Protozoan Diseases – An Overview on Protozoal Infections

One of the interesting Biology Topics is the study of animal behavior and how it is influenced by genetics and the environment.

Common Protozoan Diseases in Humans

Malaria: Causes, Symptoms, and Diagnosis

The name malaria is derived from the Italian term ‘malaria’ (mala = bad; aria = air) meaning ‘bad air’. Laveran (1880) discovered that malaria is caused by the protozoan parasite Plasmodium. He got Nobel Prize in 1907. Sir Ronald Ross, a doctor of the Indian Army, established that the malarial parasite is transmitted by the bite of a female Anopheles mosquito. In 1902 he got Nobel Prize for his discovery.

Malaria can be a fatal disease for human beings. Approximately 300 million people around the world get infected with malaria every year. More than 2 million people die every year due to malaria disease. Malaria is caused by the protozoan parasite Plasmodium. This disease spreads through the bite of an insect vector – the female Anopheles mosquito which feeds on humans blood. Male Anopheles mosquito feeds upon plant juice.

Causative Organism
Several species of protozoan parasite called Plasmodium are the causative organism of several types of malaria.

Types of Malaria and Causative Organisms:

Types of Malaria	Causative Organism	Symptomatic Distinction
1. Benign Tertian Malaria	Plasmodium vivax	Fever occurs every 48 hours.
2. Malignant Tertian Malaria	Plasmodium falciparum	Fever occurs at 24 – 48 hours intervals.
3. Mild Tertian Malaria	Plasmodium ovale	Fever occurs every 48 hours.
4. Quarantan Malaria	Plasmodium malaria	Fever occurs at 72 hours intervals.

Host of Malarial Parasite

• Definitive Host: Female Anopheles mosquito, where the sexual phase of the parasite occurs. It is also called the primary host.
• Intermediate Host: Man, where the asexual phase of the parasite occurs. It is also called the secondary host.

Mode of Infection of Malarial Parasite:

When a female Anopheles mosquito bites an infected and diseased person, the gametocyte stage of parasites gets entry into the mosquito. The sexual phase of the parasite is completed in the body of the mosquito and sporozoites are produced. The sporozoites reside in the salivary glands of the mosquito. When the infected female bites a healthy man again the sporozoites may be transmitted into a new host. Thus the disease may spread in the population.

Structural Features of Plasmodium

• Unicellular organisms with a single nucleus.
• Different morphological forms are available during different stages of the life cycle as sporozoite, trophozoite, schizont, merozoite, male and female gametocytes.
• A sporozoite is sickle-shaped with pointed terminals (10-14 µm × 1 µm) with an apical cup anteriorly, infective stage in man during the asexual phase.
• Schizont is oval in shape (42 mm diameter), transformed sporozoite after receiving nutrients from liver cells.
• Merozoite: This is round in appearance, found in RBC.
• Trophozoite: This is oval in shape, with hemozoin in the cytoplasm, containing a vacuole with a nucleus, and may be pushed peripherally forming a signet ring configuration.
• The presence of a pseudopodia-like structure may be noticed. Trophozoite may be transformed into schizont.
• Male and female gametocytes: These are formed from merozoite, oval in shape. Male gametocyte is small (9-10 µm) and female gametocyte is large (10-14 µm).
• The nucleus in the male gametocyte is large and central in location, but the nucleus in the female gametocyte is small and terminal in position.
• Male microgametocyte forms male gametes and macrogametocyte (female) forms female gamete.

Life-Cycle of Malarial Parasite

Life-cycle of Plasmodium requires two hosts for completion. Male and Female Anopheles mosquitoes are these hosts.

A. Life-Cycle in Man or Asexual Cycle or Schizogony:

When the female Anopheles mosquito bites a man, the sporozoites may enter into the blood of a man through mosquito saliva. In the blood they may invade the RBC or going into the liver they may invade the hepatic cells. In this phase of the life cycle, they may divide mitotically to produce many progeny cells called merozoites. Such divisions are called schizogony. In the human cycle, three types of schizogony are observed, namely pre-erythrocytic schizogony, erythrocytic schizogony, and exo-erythrocytic schizogony.

