Tegumentary leishmaniasis by Leishmania braziliensis complex in Cochabamba, Bolivia including the presence of L. braziliensis outlier: Tegumentary leishmaniasis in Cochabamba, Bolivia: Tegumentary leishmaniasis in Cochabamba, Bolivia.

Leishmaniasis is caused by protozoans of the Leishmania genus, which includes more than 20 species capable of infecting humans worldwide. In the Americas, the most widespread specie is L. braziliensis, present in 18 countries including Bolivia. The taxonomic position of the L. braziliensis complex has been a subject of controversy, complicated further by the recent identification of a particular subpopulation named L. braziliensis atypical or outlier. The aim of this study was to carry out a systematic analysis of the L. braziliensis complex in Bolivia and to describe the associated clinical characteristics. Forty-one strains were analyzed by sequencing an amplified 1245 bp fragment of the hsp70 gene, which allowed its identification as: 24 (59%) L. braziliensis, 16 (39%) L. braziliensis outlier, and one (2%) L. peruviana. In a dendrogram constructed, L. braziliensis and L. peruviana are grouped in the same cluster, whilst L. braziliensis outlier appears in a separate branch. Sequence alignment allowed the identification of five non-polymorphic nucleotide positions (288, 297, 642, 993, and 1213) that discriminate L. braziliensis and L. peruviana from L. braziliensis outlier. Moreover, nucleotide positions 51 and 561 enable L. peruviana to be discriminated from the other two taxa. A greater diversity was observed in L. braziliensis outlier than in L. braziliensis-L. peruviana. The 41 strains came from 32 patients with tegumentary leishmaniasis, among which 22 patients (69%) presented cutaneous lesions (11 caused by L. braziliensis and 11 by L. braziliensis outlier) and 10 patients (31%) mucocutaneous lesions (eight caused by L. braziliensis, one by L. braziliensis outlier, and one by L. peruviana). Nine patients (28%) simultaneously provided two isolates, each from a separate lesion, and in each case the same genotype was identified in both. Treatment failure was observed in six patients infected with L. braziliensis and one patient with L. peruviana.

Phenotypical and genotypical differences among Leishmania (Leishmania) amazonensis isolates that caused different clinical frames in humans and dogs: A systematic review.

Leishmania (Leishmania) amazonensis is an important etiological agent of American cutaneous leishmaniasis (ACL) in Brazil. The species causes a large spectrum of clinical manifestations in humans and dogs, ranging from cutaneous, cutaneous diffuse, mucocutaneous, and visceral involvement, however, the factors that drive the development of different disease forms by the same species are not yet fully known. In the present work, it was systematically reviewed the studies addressing phenotypic and genotypic characteristics of Leishmania (L.) amazonensis isolates causing cutaneous and visceral clinical frames in humans and dogs, comparing the results observed. For this, four research databases were searched for the following keywords: (Leishmania amazonensis AND visceral leishmaniasis) AND (tropism OR virulence OR visceralization OR adaptations OR mutation OR clinical presentation OR resistance OR survival OR wide spectrum). The results revealed that the complexity disease seems to involve the combination of genetic factors of the parasite (as modifications in molecules related to the virulence and metabolism) and also of the host's immune background and status. Nonetheless, the exact mechanism that leads to different clinical manifestations between strains of the same species is still uncertain and future studies must be developed to better elucidate this phenomenon.

Molecular Evidence of Leishmania infantum and Leishmania guyanensis in Red Howler Monkey (Alouatta seniculus) from French Guiana.

Presence of Leishmania spp. was evaluated in the blood of nine red howler monkeys (Alouatta seniculus) from a specific area of French Guiana, located in the northeast of the Amazon. The molecular detection was performed based on PCR targeting the markers 18S rRNA, kDNA and ITS2 genes, as well as rapid immunomigration tests. Two monkeys were positive for Leishmania infantum and one for Leishmania guyanensis. While L. guyanensis cutaneous leishmaniasis is common, visceral leishmaniasis (human and canine) caused by L. infantum has never been described in this area. The howler monkey proved to be a sentinel and a potential reservoir of a serious zoonosis. These results must be carefully considered by public health officials and veterinarians in the future.

