Vector-Borne Viral Infections
Leishmaniasis
Disease Burden
Leishmaniasis is caused by several species of flagellated protozoan parasites, Leishmania spp, that are transmitted by the bite of a female phlebotome sandfly. The disease is found in many areas of the world, particularly in Africa, the Mediterranean Basin, South and Central Asia, the Middle East and Latin America. Several forms of the disease exist: cutaneous (CL), mucocutaneous (MCL) and visceral (VL, also called "Kala-azar"), which, after treatment, is often followed by a dermal manifestation known as post-kala-azar dermal leishmaniasis (PKDL). The typical lesion of CL is a chronic ulcer with intense lymphoid and monocytic infiltration and granuloma formation. VL is characterized by dissemination of the parasites through the reticuloendothelial system leading to pyrexia, wasting and hepatosplenomegaly.
For many years, the public health impact of leishmaniasis has been grossly underestimated, as a substantial number of cases were never recorded. WHO estimates the worldwide prevalence to be approximately 12-15 million cases, with annual mortality of about 60 000. The size of the population at risk is about 350 million
[94]
. About 1.5 million new cases are estimated to occur annually, but only 600 000 are officially declared. In addition, deadly epidemics of VL periodically flare up but go mostly unnoticed in spite of case-fatality rates as high as 10% or more. In the 1990s Sudan suffered a crisis with an excess mortality of 100 000 deaths among people at risk. There are an estimated 200 million population at risk on the Indian subcontinent, which reports 25 000 to 40 000 cases and 200-300 deaths every year [95]. A VL elimination program is currently being launched in India, Bangladesh and Nepal [96].
The expansion of leishmaniases and the alarming rise in the number of cases is related to environmental changes such as deforestation, building of dams, new irrigation schemes and migration of non-immune people to endemic areas, which resulted in significant delay in the implementation of development programs in Saudi Arabia, Morocco, the Amazons and the tropical regions of the Andean countries. More recently, as a result of epidemiological changes, a sharp increase in the overlapping of HIV infection and visceral leishmaniasis has been observed, especially in intravenous drug users in southwestern Europe. The situation might worsen in Africa and Asia where the prevalence of HIV and Leishmania co-infections still is probably largely underestimated.
Parasitology
Leishmaniasis is a parasitic infection caused by the obligate, intracellular protozoan of the genus Leishmania, fifteen species of which can infect man. Main species in the Old World are: L major, L tropica, L donovani and L infantum and in the New World: L mexicana, L brasiliensis, L amazonensis and L guyanensis. The parasite has two life cycles, one in the alimentary canal of female phlebotome sandflies where it forms motile flagellated and elongated 'promastygotes' that are inoculated into the blood stream of a mammalian host when the female is having a blood meal; and a mostly intracellular cycle in the mammalian host where it matures inside macrophages, neutrophils and dendritic cells into tiny ovoid shape 'amastigotes' with a nucleus, a kinetoplast and a short flagellum
[97]
. The kinetoplast, which contains mitochondrial DNA, is a distinctive feature of both Leishmanias and Trypanosomias.
Recent studies have evidenced the existence of interspecific and intraspecific hybrids and recombinant genotypes among the natural populations of Leishmanias
[98]
[99]
, suggesting the formation of haploid gametes through meiosis and the generation of heterozygous progeny in the sandfly vector
[100]
.
Cutaneous (CL) and mucocutaneous (MCL) leishmaniasis in Central and South America are caused by members of the L. mexicana and L. braziliensis species, whereas CL in South and Central Asia and the Middle East is caused by L. tropica and L. major. The majority of MCL cases occur in Bolivia, Brazil and Peru, whereas 90% of CL cases occur in Afghanistan, Iran, Saudi Arabia, Syria, Brazil and Peru. VL (kala-azar), the most lethal form of the disease, is caused by L. donovani in Sudan, India, Nepal, Bangladesh and China, by L. infantum in North Africa and southern Europe, and by L. chagasi in Latin America. Transmission is most often zoonotic: the parasites (Leishmania) are transmitted from a wild animal reservoir (small rodents, dogs) by the bite of the female phlebotome sandfly. It also can be anthroponotic, the parasite being transmitted by the sandfly from an infected human host. In the Mediterranean Basin and Brazil, the main identified reservoir is the dog population, and the most effective control strategy has been the use of deltamethrin-impregnated dog collars
[101]
[102]
.
Cutaneous leishmaniasis usually leads to self-healing disease with life-long immunity against reinfection. Resolution is characterized by induction of IFN-? releasing CD4+ T cells of the Th1 phenotype whereas failure to cure is associated with elevated levels of IL-4 and IL-10 and low levels of IFN-?, a typical Th2 profile
[103]
[104]
[105]
. It was demonstrated in experimental animal models that a dominant Th1 lymphocyte response (IL-2, IFN-?) is associated with self-limited disease, whereas a dominant Th2 response (IL-4, IL-5) is linked to progressive disease.
