Reproducibility of double agar gel immunodiffusion test using stored serum and plasma from paracoccidioidomycosis patients

Background: Serological evaluation performed by double agar gel immunodiffusion test (DID) is used for diagnosis, evaluation of severity, management of paracoccidioidomycosis patients, and development of new clinical studies. For these reasons, the Botucatu Medical School of UNESP maintains a serum bank at the Experimental Research Unit with patient clinical data. This study aimed to evaluate the influence of the freeze-thaw cycle and different blood matrices on the titration of circulating antibodies. Methods: The study included 207 patients with confirmed (etiology-demonstrated) or probable (serology-demonstrated) paracoccidioidomycosis, and DID was performed with culture filtrate from Paracoccidioides brasiliensis B339 as antigen. First experiment: the antibody levels were determined in serum samples from 160 patients with the chronic form and 20 with the acute/subacute form, stored at ­80o C for more than six months. Second experiment: titers of 81 samples of serum and plasma with ethylenediaminetetraacetic acid (EDTA) or heparin, from 27 patients, were compared according to matrix and effect of storage at ­20o C for up to six months. Differences of titers higher than one dilution were considered discordant. Results: First experiment: test and retest presented concordant results in serum stored for up to three years, and discordant titers in low incidence in storage for four to six years but high incidence when stored for more than six years, including conversion from reagent test to non-reagent retest. Second experiment: serum, plasma-EDTA and plasma-heparin samples showed concordant titers, presenting direct correlation, with no interference of storage for up to six months. Conclusions: Storage at ­80o C for up to six years has no or little influence on the serum titers determined by DID, permitting its safe use in studies depending on this parameter. The concordant titrations in different blood matrices demonstrated that the plasma can be used for immunodiffusion test in paracoccidioidomycosis, with stability for at least six months after storage at ­20o C.(AU)

Bio-efficacy of iron and zinc fortified wheat flour along with bio-assessment of its hepatic and renal toxic potential / Bioeficácia da farinha de trigo fortificada com ferro e zinco e bioavaliação do seu potencial tóxico hepático e renal

Study was planned to assess the bio-efficiency along with toxicity of iron and zinc fortified whole wheat flour in Sprague dawley albino rats. Whole wheat flour was fortified with different dosage of sodium iron EDTA (NaFeEDTA), ferrous sulphate (FeSO4), zinc oxide (ZnO) and zinc sulphate (ZnSO4). The rats (n=3) in each group were fed on fortified wheat flour for 2 months. Liver biomarkers including alkaline phosphatase (ALP), alanine transaminase (ALT), aspartate aminotransferase (AST) and bilirubin were recorded from serum samples. Increased concentration of ZnSO4 affected the liver biomarkers to be highest among all whereas, bilirubin levels were less than the rats fed on control diet. The above mentioned fortificants have negligible effect on renal biomarkers including creatinine and urea. Moreover, hematological parameters were also checked and reportedly, sodium iron EDTA fed rats presented highest amount of hemoglobin, iron and total iron binding capacity. Highest zinc level was observed in rats fed on whole wheat flour fortified with 60mg/Kg Zinc oxide. Microscopic observation of liver tissue depicted that rats fed on iron and zinc fortified wheat flour have more toxic effects whereas, histopathology presentation of kidney tissue has least toxic impact. It has been concluded that mandatory fortification of wheat flour with iron and zinc may cause increased serum biomarkers along with toxicity of vital organs like liver, hence fraction of wheat flour may be fortified to fulfill the requirements of deprived and vulnerable group.

O estudo foi planejado para avaliar a bioeficiência juntamente com a toxicidade da farinha de trigo integral fortificada com ferro e zinco em ratos albinos Sprague dawley. A farinha de trigo integral foi fortificada com diferentes dosagens de ferro sódico EDTA (NaFeEDTA), sulfato ferroso (FeSO4), óxido de zinco (ZnO) e sulfato de zinco (ZnSO4). Os ratos (n = 3) de cada grupo foram alimentados com farinha de trigo fortificada por 2 meses. Biomarcadores hepáticos incluindo fosfatase alcalina (ALP), alanina transaminase (ALT), aspartato aminotransferase (AST) e bilirrubina foram registrados a partir de amostras de soro. O aumento da concentração de ZnSO4 afetou os biomarcadores hepáticos como sendo os mais altos entre todos, enquanto os níveis de bilirrubina foram menores do que os ratos alimentados com dieta controle. Os fortificantes mencionados acima têm efeito insignificante nos biomarcadores renais, incluindo creatinina e ureia. Além disso, os parâmetros hematológicos também foram verificados e, segundo relatos, os ratos alimentados com EDTA de sódio e ferro apresentaram maior quantidade de hemoglobina, ferro e capacidade total de ligação de ferro. O maior nível de zinco foi observado em ratos alimentados com farinha de trigo integral fortificada com 60 mg/Kg de óxido de zinco. A observação microscópica do tecido hepático mostrou que ratos alimentados com farinha de trigo fortificada com ferro e zinco têm mais efeitos tóxicos, enquanto a apresentação histopatológica do tecido renal tem menos impacto tóxico. Concluiu-se que a fortificação obrigatória da farinha de trigo com ferro e zinco pode causar aumento dos biomarcadores séricos juntamente com toxicidade de órgãos vitais como o fígado, portanto a fração da farinha de trigo pode ser fortificada para atender os requisitos do grupo carente e vulnerável.

Effects of chelating calcium in cryopreservation extender on frozen-thawed dog semen / [Efeitos da quelação de cálcio no meio de criopreservação no sêmen descongelado de cão]

We evaluated the effect of reducing free calcium in the cryopreservation medium, using the calcium chelator ethylene diamine tetracetic acid (EDTA) at 0.3% and 0.5% concentrations. Three male mixed breed dogs were subjected to semen collection by digital manipulation (n=16). Each ejaculate was divided in three aliquots, and each one was diluted in TRIS-glucose-egg yolk extender with 6% glycerol and 0.5% Equex STM Paste® (TGE, control); and added with 0.3% EDTA (EDTA 0.3) or 0.5% EDTA (EDTA 0.5). Calcium concentration reduced in EDTA 0.3 and all the calcium ions were chelated in EDTA 0.5. The EDTA addition did not affect sperm morphology or plasma membrane integrity; however, by removing all free calcium (EDTA 0.5), the sperm motility reduced (64.7% in TGE and 45% in EDTA 0.5; p<0.05). Acrosome integrity and sperm binding ability were not improved by calcium chelation. The failure to prevent the premature AR may explain why sperm longevity was not affected by calcium removal. Thus, the partial or complete calcium removal, through EDTA addition, is not able to prevent acrosomal damage or premature acrosomal reaction, and therefore does not improve the dog sperm binding ability.(AU)

