Phlebotomus perniciosus is a major vector of Leishmania infantum in the Mediterranean. While the seroprevalence of leishmaniosis in Spanish dogs and cats has been studied, data on the exposure of cats to P. perniciosus bites under natural conditions without repellents is limited. Stray cats could serve as sentinels for L. infantum and P. perniciosus exposure. This study analyzed sera from 204 apparently healthy stray cats, collected from January 2021 to January 2022, for antibodies against P. perniciosus saliva and L. infantum parasites. Anti-sand fly antibodies were detected in 40.69% of cats using an ELISA with the recombinant salivary protein SP03B of P. perniciosus. Seroprevalence of L. infantum infection was 23.52% by Western blot and 27.41% by ELISA, with an overall seroprevalence of 40.69% (95% CI 34.18-47.54%). This is the first assessment of antibody response to P. perniciosus saliva and L. infantum in naturally exposed stray cats in Spain. Further research is needed to examine the salivary antigens recognized by cats and to explore the relationship between P. perniciosus exposure and L. infantum infection severity in cats.

• Klíčová slova
• Cat, ELISA, Leishmania infantum, Phlebotomus perniciosus, serology, western blotting,
• MeSH
• ELISA veterinární   MeSH
• hmyz - vektory parazitologie   MeSH
• kočky MeSH
• Leishmania infantum * imunologie   MeSH
• leishmanióza viscerální * veterinární   epidemiologie   imunologie   MeSH
• nemoci koček * epidemiologie   parazitologie   imunologie   MeSH
• Phlebotomus * parazitologie   imunologie   MeSH
• protilátky protozoální * krev   MeSH
• séroepidemiologické studie MeSH
• zvířata MeSH
• Check Tag
• kočky MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Španělsko epidemiologie   MeSH
• Názvy látek
• protilátky protozoální * MeSH

Equids may be infected by zoonotic Leishmania spp., including Leishmania infantum, in regions where canine leishmaniasis (CanL) is endemic, and Leishmania martiniquensis, which has been reported in horses from Central Europe. This study was designed to evaluate the occurrence of both Leishmania spp. among equids living in CanL endemic areas of Italy, as well as to identify dipteran vectors from the same habitats. From March to October 2023, blood, serum and tissue samples from skin lesions were collected from equids (n = 98; n = 56 donkeys and n = 42 horses) living in Italy, as well as sand flies and biting midges. Blood samples (n = 98) and skin lesions (n = 56) were tested for Leishmania spp. by conventional and real time PCRs and sera were tested by immunofluorescence antibody tests (IFAT) for both L. infantum and L. martiniquensis. Insects were morphologically identified, and female specimens (n = 268 sand flies, n = 7 biting midges) analyzed for Leishmania DNA, as well as engorged sand flies (n = 16) for blood-meal detection. Two animals with skin lesions (i.e., one donkey and one horse) scored positive for Leishmania spp. DNA, and 19 animals (i.e., 19.4%; n = 13 donkeys and n = 6 horses) were seropositive for L. infantum, with five of them also for L. martiniquensis. Most seropositive animals had no dermatological lesions (i.e., 68.4%) while both animals molecularly positive for Leishmania spp. scored seronegative. Of the 356 sand flies collected, 12 females (i.e., n = 8 Sergentomyia minuta; n = 3 Phlebotomus perniciosus, n = 1 Phlebotomus perfiliewi) were positive for Leishmania spp. DNA, and one out of seven biting midges collected was DNA-positive for L. infantum. Moreover, engorged sand flies scored positive for human and equine DNA. Data suggest that equids living in CanL endemic areas are exposed to Leishmania spp., but their role in the circulation of the parasite needs further investigations.