Natural cutaneous leishmaniasis among dogs in Panama.

A search for cutaneous leishmaniasis among dogs was conducted in several forest settlements of central Panama from 1968 to 1973. A total of 11 (3.3%) of 333 animals examined was found infected and, in 9 of these, parasites were isolated in culture and characterized as Leishmania braziliensis. Infected dogs occurred in three settlements, one of which was free of human leishmaniasis. In the latter case, infections were acquired in the nearby forest during hunting excursions. Ulcerated skin lesions primarily on the lower aspect of ears, or depigmentation and inflammation of the nostrils were manifested, with persistence as long as 45 months. The parasites did not disseminate from the lesion to viscera or other areas of the skin. The dog may serve as an incidental reservoir host of human leishmaniasis and/or a liaison of the infection between the jungle and forest settlements in the Republic of Panama.

Leishmania braziliensis in the Panamanian two-toed sloth, Choloepus hoffmanni.

A total of 498 two-toed sloths, Choloepus hoffmanni, collected in central Panama was examined for Leishmania braziliensis over a 10-year period. Isolations of the parasite from 96 (19.3%) of the animals were confirmed by culture and inoculation of golden hamsters. Improved culture techniques developed toward the end of the study assisted in determining a greater prevalence of the disease. Infectins were completely cryptic in all animals, and the parasite was isolated from skin, blood, liver, spleen, bone marrow and lung tissues. Sloths maintained under seminatural conditions remained infected up to 23 months, the longest period of survival. This edentate, considered the principal reservoir host of L. braziliensis in Panama, showed infection rates from 0-59.4% in various communities, which appeared to correlate with the parasite prevalence in the indigenous human populations.

Susceptibility of Aotus trivirgatus to Leishmania braziliensis and L. mexicana.

Two Aotus trivirgatus (owl monkeys) were infected experimentally with Leishmania braziliensis and two with L. mexicana strains of Panamanian origin in a pilot study to determine the susceptibility and the course of infection of cutaneous leishmaniasis in this primate species. Montenegro skin tests performed on all animals prior to parasite inoculation were negative. A standardized inoculum of promastigotes was injected intradermally on the nose of each monkey. All of the animals developed infections which lasted from 3.5 to 8.5 months. Depigmentation developed at the site of the inoculation in all of the subjects. The severity of the resulting lesions was greater in the animals infected with L. braziliensis. Positive skin tests developed in three A. trivirgatus at days 62, 76, and 139 postinoculation, respectively. An explanation for the negative skin test in the fourth animal is discussed.

Canine American cutaneous leishmaniasis: a clinical and immunological study in dogs naturally infected with Leishmania braziliensis braziliensis in an endemic area of Rio de Janeiro, Brazil.

Clinical and immunological findings from 35 dogs infected with Leishmania braziliensis braziliensis are described. The majority of the dogs had ulcerated single lesions on the ears. Sera from all infected dogs showed detectable Leishmania-induced antibodies using an indirect fluorescent antibody test. Antimonial therapy resulted in prompt healing of the lesions in 80.9% of the animals followed by a significant reduction in the anti-Leishmania antibody titers. However, treatment follow-up showed recurrences at the site of the primary lesion in 42.8% of the cases. These data were correlated with a persistence of the parasite in clinically healed lesions as well as with a negative intradermal test (leishmanin-delayed type hypersensitivity) observed in all animals but one.

Distribution and etiology of leishmaniasis in Colombia.

A total of 340 Leishmania strains, isolated from humans, animals, and sand flies from various regions of Colombia, were examined by isozyme electrophoresis. Seven different Leishmania species were identified. Leishmania panamensis and L. braziliensis were the most common, representing 53.8% and 30.3% of the total, respectively. Isolation rates of the other species were as follows: L. chagasi, 9.4%; L. guyanensis, 2.6%; L. amazonensis, 1.8%; L. mexicana, 0.8%; and a new species requiring additional study, 1.2%. Statistical analyses of representative L. panamensis and L. braziliensis isolates indicated that the populations of these 2 species are genetically very similar. L. panamensis may have a continuous distribution in Colombia west of the eastern Andes Mountains and L. braziliensis may have a continuous distribution east of the western Andes Mountains. Information is given on disease manifestations of the parasites in human hosts and on isolation records from sand flies and animals.

