The effect of parasitization by Leishmania mexicana mexicana on macrophage function in vitro

Macrophages infected with amastigotes of Leishmania mexicana mexicana as compared to normal macrophages show decreased migration both randomly and through a 5 microns pore in response to a known chemotaxin, an increased ability to pinocytose and an increased bactericidal ability. Unless very heavily parasitized their ability to phagocytose is unaltered. Parasitized macrophages are unaltered in their ability to secrete extracellularly lysosomal enzymes, prostaglandins and lysozyme in response to known stimuli, or to kill target cells in an antibody dependent cell mediated cytotoxicity assay.     

Amine production by Leishmania mexicana

Growing promastigotes of Leishmania mexicana mexicana excreted large amounts of urea and ammonia into the culture medium. Both promastigotes and amastigotes in short-term, high-density cultures also produced urea and ammonia; the excretion rate was higher in promastigotes. Putrescine was excreted by growing promastigotes but spermine and spermidine excretion apparently did not occur. Both promastigote and amastigote cell extracts contained putrescine and spermidine. Both polyamines, but especially putrescine, were present at higher concentrations in promastigotes than amastigotes. Trace amounts of spermine were found in the promastigote extracts but none was detected in the amastigotes.     

Tissue eosinophilia and Leishmania mexicana mexicana eosinophil interactions in murine cutaneous leishmaniasis

Summary Outbred albino mice were infected subcutaneously with 10⁶ amastigotes of Leishmania mexicana mexicana and the subsequent lesions were evaluated by light and electron microscopy at various intervals after infection. The animals developed persistent nodules and a spectrum of lesions of variable size which was correlated with the host's ability to control the parasite in the tissue. During the acute phase of the disease the histopathological results showed an accumulation of granulocytes, some mononuclear phagocytes and a predominance of eosinophils as compared to other cell types. In this early acute phase, eosinophils were found in the tissue together with normal and degranulating mast cells. In the granulomatous inflammatory reaction of the chronic phases, there was infiltration of granulocytes parallel to parasite multiplication and the formation of parasitized vacuolated macrophages. The number of eosinophils was consistently greater than neutrophils, regardless of lesion type or number of parasites present in the tissue. During the acute reaction, the granulocytes apparently destroyed many parasites; however, there was an unvaryingly low level of phagocytosis of amastigotes during the chronic stages by both eosinophils and neutrophils. Neutrophils seemed to be more effective than eosinophils in the killing of ingested parasites. A close association between eosinophils and parasitized macrophages was seen in the chronic lesions; thus, eosinophils might contribute to parasite destruction through co-operation with macrophages.     

Genetic analysis of Leishmania mexicana populations from Texas, Latin America, and the Caribbean

A genetic analysis using enzyme data of 72 Leishmania mexicana isolated from hosts in Texas, Latin America, and the Caribbean is presented. All isolates from each country were combined and considered as local populations. Allomorph (allele determined by electrophoresis) frequencies for 20 enzyme (loci) were calculated and 7 populations (Texas, Mexico, Belize, Guatemala, Ecuador [EC], Venezuela, and the Dominican Republic [DR]) were compared pairwise in the statistic of genetic identity (I) (level of genetic similarity). All populations were found to be genetically similar with a mean I value for all comparisons of 0.890 +/- 0.067. When DR was included as one of the pair compared, I = 0.811 +/- 0.034. Among comparisons that include EC (excluding EC vs. DR), I = 0.875 +/- 0.026. The mean I for the other comparisons was less than 0.9. The data indicate that the DR population is divergent enough from the others that it can be considered at the subspecies/incipient species level of evolutionary divergence; the EC population is, to a lesser extent, distinct from the others, and the other 5 represent geographic populations of 1 widely distributed species. A diagrammatic representation of the allomorphs among the 72 isolates is included. There were some allomorph/geographical (or local) population relationships noted.     

Accidental Leishmania mexicana infection in an immunosuppressed laboratory technician

A 30 year-old female laboratory technician under immunosuppressive treatment because of systemic lupus erythematosus (SLE) developed cutaneous leishmaniasis 8 months after accidental percutaneous inoculation of amastigote culture forms of Leishmania mexicana . Leishmania -specific PCR and restriction analysis patterns were identical for both the laboratory strain and the clinical specimen. The lesion was resistant to local paromomycin and oral ketoconazole, but responded to local application of meglumine antimonate. No signs of dissemination or visceralization occurred during the 5-month period of observation. However, a future recurrence cannot be excluded since a persistent infection even after clinical cure has always to be considered in leishmaniasis. Patients under immunosuppressive therapy are possibly at risk of clinical relapse or disseminating infection although there is no experience with regard to leishmaniasis mexicana . Serious infection may require interferon gamma as part of the treatment which may contribute to deterioration of concomitant diseases like SLE. In any case, the exposure of immunodeficient laboratory workers to Leishmania spp . should be avoided.     