• MeSH
• Ceratopogonidae parazitologie   MeSH
• endemické nemoci veterinární   MeSH
• Equidae * parazitologie   MeSH
• hmyz - vektory * parazitologie   MeSH
• koně parazitologie   MeSH
• Leishmania infantum izolace a purifikace   genetika   MeSH
• Leishmania * izolace a purifikace   genetika   klasifikace   MeSH
• leishmanióza * veterinární   epidemiologie   parazitologie   přenos   MeSH
• nemoci koní parazitologie   epidemiologie   MeSH
• nemoci psů * parazitologie   epidemiologie   přenos   MeSH
• psi MeSH
• Psychodidae parazitologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• psi MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Itálie epidemiologie   MeSH

INTRODUCTION: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

• Klíčová slova
• Leishmania, PCR, asymptomatic, direct agglutination test, rK39, risk factors, visceral leishmaniasis,
• Publikační typ
• časopisecké články MeSH

Antibodies against Phlebotomus perniciosus sandfly salivary gland homogenate (SGH) and recombinant protein rSP03B, sandfly-borne Toscana virus (TOSV), Sandfly Fever Sicilian virus (SFSV) and Leishmania, as well as DNA of the latter parasite, were investigated in 670 blood samples from 575 human donors in Murcia Region, southeast Spain, in 2017 and 2018. The estimated SGH and rSP03B seroprevalences were 69% and 88%, respectively, although correlation between test results was relatively low (ρ = 0.39). Similarly, TOSV, SFSV and Leishmania seroprevalences were 26%, 0% and 1%, respectively, and Leishmania PCR prevalence was 2%. Prevalences were significantly greater in 2017, overdispersed and not spatially related to each other although both were positively associated with SGH but not to rSP03B antibody optical densities, questioning the value of the latter as a diagnostic marker for these infections in humans.

• Klíčová slova
• Leishmania infantum, anti-saliva antibodies, blood donors, sandflies, sandfly fever sicilian virus, toscana virus,
• MeSH
• dárci krve MeSH
• Leishmania infantum * MeSH
• leishmanióza * parazitologie   veterinární   MeSH
• lidé MeSH
• Phlebotomus * parazitologie   MeSH
• protilátky MeSH
• Psychodidae * MeSH
• rekombinantní proteiny MeSH
• virus horečky pappataci * genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Španělsko epidemiologie   MeSH
• Názvy látek
• protilátky MeSH
• rekombinantní proteiny MeSH

BACKGROUND: In onchocerciasis endemic areas in Africa, heterogenous biting rates by blackfly vectors on humans are assumed to partially explain age- and sex-dependent infection patterns with Onchocerca volvulus. To underpin these assumptions and further improve predictions made by onchocerciasis transmission models, demographic patterns in antibody responses to salivary antigens of Simulium damnosum s.l. are evaluated as a measure of blackfly exposure. METHODOLOGY/PRINCIPAL FINDINGS: Recently developed IgG and IgM anti-saliva immunoassays for S. damnosum s.l. were applied to blood samples collected from residents in four onchocerciasis endemic villages in Ghana. Demographic patterns in antibody levels according to village, sex and age were explored by fitting generalized linear models. Antibody levels varied between villages but showed consistent patterns with age and sex. Both IgG and IgM responses declined with increasing age. IgG responses were generally lower in males than in females and exhibited a steeper decline in adult males than in adult females. No sex-specific difference was observed in IgM responses. CONCLUSIONS/SIGNIFICANCE: The decline in age-specific antibody patterns suggested development of immunotolerance or desensitization to blackfly saliva antigen in response to persistent exposure. The variation between sexes, and between adults and youngsters may reflect differences in behaviour influencing cumulative exposure. These measures of antibody acquisition and decay could be incorporated into onchocerciasis transmission models towards informing onchocerciasis control, elimination, and surveillance.

• MeSH
• dítě MeSH
• dospělí MeSH
• hmyz - vektory imunologie   parazitologie   MeSH
• imunoglobulin G krev   MeSH
• imunoglobulin M krev   MeSH
• kousnutí a bodnutí hmyzem epidemiologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• Onchocerca volvulus růst a vývoj   MeSH
• onchocerkóza epidemiologie   přenos   MeSH
• předškolní dítě MeSH
• protilátky krev   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Simuliidae imunologie   parazitologie   MeSH
• sliny imunologie   MeSH
• zvířata MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• imunoglobulin G MeSH
• imunoglobulin M MeSH
• protilátky MeSH

