Sand Fly Bites Research Articles

Distinct neutrophil effector functions in response to different isolates of Leishmania aethiopica.

In Ethiopia, cutaneous leishmaniasis is mainly caused by Leishmania (L.) aethiopica parasites and presents in three main clinical forms. It is still not clear if the host immune response plays a role in the development of these different presentations. Since neutrophils are likely to be one of the first immune cells present at the site of the sand fly bite, we set up an in vitro model of infection of neutrophils with L. aethiopica and assessed some of the main neutrophil effector functions: association with and internalisation of parasites, apoptosis and ROS production. We used three freshly isolated clinical isolates and one isolate that has been kept in culture for decades. Our results showed by flow cytometry that all four L. aethiopica isolates had the ability to associate with neutrophils. The three clinical isolates of L. aethiopica associated more efficiently with neutrophils than the long-term cultured L. aethiopica. At 18h, two distinct populations of neutrophils were identified that associated with L. aethiopica, CD15high and CD15low neutrophils. Confocal microscopy demonstrated that all isolates can be internalised. Our results also showed that all parasites induced apoptosis in L. aethiopica-associated neutrophils. Moreover, our results showed that after 2h, L. aethiopica-associated neutrophils upregulated their production of ROS, but to a greater extent with the long-term cultured L. aethiopica. After 18h of incubation, CD15lowparasite+ showed an impaired ability to produce ROS compared to CD15highparasite+. Using this in vitro model, our results show that different L. aethiopica parasite isolates, most notably long-term cultured parasites, had differential effects on neutrophil effector functions.

Cutaneous leishmaniasis in British troops following jungle training in Belize: Cumulative incidence and potential risk practices

BackgroundBritish soldiers undergoing jungle training in Belize typically experience a relatively low risk of developing cutaneous leishmaniasis. However, an uncharacteristically large outbreak of cutaneous leishmaniasis occurred in 2022. This study aimed to determine the cumulative incidence of the disease and highlight potential shortcomings in personal protective measures to mitigate exposure to sand fly vector bites. MethodsA retrospective analysis was conducted on medical records of cutaneous leishmaniasis cases between 2005 and 2022, as well as on questionnaire responses regarding personal protective measures administered to cutaneous leishmaniasis cases in 2022. Data were sourced from Defence Public Health Unit, Military Environmental Health Department and British Army Training Support Unit Belize. ResultsEighty-one confirmed clinical cutaneous leishmaniasis cases were recorded between 2005 and 2022, with a substantial peak (38 cases) in 2022. Most cases occurred during the wet season. Pre-2022, the median cumulative incidence per 8-week deployment was 0.90 % (Q1–Q3: 0.34 %–1.34 %), with an annual variation of 0.2 % to 2.0 %. In 2022, the cumulative incidence spiked to 4.22 %, associated with a risk ratio of 5.3 (95 % C.I.s, 3.41, 8.16), and rising to a cumulative incidence of 7.3 % in a single unit of 450 men (33 cases) in late 2022. These values are significantly higher than the median cumulative incidence of all previous years, and to published reports for other cutaneous leishmaniasis -endemic regions. Troop responses identified limitations in the supply of optimal equipment, and in sand fly bite and leishmaniasis risk avoidance information provided by the pre-deployment health education programme. Compliance with health education advise was also suboptimal, with irregular use of insect repellents, protective clothing / head netting, and insecticide-treated hammocks. ConclusionsThe reasons behind the unusually high numbers of cutaneous leishmaniasis cases and cumulative incidence in 2022 remain unclear, emphasising the need to improve personal protective measures provision and implement a comprehensive health education programme for troops undergoing jungle training in Belize.

