First official report of bed bug (Hemiptera, Cimicidae) infestations in Algeria.

Background: Bed bugs are hematophagous insects with a long history of presence in human communities. Over the last three decades, infestations by bed bugs in human dwellings have drastically increased, leading to a rise in bed bug concerns. Nevertheless, very little is known about the bed bug species and their population diversity in Algeria. Method: A pilot entomological inventory was performed in May 2019 in Tizi Ouzou, in northern Algeria. The gathered bed bug specimens were identified by morphological and molecular approaches, followed by neighbor-joining and network phylogenetic analyses. Results: A total of seven out of 12 requested locations were allowed to inspect for bed bug infestation. Of these, three locations were found with active bed bug infestations. A total of 145 specimens belonging to different life stages [egg (21), nymph (74), adult male (17), and female (33)] were collected and analyzed using morphological and molecular approaches. The adult specimens were identified as Cimex lectularius according to specific morphological criteria, most importantly the pronotum laterally expanded with more flattened extreme margins. Morphological identification of the adults was confirmed further by conventional PCR targeting 450 bp fragment of the COI gene. All the nymphs and eggs were also molecularly identified as C. lectularius. Neighbor-Joining phylogenetic tree reconstructed with the collected specimens provides clues on the presence of two closely phylogenetic groups. The first one gathers our samples of Algeria with previously reported COI haplotype sequences from Asian, European, and North American countries. The second group encompasses a lesser-documented haplotype reported in Europe and Central America. These findings were further confirmed by network analysis. Conclusions: These results provide evidence of established C. lectularius infestation in Algeria and its potential dispersal capacity by travelers or immigrants and will help future management of these ectoparasites.

An annotated wing interferential pattern dataset of dipteran insects of medical interest for deep learning.

Several Diptera species are known to transmit pathogens of medical and veterinary interest. However, identifying these species using conventional methods can be time-consuming, labor-intensive, or expensive. A computer vision-based system that uses Wing interferential patterns (WIPs) to identify these insects could solve this problem. This study introduces a dataset for training and evaluating a recognition system for dipteran insects of medical and veterinary importance using WIPs. The dataset includes pictures of Culicidae, Calliphoridae, Muscidae, Tabanidae, Ceratopogonidae, and Psychodidae. The dataset is complemented by previously published datasets of Glossinidae and some Culicidae members. The new dataset contains 2,399 pictures of 18 genera, with each genus documented by a variable number of species and annotated as a class. The dataset covers species variation, with some genera having up to 300 samples.

Species identification of phlebotomine sandflies using deep learning and wing interferential pattern (WIP).

Sandflies (Diptera; Psychodidae) are medical and veterinary vectors that transmit diverse parasitic, viral, and bacterial pathogens. Their identification has always been challenging, particularly at the specific and sub-specific levels, because it relies on examining minute and mostly internal structures. Here, to circumvent such limitations, we have evaluated the accuracy and reliability of Wing Interferential Patterns (WIPs) generated on the surface of sandfly wings in conjunction with deep learning (DL) procedures to assign specimens at various taxonomic levels. Our dataset proves that the method can accurately identify sandflies over other dipteran insects at the family, genus, subgenus, and species level with an accuracy higher than 77.0%, regardless of the taxonomic level challenged. This approach does not require inspection of internal organs to address identification, does not rely on identification keys, and can be implemented under field or near-field conditions, showing promise for sandfly pro-active and passive entomological surveys in an era of scarcity in medical entomologists.

Reconstructing the post-glacial spread of the sand fly Phlebotomus mascittii Grassi, 1908 (Diptera: Psychodidae) in Europe.

Phlebotomine sand flies (Diptera: Phlebotominae) are the principal vectors of Leishmania spp. (Kinetoplastida: Trypanosomatidae). In Central Europe, Phlebotomus mascittii is the predominant species, but largely understudied. To better understand factors driving its current distribution, we infer patterns of genetic diversity by testing for signals of population expansion based on two mitochondrial genes and model current and past climate and habitat suitability for seven post-glacial maximum periods, taking 19 climatic variables into account. Consequently, we elucidate their connections by environmental-geographical network analysis. Most analyzed populations share a main haplotype tracing back to a single glacial maximum refuge area on the Mediterranean coasts of South France, which is supported by network analysis. The rapid range expansion of Ph. mascittii likely started in the early mid-Holocene epoch until today and its spread possibly followed two routes. The first one was through northern France to Germany and then Belgium, and the second across the Ligurian coast through present-day Slovenia to Austria, toward the northern Balkans. Here we present a combined approach to reveal glacial refugia and post-glacial spread of Ph. mascittii and observed discrepancies between the modelled and the current known distribution might reveal yet overlooked populations and potential further spread.