Avaliou-se o efeito da redução do cálcio livre no meio de congelamento, usando-se o quelante de cálcio etilenodiaminotetracético (EDTA) a 0,3% e 0,5%. Três cães machos sem raça definida foram submetidos à coleta de sêmen por manipulação digital (n=16). Cada ejaculado foi diluído em diluidor controle com TRIS-glicose - gema de ovo (TGE, controle), ou em diluidor TGE enriquecido com 0,3% (EDTA 0,3) ou 0,5% de EDTA (EDTA 0,5). A concentração de cálcio reduziu no meio EDTA 0,3, e todos os íons de cálcio foram quelados no meio EDTA 0,5. A adição do EDTA e a consequente quelação do cálcio não afetaram a morfologia espermática ou a integridade da membrana plasmática, no entanto, ao remover todo o cálcio do meio (EDTA 0,5), a motilidade espermática se reduziu (64,7% no TGE e 45% no EDTA 0,5; P<0,05). A integridade do acrossoma e a capacidade de ligação do espermatozoide não melhoraram com a quelação do cálcio. Apesar da influência da concentração de cálcio sobre a motilidade espermática após o descongelamento, a falha em prever a reação acrossomal prematura pode explicar por que a longevidade espermática não foi afetada pela remoção do cálcio no meio. Dessa forma, a remoção parcial ou total do cálcio, por meio da adição de EDTA, não é capaz de prevenir o dano no acrossoma ou a reação acrossomal prematura e, portanto, não aumenta a capacidade do espermatozoide de se ligar ao oócito.(AU)

The epiphyseal plate closure phenomena for male and female Japanese quail ( Coturnix coturnix japonica ): histological and biochemical alterations

This study was conducted to determine the effect of sex on ossification processes in quail by determining the exact closure timing of the epiphyseal plate in the proximal region of the femur. This was done by investigating the histological and biochemical parameters affecting the osteogenesis process that takes place following quail hatching to observe if any variation existed between males and females in this regard. For this purpose, blood samples were collected from six male and female specimens via IV catheters every week for the first 42 days that followed hatching. The samples were transferred into serum tubes, and PTH (PTH), triiodothyronine (T3), thyroxine (T4), inorganic phosphorus, calcium, and vitamin D values in the samples, which are known to have an impact on ossification, were analyzed. The specimens from which the blood samples were collected were then euthanized, and histological cut-sections that covered the epiphyseal growth plate were collected, along with the bone sections of the proximal regions of the right femur. Considering decalcification, these histological sections were kept in an ethylene diamine tetra acetic acid (EDTA) solution. Routine histological examinations were then conducted on these sections, after which they were embedded in paraffin. Crossman's modified triple staining method was used to prepare them, and Wilcoxon Signed Ranks Test was used to statistically evaluate whether the inspected biochemical parameters played a role in the ossification process of quail and whether a statistical difference existed between sexes in this regard. The findings of our study revealed that poultry animals also have five zones in the epiphyseal plate as do mammals, and they have calcified cartilage areas. The findings also indicate that, while ossification starts to occur both in male and female quail specimens, calcification occurs more frequently in females. It was determined that the proximal epiphyseal plate regions of the femur close at the end of the sixth week in both sexes of quail, although the calcification and ossification are more advanced in some females compared with males.(AU)

Use of EDTA in the treatment of local tissue damage caused by the Bothrops alternatus venom / Utilização do EDTA no tratamento da lesão tecidual local causado pelo veneno de Bothrops alternatus

Twelve adult rabbits were distributed in three groups and received on the femoral biceps region, via intradermal injection (ID), 25µg of Bothrops alternatus venom dissolved in NaCl 0.9% and diluted in 0.25mL of phosphate buffered saline (PBS). Thirty minutes later, the group G1 received 0.25mL of phosphate buffered saline (PBS) ID while to G2 and G3 25mg of ethylenediamine tetraacetic acid (EDTA) dissolved in 0.25mL of PBS were administered via intramuscular (IM) and intravenous (IV) injection, respectively. Evaluations included local lesion and blood profile of all animals, before (time zero) and at 1, 2, 3, 4, 5, 6, 12, 18 and 24h after venom administration. All animal treated with PBS (G1) and EDTA IV (G3) presented increase of nociceptive stimuli at the site of inoculation of the venom, followed by moderate edema that persisted for 24h. Animals treated with IM EDTA (G2) only manifested increase of nociceptive stimuli at the site of injection 1h after treatment with discrete local edema between 12 and 24h. In relation to the local hemorrhagic halo no differences were found amongst the studied groups. Blood profile revealed significant decrease of segmented neutrophils in all groups. There was also increase in triglycerides and decrease in total protein and albumin in all groups. The local lesion was not altered by the treatments.(AU)

Doze coelhos adultos, distribuídos em três grupos, receberam, na região de bíceps femoral, por via intradérmica (ID), 25µg de veneno de Bothrops alternatus, dissolvidos em NaCl 09%, diluído em 0,25mL de tampão salina fosfato (PBS). Trinta minutos após o desafio, o grupo G1 recebeu 0,25mL de (PBS) ID, e os grupos G2 e G3 receberam 25mg de ácido etilenodiamino tetra-acético (EDTA), dissolvidos em mL de PBS por via intramuscular (IM) e intravenosa (IV), respectivamente. Foram avaliados lesão local e perfil sanguíneo de todos os animais, antes - tempo zero, e à uma, às duas, três, quatro, cinco, seis, 12, 18 e 24 horas após a injeção do veneno. Tanto os animais tratados com PBS (G1) como os animais tratados com EDTA IV (G3) apresentaram aumento do estímulo nociceptivo no local da administração do veneno, seguido por moderado edema, que perdurou por 24h. Os animais tratados com EDTA IM (G2) somente manifestaram aumento do estímulo nociceptivo local uma hora após tratamento e discreto edema local entre 12 e 24 horas. Em relação ao halo hemorrágico, não houve diferença entre os três grupos estudados. No perfil hematológico, observou-se diminuição significativa dos neutrófilos segmentados nos três grupos estudados. Da mesma forma, houve aumento dos triglicerídeos e diminuição da proteína total e albumina em todos os grupos. Conclui-se que a lesão local não foi alterada pelos tratamentos.(AU)