Leishmaniasis in Humans and Animals: A One Health Approach for Surveillance, Prevention and Control in a Changing World

Leishmaniasis is classified as a neglected tropical disease (NTD), caused by protozoan parasites of the genus Leishmania, which are transmitted to humans and other animals through the bite of infected female phlebotomine sandflies. There are three forms of the disease: cutaneous leishmaniasis (CL) manifested by ulcers and scars; systemic or visceral leishmaniasis (VL), which can lead to life-threatening complications if left untreated; and mucocutaneous leishmaniasis (MCL), which can destroy the mucous membranes of the nose, mouth and throat. Human leishmaniasis is endemic in many countries across Africa, Asia, Southern Europe, the Middle East, and Central and South America. The interconnection of environmental, animal and human health underlies the spread of the Leishmania parasite. Environmental disruptions, such as climate change, deforestation or urbanisation, but also globalisation and migration, significantly affect the distribution and abundance of sand fly vectors and reservoir hosts. Climate change alters the breeding patterns of sandflies and expands their geographic range; deforestation and misuse of large areas disrupt ecosystems, leading to increased human-vector contact; and urbanisation increases the potential for contact between parties, particularly in densely populated areas. Migration of humans and animals, either through natural migration or, for example, the pet trade and breeding, can facilitate the spread of Leishmania parasites. In addition, socio-economic factors, including poverty and lack of access to healthcare, increase the burden of leishmaniasis in vulnerable populations. Due to this multitude of reasons, the geographic distribution of sandflies has expanded to higher latitudes and altitudes in recent years, with a consequent increase in disease burden. Indeed, despite ongoing challenges in the surveillance systems, data from the last available year have shown an increase in many cases in both humans and dogs. This perspective explores the interconnected factors influencing the spread of leishmaniasis worldwide and the epidemiology of the disease. In addition, it illustrates the importance of integrated strategies in a One Health approach: surveillance, prevention and control of vectors, animals and humans.

Prevalence of Leishmania infantum in felines from Heraklion region, Crete

Greece is recognized as an endemic region for various species of Leishmania, a genus of parasitic protozoa transmitted through the bites of infected female sandflies. This indicates that the country serves as a natural reservoir for this parasite, infecting both humans and animals. This study focuses on Heraklion, Crete, one of Greece's largest cities, where there is currently no available data on the prevalence of leishmaniosis in the feline population. While cases of canine leishmaniosis have been documented in various parts of the island, there have been no reported cases of feline leishmaniosis in Heraklion. This study primarily investigates the prevalence of leishmaniosis in cats as a means of detection in this species in Heraklion. To accomplish this, between 2018 – 2023, EDTA-blood samples and conjunctival swab samples were collected from a total of 253 feral cats. The cats were divided into two groups, depending on the location they lived (i.e., areas out of the city center or close to/in the city center): the initial group consisted of 112 cats, while the latter group comprised 141 cats. Both groups encompassed cats of various ages, habitats, and both genders. The samples were stored at -18 °C and analyzed with TaqMan real-time quantitative PCR for the presence of Leishmania infantum DNA.Out of the cat population included in the study, 15 were tested positive for FeL (5.93%). Despite the lower prevalence of leishmaniosis compared to dogs, it should be taken into account, especially for differential diagnosis. In conclusion, to the best of our knowledge, this is the first study that confirms the presence of FeL in felines in Heraklion.

Cutaneous Leishmaniasis in Dr. Ersin Arslan Training and Research Hospital After Migration and During the Pandemic (2019-2022).

