Leishmaniasis epidemiology in endemic areas of metropolitan France and its overseas territories from 1998 to 2020.

BACKGROUND: In France, leishmaniasis is endemic in the Mediterranean region, in French Guiana and to a lesser extent, in the French West Indies. This study wanted to provide an updated picture of leishmaniasis epidemiology in metropolitan France and in its overseas territories. METHODOLOGY/PRINCIPAL FINDINGS: Leishmaniasis cases were collected by passive notification to the French National Reference Centre for Leishmaniases (NRCL) in Montpellier from 1998 to 2020 and at the associated Centre in Cayenne (French Guiana) from 2003 to 2020. In metropolitan France, 517 autochthonous leishmaniasis cases, mostly visceral forms due to Leishmania infantum (79%), and 1725 imported cases (French Guiana excluded), mainly cutaneous leishmaniasis from Maghreb, were recorded. A slight decrease of autochthonous cases was observed during the survey period, from 0.48 cases/100,000 inhabitants per year in 1999 (highest value) to 0.1 cases/100,000 inhabitants per year in 2017 (lowest value). Conversely, imported cases increased over time (from 59.7 in the 2000s to 94.5 in the 2010s). In French Guiana, 4126 cutaneous and mucocutaneous leishmaniasis cases were reported from 2003 to 2020. The mean incidence was 103.3 cases per 100,000 inhabitants/year but varied in function of the year (from 198 in 2004 to 54 in 2006). In Guadeloupe and Martinique (French West Indies), only sporadic cases were reported. CONCLUSIONS/SIGNIFICANCE: Because of concerns about disease expansion and outbreaks in other Southern Europe countries, and leishmaniasis monitoring by the NRCL should be continued and associated with a more active surveillance.

Autochthonous and imported tegumentary leishmaniasis in Catalonia (Spain): Aetiological evolution in the last four decades and usefulness of different typing approaches based on biochemical, molecular and proteomic markers.

Leishmaniasis is a transmissible disease caused by Leishmania protozoa. Spain is endemic for both visceral and cutaneous leishmaniasis, the autochthonous aetiological agent being Leishmania infantum. Around the world, the L. donovani complex is associated with visceral symptoms, while any species of the Leishmania or Viannia subgenera affecting human can produce tegumentary forms. In a context of growing numbers of imported cases, associated with globalisation, the aim of this study was to analyse the aetiological evolution of human tegumentary leishmaniasis in a region of Spain (Catalonia). Fifty-six Leishmania strains, isolated from 1981 to 2018, were analysed using MLEE, gene sequencing (hsp70, rpoIILS, fh and ITS2) and MALDI-TOF. The utility of these different analytical methods was compared. The results showed an increase in leishmaniasis over the two last decades, particularly imported cases, which represented 39% of all cases studied. Leishmania infantum, L. major, L. tropica, L. braziliensis, L. guyanensis and L. panamensis were identified. The combination of molecular and enzymatic methods allowed the identification of 29 different strain types (A to AC). Strain diversity was higher in L. (Viannia), whilst the different L. major types were relatable with geo-temporal data. Among the autochthonous cases, type C prevailed throughout the studied period (39%). Minor types generally appeared within a short time interval. While all the techniques provided identical identification at the species complex level, MALDI-TOF and rpoIILS or fh sequencing would be the most suitable identification tools for clinical practice, and the tandem hsp70-ITS2 could substitute MLEE in the epidemiological field.

The Leishmania donovani species complex: A new insight into taxonomy☆.

Among the 20 or so Leishmania spp. described as pathogenic for humans, those of the Leishmania donovani complex are the exclusive causative agents of systemic and fatal visceral leishmaniasis. Although well studied, the complex is taxonomically controversial, which hampers clinical and epidemiological research. In this work, we analysed 56 Leishmania strains previously identified as L. donovani, Leishmania archibaldi or Leishmania infantum, isolated from humans, dogs and sandfly vectors throughout their distribution area. The strains were submitted to biochemical and genetic analyses and the resulting data were compared for congruence. Our results show: i) a partial concordance between biochemical and genetic-based data, ii) very limited genetic variability within the L. donovani complex, iii) footprints of frequent genetic exchange along an east-west gradient, marked by a widespread diffusion of alleles across the geographical range, and iv) a large-scale geographical spreading of a few genotypes. From a taxonomic point of view, considering the absence of relevant terminology in existing classes, the L. donovani complex could be treated as a single entity.

Analysis of genetic polymorphisms and tropism in East African Leishmania donovani by Amplified Fragment Length Polymorphism and kDNA minicircle sequencing.

