The early inhibition of the COX-2 pathway in viperid phospholipase A2-induced skeletal muscle myotoxicity accelerates the tissue regeneration.

The administration of non-steroidal anti-inflammatory drugs in the treatment of injury and muscle regeneration is still contradictory in effectiveness, especially regarding the timing of their administration. This can interfere with the production of prostaglandins originating from inflammatory isoform cyclooxygenase-2 (COX-2), which is essential to modulate tissue regeneration. The phospholipases A2 (PLA2) from viperid venoms cause myotoxicity, therefore constituting a tool for the study of supportive therapies to improve skeletal muscle regeneration. This study investigated the effect of early administration of lumiracoxib (selective inhibitor of COX-2) on the degeneration and regeneration stages of skeletal muscle after injury induced by a myotoxic PLA2. After 30 min and 48 h of intramuscular injection of PLA2, mice received lumiracoxib orally and histological, functional, and transcriptional parameters of muscle were evaluated from 6 h to 21 days. Inhibition of COX-2 in the early periods of PLA2-induced muscle degeneration reduced leukocyte influx, edema, and tissue damage. After the second administration of lumiracoxib, in regenerative stage, muscle showed increase in number of basophilic fibers, reduction in fibrosis content and advanced recovery of functionality characterized by the presence of fast type II fibers. The expression of Pax7 and myogenin were increased, indicating a great capacity for storing satellite cells and advanced mature state of tissue. Our data reveals a distinct role of COX-2-derived products during muscle degeneration and regeneration, in which early administration of lumiracoxib was a therapeutic strategy to modulate the effects of prostaglandins, providing a breakthrough in muscle tissue regeneration induced by a myotoxic PLA2.

Insights on the structure of native CNF, an endogenous phospholipase A2 inhibitor from Crotalus durissus terrificus, the South American rattlesnake.

Several snake species possess endogenous phospholipase A2 inhibitors (sbPLIs) in their blood plasma, the primary role of which is protection against an eventual presence of toxic phospholipase A2 (PLA2) from their venom glands in the circulation. These inhibitors have an oligomeric structure of, at least, three subunits and have been categorized into three classes (α, ß and γ) based on their structural features. SbγPLIs have been further subdivided into two subclasses according to their hetero or homomeric nature, respectively. Despite the considerable number of sbγPLIs described, their structures and mechanisms of action are still not fully understood. In the present study, we focused on the native structure of CNF, a homomeric sbγPLI from Crotalus durissus terrificus, the South American rattlesnake. Based on the results of different biochemical and biophysical experiments, we concluded that, while the native inhibitor occurs as a mixture of oligomers, tetrameric arrangement appears to be the predominant quaternary structure. The inhibitory activity of CNF is most likely associated with this oligomeric conformation. In addition, we suggest that the CNF tetramer has a spherical shape and that tyrosinyl residues could play an important role in the oligomerization. The carbohydrate moiety, which is present in most sbγPLIs, is not essential for the inhibitory activity, oligomerization or complex formation of the CNF with the target PLA2. A minor component, comprising no more than 16% of the sample, was identified in the CNF preparations. The amino-terminal sequence of that component is similar to the B subunits of the heteromeric sbγPLIs; however, the role played by such molecule in the functionality of the CNF, if any, remains to be determined.

The PLA2 inhibitor from Crotalus durissus terrificus blood plasma (CNF) inhibits group III-PLA2 from honeybee venom.

Crotalus neutralizing factor (CNF) is an endogenous glycoprotein from Crotalus durissus terrificus snake blood that inhibits secretory phospholipases A2 (sPLA2) from the Viperid but not from Elapid venoms (subgroups IA and IIA, respectively). In the present study, we demonstrated that CNF can inhibit group III-PLA2 from bee venom by forming a stable enzyme-inhibitor complex. This finding opens up new possibilities for the potential use of CNF and/or CNF-based derivatives in the therapeutics of bee stings.

Abundance of Leishmania (Kinetoplastida: Trypanosomatidae) vectors in Baldim, a municipality in the Espinhaço Range Biosphere Reserve in Brazil.