Cutaneous leishmaniasis is a parasitic skin disease transmitted by the bite of sandflies. In our region, which is endemic for this disease, there has been a great migration from a much more endemic region and population movements from our area to Türkiye and abroad. Afterward, a pandemic was experienced. Due to these two extraordinary events and the possible epidemic potential in our region, it is useful to follow-up on the disease. We aimed to contribute to the evaluation of the disease in these processes by analyzing the data of our laboratory in recent years. Between January 2019 and December 2022, samples from patients who came to our laboratory with suspected cutaneous leishmaniasis were taken, stained and examined under a microscope. Patients were evaluated in terms of age, gender, nationality, place of residence, lesion site and duration. Out of the 144 examined cases, 64 (44.4%) were positive for cutaneous leishmaniasis. Among these positive cases, 40 (62.5%) were women, 24 (37.5%) were men, and 54 (84.3%) belonged to the 0-9 age group. Of those who tested positive, 54 (84.3%) were Turkish citizens and 23 (35.9%) were Syrian citizens. Fifty-four (84.3%) patients had only single lesion. While the number of applications and positivity rates remained within normal levels in 2019 and 2020, a significant decrease was observed in both from 2021 and 2022. Cutaneous leishmaniasis is carried by migration, decreases in large-scale isolations such as pandemics, and its spread can be prevented with correct diagnosis and treatment. Although the number of patients may change over time and place, cutaneous leishmaniasis is a disease that threatens the health of societies and should always be monitored.

Astrocytes Can Be Key Players Against Cerebral Leishmaniasis: InVitro Co-Culture Model for the Assessment of Infection.

Leishmaniasis is a neglected tropical disease, caused by protozoan parasites of Leishmania (L.), and is transmitted by bite of phlebotomine sandflies. There are several studies on central nervous system infection to indicate that Leishmania can cross the blood-brain barrier, resulting in neurological manifestations, known as "cerebral leishmaniasis." This study highlighted the notions: (i) polarisation of bone marrow-derived macrophages (BMDM) incubated following stimulation with lipopolysaccharide (LPS) or soluble Leishmania antigen (SLA), (ii) quantification of parasites within co-culture of Leishmania-infected macrophages, and astrocytes, and (iii) effect of interferon-gamma (IFN-γ) on the infection rate of co-culture populations. Accordingly, 83% of overall macrophage population was identified on day 7 for CD11b and F4/80 macrophage markers. Flow cytometry analysis revealed significant increases in CD11b and F4/80 surface markers in LPS and SLA-stimulated BMDMs at 24 h, compared to untreated cells. TNF-α levels increased significantly in both LPS and SLA-treated BMDMs after 48 h. Additionally, SLA treatment induced a more elongated, spindle-like shape in the cells, indicative of M2 macrophage polarisation over the M1 phenotype. When non-infected astrocytes with/without stimulation with IFN-γ before co-culture, gp63 FITC-labelled parasite populations (%) in co-culture decreased to 25% at 72 h, thus indicating a lower infection rate in a time-dependent manner. IFN-γ and IL-6 levels significantly increased to 71.66 ± 3.51 and 184 ± 14.42 pg/mL, resulting in the inflammatory response in the co-culture system at 48 h (p ≤ 0.0001), when compared to the control (30 ± 2.52 pg/mL for IFN-γ and 8.66 ± 2.37 pg/mL for IL-6) at 0 h of the incubation. It is the first study to emphasize the communication between Leishmania-infected macrophages and astrocytes regarding Leishmania parasite load. The results suggest that astrocytes can lead to the reduction in Leishmania parasites, thereby controlling the incidence of cerebral leishmaniasis.

Effects of Amphotericin B-Conjugated Functionalized Carbon Nanoparticles in the Treatment of Cutaneous Leishmaniasis.