Use of EDTA in the treatment of local tissue damage caused by the Bothrops alternatus venom / Utilização do EDTA no tratamento da lesão tecidual local causado pelo veneno de Bothrops alternatus

Twelve adult rabbits were distributed in three groups and received on the femoral biceps region, via intradermal injection (ID), 25µg of Bothrops alternatus venom dissolved in NaCl 0.9% and diluted in 0.25mL of phosphate buffered saline (PBS). Thirty minutes later, the group G1 received 0.25mL of phosphate buffered saline (PBS) ID while to G2 and G3 25mg of ethylenediamine tetraacetic acid (EDTA) dissolved in 0.25mL of PBS were administered via intramuscular (IM) and intravenous (IV) injection, respectively. Evaluations included local lesion and blood profile of all animals, before (time zero) and at 1, 2, 3, 4, 5, 6, 12, 18 and 24h after venom administration. All animal treated with PBS (G1) and EDTA IV (G3) presented increase of nociceptive stimuli at the site of inoculation of the venom, followed by moderate edema that persisted for 24h. Animals treated with IM EDTA (G2) only manifested increase of nociceptive stimuli at the site of injection 1h after treatment with discrete local edema between 12 and 24h. In relation to the local hemorrhagic halo no differences were found amongst the studied groups. Blood profile revealed significant decrease of segmented neutrophils in all groups. There was also increase in triglycerides and decrease in total protein and albumin in all groups. The local lesion was not altered by the treatments.(AU)

Doze coelhos adultos, distribuídos em três grupos, receberam, na região de bíceps femoral, por via intradérmica (ID), 25µg de veneno de Bothrops alternatus, dissolvidos em NaCl 09%, diluído em 0,25mL de tampão salina fosfato (PBS). Trinta minutos após o desafio, o grupo G1 recebeu 0,25mL de (PBS) ID, e os grupos G2 e G3 receberam 25mg de ácido etilenodiamino tetra-acético (EDTA), dissolvidos em mL de PBS por via intramuscular (IM) e intravenosa (IV), respectivamente. Foram avaliados lesão local e perfil sanguíneo de todos os animais, antes - tempo zero, e à uma, às duas, três, quatro, cinco, seis, 12, 18 e 24 horas após a injeção do veneno. Tanto os animais tratados com PBS (G1) como os animais tratados com EDTA IV (G3) apresentaram aumento do estímulo nociceptivo no local da administração do veneno, seguido por moderado edema, que perdurou por 24h. Os animais tratados com EDTA IM (G2) somente manifestaram aumento do estímulo nociceptivo local uma hora após tratamento e discreto edema local entre 12 e 24 horas. Em relação ao halo hemorrágico, não houve diferença entre os três grupos estudados. No perfil hematológico, observou-se diminuição significativa dos neutrófilos segmentados nos três grupos estudados. Da mesma forma, houve aumento dos triglicerídeos e diminuição da proteína total e albumina em todos os grupos. Conclui-se que a lesão local não foi alterada pelos tratamentos.(AU)

Heavy metal poisoning in a cockatiel (Nymphicus hollandicus)

Background: In recent decades the demand for unconventional pets has been relatively increasing, a situation that increasingly causes veterinarians to encounter these animals in medical and surgical practice. Of these animals, the birds stand out. Animals of the order Psittaciform are known as very curious and active creatures that have the tendency to chew objects in their environment. Among the several occurrences that lead this animal to attend the veterinary clinic, we highlight the poisoning by heavy metals, especially lead poisoning (Pb) and zinc (Zn). The objective of this work was to report a case of heavy metal intoxication in cockatiel (Nymphicus hollandicus).Case: A cockatiel was taken to the veterinarian with a history of apathy, motor incoordination, exacerbated water consumption and regurgitation. Complete anamnesis was instrumental in directing suspected heavy metal intoxication. Radiopaque particles were visualized through radiographic examination, suggesting heavy metal intoxication. The diagnosis was concluded through complementary examinations since the clinical symptoms are nonspecific. The treatment was intended to provide emergency intervention, avoid further absorption, use of antidotes, provide supportive measures and provide guidance to the owner. It can be concluded that the diagnosis and treatment were successful.Discussion: Metal poisoning can kill birds. The veterinarian should always seek the literature in order to perform the best support and treatment. For this, detailed history and detailed medical history must be taken into account, since the time of ingestion and the type of metal interfere with the therapeutic conduct. The use of imaging tests such as x-rays and ultrasound are essential to assist the clinician, especially in cases where the tutor does not know whether or not the animal has ingested an object.[...]

Heavy metal poisoning in a cockatiel (Nymphicus hollandicus)

Background: In recent decades the demand for unconventional pets has been relatively increasing, a situation that increasingly causes veterinarians to encounter these animals in medical and surgical practice. Of these animals, the birds stand out. Animals of the order Psittaciform are known as very curious and active creatures that have the tendency to chew objects in their environment. Among the several occurrences that lead this animal to attend the veterinary clinic, we highlight the poisoning by heavy metals, especially lead poisoning (Pb) and zinc (Zn). The objective of this work was to report a case of heavy metal intoxication in cockatiel (Nymphicus hollandicus).Case: A cockatiel was taken to the veterinarian with a history of apathy, motor incoordination, exacerbated water consumption and regurgitation. Complete anamnesis was instrumental in directing suspected heavy metal intoxication. Radiopaque particles were visualized through radiographic examination, suggesting heavy metal intoxication. The diagnosis was concluded through complementary examinations since the clinical symptoms are nonspecific. The treatment was intended to provide emergency intervention, avoid further absorption, use of antidotes, provide supportive measures and provide guidance to the owner. It can be concluded that the diagnosis and treatment were successful.Discussion: Metal poisoning can kill birds. The veterinarian should always seek the literature in order to perform the best support and treatment. For this, detailed history and detailed medical history must be taken into account, since the time of ingestion and the type of metal interfere with the therapeutic conduct. The use of imaging tests such as x-rays and ultrasound are essential to assist the clinician, especially in cases where the tutor does not know whether or not the animal has ingested an object.[...](AU)

Rumen parameters and passage rate in cattle fed diets based on sugarcane hydrolyzed with calcium oxide / Parâmetros ruminais e taxa de passagem em bovinos alimentados com rações baseadas em cana de açúcar hidrolisada com óxido de cálcio