Leishmaniasis is a zoonoses caused by protozoan parasites belonging to the Leishmania genus that is transmitted to humans through the bite of previously infected phlebotomine sandflies (Diptera:Psychodidae:Phlebotominae). In Brazil, 2 types of leishmaniasis are endemic, tegumentary leishmaniasis (TL) and visceral leishmaniasis (VL). In Baldim, a municipality in the Espinhaço Range Biospere Reserve (ERBR), 5 autochthonous cases of TL, and one autochthonous case of VL were reported from 2017 to 2022. Owing to the lack of data on entomological fauna, we surveyed the occurrence and distribution of phlebotomine sandflies that could act as Leishmania vectors, as well as natural Leishmania infection. During 1 year, 918 sandfly specimens belonging to 12 species were collected. The predominant species was Lutzomyia longipalpis (Lutz & Neiva, 1912) (32.0%), a proven VL agent, followed by Evandromyia evandroi (Costa Lima & Antunes, 1936) (20.7%), Evandromyia cortelezzii (Brèthes, 1923) (17.0%), Nyssomyia whitmani (Antunes & Coutinho, 1939) (12.2%), and Nyssomyia intermedia (Lutz & Neiva, 1912) (10.5%), all putative or proven TL agents. Seasonal fluctuations and endo-exophilic behavior were delineated for the main phlebotomine sandflies. The population of Lu. longipalpis was higher during the coolest and driest months, although different profiles were noted for the other species. Natural Leishmania infection was not detected in any of the 197 phlebotomine sandfly females analyzed. The results were compared with those previously obtained in other municipalities belonging to the ERBR. The presence of Leishmania vectors and human cases of leishmaniasis indicate a risk of Leishmania transmission in the region.

Mapping possible interaction sites for crotoxin in CNF, a gamma PLA2 inhibitor from Crotalus durissus terrificus rattle snake, using SPOT synthesis.

Phospholipases A2 (PLA2s) are main components of snake venoms. Several snake species possess endogenous PLA2 inhibitors in their circulating blood, which are generally known as sbPLIs (an acronym for snake blood phospholipase A2inhibitors). The sbPLIs are categorized in three classes (alpha, beta or gamma) depending on the existence of distinguishing protein domains in their structure. The Crotalus durrissus terrificus venom has a highly neurotoxic PLA2 - crotoxin (CTX) - in its composition and the self-protection of the snake is mainly ensured by a sbγPLI named CNF (standing for Crotalusneutralizing factor). In an attempt to find smaller molecules able to inhibit the catalytic activity of CTX, in the present study we used linear peptide arrays to identify CNF segments possibly involved in the interaction with the toxin. Five reacting segments were identified as possible interacting regions. The target peptides were synthesized and located in the in silico CNF structure. Although all of them are exposed to the solvent, high concentrations were needed to inhibit the PLA2 activity of the whole venom or CTX. Limitations of the methodology employed and particular characteristics of CTX inhibition by CNF are discussed.

Effects of inhibition of 5-lipoxygenase and 12-lipoxygenase pathways on skeletal muscle fiber regeneration.

We investigated the effect of inhibition of 5-lipoxigenase (LOX) and 12-LOX pathways on the regeneration of skeletal muscle fibers after injury induced by a myotoxin (MTX) phospholipase A2 from snake venom in an in vivo experimental model. Gastrocnemius muscles of mice injected with MTX presented an increase in 5-LOX protein expression, while 12-LOX was found to be a constitutive protein of skeletal muscle. Animals that received oral treatments with 5-LOX inhibitor MK886 or 12-LOX inhibitor baicalein 30 min and 48 h after MTX-induced muscle injury showed a reduction in the inflammatory process characterized by a significant decrease of cell influx and injured fibers in the degenerative phase (6 and 24 h after injury). In the beginning of the regeneration process (3 days), mice that received MK886 showed fewer new basophilic fibers, suggesting fewer proliferative events and myogenic cell fusion. Furthermore, in the progression of tissue regeneration (14-21 days), the mice treated with 5-LOX inhibitor presented a lower quantity of central nucleus fibers and small-caliber fibers, culminating in a muscle that is more resistant to the stimulus of fatigue during muscle regeneration with a predominance of slow fibers. In contrast, animals early treated with the 12-LOX inhibitor presented functional fibers with higher diameters, less resistant to fatigue and predominance of fast heavy-chain myosin fibers as observed in control animals. These effects were accompanied by an earlier expression of myogenic factor MyoD. Our results suggest that both 5-LOX and 12-LOX pathways represent potential therapeutic targets for muscle regeneration. It appears that inhibition of the 5-LOX pathway represses only the degenerative process by reducing tissue inflammation levels. Meanwhile, inhibition of the 12-LOX pathway also favors the anticipation of maturation and earlier recovery of muscle fiber activity function after injury.

