A deep learning-based model for detecting Leishmania amastigotes in microscopic slides: a new approach to telemedicine.

BACKGROUND: Leishmaniasis, an illness caused by protozoa, accounts for a substantial number of human fatalities globally, thereby emerging as one of the most fatal parasitic diseases. The conventional methods employed for detecting the Leishmania parasite through microscopy are not only time-consuming but also susceptible to errors. Therefore, the main objective of this study is to develop a model based on deep learning, a subfield of artificial intelligence, that could facilitate automated diagnosis of leishmaniasis. METHODS: In this research, we introduce LeishFuNet, a deep learning framework designed for detecting Leishmania parasites in microscopic images. To enhance the performance of our model through same-domain transfer learning, we initially train four distinct models: VGG19, ResNet50, MobileNetV2, and DenseNet 169 on a dataset related to another infectious disease, COVID-19. These trained models are then utilized as new pre-trained models and fine-tuned on a set of 292 self-collected high-resolution microscopic images, consisting of 138 positive cases and 154 negative cases. The final prediction is generated through the fusion of information analyzed by these pre-trained models. Grad-CAM, an explainable artificial intelligence technique, is implemented to demonstrate the model's interpretability. RESULTS: The final results of utilizing our model for detecting amastigotes in microscopic images are as follows: accuracy of 98.95 1.4%, specificity of 98 2.67%, sensitivity of 100%, precision of 97.91 2.77%, F1-score of 98.92 1.43%, and Area Under Receiver Operating Characteristic Curve of 99 1.33. CONCLUSION: The newly devised system is precise, swift, user-friendly, and economical, thus indicating the potential of deep learning as a substitute for the prevailing leishmanial diagnostic techniques.

Leishmaniasis in otorhinolaryngology: an emerging disease and its relationship with anti-tumor necrosis factor-alpha drugs.

PURPOSE: To investigate the clinical manifestations, management and outcomes of Leishmania lesions in the ear-nose-throat (ENT) region, and its relationship with tumor necrosis factor (TNF)-α blocking drugs. METHODS: Single-center retrospective observational study. Patients diagnosed with cutaneous and mucosal leishmaniasis in the otorhinolaryngologic area at a tertiary referral center over a period of 8 years. RESULTS: Three cases of Leishmania lesions in the ear and two in the nose were encountered at our institution. All patients were under treatment with TNF-α blocking drugs. Diagnosis was challenging, and it was important to have a clinical suspicion in order to use accurate detection techniques. All patients received systemic treatment and achieved a complete resolution of the lesions. CONCLUSIONS: With the increasing use of biologic treatments like TNF-α blockers, this type of infection will be increasingly frequent in endemic areas and also worldwide. It is important to include leishmaniasis in the differential diagnosis of inflammatory/infectious lesions in the ENT region.

Unilateral keratitis secondary to Leishmania spp. infection in a horse: Clinical signs and successful topical therapy.

Dermatological clinical signs have been seldom reported in the literature secondary to equine leishmaniasis. This case depicts the clinical signs, treatment, and outcome of a young horse with a pink, elevated lesion on the ventromedial quadrant of the cornea. A corneal cytology was performed and revealed the presence of leishmania amastigotes reaching the diagnosis of keratitis secondary to leishmania. Surgical resection was recommended but the owner declined the procedure, and the lesion was treated with a topical antimonial for 6 weeks. The lesion reduced remarkably during the first weeks of treatment. The patient had not shown recurrence of the lesion for 2 years since the treatment was started. Leishmania spp. can be responsible for ocular surface abnormalities such as keratitis. Corneal cytology is an inexpensive diagnostic method that should be considered when ocular surface abnormalities are identified in horses in endemic areas.

Synchrotron-Infrared Microspectroscopy of Live Leishmania major Infected Macrophages and Isolated Promastigotes and Amastigotes.

