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High Dose

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Scientific papers - High dose - page 5

John Wilesmith's research (50) postulates that BSE will erupt in those adult cows who were fed brands of concentrates as young calves which contained particularly high percentages of MBM protein. However, as this MBM was processed via the continuous-flow rendering system, which did not employ solvent extraction, such feed would have retained the tallow fat fraction that carries any fat-bound phosmet contaminants that may be present. This MBM therefore presents a more severe threat of toxicological hazard to young calves than if fed to adult cattle. For, much like all species of mammalian infant, young calves possess immature enzymic detoxification systems (51) that are incapable of efficient detoxification of chemical pollutants that enter their body systems. An even greater degree of 'cliemical susceptibility' would apply to the context of the calf fetus that has been exposed to phosmet via a placental intake of phosmet-contaminated lipid; originating either from a source of direct phosmet treatment upon the mother cow or via ingestion of phosmet-contaminated MBM concentrates by the mother cow. For phosmet has been demonstrated to cross the placenta (34,52) and, following a 20 mg/kg application onto pregnant rats (52), has been shown to concentrate at nearly two-fold greater concentration of 380 ng/g in the liver of the fetus as opposed to the 215 ng/g concentration found in the liver of the treated mother itself. These findings add strong support to this theory whose central tenet proposes 'in utero' exposure to phosmet as the most likely developmental stage for initiation of the BSE prion.

Studies have demonstrated teratogenic complications such as hydrocephalus (53) following exposure to 1.5 mg/kg oral daily doses of phosmet during the critical 'window' period of gestation. Another lab formulation of an organo-phthalimido-phosphorus (similar to phosmet) has produced severe teratogenic complications at low doses (54), whilst high doses of the warblecide 'trichlorphon' used upon pregnant pigs between 55 and 70 days post conception has produced hyperplasia of the cerebellum accompanied by severe ataxia and tremors in the offspring piglets (55).

Considering all four of 'Hassall's postulates' for bioconcentration have been satisfied in relation to exposure of the UK bovine to systemic phosmet during the 1980s, then, despite the absence of any direct analytical evidence relating to this phenomenon, it would still seem reasonable to assume that bioconcentration probably occurred in the UK context of compulsory high-dose systemic usage.

Experimental evidence fo bioconcentration of organophosphorus compounds

Two separate trials involved feeding kestrels (37) and amphibians (36) upon frogs and tadpoles maintained in water that was deliberately contaminated with a range of doses of the 0Ps parathion, malathion, acephate and fenthion. Both trials demonstrated that significant degrees of bioconcentration can take place with 0Ps in specific 'in vivo' contexts.

In one of the trials (36), the untreated amphibians bioconcentrated fenthion and parathion from treated tadpoles to an average order of 62 and 64 times respectively.

A report (38) from the wild demonstrates how predatory birds, such as barn owls, eating prey poisoned by the OP warblecide famphur, bioconcentrated the chemical to the extent that they developed secondary poisoning. Another report (39) demonstrates how the widespread deaths of Swanson's hawks as a result of monocrotophos poisoning in Argentina may have resulted from secondary poisoning after the hawks had ingested grasshoppers that were contaminated with this OP after its application onto lucerne.

Both of these field incidents and 'in vivo' trials thus demonstrate how sufficient quantities of lipophilic 0Ps can pass through the digestive tract without undergoing hydrolysis and degradation. Once 0Ps have successfully reached the storage fats, significant degrees of bioconcentration can take place that could have catastrophic toxicological consequences in certain contexts.

Toxic impurities can exacerbate the bioconcentration factor

It should also be noted that common impurities such as OOS-trimethyl phosphorodithioate (42,56) and N-chloromethylphthalimide (42,57) that are found in association with technical formulations of systemic phosmet (the former is found in fenthion also) will actually inhibit crucial hydrolases (46 [p. 83]) such as the A esterase, carboxylesterase, providing the impurity is present at a sufficiently high percentage proportion of the compound. This would have the overall negative effect of preventing degradation of the OP compound itself. Some specific types of 'neuropathic' OP parent compound also inhibit hydrolases (58), thus inactivating their own degradation with disastrous consequences for the individual contaminated with such a compound.

Bioconcentration of low levels of systemic phosmet used upon pigs as a part causalfactorfor the low incidences of endemic BSE in Switzerland and France

In Switzerland and France, countries suffering a relatively low incidence of endemic BSE, systemic phosmet is only licensed for use upon pigs. Because of the lipophilic nature of systemic phosmet, the chemical must theoretically bind into the fat of the treated pigs and then further bioconcentrate into the fat of any animals ingesting that contaminated fat on a regular basis.

Pig fat is included in virtually all brands of cattle concentrated feeds in Switzerland, incorporated at rates of 1.5%-10% of the total ingredients (Personal communication; Dr D Guidon, Swiss Federal Research Station for Animal Production, CH-1725 Posieux, Switzerland). Various forms of straight and protected fat, etc. are also fed.

In France, 20 out of the 26 cases of reported BSE have occurred within the C6te d'Amour region of Brittany, which is also home to the most intensive centre of pig production in Europe. Systemic phosmet was intensively used on the pigs here, and the rendered pig offal was recycled back into the dairy herds of this same vicinity via the MBM ingredient that was milled up into concentrates by the local animal feed compounders. The relatively high incidence of BSE in this locality may be explained by the use of phosmet in these intensive pig units, and the consequent potential for its bioconcentration in cattle. NB, pigs do not develop TSE after exposure to phosmet themselves because they are treated at the 2 mg/kg low-dose rate, whilst pigs, as a species, do not carry the TSE susceptible types of PrP genotype.