Phlebotomus argentipes is a predominant vector of Leishmania donovani, the protozoan parasite causing visceral leishmaniasis in the Indian subcontinent. In hosts bitten by P. argentipes, sand fly saliva elicits the production of specific anti-salivary protein antibodies. Here, we have utilised these antibodies as markers of human exposure to P. argentipes in a visceral leishmaniasis endemic area in Pabna district, Bangladesh. The use of whole salivary gland homogenate as an antigen to detect these antibodies has several limitations, therefore it is being superseded by the use of specific recombinant salivary proteins. We have identified three major P. argentipes salivary antigenic proteins recognised by sera of bitten humans, expressed them in a recombinant form (rPagSP04, rPagSP05 and rPagSP06) and tested their applicability in ELISA and immunoblot. One of them, PpSP32-like protein rPagSP06, was identified as the most promising antigen, showing highest resemblance and correlation with the IgG response to P. argentipes salivary gland homogenate. Furthermore, we have validated the applicability of rPagSP06 in a large cohort of 585 individuals and obtained a high correlation coefficient for anti-rPagSP06 and anti-P. argentipes saliva IgG responses. The anti-rPagSP06 and anti-P. argentipes salivary gland homogenate IgG responses followed a similar right-skewed distribution. This is the first report of screening human sera for anti-P. argentipes saliva antibodies using recombinant salivary protein. The rPagSP06 was proven to be a valid antigen for screening human sera for exposure to P. argentipes bites in a visceral leishmaniasis endemic area.

• Klíčová slova
• Bangladesh, IgG antibodies, Leishmania donovani, Marker of exposure, Phlebotomus argentipes, Salivary glands,
• MeSH
• hmyzí proteiny * imunologie   MeSH
• kousnutí a bodnutí epidemiologie   MeSH
• Leishmania donovani MeSH
• lidé MeSH
• Phlebotomus * MeSH
• slinné proteiny a peptidy * imunologie   MeSH
• sliny MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Bangladéš epidemiologie   MeSH
• Názvy látek
• hmyzí proteiny * MeSH
• slinné proteiny a peptidy * MeSH

BACKGROUND: Simulium damnosum sensu lato (s.l.) blackflies transmit Onchocerca volvulus, a filarial nematode that causes human onchocerciasis. Human landing catches (HLCs) is currently the sole method used to estimate blackfly biting rates but is labour-intensive and questionable on ethical grounds. A potential alternative is to measure host antibodies to vector saliva deposited during bloodfeeding. In this study, immunoassays to quantify human antibody responses to S. damnosum s.l. saliva were developed, and the salivary proteome of S. damnosum s.l. was investigated. METHODOLOGY/PRINCIPAL FINDINGS: Blood samples from people living in onchocerciasis-endemic areas in Ghana were collected during the wet season; samples from people living in Accra, a blackfly-free area, were considered negative controls and compared to samples from blackfly-free locations in Sudan. Blackflies were collected by HLCs and dissected to extract their salivary glands. An ELISA measuring anti-S. damnosum s.l. salivary IgG and IgM was optimized and used to quantify the humoral immune response of 958 individuals. Both immunoassays differentiated negative controls from endemic participants. Salivary proteins were separated by gel-electrophoresis, and antigenic proteins visualized by immunoblot. Liquid chromatography mass spectrometry (LC-MS/MS) was performed to characterize the proteome of S. damnosum s.l. salivary glands. Several antigenic proteins were recognized, with the major ones located around 15 and 40 kDa. LC-MS/MS identified the presence of antigen 5-related protein, apyrase/nucleotidase, and hyaluronidase. CONCLUSIONS/SIGNIFICANCE: This study validated for the first time human immunoassays that quantify humoral immune responses as potential markers of exposure to blackfly bites. These assays have the potential to facilitate understanding patterns of exposure as well as evaluating the impact of vector control on biting rates. Future studies need to investigate seasonal fluctuations of these antibody responses, potential cross-reactions with other bloodsucking arthropods, and thoroughly identify the most immunogenic proteins.

