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Biomedical subjects

E R James

Publications and source records attributed to E R James.

At least 55 records · Page 3Linked to original sources

Pathology associated with vaccination against Schistosoma mansoni in mice using cryopreserved radiation-attenuated schistosomula.

Twenty-one mice were injected intramuscularly with 2000 Schistosoma mansoni schistosomula irradiated at 20 krad and cryopreserved; three mice were killed on each of days 0, 2, 5, 9, 19, 28 and 44 days after infection and muscle from the site of injection in the left hind leg, the lungs and livers removed for histological examination. Schistosomula were seen in sections from the leg muscle from days 0 to 19 inclusive, in the lungs from day 2 to day 28 inclusive and in the livers from days 9 to 28 inclusive. Most schistosomula were seen in sections of the leg muscle with considerably fewer parasites occurring in the lungs and especially the livers. Granulomatous reactions comprising eosinophils, polymorphs, plasma cells and macrophages were first seen in the leg muscle on day 2, in the lungs on day 5 and in the liver on day 19. The peak inflammatory reactions appeared to occur between days 5 and 9, 9 and 19 and 28 and 44 respectively in the three tissues. The pathology is discussed in relation to the dose of irradiation required to attenuate the schistosomula for optimal immunogenicity.

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Adoptive transfer of vaccine-induced resistance to Leishmania donovani.

BALB/c mice were immunized with three subcutaneous injections combining killed parasites and glucan, or were untreated. Spleen cells were transferred to syngeneic recipients. Mice which received 5 X 10(8) spleen cells from vaccinated donors demonstrated significant protection against Leishmania donovani challenge as compared to untreated mice receiving immune sera, or mice which received untreated spleen cells.

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Cryopreservation of Plasmodium chabaudi. II. Cooling and warming rates.

Various cooling and warming rates were investigated to determine the optimum conditions for cryopreserving the intraerythrocytic stages of Plasmodium chabaudi. Infected blood, equilibrated in 10% v/v glycerol at 37 degrees C or in 15% v/v Me2SO at 0 degree C for 10 min, was cryopreserved using cooling rates between 1 and 5100 degrees C min-1. After overnight storage in liquid nitrogen the samples were warmed at 12,000 degrees C min-1. Warming rates between 1 and 12,000 degrees C min-1 were investigated using samples previously cooled at 3600 degrees C min-1. After thawing, the glycerol and Me2SO were removed by dilution in 15% v/v glucose-supplemented phosphate-buffered saline. Survival was assayed by inoculation of groups of five mice each with 10(6) infected cells and the time taken to reach a level of 2% parasitemia estimated. The optimum cooling rate was 3600 degrees C min-1 for parasites frozen using either 10% glycerol or 15% Me2SO; the pre-2% patent periods were 0.90 and 1.01 days above control values (representing survival levels of 21 and 17.5%, respectively). The optimum warming rate was 12,000 degrees C min-1; the pre-2% patent periods were 1.01 and 1.32 days above control values, respectively (18 and 10% survival), for glycerol and Me2SO. With ethanediol (5% v/v) and sucrose (15% w/v) as cryoprotectants the optimum warming rates were also 12,000 degrees C min-1 while the optimum cooling rates were 330 and 3600 degrees C min-1, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

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Schistosoma mansoni: interactive effects of irradiation and cryopreservation on parasite maturation and immunization of mice.

Mechanically transformed schistosomula of Schistosoma mansoni were irradiated with levels of 60Co irradiation between 2.5 and 54 krad, cryopreserved by the two-step addition of ethanediol and rapid cooling technique, and were injected intramuscularly into groups of mice which were perfused 40 days later. The schistosomula were either irradiated and then cryopreserved (IC) or cryopreserved and then irradiated in the frozen state (CI). Development into adult worms was prevented with 4 krad for IC schistosomula, but for CI schistosomula a small number of worms (1.6%) was recovered using 8.8 krad. A dose of 4 krad was sufficient to prevent development of unfrozen controls (I), but for schistosomula irradiated while exposed to ethanediol (EI), a dose of 7 krad was required. Using the different protocols, the peak levels of protection against a challenge infection were achieved with 9 (IC) and 16 krad (CI), compared to 20 krad for unfrozen schistosomula (I) reported previously. The highest level of protection (65%) was achieved with CI schistosomula. Possible interactions between the radioprotective and damaging effects of cryopreservation are discussed.

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Successful cryopreservation of Brugia pahangi third-stage larvae in liquid nitrogen.

