The iron uptake mechanisms of enteropathogenic Escherichia coli: the use of haem and haemoglobin during growth in an iron-limited environment.

The iron uptake mechanisms of enteropathogenic Escherichia coli (EPEC) were examined and compared with those of control E. coli strains. The incidence of aerobactin production was similar (39% and 37% respectively) in the two groups. The quantities of enterochelin produced by aerobactin-negative EPEC and control strains were similar, as were the quantities of enterochelin produced by aerobactin-positive EPEC and control strains. The ability to use haem or haemoglobin as an iron source in an iron-restricted environment was found in 80.4% and 60.8% of EPEC strains respectively, and in 76.6% and 56.6% of control E. coli strains. The ability of E. coli strains to use these compounds was not related to the production of enterochelin or aerobactin or to the production of haemolysins, and may be an important characteristic of bowel organisms. When growing in an iron-limited environment, the iron contained in haemoglobin was used in preference to ovotransferrin-bound iron. During periods of haemoglobin-stimulated growth, the enterochelin uptake system was shown to be fully expressed and may be involved in transport of haemoglobin-derived iron into the cell. Uptake of ovotransferrin-bound iron took place immediately upon exhaustion of haemoglobin-derived iron. The ability to use iron derived from haem compounds represents an alternative iron uptake mechanism for organisms growing in an iron-limited environment and allows greater flexibility during growth in vivo.     

A cytogenetic deletion, del(17)(q11.22q21.1), in a patient with sporadic neurofibromatosis type 1 (NF1) associated with dysmorphism and developmental delay.

We report the first visible cytogenetic deletion involving the NF1 gene in a patient with sporadic neurofibromatosis, dysmorphic features, and marked developmental delay. The combined evidence of molecular and cytogenetic techniques based on dosage reduction, hemizygosity for microsatellite markers, high resolution G banding, and FISH analysis, predicts this deletion to be approximately 7 Mb in size. Our findings highlight the importance of conducting a detailed cytogenetic and FISH analysis in patients with NF1 who have additional dysmorphic features or particularly severe learning difficulties.     

Dicentric chromosome in the bone marrow of a child with megakaryoblastic leukaemia and Down''s syndrome.

A two year old girl with Down's syndrome (constitutional karyotype: 47 + 21), presenting with pancytopenia, developed acute megakaryoblastic leukaemia (AMKL). Her bone marrow contained an abnormal clone with a novel dicentric chromosome derived from chromosomes 5 and 7 (karyotype 46, XX, -5, -7, +dic (5;7) (p 13; p 11.2), +21. This case provides further evidence for a connection between chromosome 21 and this unusual form of childhood leukaemia, and raises questions about the loss of short arm material from chromosomes 5 and 7 compared with the more usual monosomy or long arm loss.     

A further mutation of the FGFR2 tyrosine kinase domain in mild Crouzon syndrome

We report a family heterozygous for a newly identified mutation in the tyrosine kinase I domain of the FGFR2 gene (1576A > G, encoding the missense substitution Lys526Glu), associated with variable expressivity of Crouzon syndrome, including clinical nonpenetrance. Our observations expand both the clinical and molecular spectrum of this unusual subset of FGFR2 mutations.     

Functional characterization of missense mutations at codon 838 in retinal guanylate cyclase correlates with disease severity in patients with autosomal dominant cone-rod dystrophy

