Daniel Moreira de Avelar

Neosomes of tungid fleas on wild and domestic animals

Tunga is the most specialized genus among the Siphonaptera because adult females penetrate into the skin of their hosts and, after mating and fertilization, undergo hypertrophy, forming an enlarged structure known as the neosome. In humans and other warm-blooded animals, neosomes cause tungiasis, which arises due to the action of opportunistic agents. Although its effects on humans and domestic animals are well described in the literature, little is known about the impact of tungiasis on wild animals. This review focuses on the morphology, taxonomy, geographical distribution, hosts, prevalence, sites of attachment, and impact of tungid neosomes on wild and domestic animals. Because neosomes are the most characteristic form of the genus Tunga and also the form most frequently found in hosts, they are here differentiated and illustrated to aid in the identification of the 13 currently known species. Perspectives for future studies regarding the possibility of discovering other sand flea species, adaptation to new hosts, and the transfer of tungids between hosts in natural and modified habitats are also presented.

Phlebotomine Sand Fly Fauna and Leishmania Infection in the Vicinity of the Serra do Cipó National Park, a Natural Brazilian Heritage Site

In the New World, the leishmaniases are primarily transmitted to humans through the bites of Leishmania-infected Lutzomyia (Diptera: Psychodidae) phlebotomine sand flies. Any or both of two basic clinical forms of these diseases are endemic to several cities in Brazil—the American cutaneous leishmaniasis (ACL) and the American visceral leishmaniasis (AVL). The present study was conducted in the urban area of a small-sized Brazilian municipality (Jaboticatubas), in which three cases of AVL and nine of ACL have been reported in the last five years. Jaboticatubas is an important tourism hub, as it includes a major part of the Serra do Cipó National Park. Currently, no local data is available on the entomological fauna or circulating Leishmania. During the one-year period of this study, we captured 3,104 phlebotomine sand flies belonging to sixteen Lutzomyia species. In addition to identifying incriminated or suspected vectors of ACL with DNA of the etiological agent of AVL and vice versa, we also detected Leishmania DNA in unexpected Lutzomyia species. The expressive presence of vectors and natural Leishmania infection indicates favorable conditions for the spreading of leishmaniases in the vicinity of the Serra do Cipó National Park.

Leishmania, Babesia and Ehrlichia in urban pet dogs: co-infection or cross-reaction in serological methods?

INTRODUCTION The present study was designed to assess the occurrence of co-infection or cross-reaction in the serological techniques used for detecting the anti-Leishmania spp., -Babesia canis vogeli and -Ehrlichia canis antibodies in urban dogs from an area endemic to these parasites. METHODS The serum samples from dogs were tested for the Babesia canis vogeli strain Belo Horizonte antigen and Ehrlichia canis strain São Paulo by immunofluorescence antibody test (IFAT) and by anti-Leishmania immunoglobulin G (IgG) antibody detection to assess Leishmania infection. We used the following four commercial kits for canine visceral leishmaniasis: ELISA, IFAT, Dual Path Platform (DPP) (Bio Manguinhos(r)/FIOCRUZ/MS) and a rK39 RDT (Kalazar Detect Canine Rapid Test; Inbios). RESULTS Of 96 serum samples submitted to serological assays, 4 (4.2%) were positive for Leishmania as determined by ELISA; 12 (12.5%), by IFAT; 14 (14.6%) by rK39 RDT; and 20 (20.8%), by DPP. Antibodies against Ehrlichia and Babesia were detected in 23/96 (23.9%) and 30/96 (31.2%) samples, respectively. No significant association was identified between the results of tests for detecting Babesia or Ehrlichia and those for detecting Leishmania (p-value>0.05). CONCLUSIONS In the present study, we demonstrated co-infection with Ehrlichia or Babesia and Leishmania in dogs from Minas Gerais (Brazil); we also found that the serological tests that were used did not cross-react.

Epidemiological aspects of vector; parasite; and domestic reservoir in areas of recent transmission and no reported human cases of visceral leishmaniasis in Brazil.