Survey of phlebotomine sand fly fauna in a public Zoo in Brazil: Species diversity, seasonality, and host variety.

Leishmaniasis is a dynamic disease in which transmission conditions change due to environmental and human behavioral factors. Epidemiological analyses have shown modifications in the spread profile and growing urbanization of the disease, justifying the expansion of endemic areas and increasing number of cases in dogs and humans. In the city of Belo Horizonte, located in the southeastern state of Minas Gerais (Brazil), visceral leishmaniasis (VL) is endemic, with a typical urban transmission pattern, but with different regional prevalence. This study was conducted at the Zoo of the Foundation of Municipal Parks and Zoobotany of Belo Horizonte (FPMZB-BH), located in the Pampulha region, which is among the areas most severely affected by VL. This study aimed to determine the taxonomic diversity of native phlebotomine sand flies (Diptera: Psychodidae), identify climatic variables that potentially affect the phenology of these insects, and determine the blood meal sources for female phlebotomine sand flies. To achieve this, 10 mammal enclosures in the zoo were selected using the presence of possible leishmaniasis reservoirs as a selection criterion, and sampled using light traps between August 2019 and August 2021. A total of 6034 phlebotomine sand flies were collected, indicating nine species, with Lutzomyia longipalpis being the very abundant species (65.35% of the total). Of the 108 engorged phlebotomine collected females, seven samples (6.5%) were positive for blood meals from humans, marsupials, canids, and birds. Relative humidity and rainfall increased the phenology of phlebotomine sand flies, with population increases in the hottest and wettest months. The data obtained will provide guidelines for competent health agencies to implement vector control measures to reduce the risk of leishmaniasis transmission in the FPMZB-BH.

The Program for the Control of Visceral Leishmaniasis in Brazil: The Effect of the Systematic Euthanasia of Seropositive Dogs as a Single Control Action in Porteirinha, a Brazilian City with an Intense Transmission of Visceral Leishmaniasis.

BACKGROUND: Porteirinha is endemic for visceral leishmaniasis (VL), with intense disease transmission of the disease. We evaluated the impact of canine euthanasia as a single control measure on the incidence of VL in humans and canines. METHODS: A prospective observational cohort study was carried out over four years (1998-2002) in 8 of the 12 neighborhoods of the city. The dynamics of canine visceral leishmaniasis (CVL) transmission were evaluated for 2 years, before beginning the screening-culling intervention. The comparative morbidity index (CMI) was used to stratify areas with the greatest risk of CVL, and the spatial distribution of human and canine VL cases was compared using univariate and bivariate K-functions. RESULTS: Human cases conglomerated in three neighborhoods. Spatial clusters were detected for CVL in 1998, 2000, and 2001, but not in 1999, when greater spatial dispersion occurred. The screening and culling intervention reduced the number of human VL cases and decreased the incidence of CVL, mainly in neighborhoods with a high CMI. CONCLUSIONS: The systematic euthanasia of seropositive dogs was shown to be an effective control action of the Program for Control of Visceral Leishmaniasis (PCLV) in Brazil. The fundamental role of domestic dogs in the epidemiological chain of VL was reaffirmed.

Immune response and protective efficacy of S9 ribosomal protein of Streptococcus pneumoniae in a model of sepsis.

