Toxicological Assessment of Pure Lolitrem B and Ryegrass Seed Infected with the AR37 Endophyte Using Mice.

Fungal endophytes in perennial ryegrass are essential to New Zealand's pastoral system due to anti-insect effects. However, endophytes also produce compounds which can be detrimental to animals. Furthermore, as these toxins have been detected in the milk and fat of animals grazing common-toxic (containing lolitrem B) or AR37 endophyte-infected herbage they could enter the human food chain. To assess the risk to human health mice were fed for 90 days with three dose rates of lolitrem B and of AR37. Parameters indicative of animal health were measured as well as chemical, hematological and histological analysis of samples collected on day 90. Since endophyte toxin residues have been detected in milk, they could be transferred from mother to offspring via breast milk. To evaluate possible effects on reproduction two complete generations of mice were fed lolitrem B or AR37. At the dose rates given no adverse effects were observed in either study. The 100-fold safety factor to allow the use of animal data in human health assessments was applied and by considering the concentrations of lolitrem B or AR37 metabolites which could be ingested by a consumer it is highly unlikely that they pose any risk to human health.

The contribution of massive fetomaternal hemorrhage to antepartum stillbirth: a 25-year cross-sectional study.

INTRODUCTION: Fatal antepartum fetomaternal hemorrhage is a relatively uncommon clinical presentation, though one that appears quickly and without warning. The pathophysiology of this disease is unclear, and the incidence does not appear to be decreasing in line with overall antepartum mortality. This study was undertaken to analyse trends in antepartum fetal death from fetomaternal hemorrhage over a 25-year period in a single maternity hospital in Dublin, Ireland. MATERIAL AND METHODS: A cross-sectional study of 192 132 nonanomalous infants weighing 500 g or more, delivered in a single tertiary-referral university institution between 1987 and 2011. Data was compared using Fisher's exact test, univariate analysis, and Cuzick's test for trend. RESULTS: There was no decrease in the rate of fatal fetomaternal hemorrhage over the past 25 years (p = 0.29), despite a decline in overall antepartum deaths (p = 0.0049). Fetomaternal hemorrhage accounted for 4.1% (34/828) of antepartum stillbirths. A higher proportion of these stillbirths occurred at term gestations (74%; 25/34) compared with other causes (40%; 321/794; p = 0.0003). Female infants were statistically more likely to be involved than males [odds ratio (OR) 2.33, 95% confidence interval (CI) 1.08-5.47, p = 0.02). Multiple gestations were up to six times as likely to be affected as singleton pregnancies (OR 6.52, 95% CI 1.67-18.50, p = 0.005). CONCLUSIONS: Over the past 25 years there has been no reduction in rates of fatal fetomaternal hemorrhage. Female infants and multiple gestations remain at higher risk of antepartum death from fatal fetomaternal hemorrhage.

Convenient large-scale purification of yessotoxin from Protoceratium reticulatum culture and isolation of a novel furanoyessotoxin.

Yessotoxins from a large-scale culture (226 L) of Protoceratium reticulatum strain CAWD129 were harvested by filtration followed by solid-phase extraction. The extract was purified by column chromatography over basic alumina and reverse-phase flash chromatography to afford pure yessotoxin (193 mg). Isolation of yessotoxin was greatly facilitated by selection of a strain which did not produce analogues that interfered with yessotoxin isolation. In addition to yessotoxin, numerous minor yessotoxins were detected by LC-MS in other fractions. From one of these, an early eluting minor analogue with the same molecular weight as yessotoxin and a similar mass spectrometric fragmentation pattern was isolated. This analogue was identified by NMR and mass spectrometry as a novel yessotoxin analogue containing a furan ring in the side chain. This finding reveals biosynthetic flexibility of the yessotoxin pathway in P. reticulatum and confirms earlier findings of production of many minor yessotoxin analogues by this alga. Production of these analogues appeared to be a constitutive trait of P. reticulatum CAWD129.

Enzyme-linked immunosorbent assay for the detection of yessotoxin and its analogues.

Polyclonal antibodies were produced for the development of competitive enzyme-linked immunoassays for use in quantifying yessotoxins in shellfish, algal cells, and culture supernatants. Immunizing and plate coating antigens were prepared by derivatization of yessotoxin either by ozonolysis or bromination and conjugation to proteins. Two assays that were the most sensitive for yessotoxin were optimized and characterized. Cross-reactivity studies indicated that the antibodies raised have broad specificity and that binding to analogues was strongly affected by changes to the A-ring and, to a lesser extent, the K-ring regions of the toxin molecule. ELISA provides a sensitive and rapid analytical method that is suitable for screening large numbers of samples and detects all the yessotoxin analogues that the European Commission currently requires shellfish to be tested for. The assay limit of quantitation for yessotoxin in whole shellfish flesh is 75 microg/kg; therefore, assay sensitivity is sufficient to measure toxin levels well below the maximum permitted level set by the European Commission. The antibodies produced can be used in additional applications such as the immunolocalization of yessotoxins in shellfish and preparation of immunoaffinity columns.