Wing Interferential Patterns (WIPs) and machine learning for the classification of some Aedes species of medical interest.

Hematophagous insects belonging to the Aedes genus are proven vectors of viral and filarial pathogens of medical interest. Aedes albopictus is an increasingly important vector because of its rapid worldwide expansion. In the context of global climate change and the emergence of zoonotic infectious diseases, identification tools with field application are required to strengthen efforts in the entomological survey of arthropods with medical interest. Large scales and proactive entomological surveys of Aedes mosquitoes need skilled technicians and/or costly technical equipment, further puzzled by the vast amount of named species. In this study, we developed an automatic classification system of Aedes species by taking advantage of the species-specific marker displayed by Wing Interferential Patterns. A database holding 494 photomicrographs of 24 Aedes spp. from which those documented with more than ten pictures have undergone a deep learning methodology to train a convolutional neural network and test its accuracy to classify samples at the genus, subgenus, and species taxonomic levels. We recorded an accuracy of 95% at the genus level and > 85% for two (Ochlerotatus and Stegomyia) out of three subgenera tested. Lastly, eight were accurately classified among the 10 Aedes sp. that have undergone a training process with an overall accuracy of > 70%. Altogether, these results demonstrate the potential of this methodology for Aedes species identification and will represent a tool for the future implementation of large-scale entomological surveys.

Molecular characterization and genetic diversity of Wolbachia endosymbionts in bed bugs (Hemiptera; Cimicidae) collected in Paris.

PURPOSE: This study aimed to investigate the genetic diversity of Wolbachia in field-caught bed bug species in Paris areas. METHODS: The bed bug specimens were captured from various infested localities in Paris and surrounding cities. They belonged to diverse life stages, including egg, nymph, and adult. They were then identified using morphological and molecular approaches. Furthermore, Wolbachia was detected, and its genetic diversity was investigated by conventional PCR of 16S-rRNA and Wolbachia surface protein (wsp) genes. RESULTS: A total of 256 bed bug specimens belonging to various life stages [adult (183 specimens), nymph (48), and egg (25)] were captured from seven private apartments, five social apartments, three houses, two immigrant residences, and one retirement home situated in 10 districts of Paris and 8 surrounding cities. They were identified as Cimex lectularius (237 specimens) and C. hemipterus (19) using morphological and molecular approaches. The presence and diversity of Wolbachia were ascertained by targeting 16S-rRNA and wsp genes. Based on molecular analysis, 182 and 148 out of 256 processed specimens were positive by amplifying 16S-rRNA and wsp fragments, respectively. The inferred phylogenetic analysis with 16S-rRNA and wsp sequences displayed monophyletic Wolbachia strains clustering each one in three populations. The median-joining network, including the Wolbachia 16S-rRNA and wsp sequences of C. lectularius and C. hemipterous specimens, indicated a significant genetic differentiation among these populations in Paris areas which was consent with Neighbor-Joining analyses. A phylogenetic analysis of our heterogenic Wolbachia sequences with those reported from other arthropod species confirmed their belonging to supergroup F. Moreover, no difference between Wolbachia sequences from eggs, nymphs, and adults belonging to the same clade and between Wolbachia sequences of C. lectularius and C. hemipterus were observed after sequence alignment. Furthermore, no significant correlation was found between multiple geographical locations (or accomodation type) where bed bugs were collected and the genetic diversity of Wolbachia. CONCLUSIONS: We highlight a significant heterogeneity within Wolbachia symbionts detected in C. lectularius and C. hemipterus. No correlation between Wolbachia species and bed bug species (C. lectularius versus C. hemipterus), physiological stages (egg, nymph, and adult), and sampling location was recorded in this study.

Deep learning and wing interferential patterns identify Anopheles species and discriminate amongst Gambiae complex species.