Vaccination is the most promising strategy to reduce the incidence of pneumococcal infection. Although there are vaccines available, all of them are based on polysaccharide antigens (conjugated or not). In addition to their high cost, those vaccines do not cover all serotypes. To overcome these hindrances, we evaluated the immunogenicity and the protective efficacy of the S9 ribosomal protein of Streptococcus pneumoniae with the aim of developing a protein-based vaccine in the future. The gene encoding the S9 ribosomal protein was cloned in pET21-a expression vector, and the recombinant S9 protein was used to immunize mice. Significantly higher levels of anti-S9 immunoglobulin G were achieved (with predominance of immunoglobulin G1) in comparison with the control. Antibodies elicited against S. pneumoniae protein extract in rabbit recognized the recombinant S9 protein by Western blot, thus demonstrating its immunogenicity. Moreover, mice immunized with recombinant S9 protein and challenged with a virulent strain of S. pneumoniae presented a significant reduction of bacteremia after 24 h of infection as compared with the control. However, in the S9-immunized mice the onset of death was insignificantly delayed, but all of them died by the fourth day postinfection.

Impact of vector control actions in the abundance of Lutzomyia longipalpis in Montes Claros, Brazil.

Visceral leishmaniasis (VL) is endemic to many states in Brazil. To prevent further expansion of the disease, the Brazilian Ministry of Health adopted integrated measures through the Program of Surveillance and Control of Visceral Leishmaniasis (PSCVL), whose actions include the diagnosis and euthanasia of seropositive dogs (the main domestic reservoirs), the use of residual insecticides, environmental management (EM) to control vector population (mainly Lutzomyia longipalpis phlebotomine), rigorous epidemiological surveillance, and health education. The present study was conducted in areas with recent moderate VL transmission to evaluate the efficacy of vector control activities. The systematic capture of phlebotomine was performed for three consecutive days per month, from August 2015 to July 2017. The number of specimens captured was taken as a representative of the monthly insect population. A total of 38,055 phlebotomine specimens were captured and identified at the species level. Lu. longipalpis was consistently found to be the predominant species (97.7%) each month. In the first year of the study, no intervention was performed. In the second year, two cycles of chemical spraying, EM, or a combination of both were performed before and after the rainy season. All interventions, either individually or in combination, reduced the abundance of Lu. longipalpis in the study area.

Varespladib (LY315920) prevents neuromuscular blockage and myotoxicity induced by crotoxin on mouse neuromuscular preparations.

Varespladib (LY315920) is a synthetic phospholipase A2 (PLA2) inhibitor that has been demonstrating antiophidic potential against snake venoms that present PLA2 neurotoxins. In this study, we evaluate the capacity of Varespladib to inhibit the neuromuscular effects of crotoxin (CTX), the main toxic component of Crotalus durissus terrificus snake venom, and its PLA2 subunit (CB). We performed a myographic study to compare the neuromuscular effects of CTX or CB and the mixture of these substances plus Varespladib in mice phrenic nerve-diaphragm muscle preparations. CTX (5 µg/mL), CB (20 µg/mL), or toxin-inhibitor mixtures pre-incubated with different concentration ratios of Varespladib (1:0.25; 1:0.5; 1:1; w/w) were added to the preparations and maintained throughout the experimentation period. Myotoxicity was assessed by light microscopic analysis of diaphragm muscle after myographic study. CTX and CB blocked the nerve-evoked twitches, and only CTX induced histological alterations in diaphragm muscle. Pre-incubation with Varespladib abolished the muscle-paralyzing activity of CTX and CB, and also the muscle-damaging activity of CTX. These findings emphasize the clinical potential of Varespladib in mitigating the toxic effects of C. d. terrificus snakebites and as a research tool to advance the knowledge of the mechanism of action of snake toxins.

Crotalus Neutralizing Factor (CNF) inhibits the toxic effects of Crotoxin at mouse neuromuscular preparations.