Leishmaniasis in Brazil: XVI. Isolation and identification of Leishmania species from sandflies, wild mammals and man in north Para State, with particular reference to L. braziliensis guyanensis causative agent of "pian-bois".

A total of 125 wild mammals (14 different species) were examined for evidence of infection with Leishmania in an area of primary forest highly endemic for "pian-bois", due to Leishmania braziliensis guyanensis, in north Pará State, Brazil. Parasites isolated were characterized biologically, and biochemically on enzymic profiles. L. b. guyanensis was isolated from the viscera of one lesser anteater (Tamandua tetradactyla) and one opossum (Didelphis marsupialis), and the skin of one rodent (Proechimys guyannensis). The isolates were indistinguishable from 10 others previously made from the sandfly vectors Lutzomyia umbratilis (five) and Lu. whitmani (five), and nine isolates from field-workers who became infected during these studies. Leishmania mexicana amazonensis was obtained from the skin of 21 animals, including three species of opossums (D. marsupialis, Philander opossum and Metachirus nudicaudatus) and two species of rodents (proechimys guyannensis and Dasyprocta sp.). A peripylarian Leishmania isolated from the viscera of two armadillos (Dasypus novemcinctus) was shown to be different, biologically and biochemically, from L. b. guyanensis and L. m. amazonensis. Four other isolates of Leishmania, from the rodents Rhipidomys leucodactylus (one) and P. guyannensis (three) have yet to be characterized owing to their very poor growth in both hamster skin and in vitro culture: they appear closest, however, to L. braziliensis braziliensis. The complexity of Amazonian leishmaniasis is discussed, and attention drawn to the importance of edentates as reservoir hosts of some leishmanias in the New World. Whereas L. mexicana subspecies appear largely restricted to the skin of their natural hosts, subspecies of L. braziliensis are commonly found in the viscera.

A focus of mucocutaneous leishmaniasis in Três Braços, Bahia, Brazil: characterization and identification of Leishmania stocks isolated from man and dogs.

The characterization and identification to species and subspecies of 20 stocks of Leishmania isolated from the region of Três Braços, Bahia, Brazil, are described: 17 stocks were from patients and three from dogs. The following techniques were used (i) biological (growth in culture, hamster tissues and phlebotomine gut), (ii) biochemical (isoenzyme and kinetoplast DNA analysis) and (iii) immunological (using monoclonal antibodies). All except two stocks belong to the L. braziliensis complex. One of these two corresponded to L. mexicana amazonensis but the other, while clearly in the mexicana complex, showed slight differences from the L. mexicana amazonensis reference strain on isoenzyme analysis. Two stocks from different lesions in the same patient and with different growth characteristics in hamster tissues were both identified as L. braziliensis braziliensis. All the fully characterized stocks of the L. braziliensis complex were identified as L. braziliensis braziliensis. L. braziliensis guyanensis was not identified. Dog and human stocks of L. braziliensis braziliensis were indistinguishable. From these findings and other evidence, L. braziliensis braziliensis seems to be the predominant species transmitted in Três Braços.

Experimental superimposed infection of the hamster with Leishmania mexicana and L. braziliensis.

Hamsters inoculated subcutaneously with Leishmania mexicana (L. m. amazonensis or L. m. mexicana) and with L. braziliensis panamensis, either simultaneously or with an interval, in different parts of the body (right front paw and nose), showed an independent course of infection for either parasite with the typical clinical and histopathological characteristics produced by these organisms. In one group of animals, metastases from the paw (L. m. amazonensis) to the nose (L. b. panamensis) was proven and the partial transformation of the granuloma from epithelioid and giant multinuclear cells (tuberculoid) to histiocytes full of parasites (lepromatous), was observed. These two specific contrasting reactions, are confined to the place of infection and influenced by the type of parasite. The histopathological picture normally observed in hamsters as a reaction to L. mexicana s.1., occurs only occasionally in humans. It is suggested, on the basis of this animal model, that DCL in the New World is only produced by parasites of the "mexicana" complex and that both parasite characteristics and host immune status are responsible for this phenomenon.

