Chemical Composition and Acaricidal Activity of Essential Oil of Lavandula dentata L. on Engorged Females of Rhipicephalus (Boophilus) microplus (Acari: Ixodidae).

PURPOSE: Rhipicephalus (Boophilus) microplus is one the most significant ectoparasite in cattle farming in tropical and subtropical regions, causing problems to livestock health worldwide. The control of this ectoparasite primarily relies on the use of synthetic acaricides. However, the emergence of acaricide resistance has stimulated the search for new control alternatives, including phytocompounds with acaricidal and insecticidal potential. The aim of this study was to evaluate the acaricidal potential of Lavandula dentata essential oil against the engorged females of R. (B.) microplus. METHODS: Engorged females were obtained from infested bovines in dairy farms in Pernambuco, Brazil. L. dentata essential oil was extracted, and adult immersion test assays were performed using the following oil concentrations: 0.2%, 0.4%, 0.6%, 0.8%, and 1%. RESULTS: L. dentata essential oil at a concentration of 1% was lethal to all engorged females, and concentrations of 0.6% and 0.8% caused mortality of 98.6% and 99.1%, respectively. These concentrations disrupted the reproductive capacity of engorged females, reducing oviposition by more than 90% and preventing egg hatching by over 87%. CONCLUSION: The data revealed that L. dentata essential oil possesses effective pharmacological properties against R. (B.) microplus and could be used for tick control following in vivo evaluation, thus contributing to mitigating the negative impacts of synthetic acaricide use.

Filarial Nematodes in Dogs from the Northeast Region of Brazil.

PURPOSE: Medical and veterinary filarial nematodes are transmitted by blood-feeding vectors. In dogs, these parasites are mainly represented by nematodes in which microfilariae dwell in the blood (Dirofilaria spp. and Acanthocheilonema spp.) or skin (Cercopithifilaria spp. and Onchocerca lupi). The aim of this study was to determine the prevalence of these filarial infections in dogs residing in a touristic, heavily populated location in the northeastern region of Brazil. METHODS: Blood samples (n = 245) were assessed by a modified Knott test, followed by a qualitative ELISA test (SNAP® 4Dx® Plus, IDEXX Laboratory, Westbrook, Maine, USA) for the detection of antibodies against Anaplasma spp., Borrelia burgdorferi sensu lato, Ehrlichia spp. and antigens of Dirofilaria immitis. Skin samples (n = 71) were microscopically examined and molecularly assessed through a PCR targeting the 12 S rRNA gene. RESULTS: Microfilariae and antigen of D. immitis were detected simultaneously in 15 (6.1%; 95% CI = 3.7-9.8) animals. Nine animals (3.6%; 95% CI = 1.9-6.8) were D. immitis antigen positive but microfilariae negative and nine other animals (3.6%; 95% CI = 1.9-6.8) were microfilariae positive but D. immitis antigen negative. D. immitis positive dogs were found in four different municipalities. No filarioids were detected in the skin after microscopical and molecular analyses. CONCLUSION: Data from this study demonstrate that D. immitis is the main filarial nematode infecting dogs in coastal areas in northeastern Brazil. Based on the potential risk of infection in which animals are submitted, it is essential to perform tests to detect microfilariae and D. immitis antigen. Preventive measures must be adopted by using microfilaricidal compounds and anti-feeding insecticides to prevent canine infection.

Leishmaniases in a tropical area of Brazil: a spatio-temporal analysis.