We present a new and innovative identification method based on deep learning of the wing interferential patterns carried by mosquitoes of the Anopheles genus to classify and assign 20 Anopheles species, including 13 malaria vectors. We provide additional evidence that this approach can identify Anopheles spp. with an accuracy of up to 100% for ten out of 20 species. Although, this accuracy was moderate (> 65%) or weak (50%) for three and seven species. The accuracy of the process to discriminate cryptic or sibling species is also assessed on three species belonging to the Gambiae complex. Strikingly, An. gambiae, An. arabiensis and An. coluzzii, morphologically indistinguishable species belonging to the Gambiae complex, were distinguished with 100%, 100%, and 88% accuracy respectively. Therefore, this tool would help entomological surveys of malaria vectors and vector control implementation. In the future, we anticipate our method can be applied to other arthropod vector-borne diseases.

Descriptive and Geometric Morphometry of the Wings of Phlebotomus sergenti Populations in Central Morocco.

Background: Phlebotomus sergenti, the proven vector of Leishmania tropica, the causative agent of anthroponotic cutaneous leishmaniasis, is widely distributed in Morocco. Previous works using molecular markers (Internal Transcribed Spacer 2 rDNA and Cytochrome B mtDNA) hypothesized the existence of multiple closely related populations of sand fly species (cryptic species) that would exhibit distinct vectorial capacities. This work studies morphotypic diversity using traditional and geometric morphometry analyses carried out on Ph. sergenti's wings from central Morocco, where active L. tropica transmission occurs for 30 years. Methods: Descriptive characteristics (size and shape) of the right wings were measured in Ph. sergenti's specimens collected from fourteen stations in central Morocco. Both traditional and geometric morphometry methods were used to analyse geographic variations in Ph. sergenti wing's size and shape. Results: These analyses support the existence of distinct Ph. sergenti populations, enlightening significant phenotypic variations of Ph. sergenti's wings, regarding their size and shape, depending on geographic origin. In addition, traditional and geometric morphometric analyses of the wing's length, centroid size, ß, ɵ, and γ distances allowed clear discrimination of Ph. sergenti sub-populations. Conclusion: These data pinpoint the adaptative ability of Ph. sergenti to local environmental conditions. Additional studies are now required to further shed light on the genetic structure of Ph. sergenti populations in Morocco.

Bed Bugs (Hemiptera, Cimicidae): A Global Challenge for Public Health and Control Management.

Bed bugs, Cimex lectularius, and C. hemipterus are among the most common ectoparasites in human life worldwide. They feed on humans of all ages and sexes across all socioeconomic levels. Bed bugs' blood feeding is responsible for a wide range of clinical manifestations varying from minor reactions to bullous eruptions or severe allergies. In addition, they are responsible for considerable psychological distress. Therefore, diagnosis of bed bug bites and their consequence manifestations is beneficial in adapting remedies and treatment protocols advised by clinicians. So far, there is regrettably no definitive way to control these ectoparasites despite extensive efforts of public health authorities to manage them. An overview of the literature and medical documents gathered from bed bug-infested patients referred to the Parasitology and Dermatology departments of Avicenne Hospital (Bobigny, France) allowed us to document and illustrate a range of clinical disorders and psychological concerns caused by bed bugs' bites and their clinical diagnosis. We also review the available tools currently used to control the bed bugs and present potential candidate methods for their successful eradication.

Molecular Research on Vector-Borne Diseases of Medical Interest: From Bench to Application 2.0.

Infectious diseases caused by parasites (malaria, leishmaniasis, trypanosomiasis, filariasis…), viruses (chikungunya, dengue, phlebovirus, etc [...].

Seasonal Patterns of Zoonotic Cutaneous Leishmaniasis Caused by L. major and Transmitted by Phlebotomus papatasi in the North Africa Region, a Systematic Review and a Meta-Analysis

BACKGROUND: In North African countries, zoonotic cutaneous leishmaniasis (ZCL) is a seasonal disease linked to Phlebotomus papatasi, Scopoli, 1786, the primary proven vector of L. major dynamics. Even if the disease is of public health importance, studies of P. papatasi seasonal dynamics are often local and dispersed in space and time. Therefore, a detailed picture of the biology and behavior of the vector linked with climatic factors and the framework of ZCL outbreaks is still lacking at the North African countries' level. Our study aims to fill this gap via a systematic review and meta-analysis of the seasonal incidence of ZCL and the activity of P. papatasi in North African countries. We address the relationship between the seasonal number of declared ZCL cases, the seasonal dynamic of P. papatasi, and climatic variables at the North African region scale. METHODS: We selected 585 publications, dissertations, and archives data published from 1990 to July 2022. The monthly incidence data of ZCL were extracted from 15 documents and those on the seasonal dynamic of P. papatasi from 11 publications from four North African countries. RESULTS: Our analysis disclosed that for most studied sites, the highest ZCL incidence is recorded from October to February (the hibernal season of the vector), while the P. papatasi density peaks primarily during the hot season of June to September. Overall, at the North African region scale, two to four months laps are present before the apparition of the scars reminiscent of infection by L. major. CONCLUSIONS: Such analysis is of interest to regional decision-makers for planning control of ZCL in North African countries. They can also be a rationale on which future field studies combining ZCL disease incidence, vector activity, and climatic data can be built.