The prevalence of neglected tropical diseases (NTDs) is advancing at an alarming rate. The NTD leishmaniasis is now endemic in over 90 tropical and sub-tropical low socioeconomic countries. Current diagnosis for this disease involves serological assessment of infected tissue by either light microscopy, antibody tests, or culturing with in vitro or in vivo animal inoculation. Furthermore, co-infection by other pathogens can make it difficult to accurately determine Leishmania infection with light microscopy. Herein, for the first time, we demonstrate the potential of combining synchrotron Fourier-transform infrared (FTIR) microspectroscopy with powerful discrimination tools, such as partial least squares-discriminant analysis (PLS-DA), support vector machine-discriminant analysis (SVM-DA), and k-nearest neighbors (KNN), to characterize the parasitic forms of Leishmania major both isolated and within infected macrophages. For measurements performed on functional infected and uninfected macrophages in physiological solutions, the sensitivities from PLS-DA, SVM-DA, and KNN classification methods were found to be 0.923, 0.981, and 0.989, while the specificities were 0.897, 1.00, and 0.975, respectively. Cross-validated PLS-DA models on live amastigotes and promastigotes showed a sensitivity and specificity of 0.98 in the lipid region, while a specificity and sensitivity of 1.00 was achieved in the fingerprint region. The study demonstrates the potential of the FTIR technique to identify unique diagnostic bands and utilize them to generate machine learning models to predict Leishmania infection. For the first time, we examine the potential of infrared spectroscopy to study the molecular structure of parasitic forms in their native aqueous functional state, laying the groundwork for future clinical studies using more portable devices.

Identification of infection by Leishmania spp. in wild and domestic animals in Brazil: a systematic review with meta-analysis (2001-2021).

Leishmaniasis is a zoonosis caused by protozoan species of the genus Leishmania. It generates different clinical manifestations in humans and animals, and it infects multiple hosts. Leishmania parasites are transmitted by sandfly vectors. The main objective of this systematic review was to identify the host, or reservoir animal species, of Leishmania spp., with the exception of domestic dogs, that were recorded in Brazil. This review included identification of diagnostic methods, and the species of protozoan circulating in the country. For this purpose, a literature search was conducted across index journals. This study covered the period from 2001 to 2021, and 124 studies were selected. Eleven orders possible hosts were identified, including 229 mammalian species. Perissodactyla had the highest number of infected individuals (30.69%, 925/3014), with the highest occurrence in horses. In Brazil, the most commonly infected species were found to be: horses, domestic cats, rodents, and marsupials. Bats, that were infected by one or more protozoan species, were identified as potential reservoirs of Leishmania spp. Molecular tests were the most commonly used diagnostic methods (94 studies). Many studies have detected Leishmania spp. (n = 1422): Leishmania (Leishmania) infantum (n = 705), Leishmania (Viannia) braziliensis (n = 319), and Leishmania (Leishmania) amazonensis (n = 141). Recognizing the species of animals involved in the epidemiology and biological cycle of the protozoan is important, as this allows for the identification of environmental biomarkers, knowledge of Leishmania species can improve the control zoonotic leishmaniasis.

Disposable electrochemical platform based on solid-binding peptides and carbon nanomaterials: an alternative device for leishmaniasis detection.

Neglected tropical diseases are those caused by infectious agents or parasites and are considered endemic in low-income populations. These diseases also have unacceptable indicators and low investment in research, drug production, and control. Tropical diseases such as leishmaniasis are some of the main causes of morbidity and mortality around the globe. Electrochemical immunosensors are promising tools for diagnostics against these diseases. One such benefit is the possibility of assisting diagnosis in isolated regions, where laboratory infrastructure is lacking. In this work, different peptides were investigated to detect antibodies against Leishmania in human and canine serum samples. The peptides evaluated (395-KKG and 395-G) have the same recognition site but differ on their solid-binding domains, which ensure affinity to spontaneously bind to either graphene oxide (GO) or graphene quantum dots (GQD). Cyclic voltammetry and differential pulse voltammetry were employed to investigate the electrochemical behavior of each assembly step and the role of each solid-binding domain coupled to its anchoring material. The graphene affinity peptide (395-G) showed better reproducibility and selectivity when coupled to GQD. Under the optimized set of experimental conditions, negative and positive human serum samples responses were distinguished based on a cut-off value of 82.5% at a 95% confidence level. The immunosensor showed selective behavior to antibodies against Mycobacterium leprae and Mycobacterium tuberculosis, which are similar antibodies and potentially sources of false positive tests. Therefore, the use of the graphene affinity peptide as a recognition site achieved outstanding performance for the detection of Leishmania antibodies.

Detection and differentiation of Leishmania parasites in asymptomatic canine by High-Resolution Melting analysis of microsatellite fragment in ITS gene.