Nine non-lactating Holstein x Gyr cattle were fed with rations based on chopped sugarcane supplied in natura (control) or hydrolyzed (fresh matter basis) for 24 h with 1% or 2% calcium oxide (CaO), which corresponds to 3.1% or 6.2% CaO on a dry matter (DM) basis, respectively. Ruminal parameters (pH and ammonia N, acetate, propionate and butyrate concentrations) as well as the kinetics of fluid and particulate passage in the gastrointestinal tract from cattle were evaluated. A design using three 3 x 3 contemporaneous Latin Squares (LS) was adopted. The studies of kinetics of fluid and particulate passage were performed using the external markers cobalt-EDTA and chromium-mordanted-NDF, respectively. The ruminal parameters were analyzed according to a 3 x 3 LS replicated three times with repeated measures in time using mixed models that included the fixed effects of treatment (level of CaO), time of sampling and their interaction and the following random effects: LS, animal(LS), period of LS and period*animal (LS). The kinetic parameters of fluid and particulate passage were analyzed using mixed models with treatment as a fixed effect and period of LS, animal(LS) and LS as random effects. The linear and quadratic effects of the treatments were analyzed using orthogonal contrasts. Significant differences were declared at P ≤ 0.05. No treatment*time interaction was observed (P > 0.05) for any ruminal parameter. A linear effect (P = 0.0279) of CaO inclusion on ruminal pH was observed, but there were no effects (P > 0.05) of the treatments on the ammonia N, acetate, butyrate and total volatile fatty acid ruminal concentrations. There was a quadratic effect (P = 0.05) of the addition of CaO on the ruminal propionate concentration. There was no effect (P > 0.05) of the treatments on the particulate rate of passage in the rumen as well as on the mean retention time in this compartment and in the total gastrointestinal tract.(AU)

Nove fêmeas Holândês x Gir não lactantes, alocadas em três Quadrados Latinos (QL) 3 x 3 contemporâneos, foram alimentadas com rações à base de cana de açúcar picada, fornecida in natura (Controle) ou hidrolisada (base da matéria fresca) por 24 h com 1% ou 2% de óxido de cálcio (CaO), o que correspondeu, respectivamente, a 3,1% ou 6,2% de CaO com base na matéria seca (MS). O pH e as concentrações ruminais de N amoniacal, acetato, propionato e butirato foram avaliados. As cinéticas de passagem de fluidos e de partículas no trato gastrointestinal (TGI) dos animais foram estudadas utilizando, respectivamente, os indicadores externos Cobalto-EDTA e fibra em detergente neutro mordantada com Cromo. As análises estatísticas foram realizadas utilizando modelos mistos. Os parâmetros de fermentação ruminal foram analisados como medidas repetidas no tempo, sendo considerados efeitos fixos: o tratamento (nível de adição de CaO), o tempo de amostragem e sua interação, e efeitos aleatórios: QL, animal(QL), período do QL e período*animal(QL). Os parâmetros de cinética de fluidos e de partículas no TGI foram analisados considerando o tratamento como efeito fixo, e período do QL, animal(QL) e QL como efeitos aleatórios. Diferenças foram consideradas significativas quando P ≤ 0,05. Não houve interação tratamento*tempo (P > 0,05) para nenhum parâmetro ruminal. A adição de CaO promoveu incremento linear (P = 0,0279) no pH do rúmen, mas não houve efeito (P > 0,05) dos tratamentos sobre as concentrações ruminais de N amoniacal, acetato, butirato e ácidos graxos voláteis totais. Foi observado efeito quadrático (P = 0,05) da adição de CaO sobre a concentração ruminal de propionato.(AU)

Perfil hematológico de cães em diferentes concentrações de anticoagulante e bioquímica sérica em tempos diferentes de armazenamento / Hematologic profile of dogs in different concentrations of anticoagulant and serum biochemistry at different storage times

O objetivo deste estudo foi verificar as alterações hematológicas e bioquímicas em amostras sanguíneas de cães, em diferentes concentrações de anticoagulante EDTA-Na2 e diferentes tempos de armazenamento em temperatura entre 4 e 8°C por até 48 horas. Para isso, foram coletados 12mL de sangue por venopunção jugular ou cefálica de 30 cães (Canis lupus familiaris), procedentes do atendimento clínico realizados no Hospital Veterinário da UFERSA - Mossoró-RN. As amostras foram distribuídas em 4 tubos de ensaio, sendo 3 tubos contendo 50uL de EDTA-Na2 para a análise hematológica, preenchidos com 1mL, 3mL e 4mL, e o quarto tubo, para análise bioquímica, sem anticoagulante, foi acondicionado 4mL da amostra. As análises foram realizadas durante a primeira hora, 24 e 48 horas após a coleta. Posteriormente às análises, observou-se que as variáveis hematológicas se mostraram estáveis ao longo do tempo e independente da concentração de anticoagulante, sem diferenças significativas. Os resultados revelaram que a relação sangue/anticoagulante não interfere nos parâmetros do hemograma quando mantidas em condições adequadas de armazenamento e processadas em analisador hematológico Scil Vet abc™. Quanto às análises bioquímicas, submetidas às mesmas condições de armazenamento, também se mostraram estáveis por até 48 horas. Desse modo, conclui-se que as amostras sanguíneas podem ser armazenadas entre 4°C e 8°C por até 48 horas.(AU)

The objective of this study was to verify hematological and biochemical alterations in blood samples from dogs, in different concentrations of anticoagulant EDTA-Na2 and different storage times in temperature between 4 and 8°C for up to 48 hours. For this, 12mL of blood were collected by jugular or cephalic venipuncture from 30 dogs (Canis lupus familiaris), from the clinical care performed at the Veterinary Hospital of UFERSA - Mossoró-RN. The samples were distributed in 4 test tubes, 3 tubes containing 50uL of EDTA-Na2 for the hematological analysis, filled with 1mL, 3mL and 4mL, and the fourth tube, for biochemical analysis, without anticoagulant, was conditioned 4 mL of the sample. The analyzes were performed during the first hour, 24 and 48 hours after collection. Subsequent to the analyzes, it was observed that the hematological variables were stable over time and independent of the anticoagulant concentration, without significant differences. The results showed that the blood / anticoagulant ratio did not interfere in the hemogram when stored under adequate conditions of storage and processed in a hematology analyzer Scil Vet abc ™. As for the biochemical analyzes, submitted to the same storage conditions, they were also stable for up to 48 hours. Thus, it is concluded that blood samples can be stored between 4°C and 8°C for up to 48 hours.(AU)

Giant Anteater (Myrmecophaga tridactyla Linnaeus, 1758) of the Brazilian cerrado: hematology and storage effect / Tamanduás-bandeiras (Myrmecophaga tridactyla Linnaeus, 1758) do cerrado brasileiro: hematologia e efeito de estocagem