Host-parasite relationship of Leishmania mexicana mexicana and Lutzomyia abonnenci (Diptera: Psychodidae)

The life cycle of Leishmania mexicana mexicana in the gut of the sand fly, Lutzomyia abonnenci, was studied by light and electron microscopy. Development was suprapylarian with initial establishment of parasites in the bloodmeal (posterior midgut), and anterior migration of parasites to the cardia/stomodeal valve region beginning at 2.5 days post-infection. Flagellates were first observed in the esophagus at 3.5 days, in the posterior armature region of the pharynx at 5 days, and in the anterior pharynx at 7 days; but they were not detected in the cibarium or proboscis. Infection of the pylorus region of the hindgut and of the Malpighian tubules was also commonly observed. Three different morphological forms of L. m. mexicana developed in the gut: nectomonad promastigotes, short promastigotes, and paramastigotes. Nectomonads occurred primarily in the abdominal midgut after bloodmeal digestion, where they were oriented in longitudinal masses in the lumen, or interdigitated with epithelial microvilli via the flagellum. Short promastigotes found in the cardia/stomodeal valve region are described for the first time. These forms were smaller than nectomonads, showed an amplification of the kinetoplast, apposition of kinetoplast and nucleus, and were embedded in a gel-like matrix. To maintain position in the cardia, parasites commonly inserted the flagellum deep into microvilli or cytoplasm of the epithelium; adherence to the cuticular intima of the stomodeal valve was by flagellar modification and formation of hemidesmosome plaques. Paramastigotes occurred in the esophagus, were sometimes degenerated in appearance, and were attached via flagellar hemidesmosomes. Paramastigotes observed in the lumen of the pharynx were commonly degenerated and were not attached to the intima. L. m. mexicana was able to colonize the various gut habitats of Lu. abonnenci by a number of adaptations; this sand fly appears to be a suitable biological host for the parasite.     

Leishmania mexicana complex: human infections in the Republic of Panama

Parasites of the genus Leishmania responsible for human cutaneous leishmaniasis in the New World form 2 major taxonomic divisions: the Leishmania braziliensis and the L. mexicana complexes. We report the isolation and characterization of the L. mexicana complex among humans in the Republic of Panama. Characterization was based on parasite morphology, pathogenesis in infected golden hamsters, cellulose acetate isoenzyme electrophoretic mobilities, and membrane-specific monoclonal antibodies using the radioimmune binding assay technique.     

Destruction of Leishmania mexicana amazonensis promastigotes by normal human serum

Fresh normal human serum was observed to have a lethal effect on Leishmania mexicana amazonensis promastigotes obtained from laboratory-bred Lutzomyia longipalpis or on promastigotes grown in liquid culture medium, inoculated with the same isolates. Heat inactivation abolished the Leishmania lytic activity from the sera. Resistance of culture promastigotes to lysis by normal human serum was investigated in three isolates of L. m. amazonensis. Development of resistance (up to 7%) was found in only one isolate, obtained from the bone marrow in a human case of visceral leishmaniasis.     

Transplacental Transmission of Cutaneous Leishmania mexicana Strain in BALB/c Mice

The vertical transmission of leishmaniasis has been reported in species that cause visceral leishmaniasis. However, this condition has scarcely been documented in species that cause cutaneous leishmaniasis. The aim of this study was to determine experimentally whether L. mexicana is transmitted vertically. A control group of BALB/c mice and a group infected with L. mexicana were mated, the gestation was monitored, and females were killed before delivery. Four resorptions (P = 0.023) and eight fetal deaths (P = 0.010) were observed in the infected female group; furthermore, the offspring body weight of the infected group was lower than the body weight of the healthy group (P = 0.009). DNA amplification by polymerase chain reaction (PCR) revealed that all placentas and maternal spleens as well as 39 of 110 fetal spleens obtained from the offspring of infected mothers tested positive for Leishmania. In conclusion, L. mexicana is transmitted transplacentally and causes fetal death, resorption, and reduction in offspring body weight.     

Leishmania mexicana and Leishmania major: attenuation of wild-type parasites and vaccination with the attenuated lines

A method for attenuation of Leishmania species by culturing in vitro under gentamicin pressure has been used successfully with Leishmania mexicana, L. major, L. infantum, and L. donovani. The attenuated lines invaded but were unable to survive within bone marrow-derived macrophages in vitro, whereas wild-type parasites survived and multiplied. The attenuated lines of L. mexicana and L. major both failed to induce cutaneous lesions in the majority of BALB/c mice over a minimum 12-week observation period after subcutaneous injection of stationary phase parasites. The attenuated line of L. mexicana retained its properties in gentamicin-free medium over 40 subcultures. The attenuated lines of L. mexicana and L. major both induced significant protection in mice against challenge with wild-type parasites.     

