Toxoplasma gondii genotyping from free-range chickens (Gallus gallus domesticus) in a rural area of Rio Grande do Sul, Brazil / [Genotipagem de Toxoplasma gondii em galinhas domésticas em uma área rural do Rio Grande do Sul, Brasil]

Free-range chickens may ingest oocysts of T. gondii present in the environment and consequently harbor virulent strains of this parasite in different tissues, without any clinical signs. Isolation of T. gondii through bioassays on mice and cats from naturally infected chicken tissues has been described in several countries, demonstrating the importance of free-range chickens in the transmission of this parasite. The aim of this study was the genotypic characterization of T. gondii isolates obtained from naturally infected free-range chickens in a rural area of the state of Rio Grande do Sul, Brazil. Brain and heart tissue from 12 chickens seropositive for T. gondii were processed using peptic digestion technique for parasite isolation. From 12 samples subjected to mouse bioassay, nine isolates were obtained. RFLP-PCR genotypic characterization was performed using 11 genetic markers: SAG1, 5'-3'SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico. Genetic characterization of the isolates revealed the presence of five atypical genotypes according to ToxoDB (# 11, # 55, # 64, # 140 and # 163). Our results showed a wide genetic diversity of T. gondii in free-range chickens in this region.(AU)

Galinhas criadas ao ar livre podem ingerir oocistos de T. gondii presentes no ambiente e, com isso, albergar cepas virulentas desse parasita em diferentes tecidos, sem sinais clínicos. O isolamento de T. gondii por meio de bioensaios em camundongos e gatos, a partir de tecidos de galinhas naturalmente infectadas, tem sido descrito em vários países. Isso demonstra a importância das galinhas caipiras na epidemiologia desse parasita. O objetivo deste trabalho foi caracterizar genotipicamente isolados de T. gondii obtidos de galinhas caipiras naturalmente infectadas em uma área rural do município de Santa Maria, estado do Rio Grande do Sul, Brasil. Fragmentos de cérebro e de coração, de 12 galinhas soropositivas para T. gondii, foram processados pela técnica de digestão péptica para isolamento do parasita. Das 12 amostras submetidas a bioensaio com camundongos, nove isolados foram obtidos. A caracterização genotípica por RFLP-PCR foi realizada utilizando-se 11 marcadores genéticos: SAG1, 5'-3'SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 e Apico e revelou a presença de cinco genótipos atípicos de acordo com o ToxoDB (# 11, # 55, # 64, # 140 e # 163). Os resultados mostraram uma ampla diversidade genética de T. gondii em galinhas caipiras nessa região.(AU)

Toxicity assessment and screening of tetrodotoxin in the oblong blowfish (Takifugu oblongus) from the Tamil Nadu Coast of Bay of Bengal, India

Objective To assess the toxicity of the pufferfish Takifugu oblongus, from Chennai coast, Tamil Nadu, India and to detect the presence of Tetrodotoxin (TTX). Methods The toxicity was evaluated by mouse bioassay using Swiss Albino mice which were expressed in mouse units (MU). Gross anatomical features were observed which is followed by histopathology of the dead mice tissues to establish the toxicity. Instrumental analysis for the presence of tetrodotoxin was also performed through GC–MS and HPLC. Results The toxicity of ovary was the maximum with 163 MU/g and lowest toxicity was observed in skin with 75.88 MU/g. Histopathological analyses of the dead mice showed various cellular degenerations and inflammations. The amount of Tetrodotoxin detected through GC–MS and HPLC was more reliable and sensitive than the customary mouse bioassay as instrumental analyses were able to detect even nanograms of the toxin. Conclusions The present study evidently proved that Takifugu oblongus is highly toxic and consumption of the same can pose serious threat for health and possible lethality to humans.

Development of an ICR mouse bioassay for toxicity evaluation in neurotoxic poisoning toxins-contaminated shellfish / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To develop an ICR (female) mouse bioassay (MBA) for toxicity confirmation and evaluation of neurotoxins (brevetoxins)-contaminated shellfish.</p><p><b>METHODS</b>Brevetoxins (BTX-B) as a causative agent of neurotoxic shellfish poisoning (NSP) under different shellfish matrices were intraperitoneally injected at different doses into mice to study their toxic effects and to differentiate the range of lethal and sublethal dosages. Their sensitivity and specificity were analyzed with 2 competitive ELISA kits for quantitative determination of standard BTX-B and dihydroBTX-B under different shellfish matrix-diluent combinations. Detection rates of MBA and two antibody-based assays for BTX-B from field NSP-positive shellfish samples were compared.</p><p><b>RESULTS</b>BTX-B could be detected in shellfish tissues at concentration of 50-400 μg/100 g under shellfish matrix-Tween-saline media, which were appropriate to identify toxic shellfish at or above the regulatory limit (80 μg/100 g shellfish tissues). The LD50 identified was 455 mg/kg for BTX-B under general shellfish matrices (excluding oyster matrices) dissolved in Tween-saline. The presence of shellfish matrices, of oyster matrices in particular, retarded the occurrence of death and toxicity presentation in mice. Two antibody-based assays, even in the presence of different shellfish matrix-diluent combinations, showed acceptable results in quantifying BTX-B and dihydroBTX-B well below the regulatory limit.</p><p><b>CONCLUSION</b>The two ELISA analyses agree favorably (correlation coefficient, r³⋝0.96; Student's t-tests, P>0.05) with the developed bioassay.</p>