Leishmaniases are a group of tropical and neglected diseases caused by the protozoa of the genus Leishmania which are transmitted by insect sandflies. Despite of the major efforts undertaken at global level for the control of these diseases, a recent spreading to non-endemic areas been observed. In this study we analyzed the spatio-temporal distribution of Visceral Leishmaniasis (VL) and Tegumentary Leishmaniasis (TL) cases notified in a tropical region of Brazil. A retrospective evaluation was carried out using secondary data (e.g., demographics, origin, age and area of occurrence) of human patients obtained from 2009 to 2019. Choropleth maps with levels of color intensity represented the spatial distribution of VL and TL cases. Overall, 116 (10.5 ± 8.5) cases of leishmaniases were registered in the study area during the whole period. Of those, 61 (5.5 ± 5.5) and 55 (5 ± 4.3) were diagnosed as VL and TL, respectively. VL cases were recorded in 61.9% (13/21) and TL cases in 42.8% (9/21) of the municipalities. Most of the cases were predominantly notified in men from rural areas, and specifically for TL with an age range of 30-59 years old. These findings will be useful to drive strategies of control (e.g., vector control, dog surveillance and notification of suspected cases) against these neglect diseases, preventing the spreading to non-endemic areas.

The Role of Parasitoid Wasps, Ixodiphagus spp. (Hymenoptera: Encyrtidae), in Tick Control.

Species of Ixodiphagus (Hymenoptera: Encyrtidae) are parasitoid wasps whose immature forms develop inside ixodid and argasid ticks (Acari: Ixodida). Following oviposition by adult female wasps into the idiosoma of ticks, larvae hatch and start feeding on their internal contents, eventually emerging as adult wasps from the body of the dead ticks. Species of Ixodiphagus have been reported as parasitoids of 21 species of ticks distributed across 7 genera. There are at least ten species described in the genus, with Ixodiphagus hookeri being the most studied as an agent for biological control of ticks. Although attempts of tick control by means of this parasitoid largely failed, in a small-scale study 150,000 specimens of I. hookeri were released over a 1-year period in a pasture where a small cattle population was kept, resulting in an overall reduction in the number of Amblyomma variegatum ticks per animal. This review discusses current scientific information about Ixodiphagus spp., focusing on the role of this parasitoid in the control of ticks. The interactions between these wasps and the ticks' population are also discussed, focusing on the many biological and logistical challenges, with limitations of this control method for reducing tick populations under natural conditions.

The application of the FLOTAC technique for detection of helminth eggs of medical and veterinary importance in soil samples.

The FLOTAC technique was initially developed to detected infective stages of parasites in animal and human feces. Recently, its applicability has been extended by the use in pastures and vegetables for human consumption. However, its use for the detection of parasites in the soil has never been investigated. In this study, we assessed the performance of the FLOTAC and compared with centrifugal flotation (CFT) and spontaneous sedimentation (SST) techniques. A total of 50 soil samples were collected from the Metropolitan region of Recife, Northeastern Brazil. Initially, samples were standardized, and then assessed in duplicate by the FLOTAC, CFT and SST. Cohen's kappa coefficient, sensitivity, specificity, positive and negative predictive values, and accuracy of the FLOTAC, CFT and SST were assessed using the results of the combination of all three techniques as gold standard. Out of all analyses performed, in 96% (48/50), helminth eggs of parasites were detected. In particular, 96% (48/50) of samples analysed through the FLOTAC technique, 76% (38/50) for CFT and 28% (14/50) for SST were positive. Nematode eggs of the Ancylostomatidae and Ascarididae families were those mostly detected in this study. Overall, the FLOTAC presented high values of sensitivity, demonstrating that this method may also be employed for detection of parasites in soil samples. This study opens new possibilities for the use of the FLOTAC technique, highlighting its role as a potential tool for detecting environmental contamination by parasites of medical and veterinary importance.

Molecular characterization of Trypanosoma cruzi DTUs of the triatomine species in a Chagas disease endemic area.