The high prevalence of Leishmania infection was reported in dogs as the main reservoir of CanL in many locations in the old world. Detection and firmly identification of Leishmania species in asymptomatic dogs by reliable method was considered and employed. Non-invasive and non-anesthetized blood sampling in asymptomatic dogs was conducted. Nested, conventional and real-time PCR with HRM technique was performed targeting ITS-rDNA gene. 88 asymptomatic dogs were sampled from three CanL endemic provinces of Iran in 2018-2019. 23 blood samples were Leishmania positive. L. major, L. tropica and L. infantum were accurately identified for the first time with HRM targeting ITS2-microsatellite. Three samples were mixed infections. CLC software TM predictions for microsatellite ITS-rDNA were 86.93 °C: L. major, 85.76 °C: L. tropica and 86.04 °C: L. infantum. Standard strains of Leishmania species were accurately separated with almost one to 2 °C deference (L. major: 86.61 °C, L. infantum: 85.41 °C, L. tropica: 84.82 °C). Each HRM curve represents one species in a sample for helping accurate identification of Leishmania species and even mixed infection when two curves are present. Detecting parasites at primary stages in asymptomatic cases is essential using Real-time HRM. As same as mammalian Leishmania in rodents which is present at early stages and non-pathogenesis, only L. major would exist and other Leishmania disappears. This can conclude also for L. major, L. infantum and L. tropica in dogs. The role of L. major existence in canine blood should be investigated more.

Evaluation from a B-cell epitope-based chimeric protein for the serodiagnosis of tegumentary and visceral leishmaniasis.

The diagnosis of leishmaniasis presents problems due to the variable sensitivity and/or specificity of tests. In addition, high levels of anti-parasite antibodies can remain after treatment, making it difficult to conduct a prognostic follow-up of patients. In this context, it is necessary to identify new candidates to be examined for the sensitive and specific diagnosis of the disease. In the present study, four Leishmania proteins, previously shown as antigenic for tegumentary leishmaniasis (TL), were evaluated, and their linear specific B-cell epitopes were predicted and used to generate a new gene codifying chimeric protein called ChimB, which was cloned, and the recombinant version was expressed, purified, and evaluated in ELISA (Enzyme-Linked Immunosorbent Assay) to diagnose TL and visceral leishmaniasis (VL). A total of 220 human serum samples were used, and, when ChimB was used, results showed sensitivity, specificity, and positive and negative predictive values of 100% for the diagnosis of both diseases; however, when using peptides, the sensitivity values reached from 28.0% to 57.3% and specificity varied from 16.3% to 83.7%. A soluble Leishmania extract (SLA) showed sensitivity and specificity values of 30.7% and 45.9%, respectively. The area under the curve (AUC) value for ChimB was 1.0, while for synthetic peptides, this value reached between 0.502 and 0.635, whereas for SLA, the value was of 0.589. Serological assays using sera samples collected before and after treatment showed significant reductions in the anti-ChimB antibody levels after therapy, suggesting a prognostic role of this recombinant antigen. In conclusion, preliminary data suggest the use from ChimB as a potential candidate for the diagnosis and prognosis of leishmaniasis.

A machine learning-based system for detecting leishmaniasis in microscopic images.

BACKGROUND: Leishmaniasis, a disease caused by a protozoan, causes numerous deaths in humans each year. After malaria, leishmaniasis is known to be the deadliest parasitic disease globally. Direct visual detection of leishmania parasite through microscopy is the frequent method for diagnosis of this disease. However, this method is time-consuming and subject to errors. This study was aimed to develop an artificial intelligence-based algorithm for automatic diagnosis of leishmaniasis. METHODS: We used the Viola-Jones algorithm to develop a leishmania parasite detection system. The algorithm includes three procedures: feature extraction, integral image creation, and classification. Haar-like features are used as features. An integral image was used to represent an abstract of the image that significantly speeds up the algorithm. The adaBoost technique was used to select the discriminate features and to train the classifier. RESULTS: A 65% recall and 50% precision was concluded in the detection of macrophages infected with the leishmania parasite. Also, these numbers were 52% and 71%, respectively, related to amastigotes outside of macrophages. CONCLUSION: The developed system is accurate, fast, easy to use, and cost-effective. Therefore, artificial intelligence might be used as an alternative for the current leishmanial diagnosis methods.

Surveillance of leishmaniasis cases from 15 European centres, 2014 to 2019: a retrospective analysis.