A systematic review and global analysis of the seasonal activity of Phlebotomus (Paraphlebotomus) sergenti, the primary vectors of L. tropica.

BACKGROUND: Phlebotomus (Paraphlebotomus) sergenti is a widespread proven vector of Leishmania pathogens causing anthroponotic cutaneous leishmaniasis (ACL), due to L. tropica, in the old world. The activity of P. (Par.) sergenti is seasonal and sensitive to general variations in climate. Phenological data sets can thus provide a baseline for continuing investigations on P. (Par.) sergenti population dynamics that may impact future leishmaniasis transmission and control scenarios. METHODS/PRINCIPAL FINDING: A systematic review of the seasonality of P. (Par.) sergenti was undertaken globally. Six hundred eight scientific papers were identified, and data were extracted from 35 ones, with informative data on sand fly seasonal dynamics on trapping performed from 1992 to December 2021 on 63 sites from 12 countries. Morocco, Saudi Arabia, Iraq, Iran, Pakistan, Palestine, Turkey, Spain, Portugal, Italy, Cyprus, and Georgia. The data extracted from the literature survey were further normalized. Our analysis recorded that the highest P.(Par.) sergenti activity occurs during the hot and dry seasons, primarily in July and August, whatever the location studied. We noticed a relationship between the latitude of sites and sand fly presence (from early April to June) and the type of density trend, varying from a single peak to multiple peaks. On a geographical scale, P. (Par.) sergenti concentrates between 32-37° in latitude in a large interval following the longitude and the highest number of sites with high P. (Par.) sergenti activity is located at the latitude 32°. We also quoted a similar seasonal dynamic and geographic distribution with Phlebotomus (Phlebotomus) papatasi, a proven vector of L. major that causes cutaneous infection. No apparent risk for ACL occurred from December to March, at least in the years and geographic areas considered in this survey. Altogether, knowing that high P. (Par.) sergenti activity would be linked with an increased risk of leishmaniasis transmission, and our study provides information that can be used for control programs on ACL transmission. CONCLUSIONS: Despite variations, we found a relatively homogeneous pattern of P. (Par.) sergenti potential behavior in sites whose data are published. A higher risk for L. tropica transmission was identified in the June-October period. Still, such risk was not equally distributed throughout the area since density waves of adults occurred earlier and were more frequent in some territories, like Saudi Arabia.

Wing Interferential Patterns (WIPs) and machine learning, a step toward automatized tsetse (Glossina spp.) identification.

A simple method for accurately identifying Glossina spp in the field is a challenge to sustain the future elimination of Human African Trypanosomiasis (HAT) as a public health scourge, as well as for the sustainable management of African Animal Trypanosomiasis (AAT). Current methods for Glossina species identification heavily rely on a few well-trained experts. Methodologies that rely on molecular methodologies like DNA barcoding or mass spectrometry protein profiling (MALDI TOFF) haven't been thoroughly investigated for Glossina sp. Nevertheless, because they are destructive, costly, time-consuming, and expensive in infrastructure and materials, they might not be well adapted for the survey of arthropod vectors involved in the transmission of pathogens responsible for Neglected Tropical Diseases, like HAT. This study demonstrates a new type of methodology to classify Glossina species. In conjunction with a deep learning architecture, a database of Wing Interference Patterns (WIPs) representative of the Glossina species involved in the transmission of HAT and AAT was used. This database has 1766 pictures representing 23 Glossina species. This cost-effective methodology, which requires mounting wings on slides and using a commercially available microscope, demonstrates that WIPs are an excellent medium to automatically recognize Glossina species with very high accuracy.

SILVI, an open-source pipeline for T-cell epitope selection.