About 97% of the human cases of the American visceral leishmaniasis (VL) occur in Brazil. In the last few years, the disease expanded to medium- and large-sized cities, in which surveillance and control actions have been intensified, in an effort to control VL spreading. Our two-year study was conducted in Belo Horizonte, the sixth most populous city in Brazil, which is endemic for VL. We focused in two particular districts of recent transmission of the disease, with no reported human cases and submitted to minor surveillance and control actions. Our aim was to draw an epidemiological profile of the local situation concerning Lutzomyia vector, Leishmania parasites, and the main domestic reservoirs (dogs). Lutzomyia longipalpis comprised 96.5% of the total phlebotomine sand flies captured and displayed an expressive minimal infection rate by Leishmania infantum (16.7%). Positive correlations were found between the population densities of L. longipalpis, rainfall and temperature. L. infantum was also detected in the cortelezzii complex and, for the first time, in Lutzomyia lloydi. Leishmania braziliensis, an etiological agent of the American cutaneous leishmaniasis, was also identified in L. longipalpis. Among the 1408 dogs serologically tested by standard enzyme-linked and fluorescence immune assays (ELISA/IFA) 3.6% were positive for VL. L. infantum DNA and Leishmania parasites were identified in 100% and 72.5% of the seropositive dogs, respectively. The co-positivity of other diagnostic tests for VL-Leishmania-nested PCR, imprint and myeloculture-was compared to the standard serology. Both symptomatic or asymptomatic dogs displayed an equal average number of positive diagnostic tests for VL. The districts studied display favorable conditions for the rapid spreading of human infection, in terms of L. longipalpis population density, and presence of L. infantum in both vector and main reservoir.

Ectoparasites and anti-Leishmania antibodies: Association in an observational case–control study of dogs from a Brazilian endemic area

It has been proposed that the transmission of canine visceral leishmaniasis might involve the participation of mechanical vectors, including ticks of the family Ixodidae, in particular the brown dog tick Rhipicephalus sanguineus, and the cat flea Ctenocephalides felis felis. Here, the association between the infestation by R. sanguineus and C. felis felis and the occurrence of anti-Leishmania antibodies was evaluated in an observational case-control study of dogs living in a Brazilian endemic area for canine visceral leishmaniasis. Blood samples were taken once every three months for one year from 96 initially seronegative domestic dogs, and submitted to indirect immunofluorescence antibody assay. All dogs were evaluated for the presence of ticks and fleas, and the results were expressed qualitatively as infested or non-infested, irrespective of the intensity of infestation. At the end of follow-up, twenty dogs had turned seropositive, while 68 remained seronegative and 8 were excluded because of incomplete data. All the dogs were asymptomatic. The odds of infection was significantly greater (OR=3.54, CI95%=1.10-12.53) for dogs infested by C. felis felis compared to their non-infested counterparts. In contrast, the odds of infection showed no significance difference between non-infested and R. sanguineus-infested groups of dogs (OR=0.31, CI95%=0.03-1.52). This study provides further evidence for the potential role of C. felis felis in mechanically transmitting Leishmania among the canine population.

Dynamics of Ctenocephalides felis felis (Siphonaptera: Pulicidae) Infestations on Urban Dogs in Southeastern Brazil

ABSTRACT The cat flea, Ctenocephalides felis felis (Bouché, 1835), is an important ectoparasite of dogs and cats throughout the world, causing annoyance to the animals and acting as a vector of infections and a cause of allergic dermatitis in dogs and cats. Although climatic variability and seasonality are known to influence the diversity and abundance of fleas, few investigations of seasonal prevalence of cat flea infestation have involved the same group of dogs being examined regularly over an extended period. The aim of this study was to determine the effects of temperature, rainfall, and relative humidity on the infestation by C. felis felis on 88 outdoor dogs in southeastern Brazil. The dogs, which were of mixed breed, sex, and age, were examined for ectoparasites every month during the period August 2011 to July 2012, and samples of fleas were randomly collected and identified. Meteorological data, comprising mean temperature, total rainfall, and mean relative humidity, were recorded for the calendar month prior to that in which the examinations were performed. Dogs were found to be infested only with C. felis felis, with a higher prevalence in the months with lowest rainfall (July, August, and September). The data obtained in this investigation can be used in control programs in order to establish an efficient strategy for environmental management and the application of insecticides, particularly during the driest months of the year based on the seasonal pattern of infestation of dogs by C. felis felis.

Notes on the genus Tunga (Siphonaptera: Tungidae) II – neosomes, morphology, classification, and other taxonomic notes

This review focuses on the neosomes, morphology, and taxonomy of adult species of the genus Tunga, complementing the previously published data on the phylogeny, ecology, and pathogenic role. Neosomes are structures formed after penetration of adult females into the skin of hosts resulting in significant enlargement, being the most characteristic and most frequently observed form in hosts. Neosomes can be differentiated by shape, measurements, and sites of attachment to principal hosts. The taxonomic value and morphometric data of the most widely used characteristics to separate species – such as frontal curvature, head chaetotaxy, preoral internal sclerotization, ventral and dorsal genal lobes, eyes, maxillary palps, fusion of pronotum and mesonotum, metacoxae, metatarsi chaetotaxy, spermatheca (females), manubrium, basimere, telomere, and phallosome (males) – are comparatively analyzed. The sexes, individual variations, undescribed species, higher taxa, as well as a proposal for division of the genus into two subgenera (Tunga and Brevidigita) are presented (as previously given by Wang). A key for females, males, and gravid females (neosomes) also is included for identifying the 13 known species. Data on host specificity and geographical distribution may also support the identification of Tunga species because some sand fleas and their hosts may have co-evolved.

