Trophozoite of Malarial parasite

Trophozoite of Malarial parasite having chromatin and ring form of cytoplasm  in Giemsa stained smear of peripheral blood smear (PBS) as shown above picture.

Introduction of malarial parasites

Malarial parasites are  mosquito-borne infectious agent that causes disease called Malaria. Theses agents are  Plasmodium  falciparum, P. vivax, P. ovale, and P. malariae and commonly called  malarial parasites. These four species of malaria parasites infect us. All are transmitted by female Anopheles  mosquitoes.  According to report, 3.3 billion people at risk of infection, malaria remains one of the world’s most significant health problems. P. falciparum is the most common species identified (nearly 75%) followed by P. vivax (nearly 20%) responsible for causing malaria. A new emerging species, Plasmodium knowlesi is a zoonotic species that causes malaria in macaques and these are mostly of limited public health importance.

Classification of malarial parasites

Kingdom: Animalia

Sub kingdom: Ptotozoa

Phylum: Protozoa

Class: Sporozoa

Genus: Plasmodium

Species:  vivax, falciparum, ovalae and  milariae

Geographical distribution – malaria is present worldwide in tropical and subtropical areas.

Mode of Transmission

Naturally acquired infections are via the bite of infected female Anopheles mosquitoes. Malaria is also transmitted via blood transfusion, sharing of contaminated needles among IV drug abusers, and congenital transmission also has been documented.

Vector: Transmitted by over 60 species of female Anopheles mosquito. The male mosquito feeds exclusively on fruits and juices. The female needs at least 2 blood meals, before the first batch of eggs can be laid.

Life cycle of malarial parasites

Definitive host: Female Anopheles mosquito

Intermediate host: Man

The malaria parasite multiplies by division or splitting of a process known as  schizogony.

A female Anopheles mosquito  transmits a motile infective form (called the sporozoite) to  human (the secondary host), thus acting as a transmission vector. A sporozoite travels through the blood vessels to liver cells (hepatocytes), where it reproduces asexually (tissue schizogony), producing thousands of merozoites. These infect new red blood cells and initiate a series of asexual multiplication cycles (blood schizogony) that produce 8 to 24 new infective merozoites, at which point the cells burst and the infective cycle begins a new. Where as other merozoites develop into immature gametocytes, which are the precursors of male and female gametes. When a fertilized mosquito bites an infected person, gametocytes are taken up with the blood and mature in the mosquito gut. The male and female gametocytes fuse and form an ookinete—a fertilized, motile zygote. Ookinetes develop into new sporozoites that migrate to the insect’s salivary glands, ready to infect a new host. The sporozoites are injected into the skin, in the saliva, when the mosquito takes a subsequent blood meal.

Pathogenecity of Malarial parasites

Malaria infection develops via two phases: one that involves the liver (exoerythrocytic phase), and one that involves erythrocytes (erythrocytic phase). When an infected mosquito pierces a person’s skin to take a blood meal, sporozoites in the mosquito’s saliva enter the bloodstream and migrate to the liver where they infect hepatocytes, multiplying asexually and asymptomatically.

Acute Symptoms

Classical features include cyclic symptoms
Cold stage:  chills and shaking
Hot stage:     fever, headache, vomiting, seizures in children
Sweating stage:  weakness
Feel well for period of time,  then cycle repeats itself

Other Physical symptoms:

Fever: Fever can be very high from the first day. Temperatures of 40°C and higher are often observed. Fever is usually continuous or irregular. Classic periodicity may be established after some days. Hepatomegaly: The liver may be slightly tender. Splenomegaly: Splenomegaly takes many days, especially in the first attack in nonimmune children. In children from an endemic area, huge splenomegaly sometimes occurs. Anemia: Prolonged malaria can cause anemia, and malarial anemia causes significant mortality.Jaundice: With heavy parasitemia and large-scale destruction of erythrocytes, mild jaundice may occur. This jaundice subsides with the treatment of malaria. Dehydration: High fever, poor oral intake, and vomiting all contribute to dehydration.

Each disease has a distinct course

“Tertian Malaria” (P. falciparum, P. ovale and P. vivax)fever occurs every third day. “Quartan Malaria” (P. malariae)fever occurs every fourth day. P. ovale and P. vivax can cause chronic malaria, reappearing after months or years due to latent parasites in liver.

