Detecting Kala-Azar Parasite: Novel Method Developed by Researchers from University of Dhaka

In the tropical region, a neglected life-threatening disease is visceral leishmaniasis, also known as kala-azar. This disease is prevalent in approximately 60 countries worldwide, including Bangladesh. The Mymensingh belt in Bangladesh, in particular, has a higher incidence of this disease. In the country, 45 districts are in the endemic stage of kala-azar. Research indicates that around fifty million people in Bangladesh are at risk of being infected with kala-azar. Not only a disease prevalent in the sub-tropical region, it is also neglected due to various reasons, leading to its being labeled as a Neglected Tropical Disease (NTD).

In Bangladesh, especially in the Mymensingh region, the outbreak of kala-azar is most prevalent. Source: ICDDR’B.

If left untreated, the probability of mortality from kala-azar is high. The causative agents for kala-azar are the parasites Leishmania donovani and Leishmania infantum. In Bangladesh, the transmission of the former is more prominent. The parasite enters the human body through the bite of infected sandflies. The sandflies, which reside in the cracks of mud houses in rural areas, are abundant in areas where the disease is prevalent. Once inside the body, the parasites cause infection in vital organs such as the liver, spleen, and bone marrow. This can lead to hepatomegaly (enlarged liver), splenomegaly (enlarged spleen), and other complications. In severe cases, the disease can progress to a condition known as pancytopenia, where blood cells are progressively destroyed, leading to a life-threatening situation.

The key to eliminating such infectious diseases lies in early and effective identification of the causative agents. Researchers from University of Dhaka, led by Dr. Mohammad Manjurul Karim, have developed a rapid and reliable method for identifying the parasite in the patient’s urine. This method, published in the renowned PLOS Global Public Health Journal, offers a quicker and more dependable approach to detect the parasite. Samiur Rahim, Md. Mohiuddin Sharif, Md. Robed Amin, and Mohammad Tarikur Rahman were also part of the research. Dr. Karim emphasizes the importance of timely and accurate diagnosis for effective management of the disease.

Under the microscope, Leishmania donovani parasites. Source: Thermo Fisher.

Previously, diagnostic procedures involved immunochromatographic tests on blood and microscopic examination of tissue samples from the bone marrow, liver, spleen, and lymph nodes. However, these methods were less specific. The newly developed method allows for the identification of the parasite with higher specificity and efficiency.

Dr. Karim

Despite the advancements in diagnostic methods, the collection of samples and the subsequent analysis remain complex and time-consuming. The research team suggests that obtaining samples from various organs is crucial for an accurate diagnosis. However, the complexity of the sample collection process requires skilled professionals to minimize the risk of complications such as blood contamination.

The challenges in diagnosing and treating kala-azar highlight the need for ongoing research and development of innovative approaches to tackle this neglected tropical disease effectively.

Due to the complexities involved, the researchers opt to work with samples from the patient’s blood instead of bone marrow. This choice is made because when the bone marrow is affected, the parasites are also present in the patient’s blood. They extracted a specific component from the blood called Buffy Coat. In the Buffy Coat, which contains blood cells and Leishmania parasite cells, DNA is separated. Using conventional PCR methods, the DNA is then analyzed to confirm the presence of the parasite.

Professor Dr. Muhammad Manjurul Karim 

The researchers emphasize the importance of specificity in their approach. They design the primary assay in such a way that it yields amplicons (fragments of DNA) of a specific length, around 100-150 base pairs, suitable for subsequent use in real-time PCR. Notably, real-time PCR allows for both qualitative and quantitative analysis simultaneously.

They published this research in the Asia Pacific Journal of Tropical Medicine. Subsequently, they collect urine samples from kala-azar patients at Dhaka Medical College Hospital. In this endeavor, they receive assistance from Professor Dr. Robed Amin and Dr. Mahiuddin, who manage a unit dedicated to kala-azar patients at Dhaka Medical College Hospital. Utilizing the same primary assay, they work towards confirming kala-azar from samples of bone marrow, blood, and urine.