Trypanosoma cruzi is the etiological agent of Chagas disease, a neglected tropical infection with great public health importance. This protozoan has triatomine insects as vector but may also be transmitted through blood transfusion, organ transplants, ingestion of contaminated food, or congenitally. It has a heterogeneous population classified into Discrete Typing Units (DTUs), TcI-TcVI and TcBat. The aim of this study was to molecularly characterize the DTUs of T. cruzi in triatomines from a Chagas disease endemic area in Northeastern Brazil. Triatomines were collected and the gut content was microscopically analyzed to investigate the presence of trypanosomatid flagellates. In addition, digestive tracts of some specimens were dissected and molecularly analyzed through PCR for Trypanosoma spp. and sequencing. PCR positive samples were further submitted to a multiplex PCR for DTUs of T. cruzi. A total of 117 triatomines were collected, 93.16% being in intradomicile and 6.84% in peridomicile environments. Insects were identified as Panstrongylus lutzi (37.60%), Triatoma pseudomaculata (26.50%), Triatoma brasiliensis (23.08%) and Panstrongylus megistus (12.82%). The specimens herein analyzed presented infection rates by T. cruzi of 5.49% and 12.09% in parasitological and molecular examinations, respectively. Multiplex PCR screening revealed 70.59% of the TcI genotype, detected in all triatomine species identified in this study and 29.41% of the DTU TcIII/TcIV detected in P. megistus and P. lutzi. T. cruzi infect triatomines in intradomicile and peridomicile environments, which brings attention to the risk of human infections and to the importance of the implementation of surveillance and entomological control actions.

Infections by trypanosomatid (Kinetoplastida: Trypanosomatidae) in triatomines (Hemiptera: Triatominae): A spatiotemporal assessment in an endemic area for Chagas disease.

This research analysed the spatiotemporal distribution of triatomines infected by trypanosomatid parasites in an endemic region for Chagas disease, in the state of Pernambuco, Northeastern Brazil. The database included the total number of triatomines captured from intradomicile and peridomicile areas, as well as the infection rate (IR) by trypanosomatid. The Gi∗ by Getis-Ord method was used to statistically identify significant concentration clusters and the IR of triatomines by trypanosomatids. A generalized linear regression model with a binomial distribution was used to evaluate the probability of finding an IR by trypanosomatids. Overall, of 4,800 triatomines examined, trypanosomatid forms similar to Trypanosoma cruzi were detected in 10.29% of them, and the majority of positive specimens (98.17%) were collected at intradomicile. The geospatial analyses identified triatomines clusters in intradomicile and peridomicile environments. According to the logistic regression data for species (Panstrongylus lutzi, P. megistus, Triatoma brasiliensis and T. pseudomaculata), the probability of detection of T. cruzi infection remains constant in up to 50 specimens examined or more. The findings of this research revealed a scenario never studied in this area through this type of spatiotemporal analysis, which is essential to identify areas of vulnerability for the occurrence of these vectors and consequently for Chagas disease.

Lungworms in ruminants from Brazil: A retrospective epidemiological study over four decades.

Lungworms such as Dictyocaulus spp. in cattle and small ruminants, and Muellerius capillaris and Protostrongylus rufescens in small ruminants are important pathogens, causing respiratory disease in these livestock species. Despite their veterinary importance, lungworms of livestock have been poorly studied in certain regions of the world, including Brazil. Therefore, much of their epidemiology and economic impacts on production remain unknown. This article aims to provide a comprehensive overview of the historical and current data published on lungworm infection of domestic ruminants in Brazil. This review consisted of a comprehensive search of technical and scientific publications between January 1980 to December 2020, using online sources such as PubMed, Google Scholar and Scielo. Twenty-four articles published over the last 40 years reporting lungworms exclusively in cattle (n = 16), goats (n = 6) and sheep (n = 1) in Brazil were included. In addition, a study (n = 1) with both goats and sheep were also utilized. Overall, 12 studies were based only on post-mortem examination, five in the detection of specimens in fecal samples, and seven were based on fecal analysis followed by post-mortem examination. Out of all studies, 66.7% (n = 16) articles registered D. viviparus, 4.2% (n = 1) D. filaria, 8.3% (n = 2) P. rufescens, 16.7% (n = 4) M. capillaris, and 4.2% (n = 1) co-infection by D. filaria and M. capillaris. The existence of suitable environmental conditions, as well as intermediate and definitive hosts in Brazil contribute for the survival and development of these nematode species. The majority of the reports of lungworms originate from the Southern and Southeastern regions of the country, whose mild temperatures likely contribute to their occurrence. Finally, lungworms of ruminants have been reported over the past four decades in Brazil, but most of the information was obtained at post-mortem examination. Therefore, further studies to investigate epidemiological aspects in different hosts and regions of the country are needed.

