INTRODUCTION cases of leishmaniasis reported every year as

INTRODUCTION

Leishmaniasis is caused by a parasitic protozoan of genus Leishmania, belonging to family Trypanosomatidae, they are characterized by possessing a kinetoplast a unique form of mitochondrial DNA. Their primary hosts are vertebrates; commonly infects hyraxes, canids, rodents, and humans. About 21 species of Leishmania is found to be pathogenic in humans. Leishmania is transmitted to human by the bite of vector sandflies of genus Phlebotomus in the old world, and Lutzomyia in the new world.

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Leishmaniasis can occur in three main forms: (i) Cutaneous leishmaniasis (most common form) caused by L.mexicana, L.major, L.aethiopica, L.infantum, L.tropica, L.amazonensis, L.braziliensis, L.guayanensis, L.panamesis, L.peruviana,(ii) Mucocutaneous leishmaniasis caused by L.braziliensis, L.panamensis, L.guayanensis  and  (iii) Visceral leishmaniasis which is caused by L.infantum, L.donovani  and L.chagasi.

There is a fourth form known as diffuse cutaneous leishmaniasis (DCL), caused by L.aethiopica and L.amazonensis. VL is the most severe form of the disease; Post-kala-azar dermal leishmaniasis is the complication of leishmaniasis which is characterized by macular, macula-papular, nodular rash on face, torso, or other parts of the body. Ehab.et.al.,2014.

Symptoms of the visceral form of the disease include prolonged fever, progressive anemia, substantial weight loss, splenomegaly, hepatomegaly, and hypergammaglobulinemia. It is the most severe form of the disease and if left untreated can be fatal. Shyam Sundar.et.al.,2002.

Epidemiology

There are about 900,000 to 1.3 million new cases of leishmaniasis reported every year as estimated by World Health Organization, approximately 0.7-1.2  million cases are of cutaneous and 0.2-0.4 million are of visceral leishmaniasis. Lindoso.et.al.,2016.

Increasing cases of CL have been reported from non-endemic countries due to travelings, such as a trip to new places and ecotourism. Imai.et.al.,2017.

90% of VL cases have been reported from East Africa, Brazil and Indian subcontinents, whereas elevated CL cases are reported from Latin America, Middle East, and central Asia. Muccio.et.al.,2015.

Epidemiology of Indian VL shows a shift from east to west along Ganges River, a new shift has been reported in eastern Uttar Pradesh, Himachal Pradesh and Uttarakhand and these foci have now become endemic for VL. Sarnam Singh.,2014.

It accounts for 58,000 new cases every year and affecting 79 countries. It is a health problem of poor and neglected populations. Zineb Tlamcani.,2016.

It affects 6 million people worldwide in 98 countries with 0.9-1.6 million cases each year.  Disease spectrum of leishmaniasis can range from self-healing skin lesions to fatal disease affecting the visceral organs called visceral leishmaniasis (VL) commonly known as kala-azar, a Hindi term meaning black fever, this term was first used in India before which the manifestations of the disease were known by other names. Singh.et.al.,2004.

The disease is affecting people in different regions of the world but occurs predominantly in tropics and subtropics, Southern Europe and Asia, South and Central America, the Middle East. This disease affects people in nearly 88 developing and developed countries where about 350 million people are living. Mohamed.et.al.,2013.

The spread of the disease is due to changes in environment like deforestation, changing the environment, building of dams etc.  It mainly affects poor people with poor housing, and lack of financial resources.

Most of the cases are from seven main countries: India, Sudan and South Sudan, Bangladesh, Brazil, Ethiopia, Nepal. Six countries are endemic to VL in Africa: Somalia, Kenya, Uganda, Sudan and South Sudan, Ethiopia. The people living in remote and rural areas are generally affected by the disease. If left untreated it can result eventually in death.

In southern Europe, 25-70 % adults with VL have AIDS, where Leishmania take advantage of a host with the weak immune system. The presence of parasite in peripheral blood, HIV infected patients make them a source of infection for the vectors. Shyam Sundar.et.al.,2002.