Schizodeme and zymodeme characterization of Leishmania in the investigation of foci of visceral and cutaneous leishmaniasis.

Leishmania parasites were isolated from humans and canines in foci of cutaneous and visceral leishmaniasis. After in vitro cultivation the parasites were examined by the following biochemical techniques: (i) restriction analysis of kinetoplast DNA (kDNA) also known as schizodeme analysis (Morel et al., 1980); (ii) zymodeme analysis (Barret et al., 1980); by agarose gel electrophoresis and (iii) isoelectricfocusing in polyacrylamide gels. The strains of cutaneous and visceralizing leishmanias studied could be differentiated by schizodeme analysis, using the endonuclease MspI, into three complexes agreeing with those accepted for human New World leishmaniasis. In the municipality of Rio de Janeiro, isolates from a focus of cutaneous leishmaniasis were identified as L. braziliensis braziliensis and from a focus of visceral leishmaniasis were identified as L. donovani by zymodeme characterization. Identical restriction enzyme profiles of kDNA from human and canine isolates indicated that in the cutaneous focus at Jacarepaguá, Rio de Janeiro, the same strain was probably circulating in both the canine and human populations. This suggests a possible role for dogs as a reservoir host for L. braziliensis braziliensis. In addition, our results confirm the importance of dogs as reservoirs in visceral leishmaniasis. The stability of the electrophoretic patterns of restriction digest ("fingerprints") of Leishmania kDNA as well as differences in the sensitivity of the techniques used were demonstrated. Strains from widely different geographical areas as well as strains maintained in vivo and in vitro showed identical kDNA restriction patterns, while strains showing similar banding patterns by enzyme electrophoresis could be differentiated by schizodeme analysis. These results demonstrate the usefulness of an integrated biochemical approach in the identification of Leishmania.

[Mucocutaneous leishmaniasis in naturally infected dogs in Salta, Argentina]. / Leishmaniosis mucocutánea en perros naturalmente infectados en Salta, Argentina.

The objective of the present study is to describe two cases of dogs with mucocutaneous lesions caused by Leishmania spp. Both dogs presented destruction of the nasal septum, hyperemia with soft palate edema and barking alteration due to laryngeal compromise. Biopsies were taken from the lesion border and Leishmania spp. amastigotes were seen in the imprints. The dogs presented positive serology when complex soluble antigen from Leishmania mexicana was used. One of the dogs was also suspected to be infected by Trypanosoma cruzi as suggested by its positive reaction with a purified specific antigen, Ag163B6-cruzipain. Most of the studies concerning leishmaniosis in dogs have described the cutaneous form of this disease in close association with human cases of Leishmania infection instead of the mucocutaneous form described herein. The presence of dogs with mucocutaneous leishmaniosis alerts on an increase of the prevalence of this form in humans, which can cause deforming lesions, alterations of the speech and even an inadequate nutrition due to difficulties in deglutition.

MyD88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden.

Infections with Leishmania parasites of the Leishmania Viannia subgenus give rise to both localized cutaneous (CL), and metastatic leishmaniasis. Metastasizing disease forms including disseminated (DCL) and mutocutaneous (MCL) leishmaniasis result from parasitic dissemination and lesion formation at sites distal to infection and have increased inflammatory responses. The presence of Leishmania RNA virus (LRV) in L. guyanensis parasites contributes to the exacerbation of disease and impacts inflammatory responses via activation of TLR3 by the viral dsRNA. In this study we investigated other innate immune response adaptor protein modulators and demonstrated that both MyD88 and TLR9 played a crucial role in the development of Th1-dependent healing responses against L. guyanensis parasites regardless of their LRV status. The absence of MyD88- or TLR9-dependent signaling pathways resulted in increased Th2 associated cytokines (IL-4 and IL-13), which was correlated with low transcript levels of IL-12p40. The reliance of IL-12 was further confirmed in IL12AB-/- mice, which were completely susceptible to infection. Protection to L. guyanensis infection driven by MyD88- and TLR9-dependent immune responses arises independently to those induced due to high LRV burden within the parasites.