Leishmaniasis is a parasitic disease spread by the bite of an infected sandfly and caused by protozoan parasites of the genus Leishmania. Currently, there is no vaccine available for leishmaniasis in humans, and the existing chemotherapy methods face various clinical challenges. The majority of drugs are limited to a few toxic compounds, with some parasite strains developing resistance. Therefore, the discovery and development of a new anti-leishmanial compound is crucial. One promising strategy involves the use of nanoparticle delivery systems to accelerate the effectiveness of existing treatments. In this study, Amphotericin B (AmB) was incorporated into functionalized carbon nanotube (f-CNT) and evaluated for its efficacy against Leishmania major in vitro and in a BALB/c mice model. The increase in footpad thickness was measured, and real-time PCR was used to quantify the parasite load post-infection. Levels of nitric oxide and cytokines IL-4 and IFN-γ were also determined. We found that f-CNT-AmB significantly reduced the levels of promastigotes and amastigotes of the Leishmania parasite. The nanoparticle showed strong anti-leishmanial activity with an IC50 of 0.00494 ± 0.00095 mg/mL for promastigotes and EC50 of 0.00294 ± 0.00065 mg/mL for amastigotes at 72 h post-infection, without causing harm to mice macrophages. Treatment of infected BALB/c mice with f-CNT-AmB resulted in a significant decrease in cutaneous leishmania (CL) lesion size in the foot pad, as well as reduced Leishmania burden in both lymph nodes and spleen. The levels of nitric oxide and IFN-γ significantly increased in the f-CNT-AmB treated groups. Also, our results showed that the level of IL-4 significantly decreased after f-CNT-AmB treatment in comparison to other groups. In conclusion, our results demonstrate that AmB loaded into f-CNT is significantly more effective than AmB alone in inhibiting parasite propagation and promoting a shift towards a Th1 response.

Measuring the Concentration of Interleukin-6 (IL-6) and Tumor Necrosis Factor-alpha (TNF-α) in People infected with Cutaneous leishmaniasis in Tikrit

The study conducted in Salahaddeen Governorate aimed to measure the concentrations of cytokines (interleukin-6 and tumor necrosis factor-alpha) in patients infected with Baghdad boil and to compare these concentrations with those of the control group. The endemic disease of leishmaniasis manifests in three types in Iraq; cutaneous, mucocutaneous and visceral. The most common type is cutaneous leishmaniasis (CL) which causes skin sores. CL attributed to L. tropica is disseminated via the bite of an infected female sand fly. The study was conducted at Salahaddeen Teaching Hospital in Salahaddeen Governorate, Iraq, from March 1, 2021, to September 1, 2022. Detailed data were collected about age, gender, residence, location, number and type of lesions (wet or dry). Sixty clinical cases of cutaneous leishmaniasis were included in the study. The cytokine concentration of interleukin-6 (IL-6) exhibited an increase, reaching 94.7a±7.74 picograms/ml in patients infected with multiple leishmaniasis lesions (7-10 lesions). Conversely, the lowest concentration was observed at 56.3b±7.43 picograms/ml in patients with 1-3 lesions. Furthermore, the concentration of interleukin-6 reached its peak in patients with dry lesions (96.0a±10.9) compared to those with wet lesions (63.3b±12.4). An elevation in tumor necrosis factor (TNF-α) levels was observed across different age groups, with patients infected with cutaneous leishmaniasis typically falling within the age range of 11 to 40. Consistently, the highest TNF-α levels were documented annually in patients with cutaneous leishmaniasis, specifically within the age brackets of 11-20, 21-30 and 31-40, reaching 343.0a±28.3 picograms/ml, 316.7abd±41.2 picograms/ml and 295.9b±51.9 picograms/ml respectively, in comparison to the control group. Conversely, the lowest TNF-α level was noted in patients aged 41 (238.0c±42.6 picograms/ml). Moreover, TNF-α concentration was higher in patients with leishmaniasis in rural areas compared to urban areas. In rural regions, the TNF-α concentration was 436.7a±54.4 picograms/ml.

Ex Vivo Analysis of the Association of GFP-Expressing L. aethiopica and L. mexicana with Human Peripheral Blood-Derived (PBD) Leukocytes over 24 Hours.