• MeSH
• dítě MeSH
• dospělí MeSH
• ELISA MeSH
• hmyz - vektory fyziologie   MeSH
• imunoglobulin G krev   MeSH
• imunoglobulin M krev   MeSH
• kousnutí a bodnutí hmyzem epidemiologie   imunologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• onchocerkóza MeSH
• předškolní dítě MeSH
• senioři MeSH
• Simuliidae fyziologie   MeSH
• sliny * MeSH
• zvířata MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• senioři MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Ghana MeSH
• Súdán MeSH
• Názvy látek
• imunoglobulin G MeSH
• imunoglobulin M MeSH

Phlebotomus perniciosus (Diptera: Phlebotominae) is a medically and veterinary important insect vector. It transmits the unicellular parasite Leishmania infantum that multiplies intracellularly in macrophages causing life-threatening visceral diseases. Leishmania establishment in the vertebrate host is substantially influenced by immunomodulatory properties of vector saliva that are obligatorily co-injected into the feeding site. The repertoire of P. perniciosus salivary molecules has already been revealed and, subsequently, several salivary proteins have been expressed. However, their immunogenic properties have never been studied. In our study, we tested three P. perniciosus recombinant salivary proteins-an apyrase rSP01 and yellow-related proteins rSP03 and rSP03B-and showed their anti-inflammatory nature on the murine bone-marrow derived macrophages. Even in the presence of pro-inflammatory stimuli (IFN-γ and bacterial lipopolysaccharide, LPS), all three recombinant proteins inhibited nitric oxide production. Moreover, rSP03 seems to have a very strong anti-inflammatory effect since it enhanced arginase activity, increased the production of IL-10, and inhibited the production of TNF-α even in macrophages stimulated with IFN-γ and LPS. These results suggest that P. perniciosus apyrase and yellow-related proteins may serve as enhancing factors in sand fly saliva, facilitating the development of Leishmania infection along with their anti-haemostatic properties. Additionally, rSP03 and rSP03B did not elicit the delayed-type hypersensitivity response in mice pre-exposed to P. perniciosus bites (measured as visible skin reaction). The results of our study may help to understand the potential function of recombinant's native counterparts and their role in Leishmania transmission and establishment within the host.

• Klíčová slova
• Phlebotomus, apyrase, immunogenicity, macrophage polarization, sand fly saliva, yellow-related proteins,
• MeSH
• antiflogistika MeSH
• fenotyp MeSH
• makrofágy MeSH
• myši MeSH
• Phlebotomus * MeSH
• psi MeSH
• rekombinantní proteiny MeSH
• slinné proteiny a peptidy MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• psi MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antiflogistika MeSH
• rekombinantní proteiny MeSH
• slinné proteiny a peptidy MeSH

BACKGROUND: In endemic areas of zoonotic leishmaniosis caused by L. infantum, early detection of Leishmania infection in dogs is essential to control the dissemination of the parasite to humans. The aim of this study was to evaluate the serological and/or molecular diagnostic performance of minimally and non-invasive samples (conjunctiva cells (CS) and peripheral blood (PB)) for monitoring Leishmania infection/exposure to Phlebotomus perniciosus salivary antigens in dogs at the beginning and the end of sand fly seasonal activity (May and October, respectively) and to assess associated risks factors. METHODS: A total of 208 sheltered dogs from endemic areas of leishmaniosis were screened. Leishmania DNA detection in PB on filter paper and CS was performed by nested-PCR (nPCR), while the detection of anti-Leishmania antibodies was performed using IFAT and ELISA. The exposure to P. perniciosus salivary antigens (SGH, rSP01 and rSP03B + rSP01) was measured by ELISA. RESULTS: Ninety-seven (46.6%) and 116 (55.8%) of the 208 dogs were positive to Leishmania antibodies or DNA by at least one test at the beginning and end of the sand fly season, respectively. IFAT and ELISA presented a substantial agreement in the serodiagnosis of leishmaniosis. Discrepant PB nPCR results were obtained between sampling points. Leishmania DNA was detected in CS of 72 dogs at the end of the phlebotomine season. The presence of antibodies to the parasite measured by ELISA was significantly higher in dogs presenting clinical signs compatible with leishmaniosis at both sampling points. Phlebotomus perniciosus salivary antibodies were detected in 179 (86.1%) and 198 (95.2%) of the screened dogs at the beginning and end of the phlebotomine season, respectively. CONCLUSIONS: The association between ELISA positivity and clinical signs suggests its usefulness to confirm a clinical suspicion. CS nPCR seems to be an effective and non-invasive method for assessing early exposure to the parasite. PB nPCR should not be used as the sole diagnostic tool to monitor Leishmania infection. The correlation between the levels of antibodies to P. perniciosus saliva and Leishmania antibodies suggests the use of a humoral response to sand fly salivary antigens as biomarkers of L. infantum infection.