Giant Anteater (Myrmecophaga tridactyla) is a vulnerable species because of progressive habitat destruction, mostly affected by wildfires and car accidents. The increasing number of animals that are attended by wildlife rescue centres reinforces the need of knowledge about haematological parameters, especially for those that inhabit Brazilian cerrado biome. For this purpose and in order to establish reference values for this species and also to compare them with previous studies, haematological analysis of captive giant anteaters from Brazilian cerrado were performed. Moreover, the alterations of blood samples after 24 and 48 hours of storage at refrigeration temperatures (4oC) and preserved with two different EDTA concentrations (5% and 10%) were studied. Means and standard deviations of haematological parameters analysed immediately after collection were: RBC: 2,07 x106/µL ± 0,40; hematocrit: 38,08%± 5,93; haemoglobin: 11.33g/dL±2.15; MCV:186.52 fL±21.72; MCHC: 29.68g/dL±2.56; MCH: 55.08pcg±5,94; total leucocytes: 8.142/µL±2.441; neutrophils: 5.913/µL±2.168; lymphocytes: 1.460/µL±740; eosinophil: 522/µL±385; monocytes: 247/µL±176; thrombocytes: 123.458/µL±31.362 and total plasma protein: 6.23g/dL±0.49. This data shows evidence of the existence of important differences between these values and others from other areas, either from Brazil or from other South American countries. Those variations might be connected to environment, genetic, nutritional and/or management factors. Regarding the storage effect analysis, it can be concluded that in giant anteaters, haematological analysis can be performed until 24h after collection without any significant alterations on the haematological parameters, except for thrombocytes. Concerning the different EDTA concentrations, it can be concluded that there are no quantitative differences in haematological variables. Nevertheless, relevant morphologic alterations in blood cells can be observed after a 24h storage period, being most noticeable in the leucocytes. Those alterations can lead to misinterpretation of the results, interfering diagnosis, prognosis and treatment.(AU)

O tamanduá-bandeira (Myrmecophaga tridactyla) é uma espécie vulnerável devido à destruição progressiva do seu habitat natural, sendo afetado por queimadas e atropelamentos. O aumento na casuística de atendimentos de animais silvestres reforça a necessidade de se ter conhecimento dos parâmetros hematológicos, em especial para os que vivem no bioma do cerrado. Por isso, este trabalho teve por objetivos realizar o hemograma de tamanduás-bandeiras de cativeiro localizados no cerrado brasileiro, a fim de estabelecer valores de referência para essa espécie e compará-los a estudos prévios. Além disso, verificar quais alterações podem ser encontradas em amostras armazenadas por 24 e 48 horas após a colheita, em temperatura de refrigeração (4oC) e tratadas com duas concentrações distintas de EDTA (5% e 10%). A média e o desvio padrão das variáveis hematológicas encontradas nas amostras processadas logo após a colheita foram: hemácias (2,07x106/µL±0,40); volume globular (38,08%± 5,93); hemoglobina (11,33g/dL±2,15); VCM (186,52 fL±21,72); CHCM (29,68g/dL±2,56); HCM (55,08pcg±5,94); leucócitos totais (8.142/µL±2.441); neutrófilos (5.913/µL±2.168); linfócitos (1.460/µL±740); eosinófilos (522/µL±385); monócitos (247/µL±176); plaquetas (123.458/µL±31.362) e proteínas plasmáticas totais (6,23g/dL±0,49). Tais dados permitem afirmar que existem importantes diferenças entre os valores hematológicos destes em relação aos animais provenientes de outras regiões, tanto do Brasil quanto de outros países da América do Sul. Provavelmente, tais divergências estão associadas a fatores ambientais, genéticos, nutricionais e/ou de manejo. Quanto à análise das amostras estocadas, conclui-se que, em tamanduás-bandeiras, as amostras para a realização de hemograma podem ser processadas até 24 horas após a colheita, sem alteração significativa das variáveis hematológicas, com exceção das plaquetas. Com relação às duas concentrações de EDTA, pode-se inferir que não há diferença quantitativa entre ambas para as variáveis hematológicas. Contudo, é possível constatar que o EDTA promove alterações morfológicas relevantes nas células sanguíneas após 24 horas de armazenamento, sendo os leucócitos os mais afetados. Tais alterações, quando relatadas, podem induzir interpretações equivocadas, interferindo no diagnóstico, prognóstico e tratamento.(AU)

Giant Anteater (Myrmecophaga tridactyla Linnaeus, 1758) of the Brazilian cerrado: hematology and storage effect / Tamanduás-bandeiras (Myrmecophaga tridactyla Linnaeus, 1758) do cerrado brasileiro: hematologia e efeito de estocagem

Giant Anteater (Myrmecophaga tridactyla) is a vulnerable species because of progressive habitat destruction, mostly affected by wildfires and car accidents. The increasing number of animals that are attended by wildlife rescue centres reinforces the need of knowledge about haematological parameters, especially for those that inhabit Brazilian cerrado biome. For this purpose and in order to establish reference values for this species and also to compare them with previous studies, haematological analysis of captive giant anteaters from Brazilian cerrado were performed. Moreover, the alterations of blood samples after 24 and 48 hours of storage at refrigeration temperatures (4oC) and preserved with two different EDTA concentrations (5% and 10%) were studied. Means and standard deviations of haematological parameters analysed immediately after collection were: RBC: 2,07 x106/µL ± 0,40; hematocrit: 38,08%± 5,93; haemoglobin: 11.33g/dL±2.15; MCV:186.52 fL±21.72; MCHC: 29.68g/dL±2.56; MCH: 55.08pcg±5,94; total leucocytes: 8.142/µL±2.441; neutrophils: 5.913/µL±2.168; lymphocytes: 1.460/µL±740; eosinophil: 522/µL±385; monocytes: 247/µL±176; thrombocytes: 123.458/µL±31.362 and total plasma protein: 6.23g/dL±0.49. This data shows evidence of the existence of important differences between these values and others from other areas, either from Brazil or from other South American countries. Those variations might be connected to environment, genetic, nutritional and/or management factors. Regarding the storage effect analysis, it can be concluded that in giant anteaters, haematological analysis can be performed until 24h after collection without any significant alterations on the haematological parameters, except for thrombocytes. Concerning the different EDTA concentrations, it can be concluded that there are no quantitative differences in haematological variables. Nevertheless, relevant morphologic alterations in blood cells can be observed after a 24h storage period, being most noticeable in the leucocytes. Those alterations can lead to misinterpretation of the results, interfering diagnosis, prognosis and treatment.(AU)