Some risk factors for VL include immunosuppressive drugs, malnutrition and HIV co-infection. The numbers of cases of co-infections are increasing in Brazil and India where urban HIV and rural VL are coming in contact. Tavares.et.al.,2003.

The parasites exist in two forms: amastigotes form and promastigotes form.

Amastigotes are non- flagellated ovoid forms, measuring 3-5?m with a central ovoid nucleus and adjacent kinetoplast. The flagellum is not functional and does not extend beyond the body. Cytoplasm contains both rough and smooth endoplasmic reticulum. Promastigotes forms have elongated cell body. Being motile these cells bears functional flagellum. Sarnam.et.al.,2014.

Life Cycle and Transmission of Leishmania

Transmission of Leishmania occur during blood meal of infected female sand fly Phlebotomus which inoculates the promastigotes, the parasites infect the man in promastigote form. After inoculations promastigotes are taken up by phagocytic cells and become amastigotes.

Amastigotes multiply in macrophages; these intracellular amastigotes are ingested by the sand fly. Amastigotes get transformed into promastigotes in the gut of sand fly and migrate to the mouth of the fly.

Vector-mediated transmission is most common form but transmission of Leishmania occur through other methods also like transfusion mediated transmission in which parasites keep circulating in blood before the appearance of symptoms and these asymptomatic donors can be the reason for transmission. Needle sharing is also one of the common modes in some cases.  Congenital transmission infection occurs in most cases during birth. In sexual transmission, Promastigotes are observed from the culture of urine and prostatic fluid of VL patients, reports of transmission of promastigotes from man to his wife has been observed. Person-to-person transmission: this mode of transmission is possible via contact with infected fluids of patients with VL. Mohamed.et.al.,2013.

 Leishmaniasis in AIDS patients

Leishmania in HIV infected patients has emerged as a major complication of leishmaniasis. Most of the countries endemic to leishmaniasis are facing HIV epidemics which are resulting in higher rates of co-infections. Sarnam.et.al.,2006.

Leishmaniasis behaves as an opportunistic infection for AIDS. The presence of the parasite in peripheral blood makes these patients more vulnerable to the vectors.

In co-infected patients, HIV modifies the presentation of Leishmania.

VL increases the rate of onset of AIDS and decreases the life time of HIV infected person. Also, HIV increases the rate of VL to great extent. This dual response leads to the deficiency in CD4+ T cell immune response, increasing the severity of the disease. Sarnam Singh.et.al.,2014.

It has been observed that lipophosphoglycan (LPG) on L.donovani amastigote surface leads to CD4+T cell activation and leishmania antigen activates tumor necrosis factor (TNF-a) which help in HIV-1 replication. Wolday D.et.al.,1994.

The reason of Leishmaniasis being a diagnostic challenge is because of the wide spectrum of clinical manifestations that are present: skin lesions at the site of the sand fly bite (cutaneous leishmaniasis), multiple nonulcerative nodules (diffused cutaneous leishmaniasis), mucosal inflammation (mucosal leishmaniasis) and visceral infections (visceral leishmaniasis). Maurya.et.al.,2005

These signs complicate clinical diagnosis even more. For example lesions in CL can vary in their appearance (wart-like, open ulcers), severity (lesion size) and duration. Several species can affect children and adults. The first sign of infection is small erythema at the site of the bite. Parasites will cause an inflammatory reaction due to which erythema develop into an open ulcer or viceralize to spleen, liver. Richard.et.al.,2007

An incorrect diagnosis can prove fatal. As in a case in New York, a child belonging to a leishmaniasis prone area was misdiagnosed by microscopic based examination of intramacrophage and he was treated with antimonials due to which child died of fungal infection. Steven.et.al.,1996.