Leishmania parasites are transmitted to mammalian hosts through the bite of sandflies. These parasites can infect phagocytic cells (macrophages, dendritic cells, and neutrophils) and non-phagocytic cells (B cells and fibroblasts). In mice models, the disease development or resolution is linked to T cell responses involving inflammatory cytokines and the activation of macrophages with the M1/M2 phenotype. However, this mechanism does not apply to human infection where a more complex immunological response occurs. The understanding of interactions between immune cells during Leishmania infection in humans is still limited, as current infection models focus on individual cell types or late infection using controlled human infection models (CHIMs). This study investigated the early parasite infection in freshly isolated peripheral blood-derived (PBD) leukocytes over 24 h. Flow cytometer analysis is used in immunophenotyping to identify different subpopulations. The study found that among the L. aethiopicaGFP-associated leukocytes, most cells were neutrophils (55.87% ± 0.09 at 4 h) and monocytes (23.50% ± 0.05% at 24 h). B cells were 12.43% ± 0.10% at 24 h. Additionally, 10-20% of GFP+ leukocytes did not belong to the aforementioned cell types, and further investigation revealed their identity as CD4+ T cells. Data not only confirm previous findings of Leishmania infection with PBD leukocytes and association with B cells but also suggest that CD4+ T cells might influence the early-stage of infection.

Development of Amphotericin B Decorated Gold Nanoparticles as a Promising Antileishmanial Nanoconjugate.

Leishmaniasis, attributed to the protozoan parasite Leishmania, manifests in diverse clinical forms, including cutaneous, mucocutaneous, and visceral leishmaniasis; VL constitutes a significant global health menace. Prevalent in tropical and subtropical regions, this affliction disproportionately impacts individuals below the poverty threshold, transmitted through the bite of female sandflies. Existing treatments, such as pentavalent antimony, miltefosine, and Amphotericin B, exhibit limitations. Despite the emergence of liposomal Amphotericin B (AmBisome) as a promising antileishmanial agent, its utility is impeded by adverse effects, elevated production expenses, and cytotoxicity. To address these challenges, our investigation introduces a potential remedy─a citrate-coated gold Amphotericin B nanoparticle formulation. Characterized using dynamic light scattering and transmission electron microscopy, this pioneering formulation exhibited efficacy against L. donovani Ag83 promastigotes as demonstrated by MTT cell viability testing. Evaluating internal reactive oxygen species (ROS) levels and dual staining with acridine orange and ethidium bromide unveiled its consequential impact on cell death. Significantly, our study discloses this novel nanoformulation's unprecedented inhibition of the trypanothione reductase enzyme. The findings posit the citrate-coated gold Amphotericin B nanoformulation as a promising and targeted antileishmanial agent, representing potential advancements in leishmaniasis therapeutics.

Synergistic Effects of Artesunate in Combination with Amphotericin B and Miltefosine against Leishmania infantum: Potential for Dose Reduction and Enhanced Therapeutic Strategies.

Leishmaniasis is a tropical infectious disease caused by Leishmania parasites. The disease can be spread by the bite of an infected sand fly. Currently, five chemotherapeutic drugs are available in leishmaniasis treatment. However, these drugs exhibit toxicity and serious adverse effects on infected individuals, necessitating alternative treatment strategies. One such strategy involves using combinations of existing antileishmanial drugs. In this study, we evaluated the interaction between artesunate (AS) and three antileishmanial drugs-amphotericin B (AmB), miltefosine (MF), and paromomycin (PM) against Leishmania infantum. This evaluation marks the first time such an assessment has been conducted. The Chou-Talalay combination index method was employed to analyze the drug interaction. The findings revealed that the interaction between AS and AmB ranged from antagonistic to synergistic, while the interaction between AS and MF showed moderate to strong synergism. In contrast, the interaction between AS and PM resulted in an antagonistic interaction, which differs from the combinations with AmB or MF. This study provides valuable insights for developing novel drug regimens for leishmaniasis treatment, emphasizing the potential of AS and its combination with existing antileishmanial drugs. Further research is necessary to optimize drug combinations and minimize adverse effects, leading to more effective therapeutic outcomes.