1. Pre-erythrocytic Schizogony or Pre-erythrocytic Cycle:

• A female Anopheles mosquito carrying malaria-causing parasites feeds on a human and injects the parasites in the form of sporozoites into the bloodstream. The sporozoites entering the blood are transported to the liver and invade hepatocytes. The sporozoites are sickle-shaped and length 10-15 µm and breath 1 µm.
• Each sporozoite grows rapidly and develops into a schizont. The schizont undergoes asexual multiplication termed schizogony. The nucleus of the schizont divides by multiple fission to form 6-24 daughter nuclei. After some time the totally exhausted erythrocyte bursts liberating the merozoites.
• The free merozoites either enter the plasma of blood or go to the liver sinusoid and attack the fresh liver cells where repeat the same sequence.

2. Erythrocytic Schizogony or Erythrocytic Cycle:

• The erythrocytic schizogony begins when the RBCs of blood are attacked by pre-erythrocytic merozoites. The merozoites after entering into the bloodstream, feed on erythrocytes, become rounded, and modify into trophozoites.
• As the merozoites grow a vacuole appears in the center and the nucleus is pushed to one side. It gives a ring-like appearance and is known as a signet ring stage.
• The parasites ingest haemoglobin and decompose it into protein and haematin. Protein is used as food whereas unused haematin forms a toxic which is yellowish-brown pigment, called hemozoin.
• As the signet ring parasite grows, the vacuole disappears and the parasite becomes amoeboid in appearance. This stage is called the amoeboid stage. At this stage RBC develops numerous granules, the Schuffner’s granules.
• Within 48 hours trophozoites grow in size, become rounded, and almost completely fill the RBC called schizont.
• The nucleus of the schizont divides by multiple fission to form 6-24 daughter nuclei. These nuclei arrange at the periphery, while the toxic hemozoin granules accumulate at the corner of the RBC. It appears as a flower rose, the so-called rosette stage.
• The nuclei of the rosette stage are surrounded by a little cytoplasm and are developed into merozoites. With the rupture of RBC, these merozoites are liberated into the blood plasma along with toxic hemozoin that results in feelings of chill and fever, developing the first sign and symptoms of malaria.
• They reinfect fresh RBCs and continue to repeat the cycle.

3. Exo-erythrocytic Schizogony or Exo-erythrocytic Cycle:

• The cycle begins when the merozoites from the liver parenchyma cell reenter into the new liver parenchyma cell and the merozoites form schizont.
• From this schizont, numerous exoerythrocytic merozoite or metacryptozoite, or phanerozoic are produced.
• These metacryptozoites again enter the liver cell or RBC.

4. Gametogony or Formation of Gametocyte:

• Some of the merozoites, after entering the RBC neither form trophozoites nor multiply by binary fission but grow slowly and become compact bodies, the gametocytes.
• The more numerous, but small in size and with a large centrally placed nucleus are the microgametocytes, potentially male.
• The less numerous but larger in size and with a greater amount of dense cytoplasm and a small nucleus are the mega gametocytes, potentially female.
• The mature gametocyte can survive atleast 7 days in the blood. They reach the superficial blood vessels and wait for the bite of female Anopheles.

B. Life-Cycle of Mosquito:

Sexual reproduction occurs in the mosquito and therefore, this is the sexual phase of the life cycle.

Formation of Gamete:

• When the female Anopheles mosquito sucks blood from an infected malarial patient mature microgametocytes, macrogametocytes, and asexual parasites enter the stomach of the mosquito.
• During digestion asexual parasites are destroyed, however, microgametocytes and macrogametocytes come out from erythrocyte covering.

From the microgametocyte by way of exflagellation and division of the nucleus eight long flagellated bodies as microgametes may be produced within 10-15 minutes. Little change occurs in the macrogamete.

Fertilization:
The microgametes move actively to the macrogamete and one microgamete (male) penetrates the macrogamete to form Zygote.