Experiments are described which lead to the retention of infectivity of Brugia pahangi third-stage larvae after cooling to -196 degrees C. Methanol, a well documented cryoprotectant was used at a concentration of 20% (v/v). The schedule consisted of a 5 degrees C min-1 cool to an intermediate temperature of -21 degrees C and a subsequent rapid cool into liquid nitrogen. A rapid thaw of the parasites led to approximately 34% of motile cryopreserved larvae developing in multimammate rats (Mastomys natalensis) compared to unfrozen control larvae. Cooling rate and intermediate temperature were both found to be crucial variables affecting survival levels of the larvae.

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Separation of viable and non-viable Onchocerca microfilariae using an ion exchanger.

A technique has been developed for separating viable and non-viable Onchocerca lienalis microfilariae (mff) by passing a suspension of the parasites through a column of DEAE cellulose. A proportion (between 42% and 87%) of normally motile parasites, whether unfrozen or cryopreserved, passed through the cellulose columns and retained their viability. Non-motile and sluggish mff were retained by the cellulose. The proportion of viable cryopreserved mff was greater after passage through the column as assessed by motility, migration in mice and development in the insect vector. Numbers of cryopreserved mff eluted through the columns peak quickly at a relatively high level after about 20 ml of eluate had been passed through, whereas normal unfrozen parasites rose and fell in numbers more slowly. By using sodium chloride gradients and buffers of differing conductivity it was concluded that the separation of the mff is probably due to net charge changes as well as motility. It is believed that this technique offers considerable promise as a tool to provide populations of mff of more uniform viability following freezing and thawing with existing cryopreservation techniques.

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Enzyme polymorphism in Trichinella.

The isoenzyme profiles of five isolates of the supposed 'species' of Trichinella, T. nativa, T. spiralis and T. nelsoni were compared. Four enzymes (AK, PGM, MPI and GPI) gave good resolution and clearly differentiated T. Spiralis from the other two species. T. nativa and T. nelsoni had similar isoenzyme patterns; the two separate isolates of T. nativa and T. spiralis used gave similar results, thus indicating the validity and the reproducibility of the technique. The value of enzyme electrophoresis for specific and subspecific classification of Trichinella is discussed and compared with the more traditional methods of taxonomy which have failed to resolve the controversy surrounding speciation.

Electrophoresis, Starch Gel↗

Schistosoma haematobium in the baboon (Papio anubis): effect of vaccination with irradiated larvae on the subsequent infection with percutaneously applied cercariae.

Groups of five baboons were vaccinated three times at approximately six-weekly intervals at a rate of 1,000 organisms per kg of gamma-irradiated Schistosoma haematobium larvae. Five vaccines were tested: 3 and 20 Krad cercariae applied percutaneously; fresh 3 and 20 Krad mechanically transformed schistosomula injected intramuscularly; and cryopreserved 20 Krad schistosomula injected intramuscularly. These five groups and an unvaccinated control group were challenged percutaneously with 7,500 S. haematobium cercariae three months after the last vaccination. The efficacy of the vaccines was judged by faecal egg excretion, and by adult worm and tissue egg recoveries at necropsy 4.5 months after challenge. Significant protection, with 64 to 89% reductions in worm burden and parallel reductions in egg production, was achieved by all but the cryopreserved vaccine, although egg production was not significantly reduced in those female worms which did mature. Cercariae tended to give more protection than schistosomula and 20 Krad more protection than 3 Krad. No significant pathology could be detected in an additional baboon vaccinated with 20 Krad schistosomula but not challenged with cercariae. This is an encouraging result for the development of a live vaccine against S. haematobium.

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An improved technique for the cryopreservation of Onchocerca microfilariae.

Experimental describe the use of ethanediol to store Onchocerca microfilariae in liquid nitrogen. The technique involves a 2-stage incubation of the parasites at 37 degrees C and 0 degrees C in ethanediol, before rapid cooling to -196 degrees C. Viability has been assessed by motility, by migration in a proxy host and by development to the infective stage in the insect vector. A total of 71-79% of the cryopreserved microfilariae was shown to be viable compared to unfrozen controls. The technique is simple, inexpensive and very effective when compared to previous cryopreservation procedures for microfilariae and should be particularly suited for use under field conditions.

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Further observations on immunization of sheep against Schistosoma mansoni and S. bovis using irradiation-attenuated schistosomula of homologous and heterologous species.

This paper describes further characteristics of the immunization of sheep against schistosomes using live, irradiation schistosomula. Sheep immunized with a non-virulent strain of Schistosoma mattheei were protected against a more virulent strain of the same species for over a year. As there was no evidence that the irradiated parasites were able to persist this long, it was concluded that the vaccine had induced a sterile resistance. Heterologous vaccination, using irradiated S. mattheei schistosomula to immunize against S. bovis or irradiated S. mansoni schistosomula to immunize against S. mattheei, failed to induce any protection.

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