High-throughput screening of available genomic data and identification of potential antigenic candidates have promoted the development of epitope-based vaccines and therapeutics. Several immunoinformatic tools are available to predict potential epitopes and other immunogenicity-related features, yet it is still challenging and time-consuming to compare and integrate results from different algorithms. We developed the R script SILVI (short for: from in silico to in vivo), to assist in the selection of the potentially most immunogenic T-cell epitopes from Human Leukocyte Antigen (HLA)-binding prediction data. SILVI merges and compares data from available HLA-binding prediction servers, and integrates additional relevant information of predicted epitopes, namely BLASTp alignments with host proteins and physical-chemical properties. The two default criteria applied by SILVI and additional filtering allow the fast selection of the most conserved, promiscuous, strong binding T-cell epitopes. Users may adapt the script at their discretion as it is written in open-source R language. To demonstrate the workflow and present selection options, SILVI was used to integrate HLA-binding prediction results of three example proteins, from viral, bacterial and parasitic microorganisms, containing validated epitopes included in the Immune Epitope Database (IEDB), plus the Human Papillomavirus (HPV) proteome. Applying different filters on predicted IC50, hydrophobicity and mismatches with host proteins allows to significantly reduce the epitope lists with favourable sensitivity and specificity to select immunogenic epitopes. We contemplate SILVI will assist T-cell epitope selections and can be continuously refined in a community-driven manner, helping the improvement and design of peptide-based vaccines or immunotherapies. SILVI development version is available at: github.com/JoanaPissarra/SILVI2020 and https://doi.org/10.5281/zenodo.6865909.

Epidemiology of Vector-Borne Diseases 2.0.

Arthropods' vectors-those of a large variety of families, including Culicidae, Simuliidae, Psychodidae, Ixodidae, Agarsidae, Pulicidae, Glossinidae, Reduviidae, and Tabanidae [...].

Isothermal Nucleic Acid Amplification to Detect Infection Caused by Parasites of the Trypanosomatidae Family: A Literature Review and Opinion on the Laboratory to Field Applicability.

Isothermal amplification of nucleic acids has the potential to be applied in resource-limited areas for the detection of infectious agents, as it does not require complex nucleic purification steps or specific and expensive equipment and reagents to perform the reaction and read the result. Since human and animal infections by pathogens of the Tryponasomatidae family occur mainly in resource-limited areas with scant health infrastructures and personnel, detecting infections by these methodologies would hold great promise. Here, we conduct a narrative review of the literature on the application of isothermal nucleic acid amplification for Trypanosoma and Leishmania infections, which are a scourge for human health and food security. We highlight gaps and propose ways to improve them to translate these powerful technologies into real-world field applications for neglected human and animal diseases caused by Trypanosomatidae.

Diagnosis of animal trypanosomoses: proper use of current tools and future prospects.

Reliable diagnostic tools are needed to choose the appropriate treatment and proper control measures for animal trypanosomoses, some of which are pathogenic. Trypanosoma cruzi, for example, is responsible for Chagas disease in Latin America. Similarly, pathogenic animal trypanosomoses of African origin (ATAO), including a variety of Trypanosoma species and subspecies, are currently found in Africa, Latin America and Asia. ATAO limit global livestock productivity and impact food security and the welfare of domestic animals. This review focusses on implementing previously reviewed diagnostic methods, in a complex epizootiological scenario, by critically assessing diagnostic results at the individual or herd level. In most cases, a single diagnostic method applied at a given time does not unequivocally identify the various parasitological and disease statuses of a host. These include "non-infected", "asymptomatic carrier", "sick infected", "cured/not cured" and/or "multi-infected". The diversity of hosts affected by these animal trypanosomoses and their vectors (or other routes of transmission) is such that integrative, diachronic approaches are needed that combine: (i) parasite detection, (ii) DNA, RNA or antigen detection and (iii) antibody detection, along with epizootiological information. The specificity of antibody detection tests is restricted to the genus or subgenus due to cross-reactivity with other Trypanosoma spp. and Trypanosomatidae, but sensitivity is high. The DNA-based methods implemented over the last three decades have yielded higher specificity and sensitivity for active infection detection in hosts and vectors. However, no single diagnostic method can detect all active infections and/or trypanosome species or subspecies. The proposed integrative approach will improve the prevention, surveillance and monitoring of animal trypanosomoses with the available diagnostic tools. However, further developments are required to address specific gaps in diagnostic methods and the sustainable control or elimination of these diseases.

