Lutzomyia longipalpis and the eco-epidemiology of American visceral leishmaniasis, with particular reference to Brazil: a review

An historical review is given of American visceral leishmaniasis (AVL), with particular reference to the eco-epidemiology of the disease in Brazil. Following the first records of AVL in this country, in 1934, the sandfly Lutzomyia longipalpis (Lutz and Neiva, 1912) was incriminated as the principal vector. It is now generally accepted, however, that there exist a number of cryptic species under the name of Lu. longipalpis s.l. and that variations in the quantity of the vasodilatory peptide maxadilan in the saliva of flies from different populations of Lu. longipalpis s.l., may account for the variable clinical manifestations of AVL seen in different geographic regions. Distribution of AVL has been shown to extend throughout most of South and Central America, with the domestic dog serving as the principal reservoir of infection for man. However, while one hypothesis suggests that the causative parasite is Leishmania infantum, imported from Europe with the Portuguese and Spanish colonists, the demonstration of a high rate of benign, inapparent infection in foxes in Amazonian Brazil raised an opposing suggestion that the parasite is indigenous to the Americas. Recent reports of similar infections in native marsupials, and possibly rodents, tend to support this view, particularly as Lu. longipalpis is primordially a silvatic sandfly. Although effective control measures in foci of the disease will diminish the number of canine and human infections, the presence of such an enzootic in a variety of native animals will render the total eradication of AVL unlikely.

Epidemiology of visceral leishmaniasis through spatial analysis, in Belo Horizonte municipality, state of Minas Gerais, Brazil

The geographic information system approach has permitted integration between demographic, socio-economic and environmental data, providing correlation between information from several data banks. In the current work, occurrence of human and canine visceral leishmaniases and insect vectors (Lutzomyia longipalpis) as well as biogeographic information related to 9 areas that comprise the city of Belo Horizonte, Brazil, between April 2001 and March 2002 were correlated and georeferenced. By using this technique it was possible to define concentration loci of canine leishmaniasis in the following regions: East; Northeast; Northwest; West; and Venda Nova. However, as for human leishmaniasis, it was not possible to perform the same analysis. Data analysis has also shown that 84.2% of the human leishmaniasis cases were related with canine leishmaniasis cases. Concerning biogeographic (altitude, area of vegetation influence, hydrographic, and areas of poverty) analysis, only altitude showed to influence emergence of leishmaniasis cases. A number of 4673 canine leishmaniasis cases and 64 human leishmaniasis cases were georeferenced, of which 67.5 and 71.9%, respectively, were living between 780 and 880 m above the sea level. At these same altitudes, a large number of phlebotomine sand flies were collected. Therefore, we suggest control measures for leishmaniasis in the city of Belo Horizonte, giving priority to canine leishmaniasis foci and regions at altitudes between 780 and 880 m.

Phlebotominae spatial distribution asssociated with a focus of tegumentary leishmaniasis in Las Lomitas, Formosa, Argentina, 2002

Las Lomitas, Formosa, Argentina, reported 96 cases of tegumentary leishmaniasis during 2002. The urban transmission was suggested although previous outbreaks were related with floods of the Bermejo river (BR) 50 km from the village. Phlebotomine collections were performed during March 2002 to define the spatial distribution of risk, together with satellite imaginery. The phlebotomine/trap obtained was 1679.5 in the southern BR shore, 1.1 in the periruban-rural environment and 2.3 in the northern Pilcomayo river marshes. Lutzomyia neivai was the prevalent species (91.1%) among the 2393 phlebotomine captured, and it was only found in the BR traps. The other species were L. migonei (7.9%), L. cortelezzii (0.9%), and Brumptomyia guimaraesi (0.1%). The satellite images analysis indicates that the fishing spots at the BR were significantlyoverflowed during the transmission peak, consistent with fishermen recollections. This spatial restricted flood might concentrate vectors, reservoirs, and humans in high places. Therefore, both the spatial distribution of vectors and the sensor remoting data suggests that in Las Lomitas area the higher transmission risk it is still related with the gallery forest of the BR, despite of the urban residence of the cases. The surveillance and control implications of these results are discussed.