Occurrence and anatomical distribution of myiasis caused by Cochliomyia hominivorax (Diptera: Calliphoridae) in swine.

Cochliomyia hominivorax (Diptera: Calliphoridae) has been considered the most important cause of myiasis in animals in the South America with most cases being recorded in cattle. In Brazil, the risk of myiasis in swine population is underestimated, leading to difficulties in the implementation of preventive measures. The aim of this study was to report the occurrence of myiasis infesting a swine population over a one-year period and to evaluate the anatomical distribution of these larvae on the animal's body. From May 2019 to April 2020, a total of 2038 inspections were carried out on pigs reared in two farms located in the state of Pernambuco, Northeastern Brazil. Fly larvae retrieved were morphologically identified and the anatomical distribution of myiasis was graphically represented by a heat map indicating its frequency in swine based on the interpolation of data. Cochliomyia hominivorax larvae (n = 78) of second (n = 29) and third (n = 49) developmental stages were recovered in 13 animals (0.64%) out of 2038 inspections performed. The occurrence of myiasis was detected in eight anatomical sites, with higher concentration on the forehead region and on ears. Data herein obtained provide valuable information on the epidemiology and biological aspects of C. hominivorax affecting pigs. In addition, it indicates that the forehead and ears are important sites of parasitism most likely due the high frequency of skin lesions in these areas.

Prevalence and risk factors associated with infection by Eimeria spp. in goats and sheep in Northeastern Brazil.

Eimeria spp. infections lead to acute or chronic intestinal disorders in small ruminants, being responsible for important economic losses worldwide. The aim of this study was to assess the risk factors associated with Eimeria spp. infection in small ruminants of the microregion of Garanhuns, state of Pernambuco, Northeastern Brazil. Fecal samples (n = 822) were obtained from goats (n = 414) and sheep (n = 408) and evaluated by the modified Gordon and Whitlock technique. Risk factors were assessed through univariate analysis and logistical regression. Oocysts of Eimeria species were detected in 62.9% (517/822) of the animals, with 77.8% (322/414) in goats and 47.8% (195/408) in sheep. For goats, the herd size (OR = 5.52), rearing system (OR = 1.57), feeding place (OR = 2.60), absence of mineral salt in the diet (OR = 2.54), flooring type (OR = 2.83) and periodicity of cleaning (OR = 5.39) were considered risk factors. Conversely, for sheep only the herd size (OR = 3.16) and rearing system (OR = 2.45) were important factors associated with infection by Eimeria spp. Data herein obtained brings meaningful information on the epidemiology of coccidiosis in small ruminants in Northeastern Brazil. The knowledge of these risk factors is useful to contribute to the development of preventive measures, thereby, reducing the economic impact caused by these protozoa in small ruminant production.

Detection of Rickettsia felis in ectoparasites collected from domestic animals.

Ticks and fleas are arthropods widely distributed around the world involved in the transmission of various vector-borne diseases (VBDs), including Brazilian Spotted Fever (BSF), Baggio-Yoshinari Syndrome and the plague, with outstanding consequences for the public health. The aim of this study was to investigate the presence of Rickettsia spp., Borrelia spp. and Yersinia pestis in arthropods collected from dogs, cats and horses living in the state of Pernambuco, Northeastern Brazil. From January 2017 to April 2019, ectoparasites were collected, morphologically identified and molecularly analysed through PCR and sequencing. In total 401 specimens were collected from 86 animals, being 68% (n = 273) and 32% (n = 128) from rural and urban areas, respectively. The most commonly detected species were the ticks Dermacentor nitens, Amblyomma sculptum, Rhipicephalus sanguineus sensu lato (s.l.), Rhipicephalus microplus, and Amblyomma ovale, and the fleas Ctenocephalides felis and Ctenocephalides canis. DNA of Rickettsia felis was detected in D. nitens collected from horses, and C. felis, and R. sanguineus s.l. collected from dogs. All samples scored negative for Borrelia spp. and Y. pestis DNA. This study provides valuable data on ectoparasite fauna from domestic animals and identifies the circulation of a zoonotic pathogen (i.e., R. felis) in the population of the arthropods assessed. Therefore, preventive measures should be adopted in order to reduce the risk of occurrence of neglected VBD caused by this pathogen in animal and human hosts.

