerable to dryness, high temperatures, and common chemicals; exposure to heat above 50°C for just a few minutes leads to their immediate death. Notably, amoebas can pass harmlessly through the digestive tracts of flies and cockroaches, allowing these insects to serve as mechanical carriers of cysts, thereby playing a role in the indirect transmission of the parasite.

Most cases of amoebiasis in animals are asymptomatic carriers, but acute cases can also occur. Clinically, symptoms include fever, abdominal pain and distension, vomiting, and foul-smelling dysenteric bloody stools. Pathological examinations often reveal intestinal inflammation, intestinal hemorrhage with edema, ulcers, and hyperplasia; in isolated cases, liver abscesses may also develop.

Dogs infected with amoebas typically exhibit acute symptoms. Acute cases develop suddenly, leading to persistent diarrhea accompanied by mucus and blood in the stool, along with a distinct, fishy odor. In chronic cases, dogs experience intermittent diarrhea. If liver abscesses occur, they may show tenderness in the liver area, reduced appetite, or digestive issues, causing the affected dog to gradually become thinner.

Microscopic examination reveals that the lesioned tissue in the intestinal wall of affected dogs is characterized primarily by dissolution and necrosis, with only a mild inflammatory response—limited to sparse infiltration of lymphocytes and monocytes. In cases where bacterial infection is absent, neutrophils are rarely observed. Near ulcerated areas, at the interface of live tissue, and within small veins throughout the intestinal wall, Entamoeba trophozoites can be seen, often clustered together or scattered individually. These trophozoites exhibit irregularly round shapes, with abundant cytoplasm that is slightly basophilic; they lack cilia but possess prominent pseudopods. The nucleus is small and round, barely discernible, while the cell membrane remains clearly defined. Within the cytoplasm, vacuoles, phagocytosed red blood cells, and fragmented tissue debris are frequently present. Surrounding the trophozoites, an empty space is often visible—this gap results from tissue degradation caused by enzymes secreted by the parasite.

The rat intestinal amoebiasis model revealed that, in the early stages of infection, the cecum exhibited diffuse, nonspecific inflammation and ulcer formation. Meanwhile, multiple small amoebic abscesses were scattered throughout the liver, with stem cells within the abscesses showing lytic necrosis. Occasionally, trophozoites of Entamoeba histolytica were observed around the periphery of these smaller abscesses.

Domestic surveys reveal that multiple crab-eating macaque colonies across the country exhibit varying degrees of amoebic protozoan infection, with no significant differences in infection rates among different age groups. As experimental animals, crab-eating macaques hold considerable economic value. However, when infected with amoebas, they often develop clinical symptoms such as diarrhea and abdominal pain. Repeated episodes of diarrhea can easily lead to malnutrition, resulting in stunted growth, weight loss, and even hair loss—severely compromising animal health and undermining the accuracy of experimental results.

References

1. Fernando Ramos, et al. *Entamoeba histolytica* and *Entamoeba dispar*: Prevalence of Infection in a Rural Mexican Community. *Exp. Parasitol.* July 2005;110(3):327-30.
2. Jun-ichiro Takano, et al. *Entamoeba histolytica* and *Entamoeba dispar* Infections in Cynomolgus Monkeys Imported into Japan for Research [J]. *Parasitol Res*. 2005 Oct;97(3):255-7.
3. N A Jalili, P Demes, R Holková. The occurrence of protozoa in the intestinal microflora of laboratory mice [J]. BratislLekListy. 1989 Jan;90(1):42-4.
4. Young-Suk Won, et al. Microbiological contamination of laboratory mice and rats in Korea from 1999 to 2003 [J]. Exp Anim. 2006 Jan;55(1):11-6.
5. Tian Kegong et al., Experimental Animal Epidemiology [M]. Chapter 66, Volume 5, 2014:825-830