Maturation of Ookinete and Sporozoites:

• The zygote takes a fusiform shape and produces pseudopodia forming the motile vermicule, the ookinete.
• In the ookinete, the male and female nuclei unite and it may penetrate the gut epithelium to reside between the epithelium and the outer elastic membrane.
• The ookinete then becomes spherical forming an oocyst within which the nucleus undergoes meiotic and mitotic division forming many sporozoites.
• Ultimately the oocyst bursts to liberate the sporozoite into the hemocoel and disperse throughout the body of the mosquito.
• They may penetrate the acinar cells of the salivary gland and accumulate in the acinar duct.
• During the biting of a fresh host, the sporozoites may enter the bloodstream of the host.

Symptoms of Malaria

The main symptoms of malaria include headache, nausea, muscular pain, and high fever. Each malarial attack is 6 to 10 hours in duration and consists of three stages:

• Cold stage – the feeling of extreme cold and shivers.
• Hot stage – high fever, faster respiration, and heartbeat.
• Sweating stage – due to profuse sweating, the temperature of the body goes down to normal.

Even after the patient is cured of malaria, the patient feels weak and becomes anemic. Malaria may also secondarily cause enlargement of the spleen and liver. The presence of malarial parasites can be checked by blood tests.

The name Malaria was proposed by Macculoch (1827). C.L.A. Laveran (1880), a French physician, discovered the Malarial parasite Plasmodium in the blood of a malaria patient. He received Nobel Prize for this discovery in 1907. Sir Ronald Ross (1897), a British physician, confirmed that malaria is caused by a malarial parasite and the mosquito is the vector. He received Nobel Prize for this discovery in 1902.

Recently Allan Porter and his team produced a genetically engineered aquatic bacteria at the National Institute of Singapore. When mosquito larvae feed on these bacteria, they get killed by the toxic substance produced by these bacteria. This can prove to be an effective biological control method against malaria.

Central Drug Research Institute (CDRI), Lucknow has developed an anti-cerebral malarial drug called- Arteether. This drug is extracted from a herbaceous plant Artemisia annua belonging to the family Asteraceae.

Modes of Transmission of Malarial Parasite

• Transmission by the vector (Female Anopheles mosquito): Mechanically through mosquito biting transmission of sporozoites occurs from an infected man to a healthy man.
• Transmission by transfusion: if the donor is infected, during the transfusion of blood, transmission of the parasite may be possible. It is called direct transmission.
• Transplacental transmission: Foetus in the uterus may get the infection through the placenta, though it is naturally unusual.
• Transmission for drug addiction: A drug addict sometimes uses the syringe of an infected drug addict and then the second person may get infection by way of the contaminated syringe.

Clinical Manifestation of Malaria

The following are the clinical manifestation of malaria:

1. Recurring Appearance of Fever:
At regular intervals (usually 24-72 hours), the recurrence of the attack passes through three phases namely the cold stage, hot stage, and sweating stage.

• Cold Stage: At this stage, the infected person feels a severe cold. Shivering occurs with the rising of temperature in the body. It lasts from 15 minutes to 1 hour.
• Hot Stage: Cold stage is followed by the hot stage when body temperature may rise to 106°F. The patient suffered from severe headaches, pain in the body and limbs. The feeling of weakness, feeling of headache, and vomiting tendency are other associated symptoms at this stage. This stage lasts for 2-6 hours.
• Sweating Stage: At this stage, sweating occurs in the patient, and the fever comes down. It lasts for 2-3 hours. All these symptomatic expressions of the malarial patient together are called febrile paroxysms.

2. Anaemia:
Because of recurring attacks on the RBC and subsequent destruction of the blood cells, a malarial patient suffers from severe anaemia.

3. Splenomegaly:
During malarial infection, the size of the spleen is enlarged and this is called splenomegaly.

4. Enlargement of Liver:
It is another associated symptom of this disease.

5. In malignant Malaria:
The brain gets infected and capillaries may be clogged by the infected RBC. This may lead to cessation of blood supply to the brain. Because of this, it is also called cerebral malaria.

