Plasmodium gallinaceum - Molecular diagnosis (PCR)

Information 02-09-2018.

            Malaria is a disease caused by protozoa belonging to the genus Plasmodium. More than 200 species cause the disease in vertebrates, with several species involved in avian malaria. These include Plasmodium relictum, the most common cause of avian malaria, and Plasmodium gallinaceum, which infects Gallus birds, including domestic chickens.

            The unicellular parasite Plasmodium gallinaceum is a causative agent of malaria in Gallus birds and is one of the main protozoan pathogens of poultry production systems. Plasmodium gallinaceum causes poor meat quality and low egg production. In addition, the disease is often fatal with a mortality rate of up to 80 to 90%, and therefore, can contribute to a substantial economic loss for the poultry industry. The disease is highly prevalent in tropical countries, with a high transmission rate in the Southeast and South Asia. 

            Plasmodium gallinaceum was first described in 1935, and isolated from domestic chickens in Sri Lanka. It was later reported that P. gallinaceum is endemic in several wild birds of the jungle in several countries of South Asia. Although it typically causes subclinical disease with low mortality in its primary host (birds of the India jungle Gallus sonnerati), domestic chickens of European origin (Gallus gallus) are highly susceptible to P. gallinaceum, with mortality rates of up to 90%.

Clinical signs associated with avian malaria include a pale crest, green stools and lack of appetite (anorexia), although these symptoms are not necessarily specific to malaria. The disease can present a very acute course and cause death within a week, during the acute phase of the disease due to severe parasitaemia. Also, P. gallinaceum malaria can cause death during the subsequent chronic stage, in the second or third week, due to paralysis caused by the occlusion of infected brain capillaries. However, some infected adult chickens can develop asymptomatic infections, which results in an erroneous diagnosis.

Malaria is a disease transmitted by mosquitoes. While human malaria is transmitted only by mosquitoes of the genus Anopheles, it has been shown that Plasmodium gallinaceum can be transmitted through a wide range of vectors, including mosquitoes the genera Anopheles, Culex, and Aedes. In the transmission of malaria, the vector mosquito is infected by ingesting infected blood from a bird that has sexually differentiated parasites in gametocytes, and inside it will form the zygote, which will evolve into an oocyst, which will later give rise to the sporozoites. The sporozoites enter the hemolymph of the mosquito, and invade the salivary glands, where they remain until inoculated, with saliva, into the skin of the host.

Unlike mammalian species, where the life cycle of Plasmodium is composed of a short clinically silent hepatic phase, followed by a symptomatic blood phase, exoerythrocytic tissue infection in avian malaria by P. gallinaceum can occur in parallel with the erythrocytic phase, causing the resulting clinical symptoms to overlap. This is because in mammals, the injected sporozoites are transported to the liver where they are transformed into hepatic schizonts and the merozoites are produced by asexual replication. However, P. gallinaceum sporozoites infect local macrophages at the site of the mosquito bite and differentiate into cryptozoites, the first generation of exo-erythrocytic parasites in the avian host. Some of these cryptozoites infect new dermal macrophages, while others develop into metacriptozoites in the macrophages of the reticulo-endothelial system, including the Kupffer cells of the liver. These primary exoerythrocytic parasites can be differentiated into merozoites or differentiated into phanerozoites, that is, secondary exo-erythrocytic stages, which continue to replicate in the tissues. In addition, merozoites can return from the blood to the tissues and become phanerozoites again. In addition to macrophages, phanerozoites can also infect vascular endotheliums, particularly those in the brain. The pathological changes produced by these endothelial phanerozoites resemble those of severe Plasmodium falciparum malaria in humans, which is caused by the occlusion of cerebral capillaries by erythrocytes and infected immune cells. In natural infections with P. gallinaceum, the primary and secondary exoerythrocytic development lasts several weeks, after which the infection is limited to the blood.

To prevent the spread of avian malaria diseases, it is essential to have tests for the detection of parasites that are rapid, inexpensive and reliable. The most commonly used method for the diagnosis of avian malaria is the examination of blood under an optical microscope, which is also an accepted practice for the diagnosis of human infections due to malaria. The main advantages of this technique are its low cost and it allows to estimate the parasite load in infections. However, the samples must contain a sufficient amount of parasites to allow their rapid and reliable observation, while in blood samples with low levels of parasitaemia the microscopic examination of blood samples may require a lot of work and time. For this reason, several tests based on the polymerase chain reaction (PCR) for the diagnosis of P. gallinaceum have been developed in recent years. These molecular methods have allowed to increase the sensitivity and specificity of the diagnosis of avian malaria.

Tests carried out in IVAMI:

• Molecular detection of Plasmodium gallinaceum by PCR.

Recommended sample:

• Sample of blood extracted with EDTA (2 to 5ml). Optionally, samples collected in filter papers can be sent.

Conservation and shipment of the sample:

• Refrigerated (preferred) for less than 2 days.

• Frozen: more than 2 days.

• Samples deposited on filter papers can be sent at room temperature.

Delivery of results:

• Molecular detection of Plasmodium gallinaceum by PCR: 24 to 48 hours.

Cost of the test:

• Molecular detection of Plasmodium gallinaceum by PCR: Consult to ivami@ivami.com.