A Histone Deacetylase (HDAC) Inhibitor with Pleiotropic In Vitro Anti-Toxoplasma and Anti-Plasmodium Activities Controls Acute and Chronic Toxoplasma Infection in Mice.

Toxoplasmosis is a highly prevalent human disease, and virulent strains of this parasite emerge from wild biotopes. Here, we report on the potential of a histone deacetylase (HDAC) inhibitor we previously synthesized, named JF363, to act in vitro against a large panel of Toxoplasma strains, as well as against the liver and blood stages of Plasmodium parasites, the causative agents of malaria. In vivo administration of the drug significantly increases the survival of mice during the acute phase of infection by T. gondii, thus delaying its spreading. We further provide evidence of the compound's efficiency in controlling the formation of cysts in the brain of T. gondii-infected mice. A convincing docking of the JF363 compound in the active site of the five annotated ME49 T. gondii HDACs was performed by extensive sequence-structure comparison modeling. The resulting complexes show a similar mode of binding in the five paralogous structures and a quite similar prediction of affinities in the micromolar range. Altogether, these results pave the way for further development of this compound to treat acute and chronic toxoplasmosis. It also shows promise for the future development of anti-Plasmodium therapeutic interventions.

Molecular characterization of Trichomonas infections in women of Ilam City, southwestern Iran.

Trichomoniasis is a sexually transmitted infection (STI) caused by the flagellated protozoan Trichomonas vaginalis. Little information is available on the epidemiology and genetic diversity of T. vaginalis in Ilam City, southwestern Iran. A descriptive cross-sectional investigation was carried out between July 2017 and December 2018 on the suspected women patients referred to eight gynecology clinics of Ilam City for probable Trichomonas infection. They were undergone a set of clinical, parasitological, and molecular examinations. During clinical consultation, posterior vaginal fornix secretions and urine samples were gathered from the participants. For the reasons such as physical conditions and cultural and religious constraints, most of participating women, especially young girls due to their virginity, preferred to give urine samples instead of vaginal discharge. The presence of Trichomonas was diagnosed by microscopic examination and molecular detection using conventional PCR targeting ITS1-rDNA. A total of 1765 suspected individuals were examined clinically via vaginal secretions (495 specimens) and urine samples (1270 specimens). Of them, 21 (1.18%) cases, including 13 vaginal secretions and 8 urine samples, were positive for Trichomonas infection by microscopy. Slightly more than half of the patients (11/21, 52.4%) complained of vulvar itching, burning, and frequent urination. Cervical lesions, patchy erythema, and vaginal discharge were recorded in 28.6%, 23.8%, and 19% of the patients respectively. All patients with positive microscopic identification were confirmed by amplification of 450-bp fragment of ITS1-rDNA. Phylogenetic analysis revealed a high rate of genetic homogeneity in which all our isolates together with homologous sequences from China, Philippines, Austria, and USA were clustered within the same clade. A statistically significant relationship was recorded between the patients positive for trichomoniasis and the presence of chronic disease (e.g., diabetes, immune system deficiency).

Geographic distribution of Meriones shawi,Psammomys obesus, and Phlebotomus papatasi the main reservoirs and principal vector of zoonotic cutaneous leishmaniasis in the Middle East and North Africa.

Rodents play a significant role in the balance of a terrestrial ecosystem; they are considered prey for many predators like owls and snakes. However, they present a high risk to agriculture (damaging crops) and health. These rodents are the main reservoirs of some vector-borne diseases like leishmaniasis. Meriones shawi (MS) and Psammomys obesus (PO) are the primary Zoonotic cutaneous leishmaniasis (ZCL) reservoirs in the Middle East and North Africa (MENA). A review on the MS and PO at the MENA scale was explored. A database of about 1500 papers was used. 38 sites were investigated as foci for MS and 36 sites for PO, and 83 sites of Phlebotomus papatasi (Pp) in the studied region. An updated map at the regional scale and the trend of the reservoir distribution was carried out using a performing proper density analysis. In this paper, climatic conditions and habitat characteristics of these two reservoirs were reviewed. The association of rodent density with some climatic variables is another aspect explored in a case study from Tunisia in the period 2009-2015 using Pearson correlation. Lastly, the protection and control measures of the reservoir were analyzed. The high concentration of the MS, PO, and Pp can be used as an indicator to identify the high-risk area of leishmaniasis infection.