All countries affected with endemic BSE have subjected their cattle and/or pigs to a combined exposure of both direct systemic phosmet treatment as well as indirectly to the bioconcentrated residues of phosmet via feeds containing animal fat. Such fat has to be sourced from rendered-down livestock that have originated from any country (e.g. UK), where treatment with systemic phosmet has been intensive. The intensity of phosmet exposure within each country will be proportional to that country's incidence rate of endemic BSE.

MBM produced after cessation of solvent extraction favours the retention of the tallowfraction and its phosmet-bound contaminants in the feed

Analogous to the infamous DDT debacle, countries which employed both systemic phosmet treatments and continuous-flow MBM production where phosmet-contaminated fat was continually recycled, opened themselves up to a vicious circle of phosmet bioconcentration up through the farm animal food pyramid. The UK's 160 000-head BSE epidemic can be explained by their most intensive reliance upon these prerequisites.

The potential UK problem of exposing cattle to OP contaminants in tallow was activated further once solvent extraction was phased out in the MBM plants in the early 1980s (49), making way for the more lucrative 'continuous-flow' system of rendering. This cessation of solvent extraction permitted the tallow fat fraction, which contained the fat soluble phosmet contaminants derived from rendered-down pigs and cattle, to remain as a stable component of MBM. Once bound into the fat depots, both phosmet parent compound and some of its toxic metabolites are capable of evading the usual degradation processes of hydrolysis, etc. encountered in the actual rendering process of MBM as well as in the digestive tract of the animal ingesting that feed.

In places like Guernsey island where there was a high incidence of BSE (8) with a relatively small amount of direct systemic phosmet usage (applied for the voluntary control of cattle lice and mange as a 10 mg/kg treatment with a follow-up dose 14 days later), there was an intensive 'satellite' dependence upon 'continuous-flow' MBM imported from the UK. Furthermore, a high proportion of fat was required in concentrate feeds used on Guernsey to cater for the production of high-fat Channel Island milk which has to be intensively produced on the island (e.g. demanding a high input of concentrated feeds) due to the competitive demand for a limited amount of high-priced agricultural land.

Importation of UK-rendered MBM into countries suffering low levels of endemic BSE

It should also be considered how the importation of UK manufactured 'continuous flow' MBM into countries affected by low incidences of endemic BSE, not only conveyed a source of bioconcentrated phosmet but also assisted in the spread of low levels of the hypothetical phosmet-modified PrPbse. This could have been partly responsible for causing the relatively much lower incidences of BSE in these countries.

Providing that this exogenous source of PrPbse is able to resist trypsin degradation in the gastro tract and cross the gut and blood-brain barriers as an intact 'infectious' prion, then it seems possible that BSE can be reproduced within any individual recipient of these recycled prions providing their PrP genotype is predisposed to this specific new variant TSE disease.

Table Correlating criteria of phosmet usage with incidence rates of endemic BSE in various countries (includes all phosmet-using countries)

It should also be noted that cattle which were phosmet treated in the UK and later imported into countries such as Portugal and France, where they were eventually slaughtered, would have also lead to small amounts of phosmet-modified PrP/phosmetcontaminated fat entering the MBM food chain of the home reared cattle in these countries.

As can be seen from the table of data covering all of the countries that employ phosmet - as well as some of the countries that do not - the intensity of usage of systemic phosmet within each country predicts, with a fairly high degree of reliability, the incidence rate of endemic BSE within that country.

Proposed mechanisms for a phosmet-induced abnormal post-translational covalent modification of the prion protein as initiator of BSE pathogenesis

In vitro experimental challenge

Preliminary results of an 'in-vitro' experimental challenge which exposed tissue culture cells expressing PrP to low doses of phosmet support this hypothesis.

Dr Stephen Whatley of the Department of Neuroscience, Institute of Psychiatry, De Crespigny Park, London SE5 8AF, UK, challenged PrP tissue culture cells with 2 p.p.m. and 12 p.p.m. doses of phosmet. Phosmet interacted dramatically with PrP, causing PrP to traffic and distribute abnormally in the cells. In the phosmet-treated cells, three out of the four hitherto recognized abnormal characteristics (7) specific to the TSE causing pathogenic PrP isoform were invoked. The alterations involved an increase in the expression of PrP in surface cells, an accumulation of PrP in the microsomes (lysosomes and golgi, etc.) and an abnormal resistance of the membrane anchored PrP to cleavage by phospholipase C. These phosmet-induced reactions were specific to the prion protein and were proportional to the dose of phosmet introduced. This model failed to detect protease resistance in phosmet-affected PrP.

The increase of PrP at the lysosomes as well as the abnormal resistance of membrane bound PrP to phospholipase C cleavage suggests that some abnormal PrP modification has been invoked by the introduction of phosmet. Interestingly, 'in-vitro' trials with brain cells that have been affected by an abnormal scrapie PrP (PrPsc) isoform (7,8 [p. 571) demonstrates a similar abnormal pattern of PrP distribution and PrP phospholipase C resistance as invoked in these trials with phosmet. PrPsc diseased cells also demonstrate the same increase in cellular expression of PrPc (7,8 [p. 57]) as demonstrated in the phosmet-PrP trials.

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Last updated 09-Feb-2007