Use of transgenic oryzacystatin-I-expressing plants enhances recombinant protein production.

Plants are an effective and inexpensive host for the production of commercially interesting heterologous recombinant proteins. The Escherichia coli-derived glutathione reductase was transiently expressed as a recombinant model protein in the cytosol of tobacco plants using the technique of leaf agro-infiltration. Proteolytic cysteine protease activity progressively increased over time when glutathione reductase accumulated in leaves. Application of cysteine protease promoter-GUS fusions in transgenic tobacco identified a cysteine protease NtCP2 expressed in mature leaves and being stress responsive to be expressed as a consequence of agro-infiltration. Transgenic tobacco plants constitutively expressing the rice cysteine protease inhibitor oryzacystatin-I had significantly lower cysteine protease activity when compared to non-transgenic tobacco plants. Lower cysteine protease activity in transgenic plants was directly related to higher glutathione reductase activity and also higher glutathione reductase amounts in transgenic plants. Overall, our work has demonstrated as a novel aspect that transgenic tobacco plants constitutively expressing an exogenous cysteine protease inhibitor have the potential for producing more recombinant protein which is very likely due to the reduced activity of endogenous cysteine protease.

Transfer of genetic material between the chloroplast and nucleus: how is it related to stress in plants?

BACKGROUND: The presence of chloroplast-related DNA sequences in the nuclear genome is generally regarded as a relic of the process by which genes have been transferred from the chloroplast to the nucleus. The remaining chloroplast encoded genes are not identical across the plant kingdom indicating an ongoing transfer of genes from the organelle to the nucleus. SCOPE: This review focuses on the active processes by which the nuclear genome might be acquiring or removing DNA sequences from the chloroplast genome. Present knowledge of the contribution to the nuclear genome of DNA originating from the chloroplast will be reviewed. In particular, the possible effects of stressful environments on the transfer of genetic material between the chloroplast and nucleus will be considered. The significance of this research and suggestions for the future research directions to identify drivers, such as stress, of the nuclear incorporation of plastid sequences are discussed. CONCLUSIONS: The transfer to the nuclear genome of most of the protein-encoding functions for chloroplast-located proteins facilitates the control of gene expression. The continual transfer of fragments, including complete functional genes, from the chloroplast to the nucleus has been observed. However, the mechanisms by which the loss of functions and physical DNA elimination from the chloroplast genome following the transfer of those functions to the nucleus remains obscure. The frequency of polymorphism across chloroplast-related DNA fragments within a species will indicate the rate at which these DNA fragments are incorporated and removed from the chromosomes.

The effect of sepsis and short-term exposure to nitrous oxide on the bone marrow and the metabolism of vitamin B12 and folate.

It is recognised that prolonged anaesthesia with nitrous oxide (N2O) induces megaloblastic anaemia by oxidising vitamin B12. To determine whether sepsis aggravates the effect of N2O on haemopoiesis 5 patients with severe sepsis, who required surgery and were exposed to short-term (45-105 minutes) N2O anaesthesia, were studied. None had evidence of pre-operative vitamin B12 or folate deficiency. The effect of the combination of N2O anaesthesia and sepsis on DNA synthesis in bone marrow cells was assessed morphologically, and by the deoxyuridine suppression test. In 3 patients exposed to the longest duration (75-105 minutes) of N2O, addition of folinic acid and vitamin B12 partially improved the utilisation of deoxyuridine in vitro. No patient had evidence of megaloblastic haemopoiesis as judged by bone marrow morphology. It is concluded that prolonged N2O anaesthesia in patients with severe sepsis may adversely affect DNA synthesis. Although this effect did not manifest as overt megaloblastic erythropoiesis, it may be prudent to avoid N2O in such patients.

Tetanus antibodies as a marker of potential efficacy of killed polio immunization.

The use of antibodies to tetanus toxin as a marker of efficacy of parenteral immunization was examined in a randomized sample of 1212 sera representative of the total black infant population of the RSA between 24 and 35 months. All but one of these sera had protective levels of antibodies measured by enzyme-linked immunosorbent assay. In contrast, in a previous study only 59-80% had tritypic immunity to polio and 70-84% mono- or bitypic immunity. Thus, this serological marker was found to be unreliable and some possibilities for this remarkably high level of antitetanus antibodies are considered.

Diagnosis of tuberculosis by detection of mycobacterial antigen in pleural effusions and ascites.

We describe a method for the diagnosis of pleural and peritoneal tuberculosis by the detection of tuberculous antigens using an enzyme-linked immunosorbent assay. Eleven tuberculous pleural fluid and 10 tuberculous ascitic fluid samples were studied by this technique, using 10 non-tuberculous pleural fluid and 14 non-tuberculous ascitic fluid samples as controls. An absorbance value of 0.3 was found to separate the tuberculous groups from their controls to a statistically significant extent (ascitic fluid P less than 0.05; pleural fluid P less than 0.01).