• Klíčová slova
• Blood, Conjunctival cells, Dog, Exposure, L. infantum, Phlebotomus pernicious, Saliva,
• MeSH
• antigeny protozoální imunologie   MeSH
• endemické nemoci prevence a kontrola   MeSH
• hmyz - vektory parazitologie   MeSH
• hmyzí proteiny imunologie   MeSH
• imunoglobulin G krev   MeSH
• konjunktiva cytologie   parazitologie   MeSH
• kousnutí a bodnutí hmyzem MeSH
• Leishmania infantum izolace a purifikace   MeSH
• leishmanióza krev   imunologie   veterinární   MeSH
• nemoci psů parazitologie   prevence a kontrola   přenos   MeSH
• Phlebotomus parazitologie   MeSH
• protilátky protozoální krev   MeSH
• protozoální proteiny imunologie   MeSH
• psi MeSH
• rizikové faktory MeSH
• sérologické testy MeSH
• slinné proteiny a peptidy imunologie   MeSH
• zvířata MeSH
• Check Tag
• psi MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigeny protozoální MeSH
• hmyzí proteiny MeSH
• imunoglobulin G MeSH
• protilátky protozoální MeSH
• protozoální proteiny MeSH
• slinné proteiny a peptidy MeSH

BACKGROUND: Canine leishmaniosis caused by Leishmania infantum is a neglected zoonosis transmitted by sand flies like Phlebotomus perniciosus. Clinical signs and disease susceptibility vary according to various factors, including host immune response and breed. In particular, Ibizan hounds appear more resistant. This immunocompetence could be attributed to a more frequent exposure to uninfected sand flies, eliciting a stronger anti-sand fly saliva antibody response. METHODS: This study aimed to investigate the prevalence of anti-P. perniciosus saliva antibodies in Ibizan hounds and dogs of other breeds in the Leishmania-endemic area of Mallorca, Spain, and to correlate these antibody levels with clinical, immunological and parasitological parameters. Anti-sand fly saliva IgG was examined in 47 Ibizan hounds and 45 dogs of other breeds using three methods: P. perniciosus whole salivary gland homogenate (SGH) ELISA; recombinant protein rSP03B ELISA; and rSP03B rapid tests (RT). Additionally, diagnostic performance was evaluated between methods. RESULTS: Results indicate significantly higher anti-SGH antibodies (P = 0.0061) and a trend for more positive SGH ELISA and RT results in Ibizan hounds compared to other breeds. General linear model analysis also found breed to be a significant factor in SGH ELISA units and a marginally significant factor in RT result. Although infection rates were similar between groups, Ibizan hounds included significantly more IFN-γ producers (P = 0.0122) and papular dermatitis cases (P < 0.0001). Older age and L. infantum seropositivity were also considered significant factors in sand fly saliva antibody levels according to at least one test. Fair agreement was found between all three tests, with the highest value between SGH and rSP03B RT. CONCLUSIONS: To our knowledge, this is the first study elaborating the relationship between anti-P. perniciosus saliva antibodies and extensive clinical data in dogs in an endemic area. Our results suggest that Ibizan hounds experience a higher frequency of exposure to sand flies and have a stronger cellular immune response to L. infantum infection than other breed dogs. Additional sampling is needed to confirm results, but anti-P. perniciosus saliva antibodies appear to negatively correlate with susceptibility to L. infantum infection and could possibly contribute to the resistance observed in Ibizan hounds.

• Klíčová slova
• Anti-sand fly saliva antibodies, Canine leishmaniosis, Ibizan hounds, Leishmania infantum, Papular dermatitis, Phlebotomus perniciosus, rSP03B,
• MeSH
• chov MeSH
• endemické nemoci MeSH
• hmyzí proteiny imunologie   MeSH
• imunoglobulin G imunologie   MeSH
• leishmanióza imunologie   veterinární   MeSH
• náchylnost k nemoci MeSH
• nemoci psů imunologie   parazitologie   MeSH
• Phlebotomus imunologie   MeSH
• psi MeSH
• slinné proteiny a peptidy imunologie   MeSH
• sliny imunologie   MeSH
• zoonózy parazitologie   přenos   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• psi MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Španělsko MeSH
• Názvy látek
• hmyzí proteiny MeSH
• imunoglobulin G MeSH
• slinné proteiny a peptidy MeSH