Vaccines
Vector and reservoir controls may be useful under certain conditions but are not applicable in every epidemiological setting and require infrastructure and vigilance beyond the capability of many endemic countries. Vaccination, therefore, remains the best hope for control of all forms of the disease. Progress towards a vaccine however has remained very slow, even though evidence from animal studies demonstrate that protection can be achieved through immunization with purified proteins or DNA vaccines
[106]
. There is as yet no effective vaccine for prevention of any form of leishmaniasis.
Leishmanization
For centuries, in some of the hyper-endemic areas of the Middle East, the pus of an active lesion was used to inoculate young children to protect them against future lesions on the exposed parts of the body, especially the face. The practice is known as leishmanization. Leishmanization has been practiced in several countries, including Uzbekistan, the only country where the measure is still in current use. L. major promastigotes grown in culture under GMP guidelines, rather than the exsudates from active lesions, have more recently been used for inoculation as a live vaccine. Genetically manipulated parasites with attenuated virulence or high sensitivity to chemotherapy might represent the ideal form of such a live vaccine
[107]
.
Inactivated whole-cell vaccines
In recent years most clinical trials of first generation vaccines in humans have evaluated the effect of three types of vaccines: a L. amazonensis-based vaccine derived from an earlier 5-valent vaccine (BIOBRAS, Brazil), a L. mexicana-based product (Instituto Biomedicina, Venezuela) and a L. major-based preparation (Razi Vaccine and Serum Research Institute, Iran). In addition, a trivalent preparation consisting of L. brazilensis, L. guyanensis and L. amazonensis antigens was evaluated in Ecuador [23]. Bacille Calmette-Guérin (BCG) was used as the adjuvant in some versions of the Venezuelan, Ecuadorian and Iranian candidate vaccines in an attempt to improve the vaccine's ability to induce cell mediated responses
[108]
. In trials of the combination of autoclaved L. major promastigotes with BCG as adjuvant by the Razi Institute in Iran against CL and in Sudan against VL, a limited efficacy was noted in converters to positive skin reaction to leishmania antigen (leishmanin) and unexpectedly in boys
[109]
. Alum-precipitated autoclaved L. major promastigotes plus BCG have demonstrated safety and substantial immunogenicity in Phase I and II studies in Sudan and in Iran
[110]
. It is of note that treatments combining administration of antimonials and first generation leishmania vaccines in patients suffering from post-Kala-Azar dermal leishmaniasis (PKDL) have shown benefit to the patients, suggesting that even suboptimal leishmaniasis vaccines could have a role in a therapeutic setting.
Subunit vaccines
Various subunit recombinant candidate vaccines also were tested in mice and provided some degree of protection against infection. These vaccines were based on a variety of Leishmania gene products including:
- the gp63 surface antigen, a glycoprotein with protease activity,
- lipophosphoglycan, a surface glycoconjugate;
- a 46 kD promastigote antigen derived from L. amazonensis;
- the Leishmania-activated C kinase (LACK)
Protection against L. major infection in mice was provided by DNA constructs encoding a number of such Leishmania antigens, including gp63 and LACK. A prime-boost vaccination regimen against experimental VL was tested in dogs with DNA/recombinant cysteine proteinases I and II and showed partial protection
[111]
. A prime boost DNA/ modified vaccinia virus Ankara (MVA) expressing leishmania tryparedoxin peroxidase (TRYP) was safe and immunogenic in dogs
[112]
. Further dog trials are eagerly awaited. There also is evidence that a 15 kD protein antigen derived from the salivary glands of the sandfly vector could be protective in mice when given as a vaccine
[113]
. The fact that the nucleotide sequence of the 8500 identified genes in the Leishmania (L major) genome is now known
[114]
leads to hope that screening of new potential vaccine candidates will be accelerated
[115]
.
A DNA vaccine encoding leishmania antigens LACK, TSA, LmSTl1 and CPa elicited only partial protection against hind footpad challenge in mice
[116]
. However, addition of Th1-driving adjuvants such as IL-12 or oligodeoxynucelotides (CpG) to the leishmanial antigens resulted in complete protection of susceptible mice against progressive disease, whereas no protection was observed in the absence of adjuvant
[117]
. The development of a chimeric vaccine made of the three recombinant leishmanial antigens LeIF, LmSTI-1 and TSA in the form of a fusion protein (Leish-111f) combined with monophosphoryl lipid A (MPL) in squalene oil as adjuvant was launched by a group at the Infectious Disease Research Institute in Seattle (WA) with the support of the Bill and Melinda Gates Foundation
[118]
. Phase I trials of the vaccine in healthy volunteers in the USA and initial efficacy testing as a therapeutic vaccine in patients in Latin America suggest safety and immunogenicity of the vaccine [94].