Kala-azar parasites can also infect the kidneys of patients. Therefore, the researchers investigate the possibility of detecting the presence of Leishmania parasites in the patient’s urine. A study from Brazil also supports this idea, where Leishmania parasites were identified from a patient’s urine. Hence, we explored the potential of utilizing urine samples for kala-azar confirmation in Bangladesh.

Dr. Karim

In their research, they find a significant presence of Leishmania parasites in the urine of kala-azar patients. Furthermore, they attempt to confirm the presence of kala-azar parasites from the same patient’s bone marrow, blood, and urine samples. They employ both conventional PCR and real-time PCR methods simultaneously. The results demonstrate that it is feasible to confirm the presence of kala-azar parasites using samples from urine in addition to bone marrow and blood. This development is crucial as it opens up the possibility of using urine samples for kala-azar confirmation, especially in cases where bone marrow and blood samples may not be readily available.

Life cycle of the Leishmania parasite. Source: Nature Reviews Microbiology.

The researchers highlight that this new method has been applied not only in endemic regions like the Mymensingh belt but also in non-endemic regions like Dhaka and Chittagong, where individuals may not be infected with kala-azar but may suffer from other diseases such as dengue, malaria, chikungunya, or typhoid. This approach ensures that the method is specific to kala-azar and does not show sensitivity to other pathogens, making it a valuable tool in the confirmation of kala-azar in Bangladesh.

The success of a disease detection method is also evaluated by another crucial factor, namely sensitivity. This refers to how effective the method is in detecting the presence of a disease. It is observed that diseases diagnosed through bone marrow examinations and urinary tests have been successfully confirmed in patients. Therefore, it can be stated that the method is 100% effective when considering sensitivity.

In this context, bone marrow exhibits the highest sensitivity, followed by blood tests, and finally urine tests. Even if a Leishmania parasite is present in the body’s fluid with a volume of ten microliters, this innovative method can confirm the presence of kala-azar. The time required for sample collection to disease confirmation through this method is only three hours. Additionally, the cost incurred for this new method is significantly lower, ranging from six to seven taka, compared to the two thousand taka expense for kala-azar confirmation using traditional methods at Dhaka Medical College Hospital.

Therefore, it can be concluded that this newly developed method is not only highly effective in confirming kala-azar but is also cost-efficient. The research team is now considering international collaboration with African countries for sample collection, aiming for global outreach of this innovative method. They have already established communication with the University of Nairobi in Kenya, expressing their hope for collaboration.

Furthermore, policymakers in the public health sector in Bangladesh, who are involved in kala-azar confirmation, have expressed their interest in this advanced method. Although the government has already announced initiatives for kala-azar elimination, caution is advised to ensure the complete eradication not only of kala-azar but also other Neglected Tropical Diseases (NTDs) such as visceral leishmaniasis, soil-transmitted helminthiasis, etc., under the auspices of the Directorate General of Health Services (DGHS). Dr. Mohammad Manjurul Karim, the head of the research team, believes that if these diseases are eradicated, it will not only be a significant achievement in public health but will also benefit the tourism sector in Bangladesh.

In conclusion, the newly developed method demonstrates promising results in kala-azar confirmation. Its simplicity, cost-effectiveness, and potential for international collaboration make it a viable option for disease detection and elimination programs, contributing to improved public health outcomes.

Enthusiasts can refer to research papers published on this subject: 

  1. Khatun, Mahbuba, SM Sabbir Alam, Abed Hussain Khan, M. Anwar Hossain, Jalaluddin Ashraful Haq, Md Shariful Alam Jilani, Mohammad Tariqur Rahman, and Muhammad Manjurul Karim. “Novel PCR primers to diagnose visceral leishmaniasis using peripheral blood, spleen or bone marrow aspirates.” Asian Pacific journal of tropical medicine 10, no. 8 (2017): 753-759.
  2. Rahim, Samiur, Md Mohiuddin Sharif, Md Robed Amin, Mohammad Tariqur Rahman, and Muhammad Manjurul Karim. “Real Time PCR-based diagnosis of human visceral leishmaniasis using urine samples.” PLOS global public health 2, no. 12 (2022): e0000834.





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