Crotalus Neutralizing Factor (CNF) was the first phospholipase A2 inhibitor isolated from the plasma of the South American rattlesnake (Crotalus durissus terrificus). Previous biochemical and biophysical studies demonstrate an interaction of CNF with Crotoxin (CTX), the main toxic component in the venom of these snakes. CTX promotes the blockade of neuromuscular transmission by a sum of neurotoxic and myotoxic activities. However, the ability of CNF to inhibit these activities has not been shown until the present study. We performed a myographic study to compare the neuromuscular effects of CTX and the mixture CTX plus CNF in mice phrenic nerve-diaphragm muscle preparations. CTX (5 µg/mL) alone, or pre-incubated with CNF (5, 20 or 50 µg/mL) for 15 min was added to the preparations and maintained throughout the experimentation period. Myotoxicity was assessed by light microscopic analysis of diaphragm muscle after myographic study. CTX (5 µg/mL) blocked both indirectly and directly evoked twitches in neuromuscular preparations. In addition, CTX induced histological alterations in diaphragm muscle. Pre-incubation with CNF (50 µg/mL) abolished both the muscle-paralyzing and muscle-damaging activities of CTX. Therefore, the present study confirms, through functional studies, the antiophidic potential of CNF.

Evaluation of the vectorial capacity of Rhipicephalus sanguineus (Acari: Ixodidae) in the transmission of canine visceral leishmaniasis.

The vectorial capacity of Rhipicephalus sanguineus in the transmission of canine visceral leishmaniasis has been evaluated through a laboratory-controlled experiment. One healthy Leishmania-free dog and two dogs naturally infected with Leishmania were infested with R. sanguineus in various stages of development. Engorged larvae, unfed nymphs, engorged nymphs, unfed adults, engorged female adults and fed male adults were collected from the experimental animals and examined for Leishmania infection by optical microscopy, polymerase chain reaction (PCR) and parasite culture. Leishmania forms were not detected in any of the 433 smears prepared from engorged colonies nor in any of the 118 smears prepared from unfed colonies. However, one flagellate structure was identified in one of the smears. All pools of R. sanguineus that had fed on the infected dogs tested PCR-positive for Leishmania DNA, with the single exception of the pool of engorged larvae. In contrast, all pools of ticks that had fed on the Leishmania-free dog were PCR-negative. Leishmania growth was not observed in any of the tick colonies following incubation on culture medium. Considering that no Leishmania forms were identified in any of the meticulously analysed smears derived from engorged colonies of R. sanguineus, it appears somewhat unlikely that the maintenance and multiplication of Leishmania occurs within the tick.

Interaction of Biomphalaria cells in primary cultures with Schistosoma mansoni sporocysts.

INTRODUCTION: Biomphalaria snails may display varying levels of susceptibility to Schistosoma mansoni infection. We have been developing an in vitro model to study the interaction between the snail and the parasite, using tissue-derived cell cultures from Biomphalaria. METHODS: The digestive gland- and kidney-derived cells from primary cultures of resistant (B. tenagophila Taim) and susceptible (B. tenagophila HM and B. glabrata BH) strains of Biomphalaria were exposed to S. mansoni sporocysts. RESULTS: S. mansoni sporocysts were surrounded and encapsulated exclusively by cells derived from the digestive gland (DG) of B. tenagophila Taim. The process was followed by a marked decrease in the number of free sporocysts in the culture medium. The morphological characteristics of DG-derived cells in culture have been described. CONCLUSIONS: Cells derived from DG (but not SK) primary cultures of B. tenagophila Taim may participate in S. mansoni sporocyst control.

Ecology of phlebotomine sand flies in a Brazilian area with recent leishmaniasis transmission (Itaúna, in Minas Gerais state).