Lutzomyia evandroi in a New Area of Occurrence of Leishmaniasis.

PURPOSE: Sand flies are vectors of medical and veterinary concern, responsible for the transmission of Leishmania parasites. These invertebrates are widely distributed throughout the world and their abundance and diversity is dependent on several biotic and physical factors. Based on the epidemiological importance of these insects, the aim of this study was to assess the sand fly fauna of a new area of occurrence of leishmaniasis. METHODS: From October 2018 to September 2019, samplings were performed using CDC light traps at six different collection points located in a high altitude area in the municipality of Garanhuns, Northeastern Brazil. Geographic coordinates were recorded using the satellite remote sensing Global Positioning System (GPS) Garmin Etrex20. The spatial distribution of sand flies was evaluated using kernel density estimation (KDE). Engorged females (n = 12) were molecularly processed to search for Leishmania DNA. RESULTS: A total of 138 sand fly specimens were collected during the study period, with 100% (138/138) identified as Lutzomyia evandroi. Climatic conditions did not influence the number of sand flies (males and females) collected monthly, but a statistically significant difference was observed between the number of specimens and different collections points. The kernel map showed a heterogeneous spatial distribution of sand flies in the municipality of Garanhuns with a hotspot in the south of the region. CONCLUSION: Data presented herein are pivotal in filling the gap on the epidemiology of leishmaniasis in this high altitude area, serving as an alert to local health authorities. It also indicated that the putative role of L. evandroi in the transmission of Leishmania parasites should be clarified.

An Additional Asset for the FLOTAC Technique: Detection of Gastrointestinal Parasites in Vegetables.

INTRODUCTION: Gastrointestinal parasites are considered amongst the most important threats to veterinary and human health worldwide. The transmission of these parasitic diseases usually involves the accidental ingestion of oocysts, cysts, eggs, and larvae, whose detection is generally achieved by classical coproparasitological methods, based on sedimentation and/or flotation techniques. These procedures might be inefficient or display a low sensitivity when performed on food matrices. This study aimed to assess the value of the FLOTAC technique for the detection of different parasitic stages of gastrointestinal parasites of domestic animals in fresh lettuce. MATERIAL AND METHODS: Samples (n = 100) were purchased from public markets located in the metropolitan region of Recife (Brazil). RESULTS: A total of 79% scored positive for eggs and/or cysts of gastrointestinal parasites. Protozoa and nematodes were detected in 32% (32/100) and 64% (64/100) of samples, respectively, with cysts of the genus Entamoeba (32%) and eggs of nematodes of the order Strongylida (30%) being the most frequently diagnosed. CONCLUSIONS: The findings herein reported demonstrate that the FLOTAC technique can be successfully applied for recovering food-borne parasites of medical and veterinary concern in food matrices.

Cercopithifilaria bainae in Rhipicephalus sanguineus sensu lato ticks from dogs in Brazil.