Visceral Leishmaniasis with Renal Involvement: A Case Report

Visceral leishmaniasis (VL) is widespread in tropical and temperate region. It is caused by the protozoan parasite leishmania species and is transmitted through the bite of infected female sandflies. It manifests as pancytopenia, hepatosplenomegaly and fever. Here, we present a case of 20-year-old male presented with difficulty in swallowing for 2 months, abdominal distention for 30 days and shortness of breath for 20 days along with progressive renal impairment. Clinical examination revealed hepatosplenomegaly and laboratory investigations confirmed pancytopenia, nephrotic range proteinuria, hematuria and hypocomplementemia. Bone marrow aspiration and biopsy was performed which showed leishmania donovani (LD) bodies. It’suncommon presentation makes it challenging for physicians as well as pathologist to make a timely diagnosis.

Risk factors of visceral leishmaniasis in the world: a review

Visceral leishmaniasis is a zoonotic parasitic disease caused by viscerotropic Leishmania species and transmitted by bites of infected phlebotomine sandflies, which is predominantly prevalent in the Indian subcontinent, eastern Africa and South America. Currently, visceral leishmaniasis is the second most fatal parasitic disease in the world. Because of climate changes, urban development and individual conditions, there are changes in the density of visceral leishmaniasis vector sandflies and the likelihood of contact with humans, resulting in a visceral leishmaniasis transmission risk. The review summarizes natural, social and biological factors affecting the transmission of visceral leishmaniasis, so as to provide insights into formulation of targeted control measures for visceral leishmaniasis.

Clinical manifestations and diagnostic approaches in cases of canine leishmaniasis in Bulgaria.

Leishmaniasis, a parasitic disease found in many parts of southern Europe, is transmitted in both humans and canines through the bite of phlebotomine sandflies, and can present in a variety of ways, such as cutaneous, mucocutaneous, diffuse, and visceral. In Bulgaria there are endemic areas of canine leishmaniasis, with sporadic cases in humans. However, no detailed studies of the animal population and vectors have been performed. Here we describe a few clinical cases of canine visceral leishmaniasis in two districts in western Bulgaria: one endemic and one without previously detected cases in humans or dogs. Diagnosis was confirmed serologically and molecularly using both real time and conventional PCR. Specific anti-leishmanial antibodies were confirmed in three of the cases via ELISA, with 50% of them returning extremely high values. In the majority of the cases DNA fragments were detected in the skin or lymph node aspirate but not in the blood. This paper highlights the need for further studies updating the current knowledge on the epidemiology, diagnosis, and control of visceral leishmaniasis in the reservoir host population.

Serosurvey of canine leishmaniasis in five departments near an identified human clinical case in Marseille (France)

Leishmania infantum is a protozoan parasite of the family Trypanosomatidae, transmitted by the bite of phlebotomine sand flies (Diptera: Psychodidae). It is responsible for human and canine leishmaniasis in countries bordering the Mediterranean basin. Here we describe a clinical case of human cutaneous leishmaniasis in a 76-year-old female patient living in Marseille. Upon interrogation, the patient had no history of recent travel or contact with animals. The study involved clinical, serological, and molecular investigation of the occurrence of Leishmania infection in 718 dogs from five departments within a 130 km radius perimeter around Marseille. Five dogs showed signs of moderate leishmaniasis. Additionally, the serological survey of dogs revealed a global seroprevalence of 5.1%, with a significantly different prevalence in the Vaucluse department. Molecular analysis and phylogenetic studies highlighted the close relatedness of Leishmania strains between human and canine hosts with 99.6% of identity, indicating local transmission. The findings underscore the importance of serological surveillance in dogs and human. In a One Health approach, it is necessary to continue investigating Leishmania infection in all potential reservoirs, especially for zoonotic L. infantum in wildlife (red foxes, leporids, rodents, etc.) but also in dogs, cats, and equids.

Overview and Approaches for Handling of Animal Models of Leishmaniasis.