Assessment of biomedical and pharmacological activities of sea anemones Stichodactyla mertensii and Stichodactyla gigantea from Gulf of Mannar Biosphere Reserve, southeast coast of India

Cnidarians comprise an old and diverse animal phylum, and possess a wide variety of biologically active substances. Sea anemones contain a diversity of interesting biologically active compounds including some potent toxins. In the present work, the sea anemones Stichodactyla mertensii and Stichodactyla gigantea, collected from the Mandapam coast, are characterized biomedically and pharmacologically. The crude protein was obtained by using methanol and aqueous extracts. The respective protein contents of S. mertensii and S. gigantea were found to be 2.10 µg/mL and 1.87 µg/mL. The methanol and aqueous extracts of S. mertensii and S. gigantea yielded six and nine bands by SDS-PAGE on 12 percent gel. In the hemolytic assay, both extracts exhibited hemolytic effect on chicken, goat, cow and human erythrocytes ('A', 'B' and 'O'). The neurotoxic effects of these crude extracts were determined in vivo using the sea shore crab Ocypode macrocera and mortality was observed. The mouse bioassay for lethality was performed on male albino mice. The crude extract of S. mertensii showed higher lethality (58 seconds at 1 mL-dose) than that of S. gigantea (2 minutes and 10 seconds at 0.75 mL-dose). The analgesic activity test was also carried out on albino mice by Eddy's hot plate and tail-flick methods. The extracts showed moderate analgesic effect by both hot-plate and tail-flick methods. These characteristics emphasize the need for the isolation and molecular characterization of new active toxins in S. mertensii and S. gigantea.(AU)

Ciguatera Fish Poisoning on Kakeroma Island, Kagoshima Prefecture, Japan

In this paper we report two incidents of ciguatera fish poisoning (CFP) that occurred in 2008 on Kakeroma Island in Kagoshima Prefecture, Japan. A family consisting of father (42 y.o.), mother (39 y.o.), daughter (11 y.o.) and son (6 y.o.), as well as a friend of the family (male, 78 y.o.) consumed sliced flesh (sashimi) and developed typical symptoms of CPF 4-5 h later: diarrhea, vomiting, and paresthesia of the extremities. Additionally, the two male adults (42 and 78 y.o.) developed mild hypotention (84/48 and 94/40 mmHg, respectively) and bradycardia (36 and 50 bpm, respectively) and were rushed to the ER of the nearest hospital, located on an adjacent island. The implicated fish were caught together off the west coast of the island and were identified as <i>Variola louti</i> and <i>Lutjanus monostigma</i> based on analysis of 16S rRNA gene coded on mtDNA. Remnants of the implicated fish and other fish caught on the same occasion were examined by the official mouse bioassay method (MBA), which defines the minimum amount of CFP toxin needed to kill a male mouse of ddY strain of 20 g body weight within 24 h as one mouse unit (MU). A significantly high toxicity was detected in the <i>V. louti</i> (0.2 MU/g) eaten by the family and the <i>L. monostigma</i> (0.8 MU/g) eaten by the elderly man. Other specimens of <i>Lethrinus nebulosus</i>, <i>Variola albimarginata</i>, <i>Lutjanus gibbus</i> (2 specimens), <i>Aphareus rutilans</i>, and <i>Sphyraena forsteri</i> (2 specimens) were found to be nontoxic (< 0.025 MU/g).<br> The medical records regarding island inhabitants (ca. 1,500) kept at Kakeroma Tokushukai Clinic, the only medical facility on the Island, and also at Setouchi Tokushukai Hospital, a nearest hospital on an adjacent island (Amami) revealed 6 CFP outbreaks involving 13 cases between 2005 and 2008. The estimated frequency of CFP was 10.0 incidents/10,000 persons/year and the morbidity rate was 21.7 cases/10,000 persons/year. The symptoms and signs observed in the patients and the implicated fish species, <i>L. monostigma</i> and<i> V. louti</i>, were typical of CFP in this region.

Botulism

Botulism is a life threatening disorder caused by a neurotoxin produced from the anaerobic, spore-forming bacterium Clostridium botulinum. There are seven antigenically distinct types of botulinum neurotoxins (types A through G), and the human botulism is primarily caused by toxin types A, B, and E. Four clinical forms of botulism occur in humans: foodborne botulism, wound botulism, infant botulism, and adult infectious botulism. Botulism is characterized by symmetric, descending, flaccid paralysis of motor and autonomic nerves, usually beginning with the cranial nerves. Dry mouth, blurred vision, and diplopia are usually the earliest neurologic symptoms. Botulism should be suspected in a patient with an acute onset of gastrointestinal, autonomic, and cranial nerve dysfunction. Confirmation of the diagnosis of botulism depends on the detection of the toxin or the organism in the patient. The most reliable method for the detection of the toxin is the mouse inoculation test. The mainstay of treatment for severe botulism is supportive therapy with mechanical ventilation. The administration of antitoxin is the only specific pharmacologic treatment available for botulism. Botulism is a rare but potentially fatal illness, so timely recognition of the clinical symptoms plays an important role in decreasing the mortality rate.