Rhipicephalus sanguineus sensu lato (s.l.) ticks act as intermediate host for a range of canine vector-borne pathogens, including nematodes ranked in the genus Cercopithifilaria. Though being the object of several studies in the last years, information on the distribution of these parasites is still lacking. In this study, the occurrence of Cercopithifilaria spp. was investigated in on-host population of R. sanguineus s.l. collected from naturally infested dogs. Ticks (n=1906, including one larva, 294 nymphs and 1611 adults) were sampled on domestic dogs (n=155) living in the municipality of Garanhuns (northeastern Brazil). Tick collections (n=36) were performed every 8 days, from October 2015 to June 2016. Filarioid larvae detected at tick dissection were morphologically and morphometrically identified at species level. At the end of the study, only R. sanguineus s.l. ticks were collected, with the highest number in January 2016 (n=254) and the lowest in June 2016 (n=26). Out of 1906 dissected ticks, 2.68% (51/1906) harboured Cercopithifilaria bainae larvae, whose identification was molecularly confirmed, with a nucleotide identity of 99% with C. bainae. Data here reported indicate that, in the study area, R. sanguineus s.l. is the predominant tick infesting domestic dogs. Accordingly, these animals are at a high risk of C. bainae infection.

Larvae of Ixodiphagus wasps (Hymenoptera: Encyrtidae) in Rhipicephalus sanguineus sensu lato ticks (Acari: Ixodidae) from Brazil.

The biological control of ticks represents an alternative method to the chemical control, given its ecological-friendly approach. Amongst the alternatives, the use of parasitoids of the genus Ixodiphagus (Hymenoptera: Encyrtidae) has been largely investigated. The aim of this study was to document and molecularly characterize Ixodiphagus wasps in ticks from a tropical region of Brazil. From October 2015 to March 2016, Rhipicephalus sanguineus sensu lato ticks (n=1814) were collected from naturally infested dogs and Ixodiphagus larvae were detected by microscopic examination. In addition, adult wasps were obtained in the laboratory. Larvae and adults were molecularly identified as Ixodiphagus hookeri. These findings suggest that this type of parasitism deserves to be studied in local tick populations, in order to elucidate the role of these wasps as a potential alternative to chemical tick control.

Filarioids infecting dogs in northeastern Brazil.

Species of filarial nematodes belonging to the genera Dirofilaria and Acanthocheilonema are recognised as common parasites of dogs throughout the world. Recently, other filarioids featured by the presence of dermal microfilariae (e.g., Onchocerca lupi and Cercopithifilaria spp.) have been recognised in Europe. In Brazil, reports of filarioids in dogs are limited to Dirofilaria immitis, Acanthocheilonema reconditum and Cercopithifilaria bainae. To investigate the distribution of filarial infections in dogs living in an endemic region from northeastern Brazil, blood and skin samples (n=104) were microscopically (modified Knott's test and skin snip sediment examination) and molecularly evaluated. Twenty-two dogs (21.15%) were positive at microscopic and/or molecular examination for at least one filarioid species, with 21 (20.19%) animals positive for blood microfilariae at molecular and/or at microscopic examination. Microfilariae of D. immitis were detected in 12 (11.54%) animals, with co-infection of D. immitis and A. reconditum observed in four (3.85%) individuals. One animal was positive for C. bainae at both microscopic and molecular examination. Analysis of sequence obtained in the present study showed significant alignment identity with that of C. bainae from Europe. Considering that in the area of study arthropod vectors (mosquitoes, fleas and ticks) are prevalent throughout the year, preventive measures should be disposed in order to avoid the animal infestation and pathogen infection.

Prevalence of zoonotic visceral leishmaniasis in dogs in an endemic area of Brazil.

INTRODUCTION: The northeast region of Brazil is endemic for zoonotic visceral leishmaniasis (ZVL). The aim of this study was to determine the prevalence of infection in dogs in Petrolina. METHODS: Blood samples were collected from dogs (n = 600), and bone-marrow biopsy was performed in animals with positive serological test results that presented clinical signs of ZVL. The serological analyses were performed using an enzyme-linked immunosorbent assay (ELISA) (S7(r)Biogene). RESULTS: Of the 600 dogs tested, 19% (115/600) presented anti-L. infantum chagasi antibodies. CONCLUSIONS: Our data are important because canine infection is an important risk factor for the human disease.

Ecology of sand flies in a low-density residential rural area, with mixed forest/agricultural exploitation, in north-eastern Brazil.