Leishmaniasis, a disease of global relevance, results from infection with the protozoan parasite, Leishmania, which is transmitted to susceptible hosts through the bite of sand flies. Multiple forms of leishmaniasis may occur, including cutaneous, mucocutaneous, and visceral. Research with animal models remains an important approach to help define basic pathophysi- ologic processes associated with infection and disease. In this regard, mice and hamsters represent the most commonly used models. The severity of leishmaniasis in animal models depends on several factors, including genotype of the host and parasite and the dose and route of administration of the parasite to the host, and severity of outcome may range from subclinical to severe illness. This review provides basic background on leishmaniasis, relevant animal models, the pathophysiology and clinical signs in animals used as models of leishmaniasis, and general approaches to mitigate risk to personnel.

Leishmania-sand fly interactions: exploring the role of the immune response and potential strategies for Leishmaniasis control.

Leishmaniasis is a neglected tropical disease caused by protozoan parasites of the genus Leishmania, affecting millions of people worldwide. The disease is transmitted by the bite of infected female sand flies, which act as vectors and hosts for the parasites. The interaction between Leishmania parasites and sand flies is complex and dynamic, involving various factors that influence parasite development, survival and transmission. This review examines how the immune response of sand flies affects vector competence and transmission of Leishmania parasites, and what the potential strategies are to prevent or reduce infection. The review also summarizes the main findings and conclusions of the existing literature and discusses implications and recommendations for future research and practice. The study reveals that the immune response of sand flies is a key determinant of vector competence and transmission of Leishmania parasites, and that several molecular and cellular mechanisms are involved in the interaction between parasite and vector. The study also suggests that there are potential strategies for controlling leishmaniasis, such as interfering with parasite development, modulating the vector's immune response or reducing the vector population. However, the study also identifies several gaps and limitations in current knowledge and calls for more comprehensive and systematic studies on vector-parasite interaction and its impact on leishmaniasis transmission and control.

Community Conversations: A Tool in Improving Health Outcomes and Reducing the Burden of Visceral Leishmaniasis (Kala-azar) in Isiolo County, Kenya

Visceral Leishmaniasis (Kala-azar) is a neglected tropical disease (NTD) caused by Leishmania species that affects human. Transmission is through the bite of Phlebotomus sandflies. Globally, WHO ranks Kala-azar as the second largest parasitic killer after Malaria. The disease poses a threat to more than a billion people on almost every continent. Approximately 90% of Kala-azar global burden is found in Africa. Major endemicity is documented in Eastern Africa (Kenya, South Sudan, Sudan, Ethiopia, and Somalia). Accurate burden of the disease in Kenya is unknown attributable to inadequate information on disease prevalence, diagnostic challenges, and spatial distribution. Additionally, morbidity and mortality levels from the disease in the Country are unknown ascribable to a low index of suspicion by healthcare workers, diagnostic challenges, and case management. An example of diagnostic challenges is a reported outbreak of ‘unknown disease’ akin to Kala-azar in Marsabit County in May 2023 where nine people were confirmed dead and over 80 hospitalized. This study aimed to analyze community’s prioritization of causes of Kalaazar as well as community awareness of disease prevention and control. Quantitative data were collected using Participatory Epidemiology methods in purposively selected study villages. Qualitative data were collected through semi-structured interviews. Study findings indicated that 97.2% of respondents had heard about Kala-azar. Despite the awareness, a broad gap between knowledge and practice of prevention and control strategies of the disease was evident. Kala-azar was associated with human behaviour like preferences for traditional and alternative treatment approaches (45.2%), evening outdoor practices before going to bed (99.6%), sleeping outdoors (60.1%), occasional bed nets use (69.2%), and availability of domestic animals and wild animals within house yards (54.6%). Cultural beliefs and practices were noted to play a key role in augmenting the disease burden, where most cases had burn scars from traditional healers’ therapeutic burns. As a consequence, they sought medical help after onset of complications, which ranged from increased severity to fatalities. Community engagement was minimal in prevention and control. In this participatory research, we propose behaviour change communication activities for the elimination of Kala-azar as a public health problem. This article serves as a base for future studies aimed at giving voice to communities while enhancing their understanding of causes of diseases, the possibility of enhanced early diagnostic techniques, and options for prevention, control, and surveillance which in turn is translated to action.