O tamanduá-bandeira (Myrmecophaga tridactyla) é uma espécie vulnerável devido à destruição progressiva do seu habitat natural, sendo afetado por queimadas e atropelamentos. O aumento na casuística de atendimentos de animais silvestres reforça a necessidade de se ter conhecimento dos parâmetros hematológicos, em especial para os que vivem no bioma do cerrado. Por isso, este trabalho teve por objetivos realizar o hemograma de tamanduás-bandeiras de cativeiro localizados no cerrado brasileiro, a fim de estabelecer valores de referência para essa espécie e compará-los a estudos prévios. Além disso, verificar quais alterações podem ser encontradas em amostras armazenadas por 24 e 48 horas após a colheita, em temperatura de refrigeração (4oC) e tratadas com duas concentrações distintas de EDTA (5% e 10%). A média e o desvio padrão das variáveis hematológicas encontradas nas amostras processadas logo após a colheita foram: hemácias (2,07x106/µL±0,40); volume globular (38,08%± 5,93); hemoglobina (11,33g/dL±2,15); VCM (186,52 fL±21,72); CHCM (29,68g/dL±2,56); HCM (55,08pcg±5,94); leucócitos totais (8.142/µL±2.441); neutrófilos (5.913/µL±2.168); linfócitos (1.460/µL±740); eosinófilos (522/µL±385); monócitos (247/µL±176); plaquetas (123.458/µL±31.362) e proteínas plasmáticas totais (6,23g/dL±0,49). Tais dados permitem afirmar que existem importantes diferenças entre os valores hematológicos destes em relação aos animais provenientes de outras regiões, tanto do Brasil quanto de outros países da América do Sul. Provavelmente, tais divergências estão associadas a fatores ambientais, genéticos, nutricionais e/ou de manejo. Quanto à análise das amostras estocadas, conclui-se que, em tamanduás-bandeiras, as amostras para a realização de hemograma podem ser processadas até 24 horas após a colheita, sem alteração significativa das variáveis hematológicas, com exceção das plaquetas. Com relação às duas concentrações de EDTA, pode-se inferir que não há diferença quantitativa entre ambas para as variáveis hematológicas. Contudo, é possível constatar que o EDTA promove alterações morfológicas relevantes nas células sanguíneas após 24 horas de armazenamento, sendo os leucócitos os mais afetados. Tais alterações, quando relatadas, podem induzir interpretações equivocadas, interferindo no diagnóstico, prognóstico e tratamento.(AU)

Efeito metafilático do zinco injetável sobre parâmetros metabólicos e oxidativos de ovelhas no pós-parto imediato / Metaphylactic effect of injectable zinc on metabolic and oxidative parameters of sheep in the immediate postpartum

The aim of this study was to evaluate a new source of injectable organic zinc (zinc edetate) on the energy and oxidative profile in sheep during the immediate postpartum period. Twenty-six Texel breed animals were previously identified and divided into two experimental groups: the treated group (TG; n= 13) that comprised the animals that received a subcutaneous (SC) injection of 100 mg of zinc edetate (2 mL) fifteen days before the parturition expected date and the control group (CG; n=13) that comprised the animals that received 2mL of physiological solution at the same date of TG. Blood samples were collected on the parturition day for the assessment of serum fructosamine, cholesterol and triglycerides, insulin-like growth factor type 1 (IGF-1), the oxidative stress index (OSi) and blood zinc concentration. In addition to these parameters, the measurement of zinc was made in food given to the animals. There was no difference in metabolic parameters and OSi between the experimental groups (P>0.05), as well as in blood zinc concentrations (P>0.05). The parenteral zinc edentate does not change the energy and oxidative profile of sheep in immediate postpartum.(AU)

Efeito metafilático do zinco injetável sobre parâmetros metabólicos e oxidativos de ovelhas no pós-parto imediato / Metaphylactic effect of injectable zinc on metabolic and oxidative parameters of sheep in the immediate postpartum

The aim of this study was to evaluate a new source of injectable organic zinc (zinc edetate) on the energy and oxidative profile in sheep during the immediate postpartum period. Twenty-six Texel breed animals were previously identified and divided into two experimental groups: the treated group (TG; n= 13) that comprised the animals that received a subcutaneous (SC) injection of 100 mg of zinc edetate (2 mL) fifteen days before the parturition expected date and the control group (CG; n=13) that comprised the animals that received 2mL of physiological solution at the same date of TG. Blood samples were collected on the parturition day for the assessment of serum fructosamine, cholesterol and triglycerides, insulin-like growth factor type 1 (IGF-1), the oxidative stress index (OSi) and blood zinc concentration. In addition to these parameters, the measurement of zinc was made in food given to the animals. There was no difference in metabolic parameters and OSi between the experimental groups (P>0.05), as well as in blood zinc concentrations (P>0.05). The parenteral zinc edentate does not change the energy and oxidative profile of sheep in immediate postpartum.(AU)

Na2EDTA anticoagulant impaired blood samples from the teleost Piaractus mesopotamicus / Anticoagulante Na2EDTA danifica amostras de sangue do teleósteo Piaractus mesopotamicus

The present study aimed to evaluate the effects of Na heparin and Na2EDTA on blood of Piaractus mesopotamicus (360.7±42.4g, 26.4±1.0cm). Twenty fishes were sampled in two experiment trials, ten for erythrocyte fragility analysis and ten for hematologic and plasma biochemical study. The blood collected by venous-caudal puncture was fractioned and stored in anticoagulants solution: Na2EDTA 10%, Na2EDTA 3%, Na heparin 5000 IU and Na heparin 100 IU. Plasmatic levels of calcium presented in the Na2EDTA stored samples were about 80% lower than both heparin groups. Blood samples of P. mesopotamicus stored with Na2EDTA demonstrated increase in the hematocrit and MCV, and decrease in MCHC. The dose-response effect was observed in this study. The results are reinforced by the higher levels of plasmatic protein and hemolysis presented in the Na2EDTA 10% stored blood, confirming the deleterious effect of this anticoagulant treatment on the quality of blood samples. Na2EDTA is not indicated to store P. mesopotamicus blood samples, but sodium heparin at 100 IU is the most recommended anticoagulant, since this treatment presented the lower rate of alterations in the stored blood.(AU)