Polymerase chain reaction with lesion scrapping for the diagnosis of human American tegumentary leishmaniasis

The objective of this work was to compare the polymerase chain reaction (PCR) using lesion scrapping with other conventional techniques for the diagnosis of the American tegumentary leishmaniasis (ATL). For this, patients with cutaneous lesions suspected to be ATL were studied. The DNA was amplified with the MP1L/MP3H primers. From the 156 studied patients, 79 (50.6%) presented positive parasite direct search (PD), 81 (51.9%) had positive Montenegro skin test (MST), and 90 (57.7%) presented PD and/or MST positive. The PCR was positive in all of the positive-PD patients (100% sensitivity), in 91.1% of the positive PD and/or MST patients, and in 27.3% of the patients that presented negative PD and positive MST. The PCR positivity was similar to the PD (P = 0.2482) and inferior to the MST (P = 0.0455), and to the PD/MST association (P = 0.0133). The high PCR sensitivity, and positivity in those cases where the PD was negative, highlights the importance of this technique as an auxiliary tool for the diagnosis of ATL.

Tegumentary leishmaniasis outbreak in Bella Vista City, Corrientes, Argentina during 2003

Bella Vista City, Corrientes, Argentina, reported an epidemic outbreak of tegumentary leishmaniasis during 2003. The mean age of the 31 cases was 25.0 ± 13.7 years old, with a sex ratio male:female 1.8, and without mucosal involvement. They clustered in two contiguous neighbourhoods, 96% in the periurban border and 4% in the peripheral outskirts. The transmission peak was estimated to have occurred during April 2003. Four species (3608 sand flies) were captured in nine sites: Lutzomyia neivai (90.1%), Lu. pessoai (8.9%), Lu. migonei (0.8 %), and Brumptomyia avellari (0.2 %). The outskirts/rural capture ratio of Lu. neivai was up to 3, and the outskirts/periurban up to 200. Therefore, the 'urban' transmission in this southernmost known focus is still an ecotone-border associated risk. The changes in human distribution or activities, patches of the secondary vegetation, periurban streams, rainfall of the previous year, and river period floods could all contribute to 'urban' outbreaks in the region. Tegumentary leishmaniasis risk should be assessed for any project that involves changes in land use throughout an endemic area.

Single and concomitant experimental infectionsby Endotrypanum spp. and Leishmania (Viannia) guyanensis (Kinetoplastida: Trypanosomatidae) in the Neotropical sand fly Lutzomyia longipalpis (Diptera: Psychodidae)

Lutzomyia longipalpis females received single and mixed infections with Endotrypanum and Leishmania. Two biological parameters were analyzed: the percentage of infected females and the distribution of flagellates in the gut of the females. The principal comparisons were performed between (1) two strains of Endotrypanum, (2) cloned versus primary sample of one strain of Endotrypanum, (3) Endotrypanum versus Leishmania guyanensis, and (4) the pattern of flagellates behaviour by optical microscopy in females with single or mixed infection versus the identification of parasites isolated from digestive tracts by isoenzyme electrophoresis. Flagellates of Endotrypanum showed distinct patterns of infection suggesting that there is variation between and within strains. The distribution of Endotrypanum and L. guyanensis differed significantly in relation to the colonization of the stomodeal valve. In co-infection with L. guyanensis, a large number of flagellates were seen to be plentifully infecting the stomodeal valve in significantly more specimens than in females infected by Endotrypanum only. However, the electrophoretic profiles of isoenzymes of parasites recovered from all co-infected specimens corresponded to Endotrypanum. This suggests that the mere correlation sand fly infection-biochemical analysis of isolates may induce parasitological incorrect consideration.

Abundance of Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae) and urban transmission of visceral leishmaniasis in Campo Grande, state of Mato Grosso do Sul, Brazil