Leishmania mexicana: destruction of isolated amastigotes by amino acid esters

Amino acid esters can destroy intracellular as well as isolated amastigotes of Leishmania mexicana amazonensis. In the present study we examined, using a tetrazolium reduction assay, the toxicity of the esters for amastigotes isolated from mouse lesions. Parasite killing by the "prototype" compound L-leucine methyl ester at 1 mM concentration and at pH 7.3 took place within 15-30 min. Time-lapse cinematographic observations showed that the amastigotes rounded up and became less phase-dense before they rapidly broke down. Ammonium chloride, ethylamine or monensin, known to raise the intracellular pH, reduced the sensitivity of the amastigotes to L-Leu-OMe. This finding suggests that an acidified compartment is involved in the destruction of the parasites. The leishmanicidal activity of a series of L-amino acid esters was also investigated. The ED50 (concentration for half maximal effect) for methyl esters was (in mM): Leu (0.62), Trp (0.96), Met (1.13), Glu (2.0), Phe (2.5), and Tyr (3.8). In contrast, the methyl esters of Ile, Val, Ala, beta Ala, Gly, Ser, His, and Pro were either inactive or weakly active at 15 mM. Benzyl esters were more active than their methyl homologs: the ED50 of the benzyl esters of Leu, Val, Ile, Gly, Ala, beta Ala, and Pro were, respectively, 0.07, 0.20, 0.22, 0.88, 1.5, 2.3, and 6.7 mM. Ranks of leishmanicidal activity may reflect differences in the rates of ester uptake and trapping by the amastigotes, in the specificity of the relevant hydrolytic enzyme(s), in the accumulation and metabolic fate of the released amino acids, or in the toxicity of the amino acid or alcohol released within the amastigotes.     

The histopathology of cutaneous leishmaniasis due to Leishmania (Leishmania) mexicana in the Yucatan peninsula, Mexico

Localized Cutaneous Leishmaniasis (LCL) known as "chiclero's ulcer" in southeast Mexico, was described by SEIDELIN in 1912. Since then the sylvatic region of the Yucatan peninsula has been documented as an endemic focus of LCL. This study of 73 biopsies from parasitological confirmed lesions of LCL cases of Leishmania (Leishmania) mexicana infection was undertaken: 1) to examine host response at tissue level; and 2) to relate manifestations of this response to some characteristics of clinical presentation. Based on Magalh?es' classification we found that the most common pattern in our LCL cases caused by L. (L.) mexicana was predominantly characterized by the presence of unorganized granuloma without necrosis, (43.8%). Another important finding to be highlighted is the fact that in 50/73 (68.5%) parasite identification was positive. There was direct relation between the size of the lesion and time of evolution (rs = 0.3079, p = 0.03), and inverse correlation between size of the lesion and abundance of amastigotes (rs = -0.2467, p = 0.03). In view of the complexity of clinical and histopathological findings, cell-mediated immune response of the disease related to clinical and histopathological features, as so genetic background should be studied.     

Monocyte cytokine and costimulatory molecule expression in patients infected with Leishmania mexicana

Leishmania mexicana causes localized and diffuse cutaneous leishmaniasis. Patients with localized cutaneous leishmaniasis (LCL) develop a benign disease, whereas patients with diffuse cutaneous leishmaniasis (DCL) suffer from a progressive disease associated with anergy of the cellular response towards Leishmania antigens. We evaluated the production of the interleukins (IL) IL-12, IL-15, IL-18 and tumour necrosis factor-alpha (TNF-alpha) and the expression of the costimulatory molecules CD40, B7-1 and B7-2 in monocytes from LCL and DCL patients, stimulated in vitro with Leishmania mexicana lipophosphoglycan (LPG) for 18 h. LCL monocytes significantly increased TNF-alpha, IL-15 and IL-18 production, and this increase was associated with reduced amounts of IL-12. DCL monocytes produced no IL-15 or IL-18 and showed a decreasing tendency of TNF-alpha and IL-12 production as the severity of the disease increased. No difference was observed in the expression of CD40 and B7-1 between both groups of patients, yet B7-2 expression was significantly augmented in DCL patients. It remains to be established if this elevated B7-2 expression in DCL patients is cause or consequence of the Th2-type immune response that characterizes these patients. These data suggest that the diminished ability of the monocytes from DCL patients to produce cell-activating innate proinflammatory cytokines when stimulated with LPG is a possible cause for disease progression.