Three different mutations in codon 838 of GUCY2D, the gene for retinal guanylate cyclase 1, have been linked to autosomal dominant cone-rod dystrophy at the CORD6 locus. To examine the relationship between enzyme activity and disease severity, the three disease-causing substitutions (R838C, R838H and R838S) and four artificial mutations (R838A, R838E, R838L and R838K) were generated. Assay of GCAP1-stimulated cyclase activity in vitro shows that, compared with wild-type, R838E, R838L and R838K possess only low activity, whereas R838A, R838C, R838H and R838S have activity equal or superior to wild-type at low Ca(2+) concentrations. These four latter mutants showed a higher apparent affinity for GCAP1 than did wild-type. The Ca(2+) sensitivity of the GCAP1 activation was also altered with marked residual activity at high Ca(2+), the effect increasing: wild-type < R838C < R838H < R838A < R838S. Within the photoreceptor, this would result in a failure to inactivate cyclase activity at high physiological Ca(2+ )concentrations. Amongst the three disease-associated mutations, the effect correlates directly with disease severity. The wild-type and R838H mutant displayed a difference in pH sensitivity, with the mutant showing a higher specific activity with pH > 6.0. Site 838 is in the dimerization domain that forms a coiled-coil in the active protein. A computer-aided structure prediction of this region indicates that R838 in the wild-type breaks the structure at four helical turns, and there is an increasing tendency for the structure to continue for further turns in the order R838C < R838H,S,K < R838E < R838A < R838L.     

Chemical change and energy output during muscular contraction

1. The production of heat and (internal) work and the changes in the amount of phosphocreatine (PCr), ATP, inorganic phosphate (Pi) and sometimes lactate have been measured from moment to moment during and after tetanic isometric contractions of isolated frog muscles at 0° C.

2. Heat production was measured by thermopiles and a novel apparatus was employed for freezing the muscles rapidly at a chosen instant so as to halt the chemical processes before analysis.

3. Using unpoisoned muscles in oxygen, it was shown that neither oxidative recovery processes nor glycolytic ones led to appreciable restitution of PCr or ATP during 15 sec of contraction. However, clear signs of recovery processes could be seen within a minute. In our preparations artificial `ageing' by storage at low temperature did not interfere with the capacity for glycolysis.

4. Our clearest result was that the break-down of PCr was not nearly large enough to account for the rapid heat production during the first few sec of contraction. By the end of a 15 sec tetanus as much as 10 mcal/g remained unaccounted for.

5. The source of this heat is not clear. At no time is there any sign of net break-down of ATP; indeed there appears to be a slight increase of ATP in the stimulated muscle.

6. Break-down of PCr continues both during relaxation and during the minute following, while the muscle is at rest. Thus during contraction there is heat production without PCr break-down, while subsequently there is PCr break-down without heat production.

     

Immunoglobulin G subisotype responses of pneumonic and healthy,exposed foals and adult horses to Rhodococcus equi virulence-associated proteins

Rhodococcus equi causes severe pyogranulomatous pneumonia in foals and in immunocompromised humans. Replication of virulent isolates within macrophages correlates with the presence of a large plasmid which encodes a family of seven virulence-associated proteins (VapA and VapC to VapH), whose functions are unknown. Although cell-mediated immunity is thought to be crucial in eliminating R. equi infection, antibody partially protects foals. The antibody response to both VapA and VapC was similar in six adult horses and six naturally exposed but healthy foals, as well as in eight foals with R. equi pneumonia. The immunoglobulin G (IgG) subisotype response of pneumonic foals to Vap proteins was significantly IgGb biased and also had a trend toward higher IgGT association compared to the isotype association of antibody in adult horses and healthy exposed foals. This suggests that in horses, IgGb and IgGT are Th2 isotypes and IgGa is a Th1 isotype. Furthermore, it suggests that foals which develop R. equi pneumonia have a Th2-biased, ineffective immune response whereas foals which become immune develop a Th1-biased immune response. Pneumonic foals had significantly more antibody to VapD and VapE than did healthy exposed foals. This may indicate a difference in the expression of these two Vap proteins during persistent infection. Alternatively, in pneumonic foals the deviation of the immune response toward VapD and VapE may reflect a bias unfavorable to R. equi resistance. These data indicate possible age-related differences in the equine immune response affecting Th1-Th2 bias as well as antibody specificity bias, which together favor the susceptibility of foals to R. equi pneumonia.