Diagnosis

The early diagnosis of leishmaniasis is important in order to avoid serious damage or death of the patient. Conventional diagnosis relies on microscopical examination of amastigotes in aspirates from tissues. However this procedure is not comfortable for the patients and the method of isolating parasite from culture is also time-consuming, expensive and difficult. Schallig.et.al.,2002 .

The diagnosis of the disease is complex because its clinical symptoms are common with other commonly occurring diseases like tuberculosis, typhoid, and malaria. Many of these diseases can occur as co-infection. Sundar.et.al.,2002 .

Many primary and secondary skin conditions are diagnosed as leishmaniasis in epidemic countries and misdiagnosed as another disease in nonendemic countries. Singh.et.al.,2006.

In Leishmania, HIV co-infection diagnosis is further more difficult because the clinical manifestations of the disease which occur in leishmaniasis do not develop in immunocompetent patients and these co-infected patients do not respond to antileishmanial treatments.  Deniau.et.al.,2003.

Early detection of infected animals even before the appearance of disease is important because some asymptomatic animal can serve infectious to the vector although they have lower risk than symptomatic ones. Maia.et.al.,2008.

Diagnostic tests play a major role in the screening of asymptomatic infections, screening of the patients, epidemiological studies. A diagnostic test should make a difference between asymptomatic infections and acute disease. Srivastava.et al.,2011.

Routine diagnosis of leishmaniasis depends upon the microscopical examination of amastigotes in aspirates from lymph nodes, bone marrow, skin, liver, and on culturing parasites from these sites. However, this procedure is uncomfortable for patients, difficult and economical.                             Mugasa.et.al.,2010.

But it presents low sensitivity due to few numbers of parasites in lesion areas.  Costa.et.al.,2016.

Early diagnosis and treatment serve as important aspects for controlling leishmaniasis.

The diagnosis of the disease is based on laboratory testing and clinical features. A number of diagnostic methods have been produced with huge variations in accuracy of diagnosis, including parasitological examination (histopathology, microscopy and parasite culture) and with serology and molecular diagnostics. Vries.et.al.,2015

Diagnosis relies mainly on histological examination of skin biopsy and serology. Histology can be made easy by immunohistochemistry, intracellular amastigotes are difficult to identify in sections containing few number of parasites.

Montenegro skin test (MST) is another method used which measures delayed-type hypersensitivity (DTH) reactions to leishmania antigen injected intradermally, this procedure requires a trained technician who can assure interpretation of results, the concentration of injected antigen and quantity. Antonio.et.al.,2014.

MST cannot distinguish patients with acute symptoms from those under treatment or cure. Viana et al., 2011.

A serological test is more beneficial because of them being less invasive and easy to perform, but their sensitivity is low as they present low titers of antibody and thus noninfected individuals in endemic areas can give false positive results. Malchiodi.et.al.,1994.

Techniques based on polymerase chain reactions (PCR) have been used recently with high specificity sensitivity. PCR based methods can be applied to many different types of biological samples like serum, blood, and bone marrow. Fraga.et.al.,2010.

The use of PCR techniques for identification helps in detecting parasites from different sources of samples including urine. This method has been proved successful for detecting DNA from both humans and canines Fisa.et.al.,2008.

Direct agglutination test (DAT) based on whole promastigotes of L.donovani and immunochromatographic test (ICT) based on rK39 are used in the primary diagnosis of VL because they are simple and easily applicable in field settings.Srividya G.et.al.,2012.

The disadvantage of this method is that it cannot differentiate relapsed from active VL cases. Salotra.et.al.,2006.

The aim of this paper is to highlight some of the problems with diagnosis of leishmaniasis and focus on what can be the most successful methods for developing fast, economical, specific and sensitive diagnostic tests

 

Several methods used for diagnosis are:

•    Microscopic examination of parasites in the tissue of relevance.

•    Antileishmanial antibody detection in patient’s sera.

•    Detection of parasite antigen in patient sera, urine or blood samples.

•    Detection of parasite DNA in tissues.

•    Detection of cell-mediated immunity.

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