O presente estudo teve por objetivo avaliar os efeitos da heparina sódica e Na2EDTA no sangue de Piaractus mesopotamicus (360,7±42,4g, 26,4±1,0cm). Foram amostrados vinte peixes em dois ensaios experimentais, sendo dez peixes utilizados para análise da fragilidade dos eritrócitos e dez peixes para análise dos parâmetros hematológicos e estudo bioquímico do plasma. O sangue coletado por punção veno-caudal foi aliquotado e armazenado em diferentes soluções anticoagulantes: Na2EDTA 10%, Na2EDTA 3%, heparina sódica 5.000 UI e heparina sódica 100 UI. Níveis plasmáticos de cálcio apresentados nas amostras armazenados em Na2EDTA diminuíram cerca de 80% em relação aos dois grupos armazenados com heparina. Amostras de sangue de pacus armazenados com Na2EDTA demonstraram aumento do hematócrito e VCM, e diminuição na CHCM. O efeito dose-resposta foi observado neste estudo. Estes resultados são reforçados pelos níveis mais elevados de proteína plasmática e hemólise apresentado no sangue armazenado com Na2EDTA 10%, o que confirma o efeito deletério desse tratamento anticoagulante na qualidade de amostras de sangue. Na2EDTA não é indicada para armazenar amostras de sangue de P. mesopotamicus, e heparina sódica a 100 UI é o anticoagulante mais recomendado, uma vez que este tratamento apresentou a menor taxa de alterações no sangue armazenado.(AU)

Bioquimico plasmático de cães: efeitos dos diferentes anticoagulantes em comparação com o soro / Plasmatic biochemical of dogs: effects of different anticoagulants compared to serum

Na rotina clínica veterinária são utilizados diferentes tipos de anticoagulantes, escolhidos de acordo com o exame a ser solicitado. Animais agitados, agressivos, de difícil contenção e de pequeno porte podem ser fatores limitantes quando se deseja aliquotar a amostra em tubos com diferentes anticoagulantes. Assim, esse trabalho teve como objetivo avaliar os efeitos dos anticoagulantes EDTA, Citrato de Sódio e Fluoreto associado ao EDTA, nos resultados dos exames bioquímicos de rotina realizados em cães e compará-los com o soro. Verificou-se que o plasma obtido com o anticoagulante EDTA resultou em alterações nas dosagens de glicose, colesterol e cálcio. Amostras obtidas com Citrato de Sódio tiveram resultados inferiores ao do soro em todos os parâmetros avaliados, com exceção da ALT. O anticoagulante Fluoreto associado ao EDTA, apenas nas dosagens de uréia e glicose, não resultou em alterações estatísticas quando comparado ao soro.

Different types of anticoagulants are used in the routine of a veterinary clinic, which are chosen based on the exam to be performed. Agitated, aggressive, of hard contention and small size animals can be a limiting factor when it is wished to separate samples into tubes with different anticoagulants. Therefore, this study had as the objective evaluate the effects of anticoagulants: EDTA, sodium citrate and sodium fluorite associated with EDTA, in the results of routine plasma biochemistry performed in dogs and compare it with serum. Blood samples of 10 clinically healthy dogs were collected and placed in different tubes with each anticoagulant and one without to obtain serum. Albumin, alanine aminotransferase (ALT), calcium, total cholesterol, creatine, phosphorus, glucose, total protein (PT) and urea concentrations were measured by spectophotometric. It was found that plasma obtained with EDTA anticoagulant resulted in dosage alterations for glucose, cholesterol and calcium. Samples obtained with sodium citrate presented results lower than serum in all analyzed parameters, except for ALT. The anticoagulant sodium fluorite with EDTA only didn’t result in statistical differences when compared to serum in the parameters of urea and glucose. Thus, serum is the most trustworthy sample to determine the biochemical profile of dogs, however some biochemical’s can be performed with plasma with EDTA such as creatine, ALT, urea, albumin, PT and phosphorus.

Bioquimico plasmático de cães: efeitos dos diferentes anticoagulantes em comparação com o soro / Plasmatic biochemical of dogs: effects of different anticoagulants compared to serum

Na rotina clínica veterinária são utilizados diferentes tipos de anticoagulantes, escolhidos de acordo com o exame a ser solicitado. Animais agitados, agressivos, de difícil contenção e de pequeno porte podem ser fatores limitantes quando se deseja aliquotar a amostra em tubos com diferentes anticoagulantes. Assim, esse trabalho teve como objetivo avaliar os efeitos dos anticoagulantes EDTA, Citrato de Sódio e Fluoreto associado ao EDTA, nos resultados dos exames bioquímicos de rotina realizados em cães e compará-los com o soro. Verificou-se que o plasma obtido com o anticoagulante EDTA resultou em alterações nas dosagens de glicose, colesterol e cálcio. Amostras obtidas com Citrato de Sódio tiveram resultados inferiores ao do soro em todos os parâmetros avaliados, com exceção da ALT. O anticoagulante Fluoreto associado ao EDTA, apenas nas dosagens de uréia e glicose, não resultou em alterações estatísticas quando comparado ao soro.(AU)

Different types of anticoagulants are used in the routine of a veterinary clinic, which are chosen based on the exam to be performed. Agitated, aggressive, of hard contention and small size animals can be a limiting factor when it is wished to separate samples into tubes with different anticoagulants. Therefore, this study had as the objective evaluate the effects of anticoagulants: EDTA, sodium citrate and sodium fluorite associated with EDTA, in the results of routine plasma biochemistry performed in dogs and compare it with serum. Blood samples of 10 clinically healthy dogs were collected and placed in different tubes with each anticoagulant and one without to obtain serum. Albumin, alanine aminotransferase (ALT), calcium, total cholesterol, creatine, phosphorus, glucose, total protein (PT) and urea concentrations were measured by spectophotometric. It was found that plasma obtained with EDTA anticoagulant resulted in dosage alterations for glucose, cholesterol and calcium. Samples obtained with sodium citrate presented results lower than serum in all analyzed parameters, except for ALT. The anticoagulant sodium fluorite with EDTA only didnt result in statistical differences when compared to serum in the parameters of urea and glucose. Thus, serum is the most trustworthy sample to determine the biochemical profile of dogs, however some biochemicals can be performed with plasma with EDTA such as creatine, ALT, urea, albumin, PT and phosphorus.(AU)

Use of otolith strontium: calcium and zinc:calcium ratios as an indicator of the habitat of Percophis brasiliensis Quoy & Gaimard, 1825 in the southwestern Atlantic Ocean