Laboratory Diagnosis of malarial parasites

• Demonstration of Parasite by Microscopy

Diagnosis of malaria can be made by demonstration of malarial parasite in the blood. Thick smears:  It is recommended that 200 oil immersion fields should be examined before a thick film is declared negative.Thin smears: for detecting the parasites and determining the species.

• Quantitative Buffy Coat, Smear

RBC containing malaria parasites are less dense and concentrate just below the buffy coat of leucocytes at the top of the erythrocytic column. Pre-coating of the tube with acridine orange induces a fluorescence on the parasites, which can be visualized under oil immersion.The nucleus of the parasite appears as fluorescing greenish- yellow against red background.

• Micro concentration Technique

Blood sample is collected in microhematocrit tube and centrifuged at high speed. The sediment is mixed with normal serum and smear is prepared. Though it increases the positivity rate, it changes the morphology of the parasite.

• Culture of Malaria Parasites

Several culture lines have been established from blood of infected Aotus monkey or directly from human patients. Schizogony proceeds normally in culture. Gametocytes are formed infrequently. Plasmodium retain their infectivity in culture.

• Serodiagnosis

Serodiagnosis is not helpful in clinical diagnosis because they will not differentiate between an active and past infection. It is used mainly for sero epidemiological survey and to identify the infected donors in transfusion malaria. The tests used are indirect hemagglutination (IHA), indirect fluorescent antibody test (IFA), and enzyme-linked immunosorbent assay (ELISA).

•  Rapid Antigen Detection Tests

Based on the detection of antigens using immunochromatographic method. Have been developed in different test formats like the dipstick, card, and cassette- bearing monoclonal antibody, directed against the parasite antigens.

• Fluorescence Microscopy

Fluorescent dyes like acridine orange or benzothiocarboxy purine are used, which stain the parasites entering the RBCs but not WBCs. This is a method of differential staining. Acridine orange stains DNA as fluorescent green and cytoplasmic RNA as red.

• Molecular Diagnosis

DNA probe is a highly sensitive method for the diagnosis of malaria. It can detect less than 10 parasites/µL of blood. Polymerase chain reaction (PCR) is increasingly used now for species specification and for detection of drug resistance in malaria.

• Other Tests

Measurement of hemoglobin and packed cell volume (PCV)Total WBC and platelet count in severe falciparum malaria.

Measurement of blood glucose to detect hypoglycemia in severe falciparum malaria and patients receiving quinine Coagulation tests like measurement of anti-thrombin III level, plasma fibrinogen, fibrin degradation products (FDPs), prothrombin time (PTT), if abnormal bleeding is suspected in falciparum malaria.

Urine for free hemoglobin, if black water fever is suspected.

Blood urea and serum creatinine to monitor renal failure.

Treatment of malarial parasites

• Chloroquine (Aralen)
• Quinine sulfate (Qualaquin)
• Hydroxychloroquine (Plaquenil)
• Mefloquine combination of atovaquone and proguanil (Malarone)

Control of malaria

It can control using following methods-

• Using medicine for disease control-Medicine prepares from silicon tree, bark-quinin and trade names- Chloroqunin, Mepaquinin
• by mosquito control

It is further of three types-

1. Physical control
2. Chemical control and
3. Biological control ( Use of Gambasia fish)

References

1. Isenberg clinical microbiology procedures Handbook
   2nd edition. Vol. 2
2. Merkell and voge’s medical parasitology
   9th edition.
3. Parasitology: 12th edition
   By K. D. Chatterjee
4. District laboratory practice in Tropical countries –Part-I .
   By Monica cheesbrough.
5. Atlas of Medical Helminthology and ptotozoology -4th edn  -P.L.  Chiodini, A.H. Moody, D.W. Manser
6. Medical Parasitology by Abhay R. Satoskar, Gary L. Simon, Peter J. Hotez and Moriya Tsuji
7. Atlas of Human Parasitology, Lawrence R Ash, Thomas C. Orihel, 3 rd ed, Publisher ASCP press, Chicago.
8. Molecular Medical Parasitology. Editors: J. Joseph Marr, Timothy W. Nilsen andRichard W. Komuniecki, Publisher Academic press, an imprint of Elsevier Science.
9. Topley &  Wilsons Principle of parasitology. Editors: M.T. Parker & L.H. Collier, 8 th ed 1990, Publisher Edward Arnold publication, London.