Visceral leishmaniasis (VL), the most severe form of leishmaniasis, is caused by Leishmania donovani. In addition to fatal VL, these parasites also cause skin diseases in immune-competent and -suppressed people, post-kala azar dermal leishmaniasis (PKDL) and HIV/VL co-infections, respectively. Genetic polymorphism in 36 Ethiopian and Sudanese L. donovani strains from VL, PKDL and HIV/VL patients was examined using Amplified Fragment Length Polymorphism (AFLP), kDNA minicircle sequencing and Southern blotting. Strains were isolated from different patient tissues: in VL from lymph node, spleen or bone marrow; and in HIV/VL from skin, spleen or bone marrow. When VL and PKDL strains from the same region in Sudan were examined by Southern blotting using a DNA probe to the L. donovani 28S rRNA gene only minor differences were observed. kDNA sequence analysis distributed the strains in no particular order among four clusters (A - D), while AFLP analysis grouped the strains according to geographical origin into two major clades, Southern Ethiopia (SE) and Sudan/Northern Ethiopia (SD/NE). Strains in the latter clade were further divided into subpopulations by zymodeme, geography and year of isolation, but not by clinical symptoms. However, skin isolates showed significantly (p < 0.0001) fewer polymorphic AFLP fragments (average 10 strains = 348.6 ±â€¯8.1) than VL strains (average 26 strains = 383.5 ±â€¯3.8).

Identification of Leishmania by Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) Mass Spectrometry Using a Free Web-Based Application and a Dedicated Mass-Spectral Library.

Human leishmaniases are widespread diseases with different clinical forms caused by about 20 species within the Leishmania genus. Leishmania species identification is relevant for therapeutic management and prognosis, especially for cutaneous and mucocutaneous forms. Several methods are available to identify Leishmania species from culture, but they have not been standardized for the majority of the currently described species, with the exception of multilocus enzyme electrophoresis. Moreover, these techniques are expensive, time-consuming, and not available in all laboratories. Within the last decade, mass spectrometry (MS) has been adapted for the identification of microorganisms, including Leishmania However, no commercial reference mass-spectral database is available. In this study, a reference mass-spectral library (MSL) for Leishmania isolates, accessible through a free Web-based application (mass-spectral identification [MSI]), was constructed and tested. It includes mass-spectral data for 33 different Leishmania species, including species that infect humans, animals, and phlebotomine vectors. Four laboratories on two continents evaluated the performance of MSI using 268 samples, 231 of which were Leishmania strains. All Leishmania strains, but one, were correctly identified at least to the complex level. A risk of species misidentification within the Leishmania donovani, L. guyanensis, and L. braziliensis complexes was observed, as previously reported for other techniques. The tested application was reliable, with identification results being comparable to those obtained with reference methods but with a more favorable cost-efficiency ratio. This free online identification system relies on a scalable database and can be implemented directly in users' computers.

The Montpellier Leishmania Collection, from a Laboratory Collection to a Biological Resource Center: A 39-Year-Long Story.

We report the development of a laboratory collection of Leishmania that was initiated in 1975 and, after 39 years, has become an international Biological Resource Center (BRC-Leish, Montpellier, France, BioBank No. BB-0033-00052), which includes 6353 strains belonging to 36 Leishmania taxa. This is a retrospective analysis of the technical and organizational changes that have been adopted over time to take into account the technological advances and related modifications in the collection management and quality system. The technical improvements concerned the culture and cryopreservation techniques, strain identification by isoenzymatic and molecular techniques, data computerization and quality management to meet the changes in international standards, and in the cryogenic and microbiological safety procedures. The BRC is working toward obtaining the NF-S 96-900 certification in the coming years. Our long-term expertise in Leishmania storage and typing and collection maintenance should encourage field epidemiologists and clinical practitioners in endemic countries to secure their own strain collection with the help of the French BRC-Leish.

Comparison of Leishmania killicki (syn. L. tropica) and Leishmania tropica Population Structure in Maghreb by Microsatellite Typing.

Leishmania (L.) killicki (syn. L. tropica), which causes cutaneous leishmaniasis in Maghreb, was recently described in this region and identified as a subpopulation of L. tropica. The present genetic analysis was conducted to explore the spatio-temporal distribution of L. killicki (syn. L. tropica) and its transmission dynamics. To better understand the evolution of this parasite, its population structure was then compared with that of L. tropica populations from Morocco. In total 198 samples including 85 L. killicki (syn. L. tropica) (from Tunisia, Algeria and Libya) and 113 L. tropica specimens (all from Morocco) were tested. Theses samples were composed of 168 Leishmania strains isolated from human skin lesions, 27 DNA samples from human skin lesion biopsies, two DNA samples from Ctenodactylus gundi bone marrow and one DNA sample from a Phlebotomus sergenti female. The sample was analyzed by using MultiLocus Enzyme Electrophoresis (MLEE) and MultiLocus Microsatellite Typing (MLMT) approaches. Analysis of the MLMT data support the hypothesis that L. killicki (syn. L. tropica) belongs to the L. tropica complex, despite its strong genetic differentiation, and that it emerged from this taxon by a founder effect. Moreover, it revealed a strong structuring in L. killicki (syn. L. tropica) between Tunisia and Algeria and within the different Tunisian regions, suggesting low dispersion of L. killicki (syn. L. tropica) in space and time. Comparison of the L. tropica (exclusively from Morocco) and L. killicki (syn. L. tropica) population structures revealed distinct genetic organizations, reflecting different epidemiological cycles.