The outspread and urbanization of visceral leishmaniasis (VL) in Campo Grande, state of Mato Grosso do Sul, lead us to undertake the present study over diversity and abundance of sand flies in the urban area to compare with previous search carried out during 1999/2000, before the identification of the disease in the human population.The captures were carried out with automatic light traps, weekly, from February 2004 to February 2005 on three sites including a forested area (Zé Pereira), two peridomicilies (shelters of domestic animals and cultivation areas), and intradomicilie. In the present study 110 collections were obtained during 13 months for 1320 h of collections, resulting in 5004 specimens, 3649 males and 1355 females belonging to the 20 following species: Brumptomyia avellari, Brumptomyia sp., Bichromomyia flaviscutellata, Evandromyia lenti, E. termitophila, E. cortelezzii, E. borrouli, Lutzomyia sp., L. longipalpis, Micropygomyia quinquefer, N. antunesi, N. whitmani, Pintomyia christenseni, Pi. damascenoi, Psathyromyia aragaoi, Ps. campograndensis, Ps. hermanlenti, Ps. shannoni, Pychodopygus claustrei, and Sciopemyia sordellii. L. longipalpis was the most abundant species in the anthropic environment with 92.22% of the captures. This shows an increase of sixty times in the density of L. longipalpis compared to the last sand fly evaluation in 1999/2000. The high density of L. longipalpis in Campo Grande is the main factor of risk in transmission of the disease to human in the urban area. The capture of N. antunesi, typical specie from Amazonian region, in Mato Grosso do Sul is reported for the first time.

Leishmania (Viannia) lainsoni: occurrence of intracellular promastigote forms in vivo and in vitro

Experimental chronic (45-day-old) skin lesion in hamster hind foot induced by Leishmania (Viannia) lainsoni infection showed the presence of promastigote forms in the tissue, inside parasitophorous vacuoles, as assessed by transmission electron microscopy. Experimental in vitro interaction (24 and 48 h) between Leishmania (V.)lainsoni and J774-G8 macrophage cells also demonstrated the same profile. This morphological aspect is unusual, since in this parasite genus only amastigote forms have been described as the resistant and obligate intracellular forms.

Sensitivity and specificity of polymerase chain reaction in Giemsa-stained slides for diagnosis of visceral leishmaniasis in children

The aim of this study was to evaluate the sensitivity and specificity of polymerase chain reaction (PCR) in the detection of Leishmania DNA in archived Giemsa-stained bone marrow slides for diagnosis of visceral leishmaniasis (VL), and to compare PCR with conventional diagnostic techniques, like direct microscopy and parasite culture. Specimens of archived Giemsa-stained bone marrow slides from 91 patients with VL and from 79 controls with other diseases or conditions were studied. PCR showed the highest sensitivity (92.3%) and had good specificity (97.5%). Direct examination detected 79.1% and culture 59% of positive samples. In addition, PCR was able to detect VL in 16 of 19 patients (84.2%) with negative microscopy. PCR in Giemsa-stained bone marrow slides is a suitable tool for confirming diagnosis in patients with VL and may be useful in the diagnosis of difficult cases. Slide smears are easily stored, do not require special storage conditions such as low temperatures, and can be easily mailed to centers where PCR is available, making it an excellent option for diagnosis in the field.

Cloning and characterization of a V-ATPase subunit C from the American visceral leishmaniasis vector Lutzomyia longipalpis modulated during development and blood ingestion

Visceral leishmaniasis (VL) is a serious tropical disease that affects approximately 500 thousand people worldwide every year. In the Americas, VL is caused by the parasite Leishmania (Leishmania) infantum chagasi mainly transmitted by the bite of the sand fly vector Lutzomyia longipalpis. Despite recent advances in the study of interaction between Leishmania and sand flies, very little is known about sand fly protein expression profiles. Understanding how the expression of proteins may be affected by blood feeding and/or presence of parasite in the vector's midgut might allow us to devise new strategies for controlling the spread of leishmaniasis. In this work, we report the characterization of a vacuolar ATPase subunit C from L. longipalpis by screening of a midgut cDNA library with a 220 bp fragment identified by means of differential display reverse transcriptase-polymerase chain reaction analysis. The expression of the gene varies along insect development and is upregulated in males and bloodfed L. longipalpis, compared to unfed flies.