Anti-HTLV-III testing--a practical solution for blood transfusion services?

Screening blood donations for anti-HTLV-III with a modification to a commercially available test kit has so far been found satisfactory--the reagent cost is less than R1.00 a donation. Should all transfusion services adopt this method, South Africa's total blood donations could be tested at a cost of about R650 000, instead of R2.7 million or more, a year.

Properties of human anti-lipopolysaccharide gamma globulin: specificity and protective effects.

Blood donated to the Natal Blood Transfusion Service was screened by an enzyme-linked immunosorbent assay (ELISA) for anti-lipopolysaccharide (anti-LPS) antibodies. Plasma units with high concentrations (greater than 40 micrograms/ml) of anti-LPS IgG were pooled and fractionated to obtain a gamma globulin (lot LG-1). The binding of LG-1 antibodies to LPS prepared from 14 bacterial species and strains was found to be the highest to LPS from Shigella flexneri, Salmonella abortus equi and Salmonella typhimurium and intermediate with Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella enteritidis and Escherichia coli 026:B6. Differential absorption experiments showed that LG-1 contained a mixture of specific and cross-reacting antibodies. A large proportion of antibodies binding to Sh. flexneri LPS were mainly specific, while those binding to S. typhimurium and the other Salmonella species tested were largely cross-reactive. There was little correlation between the spectrum of activity of the LG-1 antibodies and the incidence of gram-negative bacteria in blood cultures taken from hospital patients in an area covered by the Transfusion Service. Mice treated with LG-1 prior to inoculation with P. aeruginosa were significantly protected against morbidity and mortality compared to controls.

[Nutritional muscular dystrophy in a clutch of ostrich chicks]. / Vlekspiersiekte in 'n broeisel volstruiskuikens.

Six 1 1/2-month old ostrich chickens in the Upington district of the Cape Province developed lameness. Two died and pathological examination of one of them revealed lesions identical to those of white muscle disease in the larger muscle groups. Vitamin E-selenium therapy cured the other 4. The diet of the animals consisted mainly of lucerne (alfalfa).

An ELISA procedure for detecting human anti-endotoxin antibodies in serum.

We report an ELISA method suitable for the large-scale screening of blood bank stores to identify those blood units containing high concentrations of antiendotoxin antibodies. In Natal, 8.3% of total units collected had antiendotoxin antibodies at concentrations greater than 40 micrograms/ml, values that may be therapeutically useful. We found that one technician could screen enough samples per year to produce 800 litres of such high-titre plasma.

A simple and rapid method of washing and drying micro-titre plates used in ELISA.

A simple device constructed out of perspex sheeting and capable of washing 96-well micro-titre plates used in ELISA is described. It consists of three parts: (a) the 'shower' - a tank with 96 stainless steel syringe needles through which washing fluid runs into the wells of the micro-titre plate, (b) a sump into which washing fluid is collected and (c) a sliding base plate allowing the accurate positioning of the micro-titre plate below the shower. The efficiency of this washing device was established by comparing the results from several specific ELISAs with tha obtained after washing duplicate plates by hand. The use of a specially constructed centrifuge to spin-dry plates is also described.

Metamorphosis of chronic granulocytic leukaemia, lytic bone lesions and hypercalcaemia. A case report.

Chronic granulocytic leukaemia may undergo metamorphosis characterized by diffuse granulocytic sarcomas resulting in lytic bone lesions and hypercalcaemia. A patient with this rare complication is reported.

Thrombotic thrombocytopenic purpura. A case investigated with 111In-oxine-labelled platelets.

A 34-year old woman presented with the clinical and laboratory features of thrombotic thrombocytopenic purpura (TTP). Studies with isologous platelets labelled with 111In-oxine revealed a short half-life of circulating platelets (18,5 hours) and destruction of the transfused platelets in the spleen, liver and bone marrow. There was no scientigraphic evidence of deposition of labelled platelets in the vasculature. The patient was treated with daily fresh frozen plasma transfusions, but no improvement in platelet count or serum urea level was noted. Although there was no clinical evidence of a bleeding tendency at the time, the patient had a fatal cerebrovascular haemorrhage. The findings in this case suggest that an immune type destruction of platelets may occur in TTP.

Agranulocytosis in an arthritic patient treated with levamisole. A case report.

A case of agranulocytosis in a man with nonspecific seronegative polyarthritis treated with levamisole in the form of a proprietary veterinary anthelmintic is described. At the time of presentation he had a relative monocytosis. Recovery was uneventful and was heralded and paralleled by an increase in monocytes, serum vitamin B12 and total vitamin B12 binding capacity. Levamisole induced agranulocytosis and the significance of monocytosis and serum vitamin B12 binding proteins in neutropenia are briefly reviewed.