Demographic characteristics and prevalence of asymptomatic Leishmania donovani infection in migrant workers working in an endemic area in Northwest Ethiopia.

Visceral leishmaniasis (VL), a neglected tropical disease that causes substantial morbidity and mortality, is a serious health problem in Ethiopia. Infections are caused by Leishmania (L.) donovani parasites. Most individuals remain asymptomatic, but some develop VL, which is generally fatal if not treated. We identified the area of Metema-Humera in Northwest Ethiopia as a setting in which we could follow migrant workers when they arrived in an endemic area. The demographic characteristics of this population and factors associated with their risk of asymptomatic infection are poorly characterised. We divided our cohort into individuals who visited this area for the first time (first comers, FC) and those who had already been in this area (repeat comers, RC). We followed them from the beginning (Time 1, T1) to the end of the agricultural season (Time 2, T2), performing tests for sand fly bite exposure (anti-sand fly saliva antibody ELISA) and serology for Leishmania infection (rK39 rapid diagnostic test and the direct agglutination test) at each time point and collecting information on risk factors for infection. Our results show that most migrant workers come from non-endemic areas, are male, young (median age of 20 years) and are farmers or students. At T1, >80% of them had been already exposed to sand fly bites, as shown by the presence of anti-saliva antibodies. However, due to seasonality of sand flies there was no difference in exposure between FC and RC, or between T1 and T2. The serology data showed that at T1, but not at T2, a significantly higher proportion of RC were asymptomatic. Furthermore, 28.6% of FC became asymptomatic between T1 and T2. Over the duration of this study, one FC and one RC developed VL. In multivariable logistic regression of asymptomatic infection at T1, only age and the number of visits to Metema/Humera were significantly associated with asymptomatic infection. A better understanding of the dynamics of parasite transmission and the risk factors associated with the development of asymptomatic infections and potentially VL will be essential for the development of new strategies to prevent leishmaniasis.

Neutrophil extracellular traps formation: effect of Leishmania major promastigotes and salivary gland homogenates of Phlebotomus papatasi in human neutrophil culture.

Leishmaniasis as a neglected tropical disease (NTD) is caused by the inoculation of Leishmania parasites via the bite of phlebotomine sand flies. After an infected bite, a series of innate and adaptive immune responses occurs, among which neutrophils can be mentioned as the initiators. Among the multiple functions of these fighting cells, neutrophil extracellular traps (NETs) were studied in the presence of Leishmania major promastigotes and salivary gland homogenates (SGH) of Phlebotomus papatasi alone, and in combination to mimic natural conditions of transmission. The effect of L. major and SGH on NETs formation was studied in three different groups: neutrophils + SGH (NS), neutrophils + L. major (NL), neutrophils + L. major + SGH (NLS) along with negative and positive controls in 2, 4 and 6h post-incubation. Different microscopic methods were used to visualize NETs comprising: fluorescence microscopy by Acridine Orange/ Ethidium Bromide staining, optical microscopy by Giemsa staining and scanning electron microscopy. In addition, the expression level of three different genes NE, MPO and MMP9 was evaluated by Real-Time PCR. All three microscopical methods revealed similar results, as in NS group, chromatin extrusion as a sign of NETosis, was not very evident in each three time points; but, in NL and especially NLS group, more NETosis was observed and the interaction between neutrophils and promastigotes in NL and also with saliva in NLS group, gradually increased over times. Real-time reveals that, the expression of MPO, NE and MMP9 genes increased during 2 and 4h after exposure, and then decreased at 6h in most groups. Hence, it was determined that the simultaneous presence of parasite and saliva in NLS group has a greater impact on the formation of NETs compared to NL and NS groups.