Overview of Bloodborne Organisms in Poultry
Avian blood may contain various disease agents, including viruses, bacteria, rickettsiae, protozoa, microfilariae, and rarely fungi. These organisms can be identified by microscopic examination of wet mounts, buffy coat, or blood smears or by appropriate culturing and molecular techniques. Microscopically, some are within blood cells (Plasmodium, Haemoproteus, Leucocytozoon, Isospora[Atoxoplasma], Hepatozoon, Babesia, Aegyptianella), while others are free in the plasma (Trypanosoma, microfilariae, bacteria, spirochetes). None live exclusively in the blood; most are found in tissues but are present in blood during part of their life cycle. Some, such as microfilariae and Plasmodium, may have a periodicity when numbers or stages of parasites vary with time. In such cases, examining multiple smears at intervals will increase the likelihood of obtaining a diagnosis. Seasonal variations in infection rates relate to the activity of arthropod vectors. When possible, tissue cytology is also a useful adjunct to examination of blood. Most bloodborne organisms are either uncommonly or not associated with clinical disease. However, weakened or injured birds infected with hemoprotozoa may have higher mortality and slower recovery than uninfected birds. Examination for bloodborne organisms should be included in the clinical and diagnostic procedures for any ill bird.
Thin blood smears should be made with blood directly from the bird, if possible. Anticoagulants, storage, and cooling of the blood can distort protozoal morphology and introduce artifacts. A small drop of blood can be collected using a syringe and needle. The drop should be spread on a clean glass slide to make a thin smear. A Romanowsky-type stain that gives good polychromatic coloration (eg, Giemsa stain) should be used. At least 200 oil-immersion fields (~20,000 RBC) for single smears or 100 for multiple smears from the same bird should be examined. Leucocytozoon and microfilariae are found around the periphery of smears and can be easily seen on low-power magnification.
Bloodborne organisms in plasma or WBCs are concentrated in the buffy coat. The microhematocrit tube is cut just below the buffy coat above the packed RBCs. The buffy coat should be expressed from the cut end with a small amount of plasma to make a suspension, and a thin smear prepared. Stained buffy coat smears are recommended to detect bacteria, spirochetes, and chronic Leucocytozoon, Trypanosoma, or Isospora (Atoxoplasma) infections. An excellent technique to identify low numbers of motile organisms such as spirochetes and microfilariae is direct examination of the buffy coat by darkfield or phase contrast microscopy. The buffy coat and all of the plasma should be expressed onto a glass slide and covered with a coverglass, which is depressed slightly to spread the buffy coat. The buffy coat/plasma interface should be examined with darkfield or reduced light microscopy to detect motile organisms.
To make a diagnosis of infection with an intracellular blood protozoan on a thin blood film, it first should be determined that the “parasites” in question are neither normal structure nor artifact. The following should then be determined: the host cell and whether it is normal or deformed beyond identification, whether pigment granules (hemozoin) are present or absent, and whether merogony is occurring (see Table: Characteristics of Protozoa Encountered in Avian Blood). Identification of an organism beyond genus is difficult and usually unnecessary for clinical purposes.
Serologic and molecular diagnostic methods have been developed for avian hemoparasites but are usually not available commercially. Molecular methods are very sensitive and can detect infection when parasite numbers are too low to be detected in blood or tissue smears or by histology. Hemoparasites can also be studied by subinoculation of infectious blood in birds of the same or a known susceptible avian species. Bacteria can usually be identified by blood culture and molecular methods.