Incubation Period of Malarial Parasite

Mosquito bites and entry of sporozoite into the blood do not develop the symptoms of malaria or febrile paroxysm immediately, but it takes a few days to develop the symptoms. This latent period is called the incubation period. The incubation period of different malarial parasites is as under:

Name of the Parasite	Incubation Period
Plasmodium vivax	10-14 days
P. falciparum	Do
P. ovale	10-14 days
P. malariae	18 days-6 weeks

Control of Malaria

People are trying to control malaria through all sorts of efforts. Even at Government levels different health service schemes have been taken to control the incidence and spread of malaria. These efforts are focused on two aspects To prevent the incidence of malarial infection and to Control the vector species by preventing their population growth. A drug named quinine, which is extracted from the bark of the Cinchona tree, is used to treat a person suffering from malaria. This drug kills most of the stages of the malarial parasite. There are certain other drugs (medicines) that should be taken on the advice of a doctor. Scientists are trying hard to develop an antimalarial vaccine.

1. Prevention of Incidence of Malarial Infection:
This refers to the prevention of malarial infection in men through public awareness and cautious treatment of the infected man.
(a) Prophylactic measures:

• By use of mosquito nets during sleep at night: By this measure, the mosquito is prevented from biting a man and infection may not reach the individual.
• By using mosquito repellants: Some chemicals available in the market such as Aliout, Odomos, mosquito coil, etc., may be used in the rooms of human habitation.
• People may get rid of mosquito biting and the incidence of malarial infection may be minimized.

(b) Treatment of the malaria-infected man: Whenever a person is observed to be infected by malaria, he must be treated under strict medical care. To prevent the spread of the disease in the surrounding area the infected person must be kept away from mosquito bites by any means. It is better for the person to be hospitalized immediately for proper treatment.

Healthy persons may also be rescued from malarial infection by using drug prophylaxis as per the advice of the medical practitioner. Chloroquine or Proguanil is usually taken at a specific dose at regular intervals. For Chloroquine-resistant malaria Mefloquine or Doxycycline is used for this purpose. The dose and time interval must be determined with advice from a medical practitioner. Pregnant women and children are cautiously given anti-malarial drugs.

2. Control of Vectors:
Control of the population growth or minimizing the growth of the Anopheles mosquitoes may control the incidence and spread of malaria in the population. A close study of the breeding of mosquitoes and clear knowledge of the biology of Anopheles sp. causing malaria may help in the prevention of the spread of malaria. Several effective measures to control the mosquito population are as under:

• Reduction of the breeding ground: Domestic areas, indoor sites, and surrounding areas of the houses should always be kept clean and accumulation of water in no way be allowed in the specified areas.
• Cultivation of larvae-eating fishes in drains and small ditches: Some water bodies which are breeding grounds for mosquitoes are difficult to be destroyed. In such areas, open drains and small ditches may be cultivated with fishes such as Guppy, Gambusia, Tilapia, Panchax, etc., that feed upon the mosquito larvae. This sort of practice may reduce the growth of the mosquito population.
• Use of insecticides: DDT, BHC, Ethyl Parathione, Malathione, and Dialdrin are several insecticides that may be sprayed in the household areas, and bushes to kill the adult mosquitoes. By this method, the population of mosquitoes may be diminished. Some fumigants such as carbon disulfide, methyl bromide, SO₂, etc., may also be used to destroy the mosquitoes.
• Use of oils and insecticides in water bodies: Small water bodies used by mosquitoes for their breeding may also be treated with insecticides to kill the mosquito larvae. The use of kerosene oil and petroleum in water bodies has a potential effect in destroying the larvae of the mosquito.