BackgroundSurveillance of human leishmaniasis in Europe is mostly limited to country-specific information from autochthonous infections in the southern part. As at the end of 2021, no integrated analysis has been performed for cases seen across centres in different European countries.AimTo provide a broad perspective on autochthonous and imported leishmaniasis cases in endemic and non-endemic countries in Europe.MethodsWe retrospectively collected records from cutaneous, mucosal and visceral leishmaniasis cases diagnosed in 15 centres between 2014 and 2019. Centres were located in 11 countries: Belgium, France, Germany, Italy, the Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and the United Kingdom. Data on country of infection, reason for travelling, infecting species, age and sex were analysed.ResultsWe obtained diagnostic files from 1,142 cases, of which 76%, 21% and 3% had cutaneous, visceral, and mucosal disease, respectively. Of these, 68% were men, and 32% women, with the median age of 37 years (range: 0-90) at diagnosis. Visceral leishmaniasis was mainly acquired in Europe (88%; 167/190), while cutaneous leishmaniasis was primarily imported from outside Europe (77%; 575/749). Sixty-two percent of cutaneous leishmaniasis cases from outside Europe were from the Old World, and 38% from the New World. Geographic species distribution largely confirmed known epidemiology, with notable exceptions.ConclusionsOur study confirms previous reports regarding geographic origin, species, and traveller subgroups importing leishmaniasis into Europe. We demonstrate the importance of pooling species typing data from many centres, even from areas where the aetiology is presumably known, to monitor changing epidemiology.

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.

Immunoproteomics characterization of Leishmania panamensis proteins for potential clinical diagnosis of mucosal Leishmaniasis.

Diagnosis of leishmaniasis based on antibodies detection represents a challenge due to cross-reaction of sera with other infectious agents, which co-exist in endemic areas of Leishmania sp, especially patients with Trypanosoma cruzi. This work is aimed at searching for immunogenic proteins in sera from patients with cutaneous and mucosal leishmaniasis that may be potential candidates for the development of diagnostic tests and/or vaccines that help control the infection. Total protein extracts of L. panamensis promastigotes were put in contact with sera from patients with cutaneous and mucosal leishmaniasis (immunoblots). Immunoreactive proteins were identified by mass spectrometry and bioinformatics tools. 81 proteins were identified. One of these was uniquely recognized by the sera from patients with ML but not from sera from either CL or Chagas disease patients. MS analysis of this band pointed to the putative leishmanial 3-oxoacyl-(Acylcarrierprotein) reductase.

A Nuclear Magnetic Resonance-Based Metabolomic Study to Identify Metabolite Differences between Iranian Isolates of Leishmania major and Leishmania tropica.

Background: Cutaneous leishmaniasis caused by Leishmania species (L. spp) is one of the most important parasitic diseases in humans. To gain information on the metabolite variations and biochemical pathways between L. spp, we used the comparative metabolome of metacyclic promastigotes in the Iranian isolates of L. major and L. tropica by proton nuclear magnetic resonance (1H-NMR). Methods: L. tropica and L. major were collected from three areas of Iran, namely Gonbad, Mashhad, and Bam, between 2017 and 2018, and were cultured. The metacyclic promastigote of each species was separated, and cell metabolites were extracted. 1H-NMR spectroscopy was applied, and the data were processed using ProMatab in MATLAB (version 7.8.0.347). Multivariate statistical analyses, including the principal component analysis and the orthogonal projections to latent structures discriminant analysis, were performed to identify the discriminative metabolites between the two L. spp. Metabolites with variable influences in projection values of more than one and a P value of less than 0.05 were marked as significant differences. Results: A set of metabolites were detected, and 24 significantly differentially expressed metabolites were found between the metacyclic forms of L. major and L. tropica isolates. The top differential metabolites were methionine, aspartate, betaine, and acetylcarnitine, which were increased more in L. tropica than L. major (P<0.005), whereas asparagine, 3-hydroxybutyrate, L-proline, and kynurenine were increased significantly in L. major (P<0.01). The significantly altered metabolites were involved in eight metabolic pathways. Conclusion: Metabolomics, as an invaluable technique, yielded significant metabolites, and their biochemical pathways related to the metacyclic promastigotes of L. major and L. tropica. The findings offer greater insights into parasite biology and how pathogens adapt to their hosts.

Plasmonic rK28 ELISA improves the diagnosis of canine Leishmania infection.