Infection with Leishmania (Leishmania) infantum of 0 to 18-Month-Old Children Living in a Visceral Leishmaniasis-Endemic Area in Brazil

The diagnosis of Leishmania (Leishmania) infantum infection in children from birth may serve as a reference for the early identification of cases that would progress to classical visceral leishmaniasis (VL) in endemic areas. This study prospectively evaluated newborns of mothers living in the municipality of Paracatu, Minas Gerais, Brazil. The infants were followed up at 6-month intervals by clinical examination, serological tests (immunofluorescence [IIF] and enzyme-linked immunosorbent assay with rK39 [ELISA-rK39]) and polymerase chain reaction (PCR) until they had completed 18 months of age. A total of 166 pregnant women were included to evaluate the possible transfer of antibodies or even congenital transmission. Twenty-two of the women tested positive by IIF, four by ELISA-rK39, and one by PCR. Three infants of the 25 women with some positive test results were also positive in the first test (one by IIF, one by ELISA-rK39, and the third by ELISA-rK39 and PCR). One hundred and sixty infants were included in the study; of these, 43 had at least one positive sample over time. However, agreement between tests was low. Follow-up of children with a positive result in the tests studied revealed no progression to classical disease within a period of 18 months. In contrast, two children with negative IIF, PCR, and ELISA-rK39 results developed classical VL at 9 and 12 months of age. In conclusion, a positive test result was variable and sometimes temporary and agreement between tests was low. Therefore, the early diagnosis of Leishmania infection was not associated with the early identification of cases that would progress to classical VL in the endemic area studied.

Meglumine antimoniate intralesional infiltration for localised cutaneous leishmaniasis: a single arm, open label, phase II clinical trial

BACKGROUND Cutaneous leishmaniasis (CL) is a world-wide health problem which currently lacks effective, affordable and easy to use therapy. Recently, the meglumine antimoniate (MA) intralesional infiltration was included among the acceptable therapies for New World leishmaniasis. While this approach is attractive, there is currently little evidence to support its use in Americas. OBJECTIVES The aim of this study was to provide information about effectiveness and safety of a standardised MA intralesional infiltration technique for the treatment of CL. METHODS It is a single-arm phase II clinical trial conducted at a Brazilian referral centre. CL cases with parasitological confirmation presenting a maximum of three CL-compatible skin lesions were treated with weekly MA intralesional infiltration by using a validated technique, up to a maximum of eight infiltrations. RESULTS A total of 53 patients (62 lesions) were included. Overall, patients received a median of seven infiltrations (IQR25-75% 5-8) over a median treatment period of 43 days (IQR25-75% 28-52 days). The definitive cure rate at D180 was 87% (95% CI:77-96%). The majority of adverse events were local, with mild or moderate intensity. Bacterial secondary infection of the lesion site was observed in 13% of the treated patients, beside two intensity-three adverse events (hypersensitivity reactions).

Seroprevalence and molecular characterization of Leishmania in dogs from an endemic area of zoonotic visceral leishmaniasis in Brazil

Abstract Visceral leishmaniasis (VL) can cause large-scale and tenacious epidemics with high fatality rates. Current seroprevalence and circulating Leishmania species were evaluated in dogs domiciled in the municipality of Sabará, a small historic and touristic city in the Brazilian state of Minas Gerais. A total of 3926 dogs domiciled in seven different districts of Sabará were serologically tested for canine visceral leishmaniasis (CVL) by indirect enzyme-linked immunosorbent (ELISA) and immunofluorescence (IFA) assays, in a two-years census survey (2011–2012). The average positivity rate of canine infection was 3.4%. Three additional diagnostic tests – imprint/smear direct parasitological, molecular (LnPCR) and myeloculture – were performed in a random sample of fifty seropositive dogs composed of symptomatic (39) and asymptomatic (eleven) animals. LnPCR showed 100% of positivity for Leishmania DNA in, at least, one among four tissue samples tested (mesenteric lymph node, skin, spleen and bone marrow), independently of the clinical canine group. Higher and statistically equivalent positivity rates (98% and 96%) for Leishmania DNA were found in canine lymph node and spleen. Asymptomatic dogs showed expressive positivity rates in all three additional diagnostic techniques. Leishmania infantum was confirmed as the etiological agent of CVL in Sabará.