We evaluate the simultaneous use of Sr: Ca and Zn: Ca ratios of the sagitta otolith as a potential indicator of the habitat of Percophis brasiliensis along a latitudinal gradient in the southwestern Atlantic Ocean (34-42ºS and 51-67ºW), in order to reliably identify fish stocks. Fish were collected in three sampling sites: Argentine-Uruguayan Common Fishing Zone (AUCFZ), El Rincón (ER) and San Matías Gulf (SMG). The otolith Sr:Ca and Zn:Ca ratios were determined by ICP-OES and EDTA volumetric method. The otolith Sr:Ca ratio was similar in the three sampling sites, while the Zn:Ca ratio was significantly higher in AUCFZ than in ER and SMG for all age groups. The discriminant analysis showed an association between the otolith Sr:Ca and Zn:Ca ratios from ER and SMG. Present results suggest the potential occurrence of two fish stocks of P. brasiliensis in the study area.(AU)

Evaluamos el uso simultáneo de las relaciones Sr:Ca y Zn:Ca de los otolitos sagittae como un potencial indicador de hábitat de Percophis brasiliensis a lo largo de un gradiente longitudinal el Atlántico sudoccidental (34-42ºS - 51-67ºW) con el fin de contribuir a la identificación de los stocks pesqueros. Los peces fueron capturados en tres sitios de muestreo: Zona Común de Pesca Argentina-Uruguaya (ZCPAU), El Rincón (ER) y el Golfo San Matías (GSM). Las relaciones Sr:Ca y Zn:Ca se determinaron por ICP-OES y por titulación con EDTA. La relación Sr:Ca fue similar en los tres sitios de muestreo. La relación Zn:Ca fue mayor en la ZCPAU que en las demás areas (ER y GSM) para todos los rangos de edad. El análisis discriminante mostró una asociación entre las relaciones Sr:Ca y Zn:Ca de ER y GSM. Los resultados de este trabajo sugieren la presencia de al menos dos stocks de P. brasiliensis en el aérea de estudio.(AU)

Is otolith microchemistry (Sr: Ca and Ba:Ca ratios) useful to identify Mugil curema populations in the southeastern Caribbean Sea? / A microquímica do otólito é um indicador do habitat de Mugil curema no sudeste do Mar Caribenho?

The aim of the present study was to evaluate the potential use of otolith microchemistry (Sr:Ca and Ba:Ca ratios) to identify silver mullet, Mugil curema, populations in Southeastern Caribbean Sea. Fish samples were collected in 7 areas of Nueva Esparta State (Venezuela). The otolith Sr:Ca and Ba:Ca ratios and water Sr:Ca were determined (by ICP-OES and EDTA volumetric method). Otoliths Sr:Ca and Ba:Ca ratios and Sr:Ca partition coefficient of mullets in Cubagua island (south of the State) were significantly different from ratios in La Guardia (north of the State). A discriminant analysis of otolith Sr:Ca and Ba:Ca ratios separated Cubagua Island from La Guardia values. These results suggest the existence of different mullet groups in the Southeastern Caribbean Sea. For this, the simultaneous use of Sr:Ca and Ba:Ca ratios could be a potential tool to identify populations in the study area.(AU)

O objetivo do presente estudo foi avaliar o potencial uso da microquímica do otólito (razões Sr:Ca e Ba:Ca) para identificar distintas populações de tainha, Mugil curema, no sudeste do mar caribenho. Os peixes foram coletados em 7 áreas do estado de Nueva Esparta (Venezuela). As razões Sr:Ca e Ba:Ca do otólito e a razão Sr:Ca da água foram determinadas (pelo ICP-OES e EDTA método volumétrico). As razões de Sr:Ca e Ba:Ca dos otólitos e o coeficiente de partição das tainhas da Ilha Cubagua (sul do estado) foram significativamente diferentes das razões de La Guardia (norte do estado). A análise discriminante das razões de Sr:Ca e Ba:Ca dos otólitos separa os valores da Ilha Cubagua e de La Guardia. Estes resultados sugerem a existência de diferentes grupos de Mugil curema no sudeste do mar Caribenho e que o uso simultâneo das razões Sr:Ca e Ba:Ca poderiam ser uma potencial ferramenta para identificar as populações da área de estudo.(AU)

Cytolytic and systemic toxic effects induced by the aqueous extract of the fire coral Millepora alcicornis collected in the Mexican Caribbean and detection of two types of cytolisins

Background Millepora alcicornis is a branching hydrocoral common throughout the Caribbean Sea. Like other members of this genus, this species is capable of inducing skin eruptions and blisters with severe pain after contact. In the present study, we investigated the toxicity of theM. alcicornis aqueous extract on several animal models. Considering that some cnidarian hemolysins have been associated to local tissue damage, since they also induce lysis of other cell types, we also made a partial characterization of the hemolytic activity of M. alcicornis aqueous extract. This information is important for understanding the defense mechanisms of the "fire corals".Methods The effects of pH, temperature, and some divalent cations on the hemolytic activity of the extract were assayed, followed by a zymogram analysis to detect the cytolysins and determine their approximate molecular weight. The toxicity of the aqueous extract was assayed in mice, by intravenous administration, and histopathological changes on several tissues were analyzed by light microscopy. The toxicity of the extract was also tested inArtemia salina nauplii, and the damages caused on the crustaceans were analyzed by transmission and scanning electron microscopy.Results The hemolytic activity of the hydrocoral extract was enhanced in the presence of Ca 2+ (≥2 mM), Mg 2+ (≥6 mM), and Ba2+ (≥0.1 mM); however, it was reduced in the presence of Cu2+(≥0.1 mM), Zn 2+ (≥6 mM), and EDTA (≥0.34 mM). Differences in the pH did not affect the hemolytic activity, but it was temperature-sensitive, since preincubation at ≥ 50 °C sharply reduced hemolysis. The zymogram showed the presence of two types of hemolysins: ~ 28-30 kDa proteins with phospholipase A 2 activity and ~ 200 kDa proteins that do not elicit enzymatic activity. The aqueous extract of this cnidarian was lethal to mice (LD 50 = 17 μg protein/g), and induced kidney, liver, and lung damages. Under denaturing conditions, the aqueous extract completely lost its toxic and hemolytic activities.Conclusions The results showed that the M. alcicornis aqueous extract contains two types of thermolabile hemolysins: proteins of approximately 28-30 kDa with PLA 2 activity, while the others are larger proteins of approximately 200 kDa, which do not possess PLA 2activity. Those thermolabile cytolysins, which are stable to pH changes and whose activity is calcium dependent, are capable of inducing damage in lung, kidney and liver tissues, resulting in a slow death of mice. M. alcicorniscytolysins also provoke tissue dissociation inArtemia salina nauplii that might be attributed to pore forming mechanisms.(AU)