In this study, we evaluated the performance of a new enzyme-linked immunosorbent assay (ELISA) variant known as indirect "plasmonic ELISA" (pELISA) for the detection of Leishmania spp. infection. Serum samples from 170 dogs from an area where canine leishmaniosis (CanL) is endemic and from 26 healthy dogs from a nonendemic area were tested by indirect pELISA, and the results were compared to those of an indirect ELISA (both with recombinant antigen rK28) and those of an immunochromatographic test (dual-path platform, TR-DPP®) using real-time PCR on blood samples or conjunctival swabs as the gold standard. The pELISA, indirect rK28 ELISA and the TR-DPP® immunochromatographic test presented sensitivities of 94.7%, 89.5% and 79.0% and specificities of 100%, 92.7% and 91.5%, respectively. The analysis of the results revealed that the specificity of the indirect pELISA was greater than that of the method recommended by the Ministry of Health in Brazil and may increase the feasibility of diagnosis in resource-constrained countries because it does not require sophisticated instruments to read. Thus, this method can be used as an additional tool for the detection of Leishmania spp. infection in these areas.

Noninvasive Biological Samples to Detect and Diagnose Infections due to Trypanosomatidae Parasites: A Systematic Review and Meta-Analysis.

Unicellular eukaryotes of the Trypanosomatidae family include human and animal pathogens that belong to the Trypanosoma and Leishmania genera. Diagnosis of the diseases they cause requires the sampling of body fluids (e.g., blood, lymph, peritoneal fluid, cerebrospinal fluid) or organ biopsies (e.g., bone marrow, spleen), which are mostly obtained through invasive methods. Body fluids or appendages can be alternatives to these invasive biopsies but appropriateness remains poorly studied. To further address this question, we perform a systematic review on clues evidencing the presence of parasites, genetic material, antibodies, and antigens in body secretions, appendages, or the organs or proximal tissues that produce these materials. Paper selection was based on searches in PubMed, Web of Science, WorldWideScience, SciELO, Embase, and Google. The information of each selected article (n = 333) was classified into different sections and data were extracted from 77 papers. The presence of Trypanosomatidae parasites has been tracked in most of organs or proximal tissues that produce body secretions or appendages, in naturally or experimentally infected hosts. The meta-analysis highlights the paucity of studies on human African trypanosomiasis and an absence on animal trypanosomiasis. Among the collected data high heterogeneity in terms of the I2 statistic (100%) is recorded. A high positivity is recorded for antibody and genetic material detection in urine of patients and dogs suffering leishmaniasis, and of antigens for leishmaniasis and Chagas disease. Data on conjunctival swabs can be analyzed with molecular methods solely for dogs suffering canine visceral leishmaniasis. Saliva and hair/bristles showed a pretty good positivity that support their potential to be used for leishmaniasis diagnosis. In conclusion, our study pinpoints significant gaps that need to be filled in order to properly address the interest of body secretion and hair or bristles for the diagnosis of infections caused by Leishmania and by other Trypanosomatidae parasites.

Zoonotic Leishmaniasis, Bosnia and Herzegovina.

Leishmania infantum causes potentially life-threatening disease in humans. To determine the extent of the animal reservoir for this pathogen in Bosnia and Herzegovina, we tested dogs and cats. We found that a large proportion of dogs were exposed to or infected with L. infantum, indicating endemicity in dogs and zoonotic risk for humans.

Recombinant Leishmania eukaryotic elongation factor-1 beta protein: A potential diagnostic antigen to detect tegumentary and visceral leishmaniasis in dogs and humans.

The laboratorial diagnosis of leishmaniasis is based on parasitological methods, which are invasive, present high cost, require laboratorial infrastructure and/or trained professionals; as well as by immunological methods, which usually present variable sensitivity and/or specificity, such as when they are applied to identify asymptomatic cases and/or mammalian hosts presenting low levels of antileishmanial antibodies. As consequence, new studies aiming to identify more refined antigens to diagnose visceral (VL) and tegumentary (TL) leishmaniasis are urgently necessary. In the present work, the Leishmania eukaryotic elongation factor-1 beta (EF1b) protein, which was identified in L. infantum protein extracts by antibodies in VL patients' sera, was cloned and its recombinant version (rEF1b) was expressed, purified and tested as a diagnostic marker for VL and TL. The post-therapeutic serological follow-up was also evaluated in treated and untreated VL and TL patients, when anti-rEF1b antibody levels were measured before and after treatment. Results showed that rEF1b was highly sensitive and specific to diagnose symptomatic and asymptomatic canine VL, as well as human TL and VL. In addition, low cross-reactivity was observed when sera from healthy subjects or leishmaniasis-related diseases patients were tested. The serological follow-up showed also that rEF1b-specific antibodies declined significantly after treatment, suggesting that this protein could be also evaluated as a prognostic marker for human leishmaniasis.