Evolutionary history of Leishmania killicki (synonymous Leishmania tropica) and taxonomic implications.

BACKGROUND: The taxonomic status of Leishmania (L.) killicki, a parasite that causes chronic cutaneous leishmaniasis, is not well defined yet. Indeed, some researchers suggested that this taxon could be included in the L. tropica complex, whereas others considered it as a distinct phylogenetic complex. To try to solve this taxonomic issue we carried out a detailed study on the evolutionary history of L. killicki relative to L. tropica. METHODS: Thirty-five L. killicki and 25 L. tropica strains isolated from humans and originating from several countries were characterized using the MultiLocus Enzyme Electrophoresis (MLEE) and the MultiLocus Sequence Typing (MLST) approaches. RESULTS: The results of the genetic and phylogenetic analyses strongly support the hypothesis that L. killicki belongs to the L. tropica complex. Our data suggest that L. killicki emerged from a single founder event and that it evolved independently from L. tropica. However, they do not validate the hypothesis that L. killicki is a distinct complex. Therefore, we suggest naming this taxon L. killicki (synonymous L. tropica) until further epidemiological and phylogenetic studies justify the L. killicki denomination. CONCLUSIONS: This study provides taxonomic and phylogenetic information on L. killicki and improves our knowledge on the evolutionary history of this taxon.

Geographical distribution and epidemiological features of Old World Leishmania infantum and Leishmania donovani foci, based on the isoenzyme analysis of 2277 strains.

A series of 2277 Leishmania strains from Old World visceral leishmaniasis foci, isolated between 1973 and 2008, were studied by isoenzyme analysis. The strains were obtained from humans, domestic and wild carnivores, rodents and phlebotomine sandflies, and came from 36 countries. In all, 60 different zymodemes were identified and clustered by a phenetic analysis into 3 different groups corresponding to the typically visceralizing species L. donovani (20 zymodemes, 169 strains), L. archibaldi (3 zymodemes, 46 strains) and L. infantum (37 zymodemes, 2,062 strains). The taxonomic position of these isoenzymatic groups is discussed in view of contradictory results obtained from recent molecular studies.

Leishmania infantum infection in two captive barbary lions (Panthera leo leo).

A female barbary lion (Panthera leo leo) from the Montpellier Zoological Park (France) showing colitis, epistaxis, and lameness with pad ulcers was positive by polymerase chain reaction (PCR) for Leishmania infantum. Further indirect immunofluorescence (IFAT) tests on the banked sera from all lions of the park detected another infected but asymptomatic female, which was confirmed by PCR on ethylenediaminetetraacetic acid (EDTA) blood sample. Leishmania infantum zymodeme MON-1 was cultured from EDTA bone marrow samples sampled from this second animal. The first female was successfully treated with marbofloxacine at 2 mg/kg s.i.d. for 28 days (Marbocyl, Vetoquinol 70204 Lure, France) and allopurinol at 30 mg/kg s.i.d. for 3 mo (Allopurinol Mylan, Mylan SAS, 69800 Saint-Priest, France) and then 1 wk/mo. Both positive animals were born at the Rabat Zoological Park, Morocco, and arrived together at Montpellier in 2003. The chronicity and source of this current infection are unknown since Morocco and southern France are well-known to be enzootic for leishmaniasis.

Geographical distribution and epidemiological features of Old World cutaneous leishmaniasis foci, based on the isoenzyme analysis of 1048 strains.

A series of 1048 Leishmania strains from Old World cutaneous leishmaniasis foci, isolated between 1981 and 2005, were studied by isoenzyme analysis. The strains were obtained from humans, rodents, dogs and sandflies from 33 countries. The four typically dermotropic species, Leishmania major, L. tropica, L. aethiopica and L. killicki, were found. The viscerotropic species L. donovani and L. infantum, which can occasionally be responsible for cutaneous leishmaniasis, are not considered in this paper. Leishmania major was the least polymorphic species (12 zymodemes, 638 strains). Leishmania tropica was characterized by a complex polymorphism varying according to focus (35 zymodemes, 329 strains). Leishmania aethiopica, a species restricted to East Africa, showed a high polymorphism, in spite of a limited number of strains (23 zymodemes, 40 strains). Leishmania killicki, mainly restricted to Tunisia had a single zymodeme for 39 strains. Recently a parasite close to L. killicki (one zymodeme, two strains) was isolated in Algeria, which lead us to revise the taxonomic status of this taxon.