Cutaneous leishmaniasis caused by Leishmania braziliensis is endemic in Brazil, where Lutzomyia whitmani is the most important vector involved in the transmission to humans, particularly in the peridomestic environment. Herein, we assessed the ecology of sand flies, including Lu. whitmani, in a low-density residential rural area with mixed forest/agricultural exploitation in north-eastern Brazil, where cutaneous leishmaniasis is endemic. Particularly, we hypothesized that sand fly abundance was correlated with climatic variables. Sand fly collections were carried out monthly from August 2013 to August 2014, using seven CDC light traps, for three consecutive nights, in three kinds of environments: indoor, peridomicile and forest. Collected sand flies were identified based on morphology and females of Lu. whitmani (n=169), Lu. amazonensis (n=134) and Lu. complexa (n=21) were selected and tested by PCR for Leishmania (Viannia) spp. In total, 5167 sand flies belonging to 19 species were identified, being that Lu. choti (43.2%) was the most frequent species, followed by Lu. amazonensis (16.6%), Lu. whitmani (15.8%), Lu. sordellii (10.7%) and Lu. quinquefer (5.8%), which together represented over 90% of the collected sand flies. All females tested by PCR were negative. The number of sand flies collected daily was positively correlated with temperature and negatively correlated with rainfall and relative humidity. Furthermore, there was a positive correlation between daily number of sand flies and daily average saturation deficit. This study points out that the number of sand flies captured daily is correlated to climatic variables, including saturation deficit, which may represent a useful parameter for monitoring sand fly populations in leishmaniasis-endemic areas.

Quantification of Leishmania infantum DNA in the bone marrow, lymph node and spleen of dogs.

The aim of the present study was to quantify the parasite load of Leishmania infantum in dogs using real-time PCR (qPCR). Bone marrow, lymph node and spleen samples were taken from 24 dogs serologically positive for L. infantum that had been put down by the official epidemiological surveillance service. According to the clinical signs the dogs were classified as asymptomatic or symptomatic. After DNA extraction, the samples were subjected to qPCR to detect and quantify L. infantum DNA. Out of the 24 dogs, 12.5% (3/24) were classified as asymptomatic and 87.5% (21/24) as symptomatic. Real-time PCR detected L. infantum DNA in all the animals, in at least one biological sample. In particular, 100% of bone marrow and lymph node scored positive, whereas in spleen, the presence of DNA was detected in 95.9% (23/24). In addition, out of 24 animals, 15 were microscopically positive to amastigote forms of L. infantum in bone marrow. No statistical significant difference was found in the overall mean quantity of DNA among the different biological samples (P = 0.518). Considering each organ separately, there was 100% positivity in bone marrow and lymph nodes, while among the spleen samples, 95.9% (23/24) were positive. Regarding the different clinical groups, the overall mean parasite load varied significantly (P = 0.022). According to the results obtained, it was not possible determine which biological sample was most suitable tissue for the diagnosis, based only on the parasite load. Therefore, other characteristics such as convenience and easily of obtaining samples should be taken into consideration.

Histological changes and immunolabeling of Leishmania infantum in kidneys and urinary bladder of dogs.

The aim of this study was to assess histological alterations and perform immunolabeling of Leishmania infantum in the kidneys and urinary bladder of naturally infected dogs. Twenty-five urinary bladder and kidney samples of serologically positive animals (ELISA S7® Biogene and IFAT ≥ 1:40 - Biomanguinhos/Fiocruz) were analyzed by means of immunohistochemical and histological techniques. Cystitis was found in 44% (11/25) of the bladder samples and membranoproliferative glomerulonephritis in 92% (23/25) of the kidney samples. Immunolabeling of the parasite revealed that 32% (8/25) of the bladders and 8% (2/25) of the kidneys were positive. In conclusion, the immunohistochemical technique is a useful tool for detecting amastigote forms of L. infantum in organs of infected dogs. In addition, this was the first report of detection of amastigote forms of L. infantum in the bladders of dogs.