Tissue and blood protozoa including toxoplasmosis, Chagas disease, leishmaniasis, Babesia, Acanthamoeba, Balamuthia, and Naegleria in solid organ transplant recipients- Guidelines from the American Society of Transplantation Infectious Diseases Community of Practice.

These updated guidelines from the Infectious Diseases Community of Practice of the American Society of Transplantation review the diagnosis, prevention, and management of tissue and blood protozoal infections in the pre- and post-transplant period. Significant new developments in the field have made it necessary to divide the previous single guideline published in 2013 into two sections, with the intestinal parasites separated from this guideline devoted to tissue and blood protozoa. The current update reflects the increased focus on donor screening and risk-based recipient monitoring for parasitic infections. Increased donor testing has led to new recommendations for recipient management of Toxoplasma gondii and Trypanosoma cruzi. Molecular diagnostics have impacted the field, with access to rapid diagnostic testing for malaria and polymerase chain reaction testing for Leishmania. Changes in Babesia treatment regimens in the immunocompromised host are outlined. The risk of donor transmission of free-living amebae infection is reviewed. Changing immigration patterns and the expansion of transplant medicine in developing countries has contributed to the recognition of parasitic infections as an important threat to transplant outcomes. Medications such as benznidazole and miltefosine are now available to US prescribers as access to treatment of tissue and blood protozoa is increasingly prioritized.

A highly sensitive modified nested PCR to enhance case detection in leishmaniasis.

BACKGROUND: Human leishmaniasis is one of the major parasitic diseases with worldwide distribution. Sri Lanka is a recently established focus of leishmaniasis caused by a variant Leishmania donovani. Early case detection and management is a main approach identified for L. donovani control in the regional leishmaniasis elimination drive. Usefulness of light microscopy and in-vitro culture are limited in chronic, atypical or treated lesions though timely and accurate detection of all light microscopy/in-vitro culture negative cases of all forms of leishmaniasis is necessary for treatment. Timely treatment is important to minimize risk for death in visceral disease and undesired sequelae of long standing infection and illness on both patients and community. We described a 100% sensitive, Leishmania spp. specific modified version of a nested PCR (Mo-STNPCR) that also minimizes carry over and cross contaminations while facilitate investigation of light microscopy and in-vitro culture negative clinically suggestive cases of leishmaniasis. METHODS: Leishmania DNA was amplified using previously published P221: 5'-GGTTCCTTTCCTGATTTACG-3' and P332: 5'-GGCCGGTAAAGGCCGAATAG-3'outer primers followed by a nested reaction using P223: 5'-TCCCATCGCAACCTCGGTT-3' and P333: 5'-AAGCGGGCGCGGTGCTG-3' inner primers that by passes the requirement of tube handling between the two steps of the conventional nested PCR. Leishmania DNA was detected in a range of infected tissue material. Infected material from patients with cutaneous leishmaniasis (n = 30), visceral leishmaniasis (n = 10) and from a control group including patients with non-leishmanial skin diseases (n = 10), other systemic diseases (n = 10) and healthy individuals (n = 10) were examined with Mo-STNPCR. Results were further compared with those of light microscopy and in-vitro culture. RESULTS: Mo-STNPCR method was 100% sensitive and 100% specific for diagnosis of leishmaniasis. Light microscopy and in-vitro culture were positive in 75.0% (n = 30/40) and 72.5% (n = 29/40) samples respectively where combined results of them gave 87.5% (n = 35/40) sensitivity. Mo-STNPCR did not cross react with control samples. Furthermore, Mo-STNPCR reduces the risk of cross-contaminations and carry over contaminations since the full reaction is carried out without opening the tubes. Per patient cost was calculated as 22 USD while the same was 3 and 6 USD for light microscopy and in-vitro culture respectively. CONCLUSION: Mo-STNPCR method is a useful tool in detecting leishmaniasis in minority of cases that go undetected by first line investigations.

Travelers' tropical skin diseases: Challenges and interventions.

Tropical regions receive a significant part of the traveling population. It is very important that health professionals are familiar with the main tropical skin diseases and able to advice patients appropriately. This article reviews the main tropical diseases of travelers, with an emphasis on diagnosis, management, and prevention. Among others, cutaneous larva migrans, myiasis, tungiasis, Chagas disease, Dengue fever, African trypanosomiasis, filariasis, and leishmaniasis are discussed. Increasing awareness among travelers and health care professionals can help reduce morbidity and mortality. Continued research on new drugs and vaccines is needed to reduce the risks of tropical diseases.