Spiders of the genus Loxosceles, popularly known as Brown spiders, are considered a serious public health issue, especially in regions of hot or temperate climates, such as parts of North and South America. Although the venoms of these arachnids are complex in molecular composition, often containing proteins with distinct biochemical characteristics, the literature has primarily described a family of toxins, the Phospholipases-D (PLDs), which are highly conserved in all Loxosceles species. PLDs trigger most of the major clinical symptoms of loxoscelism i.e., dermonecrosis, thrombocytopenia, hemolysis, and acute renal failure. The key role played by PLDs in the symptomatology of loxoscelism was first described 40 years ago, when researches purified a hemolytic toxin that cleaved sphingomyelin and generated choline, and was referred to as a Sphingomyelinase-D, which was subsequently changed to Phospholipase-D when it was demonstrated that the enzyme also cleaved other cellular phospholipids. In this review, we present the information gleaned over the last 40 years about PLDs from Loxosceles venoms especially with regard to the production and characterization of recombinant isoforms. The history of obtaining these toxins is discussed, as well as their molecular organization and mechanisms of interaction with their substrates. We will address cellular biology aspects of these toxins and how they can be used in the development of drugs to address inflammatory processes and loxoscelism. Present and future aspects of loxoscelism diagnosis will be discussed, as well as their biotechnological applications and actions expected for the future in this field.

Studies were carried out on the recovery rate and cysticerci location in bovines experimentally infected with Taenia saginata eggs. Three calves of 6.5 months and one with 19 months of age were infected orally with 2 x 10(4) eggs of Taenia saginata. A fifth calf served as control. After 90 days of infection, the animals were slaughtered and organs and skeletal muscles were inspected using a slicing technique every 5 mm. From the four infected calves, 702 cysticerci were recovered, of which 570 (81.2%) were alive and 132 (18.8%) were degenerated. The recovery rate ranged from 0.01 to 1.43% with an average of 0.88%. The cysticerci presented the following anatomical distribution: hioideos muscles 02 (0.28%), kidneys 03 (0.43%), tongue 07 (1.00%), liver 12 (1.71%), lungs 15 (2.14%), diaphragm 18 (2.56%), mastication muscles 25 (3.56%), heart 49 (6.98%), anterior muscle 323 (46.00%) and posterior muscle 248 (35.33%).
Foi avaliada a taxa de recuperação e localização de cisticercos em bovinos experimentalmente infectados com ovos de Taenia saginata. Três bezerros de 6,5 meses e um adulto com 19 meses de idade foram infectados, por via oral, com 2 x 10(4) ovos de Taenia saginata. Um quinto bezerro serviu como testemunha. Após 90 dias da infecção, os animais foram abatidos. Fez-se inspeção, de todos os animais, por fatiamento de órgãos e musculatura esquelética, com intervalo entre os cortes de, no máximo, cinco milímetros. Dos quatros bezerros desafiados foram recuperados 702 cisticercos sendo 570 (81,20%) vivos e 132 (18,80%) degenerados. A taxa de recuperação foi de 0,01% a 1,43% com média de 0,88%. Os 702 cistos encontrados apresentaram a seguinte distribuição anatômica: músculos hióideos 02 (0,28%), rins 03 (0,43%), língua 07 (1,00%), fígado 12 (1,71%), pulmões 15 (2,14%), diafragma 18 (2,56%), músculos da mastigação 25 (3,56%), coração 49 (6,98%), musculatura dianteira 323 (46,00%) e musculatura traseira 248 (35,33%). Na infecção experimental os cistos encontraram-se distribuídos por toda a musculatura dos animais, não mostrando predileção pelos tecidos normalmente pesquisados pelo serviço de inspeção (língua, coração, diafragma, músculos mastigatórios). Os dados da inspeção de rotina pode não estimar a real incidência da cisticercose bovina. Os bovinos adultos são mais resistentes a infecção por ovos de Taenia saginata, apresentando menor número de cisticercos e com maior número de cistos calcificados

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Bites by the brown spider (Loxosceles spp.) are an important health problem in South America, where three species predominate (Loxosceles laeta, Loxosceles gaucho, Loxosceles intermedia). Brown spider bites (loxoscelism) induce a block of cutaneous necrosis and, less commonly, may cause fatal systemic poisoning. A variety of controversial protocols are used to treat loxoscelism, while treatment with antivenin is the only venom specific treatment. Here we studied the action of the venom as well as the response to the antivenin for Loxosceles through an experimental study that simulates bites of L. intermedia (bites of this species are the most common in Brazil). Beneficial effects are known for antivenin applied quickly (within 4 h) after envenomation. Here we wished to examine the temporal development of the brown spider bite as well as the temporal patterns of the action of the antivenin to determine the time limits for beneficial use of the antivenin after envenomation. This information is important since most patients only appear for treatment several hours after being bitten. New Zealand rabbits were experimentally exposed to the venom from brown spiders by the injection of venom from L. intermedia (2x minimum necrotic dose), followed at regular time intervals by antivenin. The use of the loxoscelic antivenin--CPPI (4 mL per animal)--minimized the effects of envenomation when applied for up to 12 h after the injection of the venom, as evaluated by cutaneous (erythrema, edema, ecchymosis and necrosis) and systemic (blood cell and platelet counts, hematimetrics and fibrinogen dosage) criteria. Also, antivenin reduced the size of the necrotic area when applied up to 48 h after envenomation. Thus, therapy with loxoscelic antivenin, CPPI, may provide beneficial results by interfering with envenomation well after the bite occurred and therefore may become an important tool for medical treatment of brown spider bites.

A chimeric protein was constructed expressing three epitopes of LiD1, a dermonecrotic toxin from the venom of Loxosceles intermedia spider. This species is responsible for a large number of accidents involving spiders in Brazil. We demonstrated that the chimeric protein (rCpLi) generated is atoxic and that antibodies previously developed in rabbits against synthetic epitopes reactive with rCpLi in ELISA and immunoblot assays. The antibody response in rabbits against the rCpLi was evaluated by ELISA and we have detected an antibody response in all immunized animals. Overlapping peptides covering the amino acid sequence of the rCpLi were synthesized on a cellulose membrane, and their recognition by rabbit anti-rCpLi serum assessed. Three different antigenic regions were identified. The percentage of inhibition of the dermonecrotic, hemorrhagic and edematogenic activities caused by the recombinant protein LiD1r in naïve rabbits was assessed by pre-incubation with anti-rCpLi antibodies. Anti-rCpLi induced good dermonecrotic and hemorrhagic protection. The levels of protection were similar to the antiboides anti-LiD1r. In summary, we have developed a polyepitope recombinant chimeric protein capable of inducing multiple responses of neutralizing antibodies in a rabbit model. This engineered protein may be a promising candidate for therapeutic serum development or vaccination.