INTRODUCTION: Leishmaniasis constitutes a serious but neglected tropical disease. Recently, socio-environmental, biological and physical changes have altered the range of leishmaniasis, causing it to spread into urban areas. In Minas Gerais, the disease is endemic, exhibiting regional differences and reaching urban centers. The purpose of this study was to evaluate entomological aspects related to the ecoepidemiology of leishmaniasis in Itaúna. METHODS: Monthly catches with HP traps were carried out from June 2017 to May 2018, in three ecological areas (urban, rural, and forest). The adaptability of the species to anthropic environments was assessed using the synanthropy index (SI). RESULTS: We collected 1306 specimens of phlebotomine sand flies. Of the species of medical importance, Lutzomyia longipalpis, the vector of Leishmania infantum, represented 90.4% of the specimens identified at species level (n=1260). Nyssomyia whitmani, the vector of Leishmania braziliensis, represented 1.6% of the total. Lu. longipalpis displayed an SI of +95.8, a value that denotes a marked preference for human environments. For Ny. whitmani, the SI was -25, expressing the tendency of this species to occupy uninhabited areas. The population of the three most numerous species captured increased with rain, high temperatures, and high relative humidity. Although captured at low numbers, Ny. whitmani showed a different profile concerning the climate variables analyzed. CONCLUSIONS: Understanding the epidemiology of the disease may assist the health services in formulating effective control measures for improving community health and contributing to the establishment of a dynamic relationship and a global awareness of the health/disease process.

Crotalus neutralising factor and its role in human leukocyte modulation.

Crotalus neutralising factor (CNF) is an endogenous γ-type phospholipase A2 (PLA2) inhibitor that inhibits the toxic action of crotoxin, a neurotoxin present in Crotalus durissus terrificus venom. However, its effects on the activation and modulation of immune cells, which play a major role in the development of inflammation, is not known. The objective of the present study was to assess the effects of CNF on human leukocyte modulation in vitro by analysing the following parameters: cell viability, phagocytic capacity, lipid droplet formation, reactive oxygen species production, nitric oxide production, p38 MAPK activation, and cytosolic PLA2 (cPLA2) gene expression. Neutrophils and peripheral blood mononuclear cells from healthy donors were isolated via the density gradient method, resuspended in RPMI medium, and incubated with RPMI (negative control), LPS, or PMA (positive control) or CNF (sample test) at a concentration of 50 µg/mL. Results showed that CNF was not toxic to human neutrophils after 48 and 72 h of incubation. CNF treatment induced an increase in PBMCs and neutrophil phagocytic capacity, as well as the formation of lipid droplets within these cells after 1 h of incubation. However, CNF did not induce the formation of reactive oxygen and nitric oxide species. Moreover, CNF induced p38 MAPK protein phosphorylation and cPLA2 gene expression in neutrophils. The data obtained herein showed that CNF action modulates human leukocytes, CNF activates important signalling pathways for human leukocytes, and it is pro-inflammatory. These findings also complement previous studies on CNF action on human peripheral blood leukocyte function.

Dogs with divergent serology for visceral leishmaniasis as sources of Leishmania infection for Lutzomyia longipalpis phlebotomine sand flies - an observational study in an endemic area in Brazil.

Visceral leishmaniasis (VL) is a neglected tropical disease, caused by Leishmania (Kinetoplastida, Trypanosomatidae) species. In Brazil, the transmission of this parasite essentially occurs through the bite of Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae) previously infected with Leishmania infantum. Aiming at preventing VL expansion over the country, integrated control actions have been implemented through a Visceral Leishmaniasis Surveillance and Control Program (VLSCP). Among the actions currently adopted by the program, the screening-culling of seropositive dogs for canine VL (CVL) is particularly polemic. Dogs with negative or divergent serology for CVL remain in their owner's domicile and are monitored by public health agents. In the present study, we determined the prevalence of CVL and analyzed the implementation of the VLSCP screening-culling action, in an area in Brazil where there has been a recent expansion of VL. Canine census surveys were conducted semiannually for two years (Aug/2015 to Feb/2017). Serological diagnosis of CVL was performed in accordance with current VLSCP protocol: immunochromatography (TR-DPP) followed by enzyme-linked immunosorbent assay (ELISA EIE). 6,667 dogs were serologically screened for CVL, of which 567 (8.5%) were positive in both tests and 641 (9.6%) had divergent results. A variable percentage (6.3% to 65.4%) of the dogs in the latter group became positive within nine months from the first result. Xenodiagnosis was conducted in canine samples belonging to any of the three possible serological statuses for CVL-positive, divergent or negative. Leishmania spp. DNA was detected in Lu. longipalpis that fed on 50.0% (5/10) of dogs with positive serology and on 29.4% (5/17) of dogs with divergent serological status for CVL. Therefore, dogs with divergent serology for CVL may be as Leishmania-infective to Lu. longipalpis as seropositive ones. Even with the adoption of euthanasia for seropositive dogs, part of the canine population will continue to serve as a source of Leishmania infection for phlebotomine sand flies.

Canine visceral leishmaniasis in area with recent Leishmania transmission: prevalence, diagnosis, and molecular identification of the infecting species.

INTRODUCTION: Canine visceral leishmaniasis (CVL) is an endemic disease in Brazil, and integrated control actions have been adopted by the Brazilian Ministry of Health to control its spread. However, the transmission profile is unknown in areas with recent CVL cases, including Itaúna, located in the Brazilian state of Minas Gerais, where the present study was carried out. METHODS: A total of 2,302 dogs from 12 neighborhoods were serologically tested for canine VL using the current diagnostic protocol adopted by the Brazilian Ministry of Health. Test positivity rate (TPR) and CVL prevalence were determined for each neighborhood. The presence of Leishmania was assessed in 60 seropositive dogs which had been recommended for euthanasia. Twenty-two of them (37%) were asymptomatic, and 38 (63%) were symptomatic for CVL. Parasitological (myeloculture and smear/imprint) and molecular (PCR) methods were employed for Leishmania detection in bone marrow, spleen, mesenteric lymph nodes, and ear skin. The infecting Leishmania species was identified by DNA sequencing. RESULTS: CVL prevalence (per 1,000 dogs) varied from 0.0-166.67, depending on the neighborhood, with a mean of 68.96 (SD 51.38). Leishmania DNA was detected in at least one tissue from all seropositive dogs, with comparable TPR among tissues. Leishmania parasites were identified in most (54/60) seropositive dogs, and the infecting parasite was identified as Leishmania infantum in all of these. CONCLUSIONS: Prevalence of CVL is a contributor to the spread of visceral leishmaniasis in Itaúna.

Association of Lutzomyia longipalpis (Diptera: Psychodidae) population density with climate variables in Montes Claros, an area of American visceral leishmaniasis transmission in the state of Minas Gerais, Brazil.

In the present paper, we evaluate the relationship between climate variables and population density of Lutzomyia longipalpis in Montes Claros, an area of active transmission of American visceral leishmaniasis (AVL) in Brazil. Entomological captures were performed in 10 selected districts of the city, between September 2002-August 2003. A total of 773 specimens of L. longipalpiswere captured in the period and the population density could be associated with local climate variables (cumulative rainfall, average temperature and relative humidity) through a mathematical linear model with a determination coefficient (Rsqr) of 0.752. Although based on an oversimplified statistical analysis, as far as the vector is concerned, this approach showed to be potentially useful as a starting point to guide control measures for AVL in Montes Claros.

Phlebotominae distribution in Janaúba, an area of transmission for visceral leishmaniasis in Brazil.

In Brazil, visceral leishmaniasis (VL) is caused by Leishmania chagasi parasites that are transmitted to man through the bites of infected females of Lutzomyia longipalpis sand flies. In order to evaluate transmission risk and to clarify the epidemiology of this tropical disease, studies focused on the vector and favorable environmental conditions are of fundamental importance. In this work, we surveyed the phlebotomine sand fly fauna in Janaúba, a Brazilian municipality that is endemic for VL. During a two-year period, entomological captures were performed monthly in 15 districts with high, moderate and low profiles of VL transmission. A total of 14,591 phlebotomine sand flies were captured (92% L. longipalpis), with a predominance of males. Most specimens were captured in the peri-domicile setting, although the number of specimens captured in the intra-domicile setting emphasises the anthropophilic behaviour of this insect. The population density of L. longipalpis was modulated by climate variations, particularly with clear increases immediately after the rainy season. However, the pattern of distribution did not coincide with the occurrence of human or canine cases of VL. This suggests that the eco-epidemiology of VL is particular to each area of transmission and must be taken into account during the design of public health control actions.