Prevention of Malaria

Malaria is an infectious disease. It can spread from an infected person to a healthy person (host) by the bite of female Anopheles mosquitoes (vectors). The only way to prevent malaria is to take precautions against mosquito bites. We can protect ourselves from the bite of mosquitoes by following methods:

• Wire gauze should be used on doors and windows of houses to prevent the entry of mosquitoes.
• Insect repellents (e.g., Odomos) should be used to prevent mosquito bites.
• One should sleep under mosquito nets.
• Mosquito larvae can be killed by sprinkling kerosene oil on large-sized water bodies. Some larvivorous fishes such as Gambusia, minnows or trouts, or birds (e.g., ducks) can be introduced into water bodies. These animals feed on mosquito larvae and hence the population of mosquitoes is checked. This is called the biological control of mosquitoes.
• Adult mosquitoes can be killed by spraying insecticides (e.g., BHC, malathion) on the walls of human dwellings. Insecticides are chemical poisons (toxicants), hence, regulation of the mosquito population by them is called chemical control. The insecticide DDT is not used in the chemical control of mosquitoes, as it persists in the environment for a much longer time and can cause toxicity by accumulating in the body of fishes, birds, and humans through food chains.
• The breeding grounds should be destroyed. Thus, the ditches, puddles, or swamps around human dwellings should be drained or filled. We should not allow the collection of water in any uncovered container such as a water tank, pot, cooler, flower pot, discarded tires (tires), etc.

Amoebiasis: Causes, Symptoms, and Diagnosis

It is a disease caused by protozoan (Entamoeba histolytica) which is very common in India. This parasite lives in the large intestine of man. Infection occurs by ingesting cysts contaminated food and water. Lambl (1859) first discovered the parasite. Losch (1875) proved its pathogenic nature.

Pathogen of Amoebiasis
Monogenetic protozoan – Entamoeba histolytica.

Life Cycle of E. histolytica
E. histolytica passes its life cycle only in one host. There are mainly two phases of development – Trophozoite and Cyst with a transitory phase of cystic form. The process of transformation from cyst to trophozoite is known as excystation and that from trophozoite to cyst is known as encystation. Both processes occur in one and the same host. The multiplication takes place only in the trophozoite phase.

Excystation

The mature quadrinucleate cyst is taken with contaminated food or drink, which passes it into the small intestine. The cyst wall is resistant to the action of gastric juice but is digested by the action of trypsin in the intestine. The excystation occurs when the cyst reaches the lower part of the ileum. Each cyst liberates a single amoeba with four nuclei and immediately forms eight amoeba by the division of nuclei with successive fission of cytoplasm. The young amoeba invades the tissue and ultimately lodges the submucous tissue of the large intestine. The trophozoite stage of the parasite multiplies in large numbers. Reproduction of trophozoite occurs simply by binary fission.

A certain number of trophozoites living in the intestinal wall are discharged into the lumen of the bowel and are transformed into small cystic forms from which the cysts are developed. The whole process of encystation takes place in few hours. A mature cyst is a quadrinucleate spherical body. The cyst varies in its size – the small race is 6-9 mm and the large race is 12-15 mm. The cysts come out with the stool of the affected person and infect new persons with contaminated food and water.

Incubation Period: 5 to several months.

Mode of Transmission of E. histolytica
Transmission of E. histolytica from man to man is affected through the encysted stage and infection occurs through the ingestion of these cysts. Fecal contamination of drinking water, vegetables, and food is the primary cause. Uncooked vegetables, fruits, and impure water are the secondary causes of infection. House flies may convey the infection while passing from feces to unprotected foodstuffs. The cysts of E. histolytica have been found in the droppings of cockroaches which also serve as a source of infection.

Symptoms of E. histolytica

• Abdominal pain with loose motion.
• Diarrhoea alternating with constipation.
• The passing of blood and mucous with dirty stool.
• Feces having cysts.

Diagonsis of E. histolytica:
Microscopical examination of stool finds cysts of E. histolytica. Serum antibody test, colonoscopy, etc.

Control of E. histolytica:
Uses of drug like emetine, tinidazole, metronidazole etc. No immunization is done.

Preventive Measures for E. histolytica:
Sanitary disposal of human excreta, safe drinking water.