Polymerase chain reaction with two molecular targets in mucosal leishmaniasis' diagnosis: a validation study

We validated the polymerase chain reaction (PCR) with a composite reference standard in 61 patients clinically suspected of having mucosal leishmaniasis, 36 of which were cases and 25 were non-cases according to this reference standard. Patient classification and test application were carried out independently by two blind observers. One pair of primers was used to amplify a fragment of 120 bp in the conserved region of kDNA and another pair was used to amplify the internal transcript spacers (ITS) rDNA. PCR showed 68.6% (95% CI 59.2-72.6) sensitivity and 92% (95% CI 78.9-97.7) specificity; positive likelihood ratio: 8.6 (95% CI 2.8-31.3) and negative likelihood ratio: 0.3 (95% CI 0.3-0.5), when kDNA molecular target was amplified. The test performed better on sensitivity using this target compared to the ITS rDNA molecular target which showed 40% (95% CI 31.5-42.3) sensitivity and 96% (95% CI 84.1-99.3) specificity; positive likelihood ratio: 10 (95% CI 2.0-58.8) and negative likelihood ratio: 0.6 (95% CI 0.6-0.8). The inter-observer agreement was excellent for both tests. Based upon results obtained and due to low performance of conventional methods for diagnosing mucosal leishmaniasis, we consider PCR with kDNA as molecular target is a useful diagnostic test and the ITS rDNA molecular target is useful when the aim is to identify species.

Phlebotomine fauna (Diptera: Psychodidae) and species abundance in an endemic area of American cutaneous leishmaniasis in southeastern Minas Gerais, Brazil

This study was undertaken to identify the phlebotomine fauna and species abundance in domiciliary and peridomiciliary (hen-house and guava-tree) environments, on a lake shore, a cultivated area of coffee and banana, and a forested area of Conceição da Aparecida municipality, southeastern the state of Minas Gerais, to provide information for the control and epidemiological surveillance of leishmaniasis in this area. The captures were carried out monthly between May 2001 and November 2002, with automatic light and Shannon traps. A total of 1444 sand flies were captured, 951 (76.5%) with automatic light traps and 493 (23.5%) with the Shannon trap. Thirteen species were captured, the most frequent being Nyssomyia whitmani (62.7%), Migonemyia migonei (21.4%), Pintomyia fischeri (6.9%), and Evandromyia lenti (3.6%). Species abundance was determined using the automatic light traps installed in the six environments. The most abundant species according to the standardized index of species abundance were Ny. whitmani (1.0) and Mg. migonei (0.82). In view of the dominance of these two species, known vectors of cutaneous leishmaniasis in other Brazilian areas, their participation in the transmission of the disease in this county is suggested. The diversity and evenness indexes in the domicile were the lowest due to the high frequency (83%) of Ny. whitmani. The capture of Lutzomyia longipalpis, rarely recorded in the south-eastern and southern regions of Minas Gerais, is also noteworthy.

Phlebotomine sand flies (Diptera: Psychodidae) in the municipality of Várzea Grande: an area of transmission of visceral leishmaniasis in the state of Mato Grosso, Brazil

Visceral leishmaniasis (VL) has been naturally transmitted in periurban areas due to the emergence and reemergence of its vectors in such areas. Aimed to further knowledge on ecological aspects affecting the occurrence of phlebotomine sand flies in VL transmission areas in the municipality of Várzea Grande, state of Mato Grosso (MT), Brazil, sand fly captures were carried out. Monthly collections of sand flies were undertaken with CDC light-traps, which were left in both intradomiciliary and peridomiciliary areas of ten residences during four consecutive days between January 2004 and June 2006. Twenty-two species of genus Lutzomyia and one of Brumptomyia were captured. The most abundant species was Lutzomyia longipalpis (65.23%), followed by L. evandroi (16.26%), L. lenti (7.69%), L. whitmani (4.92%), L. sallesi (2.34%) and L. termitophila (1.32%). The highest density of the main VL vector, L. longipalpis, was found in peridomiciliary areas, mostly males. No significant correlation was found between environment (temperature, air relative humidity and rain fall) and phlebotomine density; although a slight increase in sand fly density has been observed in the period following rainfalls, particularly L. longipalpis. No correlation was observed between distribution and density of L. longipalpis, prevalence of human VL cases and the presence of serologically positive dogs. The presence of infected dogs, increased vector density, susceptibility rate and interruption of epidemiological surveillance may raise the risk of VL transmission to man in Várzea Grande.

First visceral leishmaniasis focus in Argentina

An eight-year old boy from Posadas (27º 23'S, 55º 54'W) was diagnosed with visceral leishmaniasis (VL) during 2006. Lutzomyia longipalpis was discovered in the backyard of his house, while the spread of canine visceral leishmaniasis was confirmed in Posadas. This is the southernmost